Phytochemical,Antimicrobial,andAntioxidantProfilesof ...

Research ArticlePhytochemical Antimicrobial and Antioxidant Profiles ofDuranta erecta L Parts

Shadrack Donkor 12 Christopher Larbie 2 Gustav Komlaga 3

and Benjamin Obukowho Emikpe45

1Applied Radiation Biology Centre Radiological and Medical Sciences Research Institute Ghana Atomic Energy CommissionLegon Accra Ghana2Department of Biochemistry and Biotechnology Kwame Nkrumah University of Science and Technology (KNUST)Kumasi Ghana3Department of Pharmacognosy Kwame Nkrumah University of Science and Technology (KNUST) Kumasi Ghana4Department of Veterinary Pathology University of Ibadan Ibadan Nigeria5Department of Pathobiology School of Veterinary Medicine Kwame Nkrumah University of Science and Technology (KNUST)Kumasi Ghana

Correspondence should be addressed to Shadrack Donkor shadrachdonkor2000gmailcom

Received 18 February 2019 Revised 8 August 2019 Accepted 19 September 2019 Published 24 October 2019

Academic Editor Ziad Fajloun

Copyright copy 2019 Shadrack Donkor et al(is is an open access article distributed under the Creative CommonsAttribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

(e use of plant-based medicine is popular amongst individuals and communities in developing countries Duranta erecta hasbeen used in Africa and Asia to treat a wide range of diseases (is study evaluated the phytochemical profile and antioxidant andantimicrobial activities of D erecta to ascertain its health benefits in traditional medicine Phytochemical constituents andantimicrobial effect of the hydroethanolic extract of D erecta leaves (DRL) unripe fruits (DRU) and ripe fruits (DRR) wereinvestigated by standard methods Elemental analyses were carried out by atomic absorption spectroscopy (AAS) on the rawsample and extract FTIR and UV-VIS spectroscopy were used to identify functional groups Extracts were screened for theirpossible antioxidant activities by three tests (e total phenolic and total tannin contents were evaluated by using the FolinndashCiocalteu method Total flavonoid content was determined by the aluminium chloride colorimetric assay method(e antioxidantactivities were evaluated using the DPPH scavenging activity (e results of phytochemical screening showed the presence oftriterpenoids sterols alkaloids flavonoids saponins glycosides and tannins FTIR analysis revealed the presence of alcoholsphenols alkanes aldehydes ketones aromatics aliphatic amines aromatic amines amides carboxylic acids esters nitrocompounds alkynes primary and secondary amines and alkyl halides Iron zinc and copper were also detected Total phenolicand tannin contents ranged from 220plusmn 015 to 1454plusmn 029mg gallic acid equivalent (GAE)100 g and 355plusmn 007 to1382plusmn 004mg GAE100 g respectively Total flavonoid content varied from 4176plusmn 096 to 34349plusmn 345 μg quercetin equivalent(QE)100 g (e highest DPPH scavenging activity was recorded in the methanolic fraction of the leaves (e antimicrobial assayof the extract or fractions recorded no activity against the test organisms (e outcome of this study affirmed that D erectacontains phytochemicals and bioactive compounds that could be of health benefit

1 Introduction

(e use of medicinal plants has become popular amongstindividuals and communities in developing countries (erehas been an upsurge in the use of herbal medicines becausethey are affordable accessible culturally acceptable andeffective and supposedly have lesser side effects as compared

with synthetic drugs found in Africa [1] Plants producevarious bioactive compounds which contain components oftherapeutic value making them a rich source of medicinesApproximately 25 of prescription drugs dispensed in theUnited States are derived from plant sources [2](e variouscomponents confer specific distinctiveness and properties toplants

HindawiBiochemistry Research InternationalVolume 2019 Article ID 8731595 11 pageshttpsdoiorg10115520198731595

In folk medicine plants are important source of naturalremedies used to treat many diseases Overdependence ontraditional folk medicine has brought to light the issue ofsustaining our fast-degrading forest In Ghana most of thetraditional medicine practitioners depend on our forests fortheir trade while our backyards are overgrown with orna-mental plants which are equally good source of herbalmedicine (us if we want to conserve our forest for sus-tainable development then there is the need to screen or-namental plants for biological and pharmacological propertiesby researchers(is is part of studies identifying alternative usefor ornamental plant [3]

Durantabelongs to the Verbenaceae family and itcomprises 35 species It is a native plant of Asia Africa andSouth and Central America [4] Duranta erecta (synony-mous to D repens) commonly known as ldquogolden dewdroprdquo is an upright scrambling shrub with a height of 1ndash3m It is grown as a hedge or ornamental plants in varioushomes in Ghana Phytoconstituents including coumar-inolignoids (E)-cinnamic acid (E)-p-methoxycinnamicacid and lamiide have been isolated from the genusDuranta [5 6] A number of bioactive compounds such asβ-sitosterol naringenin acteoside lamiide sucrose andraffinose have been identified and isolated from Durantarepens [7] Different parts of the plant are used to treatvariety of diseases (e fruit and leaves are used in tradi-tional folk medicine for the treatment of malaria intestinalworms and abscess and in some cases serve as vermifugeor diuretic D erecta is said to possess antitumor activityand significant antibacterial activity [8] It has numerousnatural uses including antifungal and insecticidal properties[9] (e ethyl acetate extract of leaves reportedly exhibitedsignificant antiplasmodial activity against the chloroquine-sensitive and chloroquine-resistant strains of Plasmodiumfalciparum [5] Methanolic extracts of different parts such asleaves stem and roots of D erecta exhibited antifungalproperties against Aspergillus flavus Alternaria sp Peni-cillium sp Rhizopus sp and Trichoderma sp [10] (e aimof the current study was to carry out the phytochemical andantioxidant screening ofD erecta grown as hedges in Ghanaas a justification for its use in traditional medicine

2 Materials and Methods

21 Chemicals FolinmdashCiocalteu phenol reagent (FCR)gallic acid 2 2-diphenylminus 1-picrylhydrazyl hydrate (DPPH)quercetin ascorbic acid and DMSO were from SigmaAlCl3 Na2CO3 CH3COOK HClO4 CHCl3 NaOH FeCl3and NaCl were fromMerck (Darmstadt Germany) Ethanolmethanol petroleum ether ethyl acetate nitric acid andsulphuric acid were from Janssen Chimica (Beerse Bel-gium) Nutrient agar Mueller-Hinton agar and chloram-phenicol were procured from Oxoid England All chemicalswere of analytical grade

22 Plant Collection and Authentication Plant materials(leaves ripe and unripe fruits) were collected from KNUSTcampus in October 2017 and authenticated by Dr George

Sam at the Department of Herbal Medicine Faculty ofPharmaceutical Sciences KNUST and the specimen wasdeposited in the facultyrsquos herbarium (voucher numberKNUSTHM12017L011)

23 Preparation and Extraction of PlantMaterial (e leavesand fruits of D erecta were washed under running tap waterto remove dust and other extraneous substances (ey weresubsequently dried under shade for 4 weeks(e shade-driedparts of the plant were coarsely powdered using a grinder(Waring USA) 