Research ArticlePharmacological Evaluation of Selected Medicinal Plants Usedin the Management of Oral and Skin Infections in Ebem-OhafiaDistrict Abia State Nigeria
Blessing O Oyedemi1 Sunday O Oyedemi 2 Johnson V Chibuzor1 Ifeoma I Ijeh2
Roger M Coopoosamy3 and Ayobami O Aiyegoro4
1Department of Plant Science and Biotechnology Michael Okpara University of Agriculture Umudike Nigeria2Department of Biochemistry Michael Okpara University of Agriculture Umudike Nigeria3Department of Nature Conservation and Ethnobotany Mangosuthu University of Technology PO Box 12363 JacobsDurban 4026 South Africa
4GI Microbiology and Biotechnology Unit Agricultural Research Council Animal Production Institute IrenePretoria 0062 South Africa
Correspondence should be addressed to Sunday O Oyedemi silvanusdemigmailcom
Received 24 January 2018 Accepted 26 April 2018 Published 4 July 2018
Academic Editor Carlos H G Martins
Copyright copy 2018 Blessing O Oyedemi et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited
Oral and skin infections contribute significantly to the global health challenges responsible for the current trend of increasedmorbidity and premature death The purpose of this study was to document medicinal plants used in the management of oraland skin infections in Ebem-Ohafia Local Government Area (LGA) Abia State and to characterize the in vitro antioxidant andantibacterial activity The thin layer chromatography (TLC) profiling of ten of the selected folklore medicine was carried out usinga various solvent system of different polarity index The antioxidant capacity of the plant extracts was evaluated using chemical-based methods and its antibacterial effect was investigated using disc diffusion andmicrodilutionmethods Sixty-one plant speciesbelonging to 26 families were discovered and the most frequently cited species are Euphorbiaceae (1803) Fabaceae (1147)and Asteraceae (1147) All the plant extracts showed a promising free radical scavenging activity and efficient ferric reducingantioxidant power in a concentration-dependentmanner possibly due to their richness in polyphenol with TLC profiling showingmaximum three bands of phytochemicals Also the plant extracts exhibited a mild to weak antibacterial activity against our panelof bacterial strains having MIC values ranging from 256 to gt 512 120583g mL reflected in their zone of inhibition at 10 120583gdisc Thedata obtained for Breynia nivosa (BN) Eleusine indica (EI) Cassia alata (CA) Chromolaena odorata (CO) and Acalypha hispida(AH) extracts substantiate the traditional use of these herbal remedies in the region and open the possibility for the developmentof cheaper and affordable drugs in the treatment of oral and skin infections Further studies are needed to identify active ingredientwith strong antibacterial and antioxidant capacities along with their molecular mechanisms
1 Introduction
Oral health is a state of being free from the mouth and facialpain throat cancer oral infection and sores that limit humancapacity in biting chewing smiling and speaking as wellas psychosocial well-being [1] Owing to the environmentalpollution the concentration of heavy metals such as Ni CrZn Hg and Pb in most food materials and water is rapidly
increasing which severely affects both oral and general healthcondition For example dental caries periodontitis andthroat cancer have been recognised as the most prevalentnoncommunicable diseases associated with heavy metalsglobally affecting school-aged children and a huge majorityof adults [2] In Nigeria the rate of deep periodontal pocketis between 15 and 58 (aged 15 years) while the preva-lence of unmanaged dental caries is high raising alarm for
Hindawie Scientific World JournalVolume 2018 Article ID 4757458 16 pageshttpsdoiorg10115520184757458
2 The Scientific World Journal
odontogenic infection Falsetta and colleagues [3] highlightedthat poor oral hygiene and frequent consumption of sugarscreate bacterial adhesion to tooth surfaces via mineralizationand biofilm formations caused by glucosyl and fructosyltransferases The same authors indicated an adverse effectof higher concentration of fluoride in drinking water insome developing countries that is responsible for the causeof fluorosis and dental caries regardless of age and sex Theoral cavity exposed to carcinogens that may advance into pre-cancerous lesions and malignant disorders like leukoplakiasubmucous fibrosis and dental caries resulting from theimbalance in the levels of free radicals generation and naturalantioxidants in the saliva [4] There is a growing consensusthat administration of natural antioxidants to the oral cavityreduces free radicals oxidative stress and oral inflammationtriggered by nicotine alcohol and other toxins introducedinto the mouth [5]
The skin being the largest organ in the body guards theunderlying muscles bones ligaments and internal organsinterfaces with the environment as well as protecting thebody against pathogenic microbes [6] Notwithstandinginfectious skin diseases such as skin abrasions burns acnedermatitis and sebaceous cysts have become everyday inci-dences and constitute a reoccurring health problem thataffects all human age groups [6] Over the years bacterial skindisorders such as impetigo ecthyma folliculitis eczemas andurticarias are one of the most common causes of hospitalvisits in the southeastern part of Nigeria Overcrowding mal-nutrition and humidity heat food and medication allergiescause someof these skin diseases especially among the schoolchildren in urban and rural settings [2] Microbial invasionof the skin surface forms a biofilm that confers a drugtolerance on the bacteria and capacity to escape host immuneresponse [7] The discovery of antibiotic resistance genesin several microorganisms quorum sensing and horizontalgene transfer has limited treatment of skin and soft tissueinfections and thus could pose a serious health challengein a region where there is limited access to microbiologicallaboratory facilities and antimicrobial agents [8] The globalneed for effective alternative medicines for oral and skininfections with minimal side effects and economical use isincreasing due to the rapid growth of antibiotic resistance andopportunistic infections in immunocompromised patients[9]
Plant-derived products are now gaining recognitionworldwide currently recommended for primary healthcarebut only a few plants have received rigorous scientific investi-gation Medicinal plants are natural products used since timeimmemorial for the treatment of various human diseases Allover the world the citizen and health practitioners are nowstarting to rely on herbal medicines as a substitute for scien-tifically proved therapies [10] Herbal products contained sec-ondary metabolites that are synthesized through the pentosephosphate shikimic acid and phenylpropanoid pathwaysThese compounds play a significant defensive role againstpathogenic bacteria and oxidative stress caused by abioticstress in plant [11] Currently one of the strategies used to treatskin and oral infections is the use of natural products fromplant origin perhaps due to the perception that long-term
use of western medicine induces severe complications Con-sequently many Nigerian patients in particular those livingin the rural areas have resorted to seeking relief via tradi-tional healers and herbalists who administer herbal drugsin the management of human diseases Considering therate at which the vegetation gets depleted and the currentfocus on safe complementary and alternative medicine tosynthetic drugs there is therefore the need to document theindigenous knowledge of these plants and to search for moreplants with therapeutic potential in the treatment of bacterialinfections and oxidative stress
There is a paucity of literature in traditional medicineused in the management of orofacial and skin infectionsin Nigeria Our recent ethnobotanical survey conductedin Ebem-Ohafia LGAs in the southeastern part of Nigeriarevealed sixty-one plant species commonly used in the man-agement of oral and skin infections Despite the acclaimedtraditional use of these botanicals as therapeutic agents thereis little scientific evidence to support their folkloric useTherefore this study aimed at investigating the antioxidantcapacity of these herbal remedies in terms of free radicalscavenging reducing power and total phenolic and flavonoidcontent in consonance with the TLC profiling The antibacte-rial effect on some clinically isolated bacteria associated withoral and skin infections was also investigated
2 Materials and Methods
21 Study Area Ebem-Ohafia is a town in Ohafia LGA ofAbia state in the southeastern part ofNigeria situated between5∘ 381015840N and 7∘ 501015840 East and encompasses over twenty-sixhometowns with a population of over 300 000 from thelast population census in 2006 Ebem community is a partof the tropical rainforest characterized by dry and rainyseason with a total annual rainfall of over 1400 mm and anannual temperature range of 23∘C to 32∘C The people of theregion practice subsistence farming under local governmentagencies policy The inhabitants of Ebem use herbal drugseither alone or in combination with other medicines in thetreatment of human diseases Most of the people in Ebemare rural dwellers hence they use plant-based therapies in themanagement of ailments that include oral and skin infections
22 Ethnobotanical Survey The ethnobotanical survey wasconducted between December and May 2016 using a well-structured questionnaire administered to the participantswith indigenous knowledge of plants utilized in the area Theset questions contained the local diagnosis of oral and skindiseases the name of plant samples used for the treatmentmethods of preparation and duration of treatment sideeffects and mode of administration The people interviewedconsisted of women andmen both married and unmarried atthe age of 30 to 65 with little education qualification
23 PlantCollection and Identification The leaves of Ipomoeainvolucrata (II) Acalypha hispida (AH) Breynia nivosa (BN)Jatropha curcas (JC) Chromolaena odorata (CO) Macrolo-bium macrophyllum (MM) Baphia nitida (BNI) Burkeaafricana (BA) Cassia alata (CA) and the root of Eleusine
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indica (EI) were collected after the interview The plants wereidentified by their local names and authenticated by MrOriaku Williams and Professor Gabriel Osuagwu from theDepartment of Plant Science and Biotechnology College ofNatural Sciences Michael Okpara University of AgricultureUmudike in Abia state Nigeria The vouchers specimens(VICJON 1ndash10) were prepared and deposited at the depart-mental herbarium The preparations and mode of intake ofthe herbal remedies are described in Table 1
24 Sample Preparation We collected leaves and roots ofthe ten plants that were frequently mentioned in the surveyand oven-dried at 40∘C for 72 h while the root of EI wasdried longer for seven days The dried plant materials werepulverized to a fine powder using an electric blender andstored in an airtight container for further use Thirty grams(30 g) of dried powdered materials were extracted with 150mL of 100methanol for 48 h on amechanical shaker (StuartScientific Orbital 202 SOSI Essex UK) and the extractswere filtered using Buchner funnel and Whatman Number1 filter paper The filtrate was concentrated using a rotaryevaporated at 40∘C to recover the solvent and air-dried in afume chamber to give a yield ranging from 45 to 83 g
25 Thin Layer Chromatography (TLC) Profile Thin layerchromatography is a simple method for analyzing a complexmixture of compounds based on the distance travelled Theplant extracts (1 mgmL) dissolved in methanol and spottedon the plate coated with silica gel 60 F254 as a stationaryphase The slurry was prepared by dissolving 15 g of silicagel 60 F254 in 30 mL of distilled water and immediatelypoured into the plate The plates were air-dried overnightAbout 10 120583L of the plant extracts was gently loaded onthe base of the plate (5 cm above) using the capillary tubeThe plates were allowed to develop in chromatographictanks consisting of three different solvents (mobile phase)chloroform methanol acetic acid (541) until they reach34th of the TLC plate The TLC plate was removed andallowed to dry the spots were detected by iodine vapour andretention factor (Rf) was calculated using the equation Rf= distance travelled by the componentsdistance travelled bythe solvent
26 Total Phenolic Content (TPC) Assay The total phenoliccontent present in these extracts was quantified by the FolinCiocalteu reagent (FCR) using the method of Ghaffari etal [12] Briefly 02 mL of the plant extract (2 mgmL) wasadded to the reaction mixture consisting of 1 mL of 10 vvFCR and 08 mL of Na2CO3 (0075 mgmL) to give a finalconcentration of 02 mgmL of each extract The resultingmixture was incubated at 45∘C with shaking for 15 min andthe absorbance measured at 765 nm A standard curve wasprepared by mixing methanol solution of gallic acid (02mL 0025 - 0400 mgmL) with 1 mL of 10 vv FCR andsodium carbonate (08 mL 0075 mgmL) The experimentwas carried out in triplicate and the results presented asmeanvalues with standard deviation (plusmn SD) The TPC value wasexpressed as milligrams of gallic acid equivalent (GAE) perg of dried sample It was calculated using the formula T =
CtimesVM where T is the TPC (mgg) of extract in GAE C isthe concentration of gallic acid from the calibration curve Vis the volume of the extract mL M is the dry weight (g) ofthe leaf or root powder from which the extract was obtained
27 Total Flavonoid Content (TFC) Assay The concentra-tion of flavonoids in the plant extracts was determined viaaluminum colorimetric assay method [13] Briefly 1 mL of2 wv AlCl3 was prepared in 100vv methanol added to 1mL of the sample solution A yellow colour formation afterincubation at room temperature for 1 h was measured at420 nm using an AJI-C03 UV VIS spectrophotometer Thestandard curve for TFC was obtained using quercetin as astandard drug under the same procedure described in TPCdetermination The TFC was calculated using the formulaT = CtimesVM where T is the TFC (mgg) of extract in QEC is the concentration of quercetin established from thecalibration curve V is the volume of the extract mL M isthe dry weight (g) of the leaf powder from which the extractwas obtained The TFC was present in the extracts calculatedas mgg of quercetin equivalent (QE)
28 Ferric Reducing Antioxidant Power (FRAP) Assay Theability of plant extracts to reduce ferric (Fe3+) to ferrous(Fe2+) formwas evaluated following themethod described byYen and Chen [14] with slight modification A volume of 03mL of different concentrations (0025 - 2 mgmL) from plantextract BHT ascorbic acid and rutin prepared in distilledwater was mixed with reacting mixture consisting of 25 mLof 02 M phosphate buffer (pH 66) and 25 mL of K3Fe(CN)6(1 wv) The resulting mixture was incubated at 50∘C for20 min followed by addition of 25 mL of TCA (10 wv)After vigorous shaking 25 mL of the solution was mixedwith 25 mL of distilled water and 05 mL of FeCl3 (01 wv)left incubated at room temperature for 5 min and then theabsorbance was measured at 700 nm against a blank sample(without extract)
29 Free Radical Scavenging Activity Assays The free radicalscavenging potential of plant extracts was measured in vitroby the 1 11015840-diphenyl-1-picrylhydrazyl (DPPH) described byTariq et al [15] The assay was experimented by reacting16 mL of 0135 mM DPPH dissolved in 100vv methanolwith 04 mL of various concentrations (0078 - 1 mgmL) ofmethanol crude extracts The reaction mixture was vortexedthoroughly and left in the dark at room temperature for30 min The absorbance of the mixture was measured at517 nm after 2 min The method of Re et al [16] wasadopted to determine ABTS radical scavenging activity ofthe plant extracts The ABTS radical solution was generatedby mixing two stock solutions of 7 mM ABTS and 24mM potassium persulphate in the same ratio and allowingthe solution to react for 12 h at room temperature in thedark The resulting solution was diluted with methanol toobtain an absorbance of 0706 units at 734 nm A volumeof 1 mL of various concentrations (003125 ndash 1 mgmL)of the plant extracts react with 25 mL of ABTS radicalsolution in the dark for 15 min and was later measured
4 The Scientific World Journal
Table1Medicinalplantsused
inthem
anagem
ento
fskinor
oralinfections
insoutheastern
partof
Nigeria
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Macrolobium
macrophyllum
Fabaceae
Antho
notha
macroph
ylla
Leaf
Skin
Treatin
gof
woun
dandcut
Squeezethe
leavea
ndplaceo
nthe
affectedarea
ofthes
kin
Jatro
phacurcas
Euph
orbiaceae
Barbados
nut
Leaf
Skinoral
Treatm
ento
fboils
Boiltheleave
with
waterPlace
theh
otleaveo
nthea
ffected
area
ofthes
kin
Eleusin
eind
icaPo
aceae
Yard
grass
Root
Skin
Treatm
ento
fRingw
orm
Use
ther
ootand
scrubon
affectedarea
ofthes
kin
Cassiaalata
Caesalpiniaceae
Ring
worm
bush
Leaf
Skin
Treatm
ento
fRingw
orm
Scrubtheleave
onaffectedarea
ofthe
skin
Chromolaena
odorata
Aste
raceae
Siam
weed
Leaf
Skin
Tosto
pbleeding
andrashes
Squeezethe
leavea
ndrubon
affectedarea
Psidium
guajava(L)
Myrtaceae
Guava
Leaf
Skin
Treatm
ento
fskinbu
rns
scrapessunb
urn
woun
ds
psoriasiseczem
aand
others
Crushedgu
avaleafo
rboiledleaves
are
applieddirectlyto
thea
ffected
skin
providingreliefandareb
elievedto
improveh
ealin
gBryophyllum
pinn
atum
(Lam
)Oken
Crassulaceae
Mira
cleleaf
Dogrsquosliver
Leaf
Skino
ral
Treatm
ento
fboilwo
und
sore
orcut
Crushleafandapplyjuiceo
nthew
ound
surfa
ce
Ficusa
sperifolia
Miq
Moraceae
False
thistle
Leaflatex
Skin
Treatm
ento
fskinrashes
anditches
Appliedas
poultic
etothes
kinitches
Acanthus
montany
(TA
nderson)
Acanthaceae
Bearrsquosbreeches
Stem
bark
Skin
Treatm
ento
fwou
ndsa
ndskin
diseases
Woo
dbark
iscrushedandsoaked
italcoho
landextractapp
liedto
woun
dsandskin
Alliu
msativ
umL
Liliaceae
Garlic
Bulbjuice
Skin
oral
Usedto
cond
ition
thes
kin
chew
edforc
ough
andsore
throat
Thejuice
ismixed
with
lotio
nsforskin
cond
ition
ingFreshbu
lbischew
edor
boiledin
water
anddrinkdecoctionfor
coug
h
Emiliasonchifolia
(L)
Aste
raceae
Scarlettassel
Leaf
Skin
Treatm
ento
fSkintro
ubles
abscessa
ndbruises
Crushedleaves
were
pressedandused
tocoverthe
woun
dor
bruisedsurfa
ces
Sida
acutaBu
rmf
Malvaceae
Broo
mwe
edLeaf
Skin
Treatm
ento
fjaund
icesore
ulcer
Poultic
esmadefrom
boiledleaves
are
appliedon
thea
ffected
parts
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Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Ricin
uscommun
isLinn
Eu
phorbiaceae
Castorb
ean
Leafroo
tand
seed
Skin
Treatm
ento
fskininfection
anddetoxification
Applycasto
roilon
thea
ffected
area
torelieve
skin
infection
Citru
slim
on(L)Bu
rmf
(prosp
)Ru
taceae
Limon
Leaffruit
andseed
Skin
Treatm
ento
ffluheadache
cold
andoralinfections
Mixequalquantities
ofLimon
juicea
ndho
lybasilkeepitun
derthe
suntill
becomes
thickapplythison
face
Alliu
mcepa
LLiliaceae
Onion
sLeafandbu
lbSkin
Treatm
ento
fperiodo
ntitis
andcold
Theb
ulbismacerated
andmixed
with
water
andthen
takenorally
Breynianivosa
Euph
orbiaceae
Icep
lant
Leaf
Oral
Treatin
gof
toothache
Boiltheleave
with
waterthentake
the
water
Ipom
oeainvolucrata
Con
volvulaceae
Water
convolvu
lus
Leafandste
mOral
Treatm
ento
foralinfectio
nMixtheleave
with
ugbaga
treea
ndbo
ilwith
water
anddrinkthew
ater
Burkea
afric
ana
Caesalpinioideae
Icheku
Leaf
Oral
Treatm
ento
forald
isease
Boiltheleave
with
water
andtakebut
donrsquot
swallowit
Baphianitid
aPapilio
naceae
Cam
wood
Leafor
stem
Oral
Streng
thensthe
teeth
Chew
thes
tem
orscrubtheleave
onthe
teeth
Gongronem
alatifolium
Asclepiadaceae
Utazi
Leaf
Oral
Treatm
ento
fcou
ghand
diabetes
Crushtheleafw
ithmortara
ndmixwith
ginger
orgarlica
nddrink
Anonasenegalen
sisAnn
onaceae
Ubu
ruocha
Root
Oral
Treatm
ento
fmalariaand
mou
thinfection
Boilther
ootsanddrink
Abrusp
recatoriu
sPapilio
noideae
Crabrsquoseyes
Seed
androot
Oralskin
Treatm
ento
fcou
ghskin
carew
ound
Grin
dther
ootand
applythep
oultice
onthea
ffected
skin
areaA
lsocrushedseeds
canbe
takenorally
Acanthus
montanu
sAc
anthaceae
False
thistle
Leafandroot
Oraland
Skin
Treatm
ento
ffurun
cles
lesio
nsandurinary
infections
Boilther
ootand
applythep
oultice
tothe
affectedpartof
theb
odyFo
rinfectio
ns
boilther
ootand
leaves
anddrink
Piperu
mbellatum
Piperaceae
Njamuja
Leaf
Oral
Treatm
ento
fpeptic
ulcers
Crushtheleavesa
ndsoak
inwarm
orho
twater
anddrink
Euphorbiahirta
Euph
orbiaceae
Asth
maw
eed
Who
leplant
Oral
Treatm
ento
fwou
nds
Leaves
arem
acerated
andappliedto
affectedpart
Ficusb
urkei(Miq)Miq
Moraceae
Sand
paper
Leaf
Oral
Treatm
ento
fhypertension
Leaves
areb
oiledandtakenorally
6 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Gmelina
arborea
Verbenaceae
Gmelina
Stem
andleaf
Oral
Treatm
ento
fgon
orrhea
pilesabdo
minalpains
burningsensationsfever
mou
thwo
undanddiabetes
Snake-biteandscorpion
sting
Leaves
grou
ndinto
paste
with
water
are
appliedto
theforeheadforh
eadachein
feversB
oiledste
msa
retakenorally
Lophira
alata
Ochnaceae
Ekki
Stem
andleaf
Oral
Treatm
ento
ffeverm
outh
woun
dandmalaria
Boiltheleavesa
ndste
mandtake
orally
Nau
clealatifolia
Rubiaceae
Ubu
ru-in
uStem
andleaf
Oral
Itisused
inthetreatment
ofmalariaepilepsymou
thwo
undanxietypainfever
Boilther
ootsanddrink
Annona
murica
taAnn
onaceae
Sour
sop
Leaf
Oral
Treatm
ento
fcancera
ndmou
thwo
undinfections
Crushor
boiltheleavesa
ndtake
orally
Psidium
guajava
Myrtaceae
Guava
Leaf
Oral
Treatm
ento
fperiodo
ntal
diseasebu
rnsscrapes
sunb
urnwo
undspsoria
sis
eczemaandothers
Leaves
arec
rushed
andtakenorally
Astra
galusm
embran
aceus
Fabaceae
Beiq
iStem
andleaf
Oral
Treatm
ento
fcoldup
per
respira
tory
infections
allergiesfib
romyalgia
anem
iaH
IVA
IDS
Leaves
arec
rushed
andtakenorallyand
canalso
beappliedto
skin
Buchholziacoria
cea
Capparaceae
Won
derfu
lkola
Leaf
Oral
Itisused
tradition
allyfor
treatingdiabetes
hypertensio
nrheumatism
coldcou
ghand
catarrh
Boilleaves
andtake
orally
Gnetum
afric
anum
Gnetacea
Afang
Leaf
Oral
Enlarged
spleen
sore-th
roatandcathartic
Leaves
canbe
boiledor
cooked
andtaken
orally
Anoridium
man
niAnn
onaceae
Ewuro-ig
boStem
bark
Oral
Treatm
ento
fDiarrhea
coug
hfeverrheumatism
Theb
arkiscrushedandbo
iledandthen
takenorally
Funtum
iaafric
ana
Apocyn
aceae
Akorie
mbamiri
Leafstem
root
Skin
treatmento
fcon
stipatio
nwo
undsw
eakbladder
jaun
dice
Who
leplantisb
oiledandtakenorally
Also
thec
rushed
leaves
area
ppliedto
the
affectedskin
area
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
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Zoology
Hindawiwwwhindawicom Volume 2018
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GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
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2 The Scientific World Journal
odontogenic infection Falsetta and colleagues [3] highlightedthat poor oral hygiene and frequent consumption of sugarscreate bacterial adhesion to tooth surfaces via mineralizationand biofilm formations caused by glucosyl and fructosyltransferases The same authors indicated an adverse effectof higher concentration of fluoride in drinking water insome developing countries that is responsible for the causeof fluorosis and dental caries regardless of age and sex Theoral cavity exposed to carcinogens that may advance into pre-cancerous lesions and malignant disorders like leukoplakiasubmucous fibrosis and dental caries resulting from theimbalance in the levels of free radicals generation and naturalantioxidants in the saliva [4] There is a growing consensusthat administration of natural antioxidants to the oral cavityreduces free radicals oxidative stress and oral inflammationtriggered by nicotine alcohol and other toxins introducedinto the mouth [5]
The skin being the largest organ in the body guards theunderlying muscles bones ligaments and internal organsinterfaces with the environment as well as protecting thebody against pathogenic microbes [6] Notwithstandinginfectious skin diseases such as skin abrasions burns acnedermatitis and sebaceous cysts have become everyday inci-dences and constitute a reoccurring health problem thataffects all human age groups [6] Over the years bacterial skindisorders such as impetigo ecthyma folliculitis eczemas andurticarias are one of the most common causes of hospitalvisits in the southeastern part of Nigeria Overcrowding mal-nutrition and humidity heat food and medication allergiescause someof these skin diseases especially among the schoolchildren in urban and rural settings [2] Microbial invasionof the skin surface forms a biofilm that confers a drugtolerance on the bacteria and capacity to escape host immuneresponse [7] The discovery of antibiotic resistance genesin several microorganisms quorum sensing and horizontalgene transfer has limited treatment of skin and soft tissueinfections and thus could pose a serious health challengein a region where there is limited access to microbiologicallaboratory facilities and antimicrobial agents [8] The globalneed for effective alternative medicines for oral and skininfections with minimal side effects and economical use isincreasing due to the rapid growth of antibiotic resistance andopportunistic infections in immunocompromised patients[9]
Plant-derived products are now gaining recognitionworldwide currently recommended for primary healthcarebut only a few plants have received rigorous scientific investi-gation Medicinal plants are natural products used since timeimmemorial for the treatment of various human diseases Allover the world the citizen and health practitioners are nowstarting to rely on herbal medicines as a substitute for scien-tifically proved therapies [10] Herbal products contained sec-ondary metabolites that are synthesized through the pentosephosphate shikimic acid and phenylpropanoid pathwaysThese compounds play a significant defensive role againstpathogenic bacteria and oxidative stress caused by abioticstress in plant [11] Currently one of the strategies used to treatskin and oral infections is the use of natural products fromplant origin perhaps due to the perception that long-term
use of western medicine induces severe complications Con-sequently many Nigerian patients in particular those livingin the rural areas have resorted to seeking relief via tradi-tional healers and herbalists who administer herbal drugsin the management of human diseases Considering therate at which the vegetation gets depleted and the currentfocus on safe complementary and alternative medicine tosynthetic drugs there is therefore the need to document theindigenous knowledge of these plants and to search for moreplants with therapeutic potential in the treatment of bacterialinfections and oxidative stress
There is a paucity of literature in traditional medicineused in the management of orofacial and skin infectionsin Nigeria Our recent ethnobotanical survey conductedin Ebem-Ohafia LGAs in the southeastern part of Nigeriarevealed sixty-one plant species commonly used in the man-agement of oral and skin infections Despite the acclaimedtraditional use of these botanicals as therapeutic agents thereis little scientific evidence to support their folkloric useTherefore this study aimed at investigating the antioxidantcapacity of these herbal remedies in terms of free radicalscavenging reducing power and total phenolic and flavonoidcontent in consonance with the TLC profiling The antibacte-rial effect on some clinically isolated bacteria associated withoral and skin infections was also investigated
2 Materials and Methods
21 Study Area Ebem-Ohafia is a town in Ohafia LGA ofAbia state in the southeastern part ofNigeria situated between5∘ 381015840N and 7∘ 501015840 East and encompasses over twenty-sixhometowns with a population of over 300 000 from thelast population census in 2006 Ebem community is a partof the tropical rainforest characterized by dry and rainyseason with a total annual rainfall of over 1400 mm and anannual temperature range of 23∘C to 32∘C The people of theregion practice subsistence farming under local governmentagencies policy The inhabitants of Ebem use herbal drugseither alone or in combination with other medicines in thetreatment of human diseases Most of the people in Ebemare rural dwellers hence they use plant-based therapies in themanagement of ailments that include oral and skin infections
22 Ethnobotanical Survey The ethnobotanical survey wasconducted between December and May 2016 using a well-structured questionnaire administered to the participantswith indigenous knowledge of plants utilized in the area Theset questions contained the local diagnosis of oral and skindiseases the name of plant samples used for the treatmentmethods of preparation and duration of treatment sideeffects and mode of administration The people interviewedconsisted of women andmen both married and unmarried atthe age of 30 to 65 with little education qualification
23 PlantCollection and Identification The leaves of Ipomoeainvolucrata (II) Acalypha hispida (AH) Breynia nivosa (BN)Jatropha curcas (JC) Chromolaena odorata (CO) Macrolo-bium macrophyllum (MM) Baphia nitida (BNI) Burkeaafricana (BA) Cassia alata (CA) and the root of Eleusine
The Scientific World Journal 3
indica (EI) were collected after the interview The plants wereidentified by their local names and authenticated by MrOriaku Williams and Professor Gabriel Osuagwu from theDepartment of Plant Science and Biotechnology College ofNatural Sciences Michael Okpara University of AgricultureUmudike in Abia state Nigeria The vouchers specimens(VICJON 1ndash10) were prepared and deposited at the depart-mental herbarium The preparations and mode of intake ofthe herbal remedies are described in Table 1
24 Sample Preparation We collected leaves and roots ofthe ten plants that were frequently mentioned in the surveyand oven-dried at 40∘C for 72 h while the root of EI wasdried longer for seven days The dried plant materials werepulverized to a fine powder using an electric blender andstored in an airtight container for further use Thirty grams(30 g) of dried powdered materials were extracted with 150mL of 100methanol for 48 h on amechanical shaker (StuartScientific Orbital 202 SOSI Essex UK) and the extractswere filtered using Buchner funnel and Whatman Number1 filter paper The filtrate was concentrated using a rotaryevaporated at 40∘C to recover the solvent and air-dried in afume chamber to give a yield ranging from 45 to 83 g
25 Thin Layer Chromatography (TLC) Profile Thin layerchromatography is a simple method for analyzing a complexmixture of compounds based on the distance travelled Theplant extracts (1 mgmL) dissolved in methanol and spottedon the plate coated with silica gel 60 F254 as a stationaryphase The slurry was prepared by dissolving 15 g of silicagel 60 F254 in 30 mL of distilled water and immediatelypoured into the plate The plates were air-dried overnightAbout 10 120583L of the plant extracts was gently loaded onthe base of the plate (5 cm above) using the capillary tubeThe plates were allowed to develop in chromatographictanks consisting of three different solvents (mobile phase)chloroform methanol acetic acid (541) until they reach34th of the TLC plate The TLC plate was removed andallowed to dry the spots were detected by iodine vapour andretention factor (Rf) was calculated using the equation Rf= distance travelled by the componentsdistance travelled bythe solvent
26 Total Phenolic Content (TPC) Assay The total phenoliccontent present in these extracts was quantified by the FolinCiocalteu reagent (FCR) using the method of Ghaffari etal [12] Briefly 02 mL of the plant extract (2 mgmL) wasadded to the reaction mixture consisting of 1 mL of 10 vvFCR and 08 mL of Na2CO3 (0075 mgmL) to give a finalconcentration of 02 mgmL of each extract The resultingmixture was incubated at 45∘C with shaking for 15 min andthe absorbance measured at 765 nm A standard curve wasprepared by mixing methanol solution of gallic acid (02mL 0025 - 0400 mgmL) with 1 mL of 10 vv FCR andsodium carbonate (08 mL 0075 mgmL) The experimentwas carried out in triplicate and the results presented asmeanvalues with standard deviation (plusmn SD) The TPC value wasexpressed as milligrams of gallic acid equivalent (GAE) perg of dried sample It was calculated using the formula T =
CtimesVM where T is the TPC (mgg) of extract in GAE C isthe concentration of gallic acid from the calibration curve Vis the volume of the extract mL M is the dry weight (g) ofthe leaf or root powder from which the extract was obtained
27 Total Flavonoid Content (TFC) Assay The concentra-tion of flavonoids in the plant extracts was determined viaaluminum colorimetric assay method [13] Briefly 1 mL of2 wv AlCl3 was prepared in 100vv methanol added to 1mL of the sample solution A yellow colour formation afterincubation at room temperature for 1 h was measured at420 nm using an AJI-C03 UV VIS spectrophotometer Thestandard curve for TFC was obtained using quercetin as astandard drug under the same procedure described in TPCdetermination The TFC was calculated using the formulaT = CtimesVM where T is the TFC (mgg) of extract in QEC is the concentration of quercetin established from thecalibration curve V is the volume of the extract mL M isthe dry weight (g) of the leaf powder from which the extractwas obtained The TFC was present in the extracts calculatedas mgg of quercetin equivalent (QE)
28 Ferric Reducing Antioxidant Power (FRAP) Assay Theability of plant extracts to reduce ferric (Fe3+) to ferrous(Fe2+) formwas evaluated following themethod described byYen and Chen [14] with slight modification A volume of 03mL of different concentrations (0025 - 2 mgmL) from plantextract BHT ascorbic acid and rutin prepared in distilledwater was mixed with reacting mixture consisting of 25 mLof 02 M phosphate buffer (pH 66) and 25 mL of K3Fe(CN)6(1 wv) The resulting mixture was incubated at 50∘C for20 min followed by addition of 25 mL of TCA (10 wv)After vigorous shaking 25 mL of the solution was mixedwith 25 mL of distilled water and 05 mL of FeCl3 (01 wv)left incubated at room temperature for 5 min and then theabsorbance was measured at 700 nm against a blank sample(without extract)
29 Free Radical Scavenging Activity Assays The free radicalscavenging potential of plant extracts was measured in vitroby the 1 11015840-diphenyl-1-picrylhydrazyl (DPPH) described byTariq et al [15] The assay was experimented by reacting16 mL of 0135 mM DPPH dissolved in 100vv methanolwith 04 mL of various concentrations (0078 - 1 mgmL) ofmethanol crude extracts The reaction mixture was vortexedthoroughly and left in the dark at room temperature for30 min The absorbance of the mixture was measured at517 nm after 2 min The method of Re et al [16] wasadopted to determine ABTS radical scavenging activity ofthe plant extracts The ABTS radical solution was generatedby mixing two stock solutions of 7 mM ABTS and 24mM potassium persulphate in the same ratio and allowingthe solution to react for 12 h at room temperature in thedark The resulting solution was diluted with methanol toobtain an absorbance of 0706 units at 734 nm A volumeof 1 mL of various concentrations (003125 ndash 1 mgmL)of the plant extracts react with 25 mL of ABTS radicalsolution in the dark for 15 min and was later measured
4 The Scientific World Journal
Table1Medicinalplantsused
inthem
anagem
ento
fskinor
oralinfections
insoutheastern
partof
Nigeria
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Macrolobium
macrophyllum
Fabaceae
Antho
notha
macroph
ylla
Leaf
Skin
Treatin
gof
woun
dandcut
Squeezethe
leavea
ndplaceo
nthe
affectedarea
ofthes
kin
Jatro
phacurcas
Euph
orbiaceae
Barbados
nut
Leaf
Skinoral
Treatm
ento
fboils
Boiltheleave
with
waterPlace
theh
otleaveo
nthea
ffected
area
ofthes
kin
Eleusin
eind
icaPo
aceae
Yard
grass
Root
Skin
Treatm
ento
fRingw
orm
Use
ther
ootand
scrubon
affectedarea
ofthes
kin
Cassiaalata
Caesalpiniaceae
Ring
worm
bush
Leaf
Skin
Treatm
ento
fRingw
orm
Scrubtheleave
onaffectedarea
ofthe
skin
Chromolaena
odorata
Aste
raceae
Siam
weed
Leaf
Skin
Tosto
pbleeding
andrashes
Squeezethe
leavea
ndrubon
affectedarea
Psidium
guajava(L)
Myrtaceae
Guava
Leaf
Skin
Treatm
ento
fskinbu
rns
scrapessunb
urn
woun
ds
psoriasiseczem
aand
others
Crushedgu
avaleafo
rboiledleaves
are
applieddirectlyto
thea
ffected
skin
providingreliefandareb
elievedto
improveh
ealin
gBryophyllum
pinn
atum
(Lam
)Oken
Crassulaceae
Mira
cleleaf
Dogrsquosliver
Leaf
Skino
ral
Treatm
ento
fboilwo
und
sore
orcut
Crushleafandapplyjuiceo
nthew
ound
surfa
ce
Ficusa
sperifolia
Miq
Moraceae
False
thistle
Leaflatex
Skin
Treatm
ento
fskinrashes
anditches
Appliedas
poultic
etothes
kinitches
Acanthus
montany
(TA
nderson)
Acanthaceae
Bearrsquosbreeches
Stem
bark
Skin
Treatm
ento
fwou
ndsa
ndskin
diseases
Woo
dbark
iscrushedandsoaked
italcoho
landextractapp
liedto
woun
dsandskin
Alliu
msativ
umL
Liliaceae
Garlic
Bulbjuice
Skin
oral
Usedto
cond
ition
thes
kin
chew
edforc
ough
andsore
throat
Thejuice
ismixed
with
lotio
nsforskin
cond
ition
ingFreshbu
lbischew
edor
boiledin
water
anddrinkdecoctionfor
coug
h
Emiliasonchifolia
(L)
Aste
raceae
Scarlettassel
Leaf
Skin
Treatm
ento
fSkintro
ubles
abscessa
ndbruises
Crushedleaves
were
pressedandused
tocoverthe
woun
dor
bruisedsurfa
ces
Sida
acutaBu
rmf
Malvaceae
Broo
mwe
edLeaf
Skin
Treatm
ento
fjaund
icesore
ulcer
Poultic
esmadefrom
boiledleaves
are
appliedon
thea
ffected
parts
The Scientific World Journal 5
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Ricin
uscommun
isLinn
Eu
phorbiaceae
Castorb
ean
Leafroo
tand
seed
Skin
Treatm
ento
fskininfection
anddetoxification
Applycasto
roilon
thea
ffected
area
torelieve
skin
infection
Citru
slim
on(L)Bu
rmf
(prosp
)Ru
taceae
Limon
Leaffruit
andseed
Skin
Treatm
ento
ffluheadache
cold
andoralinfections
Mixequalquantities
ofLimon
juicea
ndho
lybasilkeepitun
derthe
suntill
becomes
thickapplythison
face
Alliu
mcepa
LLiliaceae
Onion
sLeafandbu
lbSkin
Treatm
ento
fperiodo
ntitis
andcold
Theb
ulbismacerated
andmixed
with
water
andthen
takenorally
Breynianivosa
Euph
orbiaceae
Icep
lant
Leaf
Oral
Treatin
gof
toothache
Boiltheleave
with
waterthentake
the
water
Ipom
oeainvolucrata
Con
volvulaceae
Water
convolvu
lus
Leafandste
mOral
Treatm
ento
foralinfectio
nMixtheleave
with
ugbaga
treea
ndbo
ilwith
water
anddrinkthew
ater
Burkea
afric
ana
Caesalpinioideae
Icheku
Leaf
Oral
Treatm
ento
forald
isease
Boiltheleave
with
water
andtakebut
donrsquot
swallowit
Baphianitid
aPapilio
naceae
Cam
wood
Leafor
stem
Oral
Streng
thensthe
teeth
Chew
thes
tem
orscrubtheleave
onthe
teeth
Gongronem
alatifolium
Asclepiadaceae
Utazi
Leaf
Oral
Treatm
ento
fcou
ghand
diabetes
Crushtheleafw
ithmortara
ndmixwith
ginger
orgarlica
nddrink
Anonasenegalen
sisAnn
onaceae
Ubu
ruocha
Root
Oral
Treatm
ento
fmalariaand
mou
thinfection
Boilther
ootsanddrink
Abrusp
recatoriu
sPapilio
noideae
Crabrsquoseyes
Seed
androot
Oralskin
Treatm
ento
fcou
ghskin
carew
ound
Grin
dther
ootand
applythep
oultice
onthea
ffected
skin
areaA
lsocrushedseeds
canbe
takenorally
Acanthus
montanu
sAc
anthaceae
False
thistle
Leafandroot
Oraland
Skin
Treatm
ento
ffurun
cles
lesio
nsandurinary
infections
Boilther
ootand
applythep
oultice
tothe
affectedpartof
theb
odyFo
rinfectio
ns
boilther
ootand
leaves
anddrink
Piperu
mbellatum
Piperaceae
Njamuja
Leaf
Oral
Treatm
ento
fpeptic
ulcers
Crushtheleavesa
ndsoak
inwarm
orho
twater
anddrink
Euphorbiahirta
Euph
orbiaceae
Asth
maw
eed
Who
leplant
Oral
Treatm
ento
fwou
nds
Leaves
arem
acerated
andappliedto
affectedpart
Ficusb
urkei(Miq)Miq
Moraceae
Sand
paper
Leaf
Oral
Treatm
ento
fhypertension
Leaves
areb
oiledandtakenorally
6 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Gmelina
arborea
Verbenaceae
Gmelina
Stem
andleaf
Oral
Treatm
ento
fgon
orrhea
pilesabdo
minalpains
burningsensationsfever
mou
thwo
undanddiabetes
Snake-biteandscorpion
sting
Leaves
grou
ndinto
paste
with
water
are
appliedto
theforeheadforh
eadachein
feversB
oiledste
msa
retakenorally
Lophira
alata
Ochnaceae
Ekki
Stem
andleaf
Oral
Treatm
ento
ffeverm
outh
woun
dandmalaria
Boiltheleavesa
ndste
mandtake
orally
Nau
clealatifolia
Rubiaceae
Ubu
ru-in
uStem
andleaf
Oral
Itisused
inthetreatment
ofmalariaepilepsymou
thwo
undanxietypainfever
Boilther
ootsanddrink
Annona
murica
taAnn
onaceae
Sour
sop
Leaf
Oral
Treatm
ento
fcancera
ndmou
thwo
undinfections
Crushor
boiltheleavesa
ndtake
orally
Psidium
guajava
Myrtaceae
Guava
Leaf
Oral
Treatm
ento
fperiodo
ntal
diseasebu
rnsscrapes
sunb
urnwo
undspsoria
sis
eczemaandothers
Leaves
arec
rushed
andtakenorally
Astra
galusm
embran
aceus
Fabaceae
Beiq
iStem
andleaf
Oral
Treatm
ento
fcoldup
per
respira
tory
infections
allergiesfib
romyalgia
anem
iaH
IVA
IDS
Leaves
arec
rushed
andtakenorallyand
canalso
beappliedto
skin
Buchholziacoria
cea
Capparaceae
Won
derfu
lkola
Leaf
Oral
Itisused
tradition
allyfor
treatingdiabetes
hypertensio
nrheumatism
coldcou
ghand
catarrh
Boilleaves
andtake
orally
Gnetum
afric
anum
Gnetacea
Afang
Leaf
Oral
Enlarged
spleen
sore-th
roatandcathartic
Leaves
canbe
boiledor
cooked
andtaken
orally
Anoridium
man
niAnn
onaceae
Ewuro-ig
boStem
bark
Oral
Treatm
ento
fDiarrhea
coug
hfeverrheumatism
Theb
arkiscrushedandbo
iledandthen
takenorally
Funtum
iaafric
ana
Apocyn
aceae
Akorie
mbamiri
Leafstem
root
Skin
treatmento
fcon
stipatio
nwo
undsw
eakbladder
jaun
dice
Who
leplantisb
oiledandtakenorally
Also
thec
rushed
leaves
area
ppliedto
the
affectedskin
area
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
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Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
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Cell BiologyInternational Journal of
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Hindawiwwwhindawicom Volume 2018
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ArchaeaHindawiwwwhindawicom Volume 2018
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Nucleic AcidsJournal of
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Submit your manuscripts atwwwhindawicom
The Scientific World Journal 3
indica (EI) were collected after the interview The plants wereidentified by their local names and authenticated by MrOriaku Williams and Professor Gabriel Osuagwu from theDepartment of Plant Science and Biotechnology College ofNatural Sciences Michael Okpara University of AgricultureUmudike in Abia state Nigeria The vouchers specimens(VICJON 1ndash10) were prepared and deposited at the depart-mental herbarium The preparations and mode of intake ofthe herbal remedies are described in Table 1
24 Sample Preparation We collected leaves and roots ofthe ten plants that were frequently mentioned in the surveyand oven-dried at 40∘C for 72 h while the root of EI wasdried longer for seven days The dried plant materials werepulverized to a fine powder using an electric blender andstored in an airtight container for further use Thirty grams(30 g) of dried powdered materials were extracted with 150mL of 100methanol for 48 h on amechanical shaker (StuartScientific Orbital 202 SOSI Essex UK) and the extractswere filtered using Buchner funnel and Whatman Number1 filter paper The filtrate was concentrated using a rotaryevaporated at 40∘C to recover the solvent and air-dried in afume chamber to give a yield ranging from 45 to 83 g
25 Thin Layer Chromatography (TLC) Profile Thin layerchromatography is a simple method for analyzing a complexmixture of compounds based on the distance travelled Theplant extracts (1 mgmL) dissolved in methanol and spottedon the plate coated with silica gel 60 F254 as a stationaryphase The slurry was prepared by dissolving 15 g of silicagel 60 F254 in 30 mL of distilled water and immediatelypoured into the plate The plates were air-dried overnightAbout 10 120583L of the plant extracts was gently loaded onthe base of the plate (5 cm above) using the capillary tubeThe plates were allowed to develop in chromatographictanks consisting of three different solvents (mobile phase)chloroform methanol acetic acid (541) until they reach34th of the TLC plate The TLC plate was removed andallowed to dry the spots were detected by iodine vapour andretention factor (Rf) was calculated using the equation Rf= distance travelled by the componentsdistance travelled bythe solvent
26 Total Phenolic Content (TPC) Assay The total phenoliccontent present in these extracts was quantified by the FolinCiocalteu reagent (FCR) using the method of Ghaffari etal [12] Briefly 02 mL of the plant extract (2 mgmL) wasadded to the reaction mixture consisting of 1 mL of 10 vvFCR and 08 mL of Na2CO3 (0075 mgmL) to give a finalconcentration of 02 mgmL of each extract The resultingmixture was incubated at 45∘C with shaking for 15 min andthe absorbance measured at 765 nm A standard curve wasprepared by mixing methanol solution of gallic acid (02mL 0025 - 0400 mgmL) with 1 mL of 10 vv FCR andsodium carbonate (08 mL 0075 mgmL) The experimentwas carried out in triplicate and the results presented asmeanvalues with standard deviation (plusmn SD) The TPC value wasexpressed as milligrams of gallic acid equivalent (GAE) perg of dried sample It was calculated using the formula T =
CtimesVM where T is the TPC (mgg) of extract in GAE C isthe concentration of gallic acid from the calibration curve Vis the volume of the extract mL M is the dry weight (g) ofthe leaf or root powder from which the extract was obtained
27 Total Flavonoid Content (TFC) Assay The concentra-tion of flavonoids in the plant extracts was determined viaaluminum colorimetric assay method [13] Briefly 1 mL of2 wv AlCl3 was prepared in 100vv methanol added to 1mL of the sample solution A yellow colour formation afterincubation at room temperature for 1 h was measured at420 nm using an AJI-C03 UV VIS spectrophotometer Thestandard curve for TFC was obtained using quercetin as astandard drug under the same procedure described in TPCdetermination The TFC was calculated using the formulaT = CtimesVM where T is the TFC (mgg) of extract in QEC is the concentration of quercetin established from thecalibration curve V is the volume of the extract mL M isthe dry weight (g) of the leaf powder from which the extractwas obtained The TFC was present in the extracts calculatedas mgg of quercetin equivalent (QE)
28 Ferric Reducing Antioxidant Power (FRAP) Assay Theability of plant extracts to reduce ferric (Fe3+) to ferrous(Fe2+) formwas evaluated following themethod described byYen and Chen [14] with slight modification A volume of 03mL of different concentrations (0025 - 2 mgmL) from plantextract BHT ascorbic acid and rutin prepared in distilledwater was mixed with reacting mixture consisting of 25 mLof 02 M phosphate buffer (pH 66) and 25 mL of K3Fe(CN)6(1 wv) The resulting mixture was incubated at 50∘C for20 min followed by addition of 25 mL of TCA (10 wv)After vigorous shaking 25 mL of the solution was mixedwith 25 mL of distilled water and 05 mL of FeCl3 (01 wv)left incubated at room temperature for 5 min and then theabsorbance was measured at 700 nm against a blank sample(without extract)
29 Free Radical Scavenging Activity Assays The free radicalscavenging potential of plant extracts was measured in vitroby the 1 11015840-diphenyl-1-picrylhydrazyl (DPPH) described byTariq et al [15] The assay was experimented by reacting16 mL of 0135 mM DPPH dissolved in 100vv methanolwith 04 mL of various concentrations (0078 - 1 mgmL) ofmethanol crude extracts The reaction mixture was vortexedthoroughly and left in the dark at room temperature for30 min The absorbance of the mixture was measured at517 nm after 2 min The method of Re et al [16] wasadopted to determine ABTS radical scavenging activity ofthe plant extracts The ABTS radical solution was generatedby mixing two stock solutions of 7 mM ABTS and 24mM potassium persulphate in the same ratio and allowingthe solution to react for 12 h at room temperature in thedark The resulting solution was diluted with methanol toobtain an absorbance of 0706 units at 734 nm A volumeof 1 mL of various concentrations (003125 ndash 1 mgmL)of the plant extracts react with 25 mL of ABTS radicalsolution in the dark for 15 min and was later measured
4 The Scientific World Journal
Table1Medicinalplantsused
inthem
anagem
ento
fskinor
oralinfections
insoutheastern
partof
Nigeria
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Macrolobium
macrophyllum
Fabaceae
Antho
notha
macroph
ylla
Leaf
Skin
Treatin
gof
woun
dandcut
Squeezethe
leavea
ndplaceo
nthe
affectedarea
ofthes
kin
Jatro
phacurcas
Euph
orbiaceae
Barbados
nut
Leaf
Skinoral
Treatm
ento
fboils
Boiltheleave
with
waterPlace
theh
otleaveo
nthea
ffected
area
ofthes
kin
Eleusin
eind
icaPo
aceae
Yard
grass
Root
Skin
Treatm
ento
fRingw
orm
Use
ther
ootand
scrubon
affectedarea
ofthes
kin
Cassiaalata
Caesalpiniaceae
Ring
worm
bush
Leaf
Skin
Treatm
ento
fRingw
orm
Scrubtheleave
onaffectedarea
ofthe
skin
Chromolaena
odorata
Aste
raceae
Siam
weed
Leaf
Skin
Tosto
pbleeding
andrashes
Squeezethe
leavea
ndrubon
affectedarea
Psidium
guajava(L)
Myrtaceae
Guava
Leaf
Skin
Treatm
ento
fskinbu
rns
scrapessunb
urn
woun
ds
psoriasiseczem
aand
others
Crushedgu
avaleafo
rboiledleaves
are
applieddirectlyto
thea
ffected
skin
providingreliefandareb
elievedto
improveh
ealin
gBryophyllum
pinn
atum
(Lam
)Oken
Crassulaceae
Mira
cleleaf
Dogrsquosliver
Leaf
Skino
ral
Treatm
ento
fboilwo
und
sore
orcut
Crushleafandapplyjuiceo
nthew
ound
surfa
ce
Ficusa
sperifolia
Miq
Moraceae
False
thistle
Leaflatex
Skin
Treatm
ento
fskinrashes
anditches
Appliedas
poultic
etothes
kinitches
Acanthus
montany
(TA
nderson)
Acanthaceae
Bearrsquosbreeches
Stem
bark
Skin
Treatm
ento
fwou
ndsa
ndskin
diseases
Woo
dbark
iscrushedandsoaked
italcoho
landextractapp
liedto
woun
dsandskin
Alliu
msativ
umL
Liliaceae
Garlic
Bulbjuice
Skin
oral
Usedto
cond
ition
thes
kin
chew
edforc
ough
andsore
throat
Thejuice
ismixed
with
lotio
nsforskin
cond
ition
ingFreshbu
lbischew
edor
boiledin
water
anddrinkdecoctionfor
coug
h
Emiliasonchifolia
(L)
Aste
raceae
Scarlettassel
Leaf
Skin
Treatm
ento
fSkintro
ubles
abscessa
ndbruises
Crushedleaves
were
pressedandused
tocoverthe
woun
dor
bruisedsurfa
ces
Sida
acutaBu
rmf
Malvaceae
Broo
mwe
edLeaf
Skin
Treatm
ento
fjaund
icesore
ulcer
Poultic
esmadefrom
boiledleaves
are
appliedon
thea
ffected
parts
The Scientific World Journal 5
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Ricin
uscommun
isLinn
Eu
phorbiaceae
Castorb
ean
Leafroo
tand
seed
Skin
Treatm
ento
fskininfection
anddetoxification
Applycasto
roilon
thea
ffected
area
torelieve
skin
infection
Citru
slim
on(L)Bu
rmf
(prosp
)Ru
taceae
Limon
Leaffruit
andseed
Skin
Treatm
ento
ffluheadache
cold
andoralinfections
Mixequalquantities
ofLimon
juicea
ndho
lybasilkeepitun
derthe
suntill
becomes
thickapplythison
face
Alliu
mcepa
LLiliaceae
Onion
sLeafandbu
lbSkin
Treatm
ento
fperiodo
ntitis
andcold
Theb
ulbismacerated
andmixed
with
water
andthen
takenorally
Breynianivosa
Euph
orbiaceae
Icep
lant
Leaf
Oral
Treatin
gof
toothache
Boiltheleave
with
waterthentake
the
water
Ipom
oeainvolucrata
Con
volvulaceae
Water
convolvu
lus
Leafandste
mOral
Treatm
ento
foralinfectio
nMixtheleave
with
ugbaga
treea
ndbo
ilwith
water
anddrinkthew
ater
Burkea
afric
ana
Caesalpinioideae
Icheku
Leaf
Oral
Treatm
ento
forald
isease
Boiltheleave
with
water
andtakebut
donrsquot
swallowit
Baphianitid
aPapilio
naceae
Cam
wood
Leafor
stem
Oral
Streng
thensthe
teeth
Chew
thes
tem
orscrubtheleave
onthe
teeth
Gongronem
alatifolium
Asclepiadaceae
Utazi
Leaf
Oral
Treatm
ento
fcou
ghand
diabetes
Crushtheleafw
ithmortara
ndmixwith
ginger
orgarlica
nddrink
Anonasenegalen
sisAnn
onaceae
Ubu
ruocha
Root
Oral
Treatm
ento
fmalariaand
mou
thinfection
Boilther
ootsanddrink
Abrusp
recatoriu
sPapilio
noideae
Crabrsquoseyes
Seed
androot
Oralskin
Treatm
ento
fcou
ghskin
carew
ound
Grin
dther
ootand
applythep
oultice
onthea
ffected
skin
areaA
lsocrushedseeds
canbe
takenorally
Acanthus
montanu
sAc
anthaceae
False
thistle
Leafandroot
Oraland
Skin
Treatm
ento
ffurun
cles
lesio
nsandurinary
infections
Boilther
ootand
applythep
oultice
tothe
affectedpartof
theb
odyFo
rinfectio
ns
boilther
ootand
leaves
anddrink
Piperu
mbellatum
Piperaceae
Njamuja
Leaf
Oral
Treatm
ento
fpeptic
ulcers
Crushtheleavesa
ndsoak
inwarm
orho
twater
anddrink
Euphorbiahirta
Euph
orbiaceae
Asth
maw
eed
Who
leplant
Oral
Treatm
ento
fwou
nds
Leaves
arem
acerated
andappliedto
affectedpart
Ficusb
urkei(Miq)Miq
Moraceae
Sand
paper
Leaf
Oral
Treatm
ento
fhypertension
Leaves
areb
oiledandtakenorally
6 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Gmelina
arborea
Verbenaceae
Gmelina
Stem
andleaf
Oral
Treatm
ento
fgon
orrhea
pilesabdo
minalpains
burningsensationsfever
mou
thwo
undanddiabetes
Snake-biteandscorpion
sting
Leaves
grou
ndinto
paste
with
water
are
appliedto
theforeheadforh
eadachein
feversB
oiledste
msa
retakenorally
Lophira
alata
Ochnaceae
Ekki
Stem
andleaf
Oral
Treatm
ento
ffeverm
outh
woun
dandmalaria
Boiltheleavesa
ndste
mandtake
orally
Nau
clealatifolia
Rubiaceae
Ubu
ru-in
uStem
andleaf
Oral
Itisused
inthetreatment
ofmalariaepilepsymou
thwo
undanxietypainfever
Boilther
ootsanddrink
Annona
murica
taAnn
onaceae
Sour
sop
Leaf
Oral
Treatm
ento
fcancera
ndmou
thwo
undinfections
Crushor
boiltheleavesa
ndtake
orally
Psidium
guajava
Myrtaceae
Guava
Leaf
Oral
Treatm
ento
fperiodo
ntal
diseasebu
rnsscrapes
sunb
urnwo
undspsoria
sis
eczemaandothers
Leaves
arec
rushed
andtakenorally
Astra
galusm
embran
aceus
Fabaceae
Beiq
iStem
andleaf
Oral
Treatm
ento
fcoldup
per
respira
tory
infections
allergiesfib
romyalgia
anem
iaH
IVA
IDS
Leaves
arec
rushed
andtakenorallyand
canalso
beappliedto
skin
Buchholziacoria
cea
Capparaceae
Won
derfu
lkola
Leaf
Oral
Itisused
tradition
allyfor
treatingdiabetes
hypertensio
nrheumatism
coldcou
ghand
catarrh
Boilleaves
andtake
orally
Gnetum
afric
anum
Gnetacea
Afang
Leaf
Oral
Enlarged
spleen
sore-th
roatandcathartic
Leaves
canbe
boiledor
cooked
andtaken
orally
Anoridium
man
niAnn
onaceae
Ewuro-ig
boStem
bark
Oral
Treatm
ento
fDiarrhea
coug
hfeverrheumatism
Theb
arkiscrushedandbo
iledandthen
takenorally
Funtum
iaafric
ana
Apocyn
aceae
Akorie
mbamiri
Leafstem
root
Skin
treatmento
fcon
stipatio
nwo
undsw
eakbladder
jaun
dice
Who
leplantisb
oiledandtakenorally
Also
thec
rushed
leaves
area
ppliedto
the
affectedskin
area
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
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Hindawiwwwhindawicom Volume 2018
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Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
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Cell BiologyInternational Journal of
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Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
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Hindawiwwwhindawicom Volume 2018
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Hindawiwwwhindawicom Volume 2018
International Journal of
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Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
4 The Scientific World Journal
Table1Medicinalplantsused
inthem
anagem
ento
fskinor
oralinfections
insoutheastern
partof
Nigeria
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Macrolobium
macrophyllum
Fabaceae
Antho
notha
macroph
ylla
Leaf
Skin
Treatin
gof
woun
dandcut
Squeezethe
leavea
ndplaceo
nthe
affectedarea
ofthes
kin
Jatro
phacurcas
Euph
orbiaceae
Barbados
nut
Leaf
Skinoral
Treatm
ento
fboils
Boiltheleave
with
waterPlace
theh
otleaveo
nthea
ffected
area
ofthes
kin
Eleusin
eind
icaPo
aceae
Yard
grass
Root
Skin
Treatm
ento
fRingw
orm
Use
ther
ootand
scrubon
affectedarea
ofthes
kin
Cassiaalata
Caesalpiniaceae
Ring
worm
bush
Leaf
Skin
Treatm
ento
fRingw
orm
Scrubtheleave
onaffectedarea
ofthe
skin
Chromolaena
odorata
Aste
raceae
Siam
weed
Leaf
Skin
Tosto
pbleeding
andrashes
Squeezethe
leavea
ndrubon
affectedarea
Psidium
guajava(L)
Myrtaceae
Guava
Leaf
Skin
Treatm
ento
fskinbu
rns
scrapessunb
urn
woun
ds
psoriasiseczem
aand
others
Crushedgu
avaleafo
rboiledleaves
are
applieddirectlyto
thea
ffected
skin
providingreliefandareb
elievedto
improveh
ealin
gBryophyllum
pinn
atum
(Lam
)Oken
Crassulaceae
Mira
cleleaf
Dogrsquosliver
Leaf
Skino
ral
Treatm
ento
fboilwo
und
sore
orcut
Crushleafandapplyjuiceo
nthew
ound
surfa
ce
Ficusa
sperifolia
Miq
Moraceae
False
thistle
Leaflatex
Skin
Treatm
ento
fskinrashes
anditches
Appliedas
poultic
etothes
kinitches
Acanthus
montany
(TA
nderson)
Acanthaceae
Bearrsquosbreeches
Stem
bark
Skin
Treatm
ento
fwou
ndsa
ndskin
diseases
Woo
dbark
iscrushedandsoaked
italcoho
landextractapp
liedto
woun
dsandskin
Alliu
msativ
umL
Liliaceae
Garlic
Bulbjuice
Skin
oral
Usedto
cond
ition
thes
kin
chew
edforc
ough
andsore
throat
Thejuice
ismixed
with
lotio
nsforskin
cond
ition
ingFreshbu
lbischew
edor
boiledin
water
anddrinkdecoctionfor
coug
h
Emiliasonchifolia
(L)
Aste
raceae
Scarlettassel
Leaf
Skin
Treatm
ento
fSkintro
ubles
abscessa
ndbruises
Crushedleaves
were
pressedandused
tocoverthe
woun
dor
bruisedsurfa
ces
Sida
acutaBu
rmf
Malvaceae
Broo
mwe
edLeaf
Skin
Treatm
ento
fjaund
icesore
ulcer
Poultic
esmadefrom
boiledleaves
are
appliedon
thea
ffected
parts
The Scientific World Journal 5
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Ricin
uscommun
isLinn
Eu
phorbiaceae
Castorb
ean
Leafroo
tand
seed
Skin
Treatm
ento
fskininfection
anddetoxification
Applycasto
roilon
thea
ffected
area
torelieve
skin
infection
Citru
slim
on(L)Bu
rmf
(prosp
)Ru
taceae
Limon
Leaffruit
andseed
Skin
Treatm
ento
ffluheadache
cold
andoralinfections
Mixequalquantities
ofLimon
juicea
ndho
lybasilkeepitun
derthe
suntill
becomes
thickapplythison
face
Alliu
mcepa
LLiliaceae
Onion
sLeafandbu
lbSkin
Treatm
ento
fperiodo
ntitis
andcold
Theb
ulbismacerated
andmixed
with
water
andthen
takenorally
Breynianivosa
Euph
orbiaceae
Icep
lant
Leaf
Oral
Treatin
gof
toothache
Boiltheleave
with
waterthentake
the
water
Ipom
oeainvolucrata
Con
volvulaceae
Water
convolvu
lus
Leafandste
mOral
Treatm
ento
foralinfectio
nMixtheleave
with
ugbaga
treea
ndbo
ilwith
water
anddrinkthew
ater
Burkea
afric
ana
Caesalpinioideae
Icheku
Leaf
Oral
Treatm
ento
forald
isease
Boiltheleave
with
water
andtakebut
donrsquot
swallowit
Baphianitid
aPapilio
naceae
Cam
wood
Leafor
stem
Oral
Streng
thensthe
teeth
Chew
thes
tem
orscrubtheleave
onthe
teeth
Gongronem
alatifolium
Asclepiadaceae
Utazi
Leaf
Oral
Treatm
ento
fcou
ghand
diabetes
Crushtheleafw
ithmortara
ndmixwith
ginger
orgarlica
nddrink
Anonasenegalen
sisAnn
onaceae
Ubu
ruocha
Root
Oral
Treatm
ento
fmalariaand
mou
thinfection
Boilther
ootsanddrink
Abrusp
recatoriu
sPapilio
noideae
Crabrsquoseyes
Seed
androot
Oralskin
Treatm
ento
fcou
ghskin
carew
ound
Grin
dther
ootand
applythep
oultice
onthea
ffected
skin
areaA
lsocrushedseeds
canbe
takenorally
Acanthus
montanu
sAc
anthaceae
False
thistle
Leafandroot
Oraland
Skin
Treatm
ento
ffurun
cles
lesio
nsandurinary
infections
Boilther
ootand
applythep
oultice
tothe
affectedpartof
theb
odyFo
rinfectio
ns
boilther
ootand
leaves
anddrink
Piperu
mbellatum
Piperaceae
Njamuja
Leaf
Oral
Treatm
ento
fpeptic
ulcers
Crushtheleavesa
ndsoak
inwarm
orho
twater
anddrink
Euphorbiahirta
Euph
orbiaceae
Asth
maw
eed
Who
leplant
Oral
Treatm
ento
fwou
nds
Leaves
arem
acerated
andappliedto
affectedpart
Ficusb
urkei(Miq)Miq
Moraceae
Sand
paper
Leaf
Oral
Treatm
ento
fhypertension
Leaves
areb
oiledandtakenorally
6 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Gmelina
arborea
Verbenaceae
Gmelina
Stem
andleaf
Oral
Treatm
ento
fgon
orrhea
pilesabdo
minalpains
burningsensationsfever
mou
thwo
undanddiabetes
Snake-biteandscorpion
sting
Leaves
grou
ndinto
paste
with
water
are
appliedto
theforeheadforh
eadachein
feversB
oiledste
msa
retakenorally
Lophira
alata
Ochnaceae
Ekki
Stem
andleaf
Oral
Treatm
ento
ffeverm
outh
woun
dandmalaria
Boiltheleavesa
ndste
mandtake
orally
Nau
clealatifolia
Rubiaceae
Ubu
ru-in
uStem
andleaf
Oral
Itisused
inthetreatment
ofmalariaepilepsymou
thwo
undanxietypainfever
Boilther
ootsanddrink
Annona
murica
taAnn
onaceae
Sour
sop
Leaf
Oral
Treatm
ento
fcancera
ndmou
thwo
undinfections
Crushor
boiltheleavesa
ndtake
orally
Psidium
guajava
Myrtaceae
Guava
Leaf
Oral
Treatm
ento
fperiodo
ntal
diseasebu
rnsscrapes
sunb
urnwo
undspsoria
sis
eczemaandothers
Leaves
arec
rushed
andtakenorally
Astra
galusm
embran
aceus
Fabaceae
Beiq
iStem
andleaf
Oral
Treatm
ento
fcoldup
per
respira
tory
infections
allergiesfib
romyalgia
anem
iaH
IVA
IDS
Leaves
arec
rushed
andtakenorallyand
canalso
beappliedto
skin
Buchholziacoria
cea
Capparaceae
Won
derfu
lkola
Leaf
Oral
Itisused
tradition
allyfor
treatingdiabetes
hypertensio
nrheumatism
coldcou
ghand
catarrh
Boilleaves
andtake
orally
Gnetum
afric
anum
Gnetacea
Afang
Leaf
Oral
Enlarged
spleen
sore-th
roatandcathartic
Leaves
canbe
boiledor
cooked
andtaken
orally
Anoridium
man
niAnn
onaceae
Ewuro-ig
boStem
bark
Oral
Treatm
ento
fDiarrhea
coug
hfeverrheumatism
Theb
arkiscrushedandbo
iledandthen
takenorally
Funtum
iaafric
ana
Apocyn
aceae
Akorie
mbamiri
Leafstem
root
Skin
treatmento
fcon
stipatio
nwo
undsw
eakbladder
jaun
dice
Who
leplantisb
oiledandtakenorally
Also
thec
rushed
leaves
area
ppliedto
the
affectedskin
area
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
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Hindawiwwwhindawicom Volume 2018
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Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
The Scientific World Journal 5
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Ricin
uscommun
isLinn
Eu
phorbiaceae
Castorb
ean
Leafroo
tand
seed
Skin
Treatm
ento
fskininfection
anddetoxification
Applycasto
roilon
thea
ffected
area
torelieve
skin
infection
Citru
slim
on(L)Bu
rmf
(prosp
)Ru
taceae
Limon
Leaffruit
andseed
Skin
Treatm
ento
ffluheadache
cold
andoralinfections
Mixequalquantities
ofLimon
juicea
ndho
lybasilkeepitun
derthe
suntill
becomes
thickapplythison
face
Alliu
mcepa
LLiliaceae
Onion
sLeafandbu
lbSkin
Treatm
ento
fperiodo
ntitis
andcold
Theb
ulbismacerated
andmixed
with
water
andthen
takenorally
Breynianivosa
Euph
orbiaceae
Icep
lant
Leaf
Oral
Treatin
gof
toothache
Boiltheleave
with
waterthentake
the
water
Ipom
oeainvolucrata
Con
volvulaceae
Water
convolvu
lus
Leafandste
mOral
Treatm
ento
foralinfectio
nMixtheleave
with
ugbaga
treea
ndbo
ilwith
water
anddrinkthew
ater
Burkea
afric
ana
Caesalpinioideae
Icheku
Leaf
Oral
Treatm
ento
forald
isease
Boiltheleave
with
water
andtakebut
donrsquot
swallowit
Baphianitid
aPapilio
naceae
Cam
wood
Leafor
stem
Oral
Streng
thensthe
teeth
Chew
thes
tem
orscrubtheleave
onthe
teeth
Gongronem
alatifolium
Asclepiadaceae
Utazi
Leaf
Oral
Treatm
ento
fcou
ghand
diabetes
Crushtheleafw
ithmortara
ndmixwith
ginger
orgarlica
nddrink
Anonasenegalen
sisAnn
onaceae
Ubu
ruocha
Root
Oral
Treatm
ento
fmalariaand
mou
thinfection
Boilther
ootsanddrink
Abrusp
recatoriu
sPapilio
noideae
Crabrsquoseyes
Seed
androot
Oralskin
Treatm
ento
fcou
ghskin
carew
ound
Grin
dther
ootand
applythep
oultice
onthea
ffected
skin
areaA
lsocrushedseeds
canbe
takenorally
Acanthus
montanu
sAc
anthaceae
False
thistle
Leafandroot
Oraland
Skin
Treatm
ento
ffurun
cles
lesio
nsandurinary
infections
Boilther
ootand
applythep
oultice
tothe
affectedpartof
theb
odyFo
rinfectio
ns
boilther
ootand
leaves
anddrink
Piperu
mbellatum
Piperaceae
Njamuja
Leaf
Oral
Treatm
ento
fpeptic
ulcers
Crushtheleavesa
ndsoak
inwarm
orho
twater
anddrink
Euphorbiahirta
Euph
orbiaceae
Asth
maw
eed
Who
leplant
Oral
Treatm
ento
fwou
nds
Leaves
arem
acerated
andappliedto
affectedpart
Ficusb
urkei(Miq)Miq
Moraceae
Sand
paper
Leaf
Oral
Treatm
ento
fhypertension
Leaves
areb
oiledandtakenorally
6 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Gmelina
arborea
Verbenaceae
Gmelina
Stem
andleaf
Oral
Treatm
ento
fgon
orrhea
pilesabdo
minalpains
burningsensationsfever
mou
thwo
undanddiabetes
Snake-biteandscorpion
sting
Leaves
grou
ndinto
paste
with
water
are
appliedto
theforeheadforh
eadachein
feversB
oiledste
msa
retakenorally
Lophira
alata
Ochnaceae
Ekki
Stem
andleaf
Oral
Treatm
ento
ffeverm
outh
woun
dandmalaria
Boiltheleavesa
ndste
mandtake
orally
Nau
clealatifolia
Rubiaceae
Ubu
ru-in
uStem
andleaf
Oral
Itisused
inthetreatment
ofmalariaepilepsymou
thwo
undanxietypainfever
Boilther
ootsanddrink
Annona
murica
taAnn
onaceae
Sour
sop
Leaf
Oral
Treatm
ento
fcancera
ndmou
thwo
undinfections
Crushor
boiltheleavesa
ndtake
orally
Psidium
guajava
Myrtaceae
Guava
Leaf
Oral
Treatm
ento
fperiodo
ntal
diseasebu
rnsscrapes
sunb
urnwo
undspsoria
sis
eczemaandothers
Leaves
arec
rushed
andtakenorally
Astra
galusm
embran
aceus
Fabaceae
Beiq
iStem
andleaf
Oral
Treatm
ento
fcoldup
per
respira
tory
infections
allergiesfib
romyalgia
anem
iaH
IVA
IDS
Leaves
arec
rushed
andtakenorallyand
canalso
beappliedto
skin
Buchholziacoria
cea
Capparaceae
Won
derfu
lkola
Leaf
Oral
Itisused
tradition
allyfor
treatingdiabetes
hypertensio
nrheumatism
coldcou
ghand
catarrh
Boilleaves
andtake
orally
Gnetum
afric
anum
Gnetacea
Afang
Leaf
Oral
Enlarged
spleen
sore-th
roatandcathartic
Leaves
canbe
boiledor
cooked
andtaken
orally
Anoridium
man
niAnn
onaceae
Ewuro-ig
boStem
bark
Oral
Treatm
ento
fDiarrhea
coug
hfeverrheumatism
Theb
arkiscrushedandbo
iledandthen
takenorally
Funtum
iaafric
ana
Apocyn
aceae
Akorie
mbamiri
Leafstem
root
Skin
treatmento
fcon
stipatio
nwo
undsw
eakbladder
jaun
dice
Who
leplantisb
oiledandtakenorally
Also
thec
rushed
leaves
area
ppliedto
the
affectedskin
area
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
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Hindawiwwwhindawicom Volume 2018
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Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
6 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Gmelina
arborea
Verbenaceae
Gmelina
Stem
andleaf
Oral
Treatm
ento
fgon
orrhea
pilesabdo
minalpains
burningsensationsfever
mou
thwo
undanddiabetes
Snake-biteandscorpion
sting
Leaves
grou
ndinto
paste
with
water
are
appliedto
theforeheadforh
eadachein
feversB
oiledste
msa
retakenorally
Lophira
alata
Ochnaceae
Ekki
Stem
andleaf
Oral
Treatm
ento
ffeverm
outh
woun
dandmalaria
Boiltheleavesa
ndste
mandtake
orally
Nau
clealatifolia
Rubiaceae
Ubu
ru-in
uStem
andleaf
Oral
Itisused
inthetreatment
ofmalariaepilepsymou
thwo
undanxietypainfever
Boilther
ootsanddrink
Annona
murica
taAnn
onaceae
Sour
sop
Leaf
Oral
Treatm
ento
fcancera
ndmou
thwo
undinfections
Crushor
boiltheleavesa
ndtake
orally
Psidium
guajava
Myrtaceae
Guava
Leaf
Oral
Treatm
ento
fperiodo
ntal
diseasebu
rnsscrapes
sunb
urnwo
undspsoria
sis
eczemaandothers
Leaves
arec
rushed
andtakenorally
Astra
galusm
embran
aceus
Fabaceae
Beiq
iStem
andleaf
Oral
Treatm
ento
fcoldup
per
respira
tory
infections
allergiesfib
romyalgia
anem
iaH
IVA
IDS
Leaves
arec
rushed
andtakenorallyand
canalso
beappliedto
skin
Buchholziacoria
cea
Capparaceae
Won
derfu
lkola
Leaf
Oral
Itisused
tradition
allyfor
treatingdiabetes
hypertensio
nrheumatism
coldcou
ghand
catarrh
Boilleaves
andtake
orally
Gnetum
afric
anum
Gnetacea
Afang
Leaf
Oral
Enlarged
spleen
sore-th
roatandcathartic
Leaves
canbe
boiledor
cooked
andtaken
orally
Anoridium
man
niAnn
onaceae
Ewuro-ig
boStem
bark
Oral
Treatm
ento
fDiarrhea
coug
hfeverrheumatism
Theb
arkiscrushedandbo
iledandthen
takenorally
Funtum
iaafric
ana
Apocyn
aceae
Akorie
mbamiri
Leafstem
root
Skin
treatmento
fcon
stipatio
nwo
undsw
eakbladder
jaun
dice
Who
leplantisb
oiledandtakenorally
Also
thec
rushed
leaves
area
ppliedto
the
affectedskin
area
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
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GenomicsInternational Journal of
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The Scientific World Journal
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Submit your manuscripts atwwwhindawicom
The Scientific World Journal 7
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Commiphora
afric
ana
Burseraceae
Turari
dashi
Rootsfruits
Oral
Treatm
ento
fwho
oping
coug
hbron
chitis
Rootsa
remacerated
andtakenorally
Dacryodesedulis
Burseraceae
Elem
iub
eBa
rkroo
tsfruits
Skin
Treatm
ento
fJiggersSkin
diseasesE
leph
antia
sis
Theleavesa
reeatenrawwith
kolanu
tas
anantie
meticLeaf-s
apisinstilledinto
thee
arfore
ar-tr
oubleanda
leaf-decoctio
nisprepared
asa
vapo
ur-bathforfeverish
stiffn
essw
ithheadache
Combretum
mucronatum
Com
bretaceae
Fararg
eza
Rootsa
ndleaf
Skin
Treatm
ento
fskininfection
Ther
ootand
leafarem
acerated
and
appliedto
affectedskin
area
Cleomec
iliata
Capparaceae
Ekuya
Leafseeds
Skin
Treatm
ento
fCon
vulsion
wo
undssores
Seedsa
recrushedandthep
oultice
appliedto
thea
ffected
areaTh
emacerated
leaves
after
boiledaretaken
orally
Ageratum
conyzoides
Aste
raceae
Urata
Who
leplant
Skin
Treatm
ento
fwou
ndU
lcer
Sleeping
sickn
essEyew
ash
Thep
lant
ismacerated
andappliedto
affectedskin
area
Aspilia
afric
ana
Aste
raceae
Yunyun
Ka
lank
uwa
Leafflow
erSkin
Treatm
ento
fskinrashes
cleaning
sorescorneal
opacities
Thefl
ower
issqueezed
andappliedto
affectedskin
area
Centau
reaperrottetii
Aste
raceae
Danyi
Who
leplant
Skin
Treatm
ento
fskin
infectionsSyphilis
Thefl
ower
issqueezed
andap
oultice
appliedto
affectedskin
area
Chrysanthellu
mindicum
Aste
raceae
Abilereoyigi
Who
leplant
Skin
Treatm
ento
fboils
gono
rrheaJaun
dice
Thep
lant
iscrushedandthep
oultice
applieddirectlyto
thes
kinarea
Elephantopus
scaber
Aste
raceae
Elephant
foot
Leafroo
tOral
Treatm
ento
ffevercou
ghLeafandroot
areb
oiledtogether
and
takenorally
Emiliacoccinea
Aste
raceae
Odu
ndun
Leafsap
and
root
Skin
Treatm
ento
fulcerhernia
measle
sWho
leplantissqu
eezedandsaptaken
orally
Acalypha
godseffi
ana
Euph
orbiaceae
Jinwinini
Leaf
Skin
Treatm
ento
fskin
infectionsantim
icrobials
Leafismacerated
andtakenorallyas
well
asappliedto
affectedskin
area
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
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PeptidesInternational Journal of
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GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
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Nucleic AcidsJournal of
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Submit your manuscripts atwwwhindawicom
8 The Scientific World Journal
Table1Con
tinued
PlantN
ame
Family
Localn
ame
Partused
Infection
Tradition
aluses
Metho
dof
preparation
Brideliafer
rugin
eaEu
phorbiaceae
Irikirni
Leafstem
bark
root
Oral
Treatm
ento
fInsom
nia
mou
thwashgono
rrhea
Who
leplantisb
oiledandtakenorally
Euphorbiaheterophylla
Euph
orbiaceae
Egele
Leafroo
tsSkin
Treatm
ento
fSkindiseases
Leafandroot
arem
acerated
andthe
poultic
eapp
liedto
thea
ffected
skin
area
Euphorbiahirta
Euph
orbiaceae
Non
onku
rciya
odne
Who
leplant
exud
ate
Oral
Treatm
ento
fAsth
ma
coug
hshapeo
fbreasts
Thew
holeplantism
acerated
andtaken
orally
Euphorbialaterifl
ora
Euph
orbiaceae
Orow
ere
Leafexud
ate
Skin
Treatm
ento
fdermatitis
constip
ation
Theleafissqu
eezedandthep
oultice
appliedto
thea
ffected
skin
area
Cajanu
scajan
Fabaceae
Orela
Leafseed
Skin
andoral
Treatm
ento
fSmallpox
and
mou
thwash
Leafissqueezed
andappliedto
the
affectedarea
Dan
iellia
thurifera
Fabaceae
Iya
Stew
-woo
ddu
stSkin
Treatm
ento
fscabies
Theb
arkismacerated
andappliedto
the
affectedskin
area
Dialiu
mguineense
Fabaceae
Tasm
iyar
kumi
Leafbark
fruitand
twig
Oral
Treatm
ento
fBronchitis
coug
handdiuretic
Theleafand
bark
arec
rushed
andbo
iled
togetherTh
emixture
isthen
takenorally
Acacianilotica
Fabaceae
Baanigabaruwa
Fruitbark
exud
ate
Skin
Treatm
ento
fskindiseases
fung
alinfections
Thee
xudatesa
resqueezed
from
theb
ark
andapplieddirectlyto
thea
ffected
skin
area
Napoleona
imperia
lisLecythidaceae
Mabun
giTw
igs
Oral
Treatm
ento
fAsth
ma
coug
hTw
igsa
remacerated
andtakenorally
Alliu
msativ
umLiliaceae
Ayoayuu
Bulb
Oral
Treatm
ento
ffevercou
gh
asthmaantim
icrobial
Theb
ulbismacerated
andtakenorally
Hibiscus
asper
Malvaceae
Isapad
angiraa
Leaf
Oraland
skin
Treatm
ento
fcou
gh
woun
dsanddiuretic
Leafisbo
iledandtakenorally
Hibiscus
rosasin
ensis
Malvaceae
Kekekeireagu
Leafstem
flowe
rSkin
Treatm
ento
finfl
uenza
woun
dsand
diuretic
Who
leplantism
acerated
andtaken
orally
Theleafissqu
eezedandappliedin
affectedskin
area
Hibiscus
sabdariffa
Malvaceae
Sobo
gurgu
zuLeafflow
erOral
Treatm
ento
fDiuretic
scoug
handdressin
gwo
unds
Leafisbo
iledandtakenorally
Ficuseleg
ans
Moraceae
Asoro
Leaf
Skinoral
Treatm
ento
fPile
constip
ationandcraw
-craw
Leafissqueezed
andappliedto
the
affectedarea
The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
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Zoology
Hindawiwwwhindawicom Volume 2018
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PeptidesInternational Journal of
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GenomicsInternational Journal of
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The Scientific World Journal
Volume 2018
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The Scientific World Journal 9
the absorbance The percentage inhibition of DPPH orABTS radical scavenging activity by the plant extracts wascalculated as (Abscontrol ndash Abssample)(Abscontrol) times 100where Abscontrol is the absorbance of DPPH or ABTS +methanol Abssample is the absorbance of DPPH or ABTSradical + sample extractstandard Here the concentration ofthe extracts needed to decrease the absorbance of DPPH orABTS radical by 50 was calculated Rutin (Sigma-Aldrichge 94 HPLC grade) at the same working concentrations ofthe plant extracts was used as reference drug
210 Bacterial Strains All bacterial strains were culturedon nutrient agar slopes and were incubated for 24 h at37∘C before the MIC An inoculum turbidity equivalent toa 05 McFarland standard (1times107 CFUmL) was prepared innormal saline for each test organism and then diluted 1100in Mueller-Hinton broth just before injection of the platesStandard strains used in the study included S aureus and Ecoli which were a gift from Dr EO Ekundayo (Departmentof Microbiology Michael Okpara University of Agriculture)and clinically isolated strains of K pneumoniae and Pmirabilis were gotten from Department of MicrobiologyFederal Medical Centre (FMC) Umuahia Abia State
211 Antibacterial Susceptibility Using Disc Diffusion MethodThe microbial growth inhibition potential of the plantextracts was determined by using the agar disc diffusionmethod [17] The plant extracts were dissolved in 10 vvDMSO (Merck UK Ltd) to a final concentration of 2 mgmLThe nutrient agar plate was streaked with the standardizedtest bacterial inoculum over the surface evenly and 2mgdiscof the plant extracts transferred onto sterile filter disc papers(about 6 mm) containing the test compounds was placed onthe agar surface in the Petri dishes 100 120583gdisc of standardantibiotic ciprofloxacin was used as control and 100 120583LDMSOdisc served as negative control The plates were thenincubated at 37∘C for 24 h Generally the antimicrobialextracts diffuse into the agar and inhibit the growth of thetest bacteria All tests were done in duplicate and diame-ter zones of inhibition of growth were measured using ameter rule from the edge of each dish after the incubationperiod
212 Minimum Inhibitory Concentration (MIC) Assays Wefollowed the method of Smith et al [18] to determine theMIC ofmethanol crude extracts of selected botanicals againstclinical isolates of bacteria associated with skin and oralinfection A volumeof 100mLof sterileMueller-Hinton broth(Oxoid Basingstoke UK) containing 20 mgL of Ca2+ and10 mgL of Mg2+ was dispensed into a 96-well microtitreplate (Nunc 03 mL total volume per well) All antibacterialagents were dissolved in dimethyl sulphoxide (DMSO) anddiluted in Mueller-Hinton broth to give a stock solutionThen 100 120583L of the antibacterial agent stock solution (2000mgL) was serially diluted into each well and 100 mL ofthe bacterial inoculum added to each well to give a finalconcentration range of 0024 - 1 mgL All procedures wereperformed in duplicate and the plates were incubated for 18
h at 37∘C Briefly 20 mL of a 5 mgL methanol solution of 3-[45-dimethylthiazol-2-yl]-25-diphenyltetrazolium bromide(MTT) (Sigma-Aldrich Ltd South Africa) was added to eachwell and then incubated for 30 min Blue colouration indi-cated bacterial growth The MIC was recorded as the lowestconcentration at which no colour change was observed
213 Statistical Analysis Data analysis was done inMicrosoftExcel to obtain descriptive statistics Means values wereseparated by the Duncan multiple tests using SAS The dif-ferent levels of significance within the groups were analyzedusing one-way analysis of variance (ANOVA) Values wereconsidered significant at P lt005
3 Results and Discussion
31 Ethnobotanical Survey Modern drugs are expensive andsupply to a remote area may be irregular hence result-ing in overreliance of traditional methods of healing as aprimary source of healthcare [19] The rural communitiesof Ebem-Ohafia region of southeastern Nigeria depend onherbal remedies to meet their domestic and health needsDuring our ethnobotanical survey a total of 30 respondentsincluding the elders herbalists and traditional healers wereinterviewed in the study areaMajority of the people (22)wereaged 30ndash65 years dominated by males (25) and one-third(10) had a primary school education while 15 had secondaryeducation and others informal education A total of 61medic-inal plant species belonging to 26 families were identified inthe treatment of oral and skin diseases as shown in Table 1Twenty-seven of them are used in the management of oralinfections and twenty-six plants for the management of skininfection while eight plants are used in the management ofboth infections Table 2 shows the list of family number of theplant species used and their percentage occurrenceThemostrepresented plant family is the Euphorbiaceae having 1803of the plant species followed by Asteraceae and Fabaceae with1147 each [20] Euphorbiaceae is a large family of floweringplants with 300 genera and about 7500 species found usefulin the curing of some common human diseases Asteraceae isa large family of flowering plants having 32 913 species and1911 genera and has been found to possess diverse biologicaleffects Fabaceae is a family of cosmopolitan distributionwith approximately 730 genera and 19400 species richnessat a global level [20] The leaves (60) recorded the mostcommonly used plant part followed by the root (16) andthen stem bark (12) None of the respondents indicated anypossible side effects caused by any of these plants listed inTable 1 The most common methods of preparation includeinfusion decoction and maceration These herbalists ortraditional healers often collect the plants from the fieldand dry and crush them into powder form before storingin bottles to prevent patients from identifying the plantsused for their treatment Most healers refused to provideinformation without payment whereas others thought toprovide information on the plant use is disrespect of theirtraditional belief Furthermore the majority of the plantsmentioned lack scientific data to substantiate their acclaimedfolkloric use From this work it is evident that medicinal
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
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Zoology
Hindawiwwwhindawicom Volume 2018
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PeptidesInternational Journal of
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GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
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Hindawiwwwhindawicom Volume 2018
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Cell BiologyInternational Journal of
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ArchaeaHindawiwwwhindawicom Volume 2018
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Nucleic AcidsJournal of
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Submit your manuscripts atwwwhindawicom
10 The Scientific World Journal
Table 2 The list of family number of the plant species and theirpercentage occurrence
Family Number of plant species OccurrenceFabaceae 7 1147Euphorbiaceae 11 1803Asteraceae 7 1147Myrtaceae 4 656Moraceae 3 492Malvaceae 4 656Liliaceae 2 328Capparaceae 2 328Annonaceae 3 492Combretaceae 1 164Burseraceae 2 328Apocynaceae 1 164Gnetaceae 1 164Lecythidaceae 1 164Verbenaceae 1 164Ochnaceae 1 164Rubiaceae 1 164Piperaceae 1 164Acanthaceae 2 328Papilionoideae 2 328Asclepiadaceae 1 164Caesalpiniaceae 2 328Convolvulaceae 1 164Rutaceae 1 164Crassulaceae 1 164Poaceae 1 164
plants play a significant role in the primary healthcare ofthe people and conservation of the indigenous knowledge ofthis folklore medicine is crucial for the innovations of futuredrug discovery and possible herbal plants trade in the region[19] On the contrary we observed imprecise dosage lowhygiene standards hiding of healing methods and lack ofpatients record as a significant setback to traditional healthpractitioners in the region
32 Thin Layer Chromatography (TLC) Profiling Thin layerchromatography remains the most common useful techniqueby researchers as a quick and simple method to resolvethe constituents of a crude extract followed by advancedextraction methods to purify the active ingredients Previousstudies reported that specific secondary metabolites in plantexhibit a wide range of antioxidant and antibacterial proper-ties [21] TLC was formed on a glass sheet coated with silicagel After the sample is applied to the plate a solvent system(mobile phase) is drawn up via capillary action on the TLCplate at different mobility rate depending on the chemicalcompositionsThe separated spotswith different colourswerevisualized by the UV light projection onto the glass sheetThe variation of coloured spots detected in the crude extractsindicates the presence of different phytochemicals that may
be responsible for the pharmacological effects of selectedplants use in the management of oral and skin infections asacclaimed by the traditional healers We quantified the resultsby measuring the distance travelled by the plant compoundsdivided by the total distance travelled by the mobile phaseFrom Table 3 it is apparent that mobile phase 1 consistingof chloroform and ethanol in the volume ratio of 64 had abetter separation than mobile phase II containing tolueneethyl acetate and acetic acid in the volume ratio of 541 Themethanol extract of AH in bothmobile phases had the highestnumber of three bands with Rf values of 0667 0833 and0989 for mobile phase I and Rf values of 0381 0648 and0781 for mobile phase IIThe plant extracts MM BA CO JCandCAhad two bands inmobile phase I with clear separationbut had single band in mobile phase II The plant extractshaving only one band in either mobile phase suggest a poorsolvent system for compounds separation The TLC profilesprovide a characteristic fingerprint of these plants that mayalso be useful for their identification
33 Antioxidant Assays
331 TPC and TFC Phenolic compounds are secondarymetabolites distributed in fruits vegetables nuts herbsseeds and beverages [22] They are excellent scavengers offree radical that has been implicated in the pathologies ofvarious human diseases the antioxidant activity is dependenton the structure hydrophobic benzenoid rings and hydrogen-bonding potential of the phenolic hydroxyl groups In TPCassay a phenol in plant extracts loses a proton to formphenolate ion which reduces Folin Ciocalteu reagent andthe resulting changes are measured spectrophotometricallyat 765 nm [13] The content of phenol is reported in mg oftannic acid equivalent (TAE)mL of extract As shown inFigure 1 the total phenolic contents of absolute methanolcrude extract of these botanicals range from 700 to 3600mgTAEg The AH extract had the highest concentrationfollowed by CA CO MM BNI BA EI BN and JC whilethe least concentration was recorded in II extract Moreoverthe Folin Ciocalteu reagent (FCR) has been used to measureother compounds such as ascorbic acid amino acids andsugars hence it may not provide a concise amount of thephenolic compounds present in these plants Flavonoids areincreasingly consumed in a significant amount in daily dietsworldwide due to their various health benefits toman [23 24]In TFC assay aluminum ion in the reacting mixture formscomplexes with the C- 4 keto and either C- 3 or C- 5 hydroxylor with ortho hydroxyl groups in the A or B ring of flavonoidsdetected in the plant extracts [14] As shown in Figure 2the total flavonoid content was recorded in the plant extractsranging from 750 to 1500 mgQEg as extrapolated from thestandard quercetin curve The CA extract had the highestconcentration followed by BNI CO II AH BN MM JCand BA while the lowest concentration was recorded in EIextract TPC is found higher than the TFC in some of theextracts supporting the fact that most flavonoids are alsophenolics According to our results all the plant extractsare rich source of phenolic compounds The composition ofTPC and TFC registered in these plants may be difficult to
The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
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The Scientific World Journal 11
Table 3 Thin layer chromatography retention factor using mobile phase CHL ET (mobile phase I) and Tol EA AA (mobile phase II)
SN Plant Extract CHL ET (64) Tol EA AA (541)1 Baphia nitida 1 0742 Macrolobium macrophyllum 0706 0941 06953 Chromolaena odorata 0471 0647 06674 Jatropha curcas 0353 0941 06485 Cassia alata 0118 0882 06866 Ipomoea involucrata 0956 -7 Acalypha hispida 0667 0833 0989 0381 0648 07818 Breynia nivosa 0989 07629 Burkea africana 0644 1 075210 Eleusine indica 0978 0743CHL chloroform ET ethanol Tol toluene EA ethylacetate AA acetic acid
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
500
1000
1500
2000
2500
3000
3500
4000
TPC
(mgT
AE
g)
Figure 1 The total phenolic content (TPC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata TAE tannic acid equivalent
compare with others reported elsewhere due to many factorsincluding differences from species to species maturity stageplant parts and assay methods harvesting time standarddrugs and geographical location
332 Reducing Power (RP) Ferric reducing power is a globalmethod often used to assess the capacity of antioxidantsto donate either an electron or hydrogen atom to unpairedelectron [25] In reducing power assay plant extracts exertantioxidant activities via reduction of Fe3+ferric cyanidecomplex to Fe2+ (ferrous form) resulting in changing thegreen colour to blue and sometimes Perlrsquos Prussian bluecolour depending on the antioxidant potential [10] As shownin Figure 3 the reducing power of the methanol crudeextracts and the standard gallic acid rutin and ascorbic acidincreased with increase in concentration Higher absorbanceat 700 nm can be read after potassium ferricyanide reactionwith ferric chloride At 1000 120583gmL the extracts from AH
0
200
400
600
800
1000
1200
1400
1600
TFC
(mgQ
Eg)
AH BN JC CO MM EI BNI BA CAIIPlant extracts
Figure 2 The total flavonoid content (TFC) of selected plants usedin the management of oral and skin infections in southeast NigeriaII Ipomoea involucrata AH Acalypha hispida BN Breynia nivosaJC Jatropha curcas CO Chromolaena odorata MM Macrolobiummacrophyllum EI Eleusine indica BNI Baphia nitida BA Burkeaafricana CA Cassia alata QE Quercetin equivalent
BNI MM CA BA EI and BN had absorbance value of 270086 073 055 036 027 and 027 respectively while that ofAH is higher than the standard rutin (254) but comparablewell with gallic acid (275) and ascorbic acid (269) EI andBN extracts had a similar trend of ferric reducing powerthough significantly lower as compared with the standarddrugs AH extract exhibits remarkable antioxidant capacity(RP) correlating with its richness in phenolic and flavonoidcontents It is reasonable to suggest that methanol selectivelyextracts antioxidant principles from these botanicals basedon their polarities and chemical structures which are reflectedin plant extracts with high TFC and TPC having minimal RP[26] We have shown that some plant extracts in particularAH BNI MM and CA extracts may serve as an excellentcandidate to minimize skin and oral infections caused byoxidative stress as a result of imbalance of free radicalsgeneration and natural antioxidants in the saliva or woundarea According to our results the increasing RP of the plantextracts correlateswith their TPCandTFCThis study at least
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
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Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
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Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
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Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
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Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
12 The Scientific World JournalRe
duci
ng P
ower
7
00nm
625 125 250 500 10003125Concentration (gmL)
0
05
1
15
2
25
3
IIAHBNJCCOMMEI
BNIBACAGARutinVit C
Figure 3 The ferric reducing antioxidative capacity of selectedplant extracts used in the management of oral and skin infectionsin southeast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata GA gallic acid Vit C vitamin C (standard antioxidants)
in part lends credence to the ethno-therapeutic use of someplant as natural antioxidant in themanagement of skin or oralinfections in the study area
333 FRSA of Plant Extracts Skin and oral cavity exposedto an array of oxidants such as smokes from cigaretteschemicals UV light nicotine and alcohol that are directlyor indirectly leading to the production of reactive oxygenspecies (ROS) are known to be responsible for some com-mon human diseases [27] These free radicals in particularhydroxyl and superoxide anions are highly reactivemoleculesderived from the metabolism of oxygen Although they playan important role in the biochemical immunological andphysiological processes it could be deleterious to vital organsif excessively accumulated in the human body [28] Food andpharmaceutical industries discovered that medicinal plantscontaining phenol and flavonoid compounds are essentialin the development of novel drugs against stress-induceddiseases [28] In the present investigation the concentration(IC50) at which 50 of free radical scavenged was calculatedand extrapolated from the plot of percentage inhibition ver-sus plant extract concentration The herbal drugs exhibitedrobust DPPH radical scavenging activity in a concentration-dependent manner having IC50 ranging from 2025 to 710120583gmL As shown in Figure 4 MM BA EI JC AH BNIand CA extracts had a potent DPPH radical scavengingactivity corresponding to their TPC TFC and RP Theplant extracts scavenged ABTS radical by the decrease inits absorbance at 734 nm All the plant extracts exhibitedstrong ABTS radical scavenging capacities in the following
AH BN JC CO MM EI BNI BA CAIIPlant extracts
0
200
400
600
800
1000
1200
1400
1600
IC50
(g
mL)
Figure 4 DPPH free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
050
100150200250300350400450500
IC50
(g
mL)
AH BN JC CO MM EI BNI BA CAIIPlant samples
Figure 5 ABTS free radical scavenging activity of selected plantextracts used in the management of oral and skin infections insoutheast folklore medicine of Nigeria II Ipomoea involucrataAH Acalypha hispida BN Breynia nivosa JC Jatropha curcasCO Chromolaena odorata MM Macrolobium macrophyllum EIEleusine indica BNI Baphia nitida BA Burkea africana CACassiaalata
order BN gt JC gt CA gt MM gt BNI gt EI gt CO gt AH gtII gt BA having IC50 values ranging from 25 to 400 120583gmL(Figure 5) DPPH and ABTS radical scavenging activities ofthese herbal remedies displayed a positive correlation withthe working concentrations It is obvious that most of theplant extracts tested in this work possess the ability to quenchthe free radicals generation via electron or proton donationdemonstrated by the termination of radical chain reaction[28] Although both DPPH and ABTS radicals are electron-transfer based methods however it is apparent that ABTSmethod seems to be more sensitive than DPPH radical assaypossibly due to their solubility and diffusivity as shown inthe IC50 values [10] It is worth noting that BA extract withstrong DPPH radical scavenging effect had a moderate ABTS
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
Virolog y Stem Cells International
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Enzyme Research
Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
The Scientific World Journal 13
Table 4 Zone of bacterial inhibition by the selected plant extracts use in the management of oral and skin infections
Diameter of inhibition (mm)SN Plant samples K pneumoniae P mirabilis S aureus E coli1 Cassia alata 15 plusmn 141 9 plusmn 042 15 plusmn 083 13 plusmn 0802 Breynia nivosa 8 plusmn 052 17 plusmn 100 8 plusmn 008 15 plusmn 1023 Burkea africana NI 11 plusmn 084 13 plusmn 072 NI4 Baphia nitida 8 plusmn 050 13 plusmn 122 8 plusmn 051 NI5 Ipomoea involucrata 8 plusmn 052 NI 8 plusmn 050 13 plusmn 0526 Jatropha curcas NI 9 plusmn 030 9 plusmn 062 13 plusmn 0727 Acalypha hispida 21 plusmn 163 NI 11 plusmn 009 17 plusmn 1328 Chromolaena odorata 8 plusmn 042 8 plusmn 035 9 plusmn 009 15 plusmn1209 Eleusine indica 8 plusmn 042 8 plusmn 035 NI 17 plusmn 10010 Macrolobium macrophyllum NI NI 8 plusmn 004 15 plusmn 12311 Ciprofloxacin - - le16(R) ge17(S) le16(R)ge20(S)NI indicates no zone of inhibition S sensitive R resistance
radical scavenging capacity which can be explained by thepartial ionization of the extract in the test medium [29] SinceROS promotes tissue damage and epithelial inflammationvia exposure to sunlight radiation or smoke as a result theantioxidant capacity demonstrated in this work may supportthe traditional use of these plants against the influence ofinflammatory markers and oxidants in the skin and oralcavity [29] Based on the results of the three antioxidantassays it is obvious that MM AH BNI EI and CA extractshad the best FRSA indicating their importance in convertingfree radicals into more stable products and then terminatethe free radical chain reaction Our study showed that themethanol crude extracts from these plants could protectagainst free radical damage in the mouth such as humangingival periodontal tissues and certain skin infections [5]
334 Antibacterial Activity of Plant Extracts Pathogenicbacteria such as Staphylococcus aureus Escherichia coli Kleb-siella pneumoniae and Proteus mirabilis play a significantrole as the primary cause of skin and oral infections [30]Staphylococcus aureus is a part of normal skin known to bethe most problematic in the cause of skin infections as aresult of their resistance mechanisms to first-line antibiotics[30] Although the presence of this organism as a residentoral floral is controversial nonetheless MSSA and MRSAhave been isolated from oral rinse and tongue swab inpatients with oral infections [31] Escherichia coli are Gram-negative bacteria known to be notorious for their resistantmechanisms towards the existing arsenal of antibiotics Oralinfections such as edematous and erythematous and skininfections like omphalitis surgical site infections burnnecrotizing fasciitis and cellulitis at all age groups are associ-ated with enterobacterium E coli [32] Klebsiella pneumoniaeis a Gram-negative encapsulated facultative anaerobic androd-shaped bacterium found in the normal flora of themouth skin and intestines This bacterium is responsiblefor the prolonged surgical wound biliary tract wound andurinary tract infections and it could pose a serious healthrisk when resistant strains invade the throat mouth or
intestines [33] Villegas et al [34] detected a CTX-M-12 120573-lactamase producing K pneumoniae new plasmid-mediatedESBLs highly resistant to ceftazidime from a patient withhospital-acquired wound infection Proteus mirabilis is oneof the most common enterobacteria after E coli identified asGram-negativemotile and rod-shaped bacterium It is foundin clinical specimen implicated in the skin respiratory tractand bacteraemia as well as burns wounds and urinary tractinfections [35] All these pathogenic multidrug-resistancebacteria are causative agent in a periodontal lips ulcer whitesponge nevus and skin infections [36]
335 Susceptibility Test and MIC Determination The resultof disc diffusion assay showed the diameter of inhibitionof bacteria growth classified as le 15 (resistant) and ge 16(susceptible) according to CLSI guideline [37] As shown inTable 4 K pneumoniae and E coli are more susceptible tothe treatment of Acalypha hispida (AH) having a zone ofinhibition (ZI) 21 and 17mm respectively BN and EI extractsinhibit the growth of P mirabilis and E coli having the samezone of inhibition 17 mm corresponding to the report of Al-Zubairi et al [38] The data obtained for the methanol crudeextracts from AH BN and EI show their selective inhibitionof the test bacteria growth The plant extracts with ZI above13 mm against our panel of bacterial strains were selected forMICdeterminationHere themethanol crude extracts exhibitMIC values of 256 to gt 512 120583gmL against the test bacterialstrains but were less active as compared to the standardciprofloxacin 003 - 006 120583gmL (Table 5) Ciprofloxacin isan active broad-spectrum antibiotic use in the treatmentof urinary tract infections prostatitis and acute cystitis viainterfering with bacterial DNA replication and transcriptionthrough inhibition of DNA gyrase or topoisomerase [39] Allthe plant extracts demonstrated a mild to weak antibacterialactivity based on the classification of Rios and Recio [40] whoconsidered antibacterial activity of the crude plant extractsignificant (MIC lt 100 120583gmL) moderate (100 lt MIC = 512120583gmL) or weak (MIC gt 512 120583gmL) The herbal remediesshowed moderate inhibition of bacterial growth except for K
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
Virolog y Stem Cells International
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Enzyme Research
Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
14 The Scientific World Journal
Table 5 Minimum inhibitory concentration (MIC) of selectedmedicinal plants against bacterial strains associated with oral andskin infections
Plants SA KP PM ECMIC (120583gmL)
Acalypha hispida 512 256 gt512 512Cassia alata 512 256 256 256Breynia nivosa 256 gt512 256 512Eleusine indica 256 512 256 512Chromolaena odorata 512 256 256 256Ciprofloxacin 003 003 003 003SA S aureus KP K pneumoniae PM P mirabilis EC E coli
pneumoniae and P mirabilis towards BN and AH extractsrespectively BN and EI extracts were more active againstS aureus and P mirabilis CA and CO extracts were moreactive against all the test bacterial strains except S aureusPrevious work reported by Amadi et al [41] confirmed theweak antibacterial activity exhibited by BN extract againstK pneumoniae The antibacterial activity demonstrated byAH extract against our panel of bacterial strains contradictsthat of Aboaba and Yeye [42] who reported MIC value of100 mgmL against E coli S aureus and K pneumoniaeIt is interesting to note that K pneumoniae E coli and Pmirabilis that confer resistance to the extended spectrumcephalosporin and penicillin [43] are more susceptible tothe treatment with CA and CO extracts Furthermore theobserved antibacterial activity exhibited by EI CA and COextracts against the superbugs of modern day healthcare [38]is worth noting whichmay perhaps attribute to a wide varietyof secondary metabolites especially phenolics and flavonoidsthat have been reported with potent antimicrobial properties[44] Among the three extracts Eleusine indica has IC50 gt30 120583g mL towards MCF-7 HT-29 and CEM-SS humancancer cell lines suggesting that continued exposure to thisherbal remedy may alter human organs [38] The probablemechanism of action of these plants is not known but it couldbe attributed to the leakage of the cell wall or alteration in themembrane permeability or inhibition of enzyme gyraseDNAsynthesis and protein synthesis [45] Further researches areneeded to identify the active ingredients curare effect andthe molecular mechanisms of these plants
4 Conclusion
The results of this study therefore at least in part providescientific evidence for the traditional use of these medicinalplants in ethnomedicine by the dwellers of Ebem-Ohafiacommunities in the treatment of skin and oral infections Thefree radicals scavenging activity moderate to high TPC TFCand themild antibacterial properties demonstrated by certainplant extracts are worth harnessing for proper evaluation andintegration into the primary healthcare system Further frac-tionation to identify the single compound responsible for theobserved activities is highly recommended EncouraginglyBN CA CO and AH extracts with potent antioxidant andantibacterial properties did not exhibit any significant toxic
effect as reported by other researchers and thus substantiateits safe use as complementary and alternative therapy for thetreatment or prevention of human diseases
Data Availability
The data used to support the findings of this study areavailable from the corresponding author upon request
Conflicts of Interest
Nopotential conflicts of interestwere reported by the authors
Acknowledgments
The authors thank the National Research Foundation (NRF)South Africa Agency for Science and Technology Advance-ment for their financial support
References
[1] WorldHealth Organization ldquoOral healthrdquo Fact sheets 318 2012httpwwwwhointmediacentrefactsheetsfs318enindexhtml
[2] F E Dewhirst T Chen J Izard et al ldquoThe human oralmicrobiomerdquo Journal of Bacteriology vol 192 no 19 pp 5002ndash5017 2010
[3] M L Falsetta M I Klein P M Colonne et al ldquoSymbiotic rela-tionship between Streptococcus mutans and Candida albicanssynergizes virulence of plaque biofilms in vivordquo Infection andImmunity vol 82 no 5 pp 1968ndash1981 2014
[4] A Shinde J Ganu and P Naik ldquoEffect of Free Radicals ampAntioxidants on Oxidative Stress A Reviewrdquo Journal of Dentaland Allied Sciences vol 1 no 2 p 63 2012
[5] O Hershkovich I Shafat and R M Nagler ldquoAge-relatedchanges in salivary antioxidant profile possible implications fororal cancerrdquo Journals of Gerontology Series A Biological Sciencesand Medical Sciences vol 62 no 4 pp 361ndash366 2007
[6] S Jagtap S Yavankar K Pardesi and B Chopade ldquoProductionof bioemulsifier by Acinetobactersp from healthy human skinof tribal populationrdquo Indian Journal of Experimental Biologyvol 48 pp 70ndash76 2010
[7] D H Limoli A B Rockel K M Host et al ldquoCationicAntimicrobial Peptides Promote Microbial Mutagenesis andPathoadaptation inChronic InfectionsrdquoPLoS Pathogens vol 10no 4 Article ID e1004083 2014
[8] A Al-Ahmad H Ameen K Pelz et al ldquoAntibiotic resistanceand capacity for biofilm formation of different bacteria isolatedfrom endodontic infections associated with root-filled teethrdquoJournal of Endodontics vol 40 no 2 pp 223ndash230 2014
[9] F A Badria andO A Zidan ldquoNatural products for dental cariespreventionrdquo Journal of Medicinal Food vol 7 no 3 pp 381ndash3842004
[10] S O Oyedemi B O Oyedemi I I Ijeh P E Ohanyerem R MCoopoosamy and O A Aiyegoro ldquoAlpha-amylase inhibitionand antioxidative capacity of some antidiabetic plants used bythe traditional healers in Southeastern Nigeriardquo The ScientificWorld Journal vol 2017 Article ID 3592491 11 pages 2017
[11] T Vogt ldquoPhenylpropanoid biosynthesisrdquo Molecular Plant vol3 no 1 pp 2ndash20 2010
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
Virolog y Stem Cells International
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Enzyme Research
Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
The Scientific World Journal 15
[12] H Ghaffari B J Ghassam S C Nayaka K Ramachandra Kiniand H S Prakash ldquoAntioxidant and neuroprotective activitiesof Hyptis suaveolens (L) Poit against oxidative stress-inducedneurotoxicityrdquoCellular andMolecularNeurobiology vol 34 no3 pp 323ndash331 2014
[13] C Chang M Yang H Wen and J Chern ldquoEstimation of totalflavonoid content in propolis by two complementary colometricmethodsrdquo Journal of Food and Drug Analysis vol 10 no 3 pp178ndash182 2002
[14] G C Yen and H Y Chen ldquoAntioxidant activity of varioustea extracts in relation to their antimutagenicityrdquo Journal ofAgricultural and Food Chemistry vol 43 no 1 pp 27ndash32 1995
[15] A Tariq M Athar J Ara V Sultana S Ehteshamul-HaqueandM Ahmad ldquoBiochemical evaluation of antioxidant activityin extracts and polysaccharide fractions of seaweedsrdquo GlobalJournal of Environmental Science Management vol 1 no 1 pp47ndash62 2015
[16] R Re N Pellegrini A ProteggenteA Pannala M Yang andCRice-Evans ldquoAntioxidant activity applying an improved ABTSradical cation decolorization assayrdquo Free Radical Biology ampMedicine vol 26 no 9-10 pp 1231ndash1237 1999
[17] National Committee for Clinical Laboratory Standards ldquoPer-formance Standards for antimicrobial susceptibility testingrdquo 8thInformational Supplement M100 S12 National Committee forClinical Laboratory Standards Villanova Pa USA 2002
[18] E E Smith E Williamson M Zloh and S Gibbons ldquoIsopi-maric acid from Pinus nigra shows activity against multidrug-resistant and EMRSA strains of Staphylococcus aureusrdquo Phy-totherapy Research vol 19 no 6 pp 538ndash542 2005
[19] N R Farnsworth ldquoEthnopharmacology and drug develop-mentrdquo Ciba Foundation Symposium vol 185 pp 42ndash59 2007
[20] W S Judd C S Campbell E A Kellogg P F Stevens andM JDonoghue Plant Systematics A Phylogenetic Approach SinauerAssociates Sunderland Mass USA 2002
[21] M Wink Functions of Plant Secondary Metabolites and theirExploitation in Biotechnology vol 39 of Annual Plant ReviewsWiley-Blackwell London UK 2010
[22] V Cheynier ldquoPolyphenols in foods aremore complex than oftenthoughtrdquo The American Journal of Clinical Nutrition vol 81supplement 1 pp 223Sndash229S 2005
[23] P C H Hollman and M B Katan ldquoDietary flavonoidsIntake health effects and bioavailabilityrdquo Food and ChemicalToxicology vol 37 no 9-10 pp 937ndash942 1999
[24] G P Hubbard S Wolffram J A Lovegrove and J M GibbinsldquoIngestion of quercetin inhibits platelet aggregation and essen-tial components of the collagen-stimulated platelet activationpathway in humansrdquo Journal of Thrombosis and Haemostasisvol 2 no 12 pp 2138ndash2145 2004
[25] M-C Kou S-Y Chiou C-Y Weng L Wang C-T Hoand M-J Wu ldquoCurcuminoids distinctly exhibit antioxidantactivities and regulate expression of scavenger receptors andheme oxygenase-1rdquo Molecular Nutrition amp Food Research vol57 no 9 pp 1598ndash1610 2013
[26] M-Y Juan and C-C Chou ldquoEnhancement of antioxidantactivity total phenolic and flavonoid content of black soybeansby solid state fermentation with Bacillus subtilis BCRC 14715rdquoFood Microbiology vol 27 no 5 pp 586ndash591 2010
[27] S O Oyedemi and A J Afolayan ldquoIn vitro and in vivo antioxi-dant activity of aqueous leaves extract of Leonotis leonurus (L)R Brrdquo International Journal of Pharmacology vol 7 no 2 pp248ndash256 2011
[28] A Temraz and W H El-Tantawy ldquoCharacterization of antiox-idant activity of extract from Artemisia vulgarisrdquo PakistanJournal of Pharmaceutical Sciences vol 21 no 4 pp 321ndash3262008
[29] I Fridovich ldquoFundamental aspects of reactive oxygen speciesor whatrsquos the matter with oxygenrdquo Annals of the New YorkAcademy of Sciences vol 893 no 22 pp 13ndash18 1999
[30] D J Wilson R N Gonzalez and H H Das ldquoBovine mastitispathogens in New York and Pennsylvania prevalence andeffects on somatic cell count and milk productionrdquo Journal ofDairy Science vol 80 no 10 pp 2592ndash2598 1997
[31] A J Smith M S Jackson and J Bagg ldquoThe ecology ofstaphylococci in the oral cavity a reviewrdquo Journal of MedicalMicrobiology vol 50 no 11 pp 940ndash946 2001
[32] G J Moet R N Jones D J Biedenbach M G Stilwelland T R Fritsche ldquoContemporary causes of skin and softtissue infections in North America Latin America and Europereport from the SENTRY antimicrobial surveillance program(1998ndash2004)rdquo Diagnostic Microbiological Infectious Diseasesvol 57 no 1 pp 7ndash13 2007
[33] K J Ryan and C G Ray Sharris Medicinal MicrobiologyMcGraw Hill New York NY USA 4th edition 2004
[34] MVVillegasACorrea F Perez et al ldquoCTX-M-12120573-lactamasein aKlebsiella pneumoniae clinical isolate in Colombiardquo Antimi-crobial Agents and Chemotherapy vol 48 no 2 pp 629ndash6312004
[35] R Tibbetts J G Frye J Marschall D Warren and W DunneldquoDetection of KPC-2 in a clinical isolate of Proteus mirabilisand first reported description of carbapenemase resistancecaused by a KPC 120573-lactamase in P mirabilisrdquo Journal of ClinicalMicrobiology vol 46 no 9 pp 3080ndash3083 2008
[36] S M JacobsenD J Stickler H L T Mobley andM E ShirtliffldquoComplicated catheter-associated urinary tract infections dueto Escherichia coli and Proteus mirabilisrdquo Clinical MicrobiologyReviews vol 21 no 1 pp 26ndash59 2008
[37] Clinical and Laboratory Standards Institute ldquoMethods fordilution antimicrobial susceptibility tests for bacteria that growaerobically approved standardrdquo CLSI document M7-A7 Clini-cal and Laboratory Standards Institute Wayne Pa USA 2006
[38] A S Al-Zubairi A B Abdul S I Abdelwahab C Y Peng SMohan andMM Elhassan ldquoEleucine indica possesses antiox-idant antibacterial and cytotoxic propertiesrdquo Evidence-BasedComplementary and Alternative Medicine vol 2011 Article ID965370 6 pages 2011
[39] Y A Al-Soud and N A Al-Masoudi ldquoA new class of dihalo-quinolones bearingN1015840- aldehydoglycosylhydrazidesmercapto-124-triazole oxadiazoline and 120572-amino ester precursors syn-thesis and antimicrobial activityrdquo Journal of the BrazilianChem-ical Society vol 14 no 5 pp 790ndash796 2003
[40] J L Rios and M C Recio ldquoMedicinal plants and antimicrobialactivityrdquo Journal of Ethnopharmacology vol 100 no 2 pp 80ndash84 2005
[41] E S Amadi C A Oyeka R A Onyeagba O C Ugboguand I Okoli ldquoAntimicrobial screening of Breynia nivosus andAgeratum conyzoides against dental caries organismsrdquo Journalof Biological Sciences vol 7 no 2 pp 354ndash358 2007
[42] S A Aboaba and E Yeye ldquoStudies on phytochemical screeningantimicrobial and toxicity effect of the shoot system ofAcalyphasegetalis Muell Argrdquo African Journal of Pure and AppliedChemistry vol 8 no 12 pp 191ndash195 2014
[43] A Wahab S Begum A Ayub et al ldquoLuteolin and kaempferolfrom Cassia alata antimicrobial and antioxidant activity of its
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
Virolog y Stem Cells International
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Enzyme Research
Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
16 The Scientific World Journal
methanolic extractsrdquo FUUAST Journal of Biology vol 4 no 1pp 1ndash5 2014
[44] K Lewis and F M Ausubel ldquoProspects for plant-derivedantibacterialsrdquo Nature Biotechnology vol 24 no 12 pp 1504ndash1507 2006
[45] G Kapoor S Saigal and A Elongavan ldquoAction and resistancemechanisms of antibiotics A guide for cliniciansrdquo Journal ofAnaesthesiology Clinical Pharmacology vol 33 no 3 pp 300ndash305 2017
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
Virolog y Stem Cells International
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Enzyme Research
Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
Hindawiwwwhindawicom
International Journal of
Volume 2018
Zoology
Hindawiwwwhindawicom Volume 2018
Anatomy Research International
PeptidesInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Journal of Parasitology Research
GenomicsInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom
The Scientific World Journal
Volume 2018
Hindawiwwwhindawicom Volume 2018
BioinformaticsAdvances in
Marine BiologyJournal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Neuroscience Journal
Hindawiwwwhindawicom Volume 2018
BioMed Research International
Cell BiologyInternational Journal of
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Biochemistry Research International
ArchaeaHindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Genetics Research International
Hindawiwwwhindawicom Volume 2018
Advances in
Virolog y Stem Cells International
Hindawiwwwhindawicom Volume 2018
Hindawiwwwhindawicom Volume 2018
Enzyme Research
Hindawiwwwhindawicom Volume 2018
International Journal of
MicrobiologyHindawiwwwhindawicom
Nucleic AcidsJournal of
Volume 2018
Submit your manuscripts atwwwhindawicom
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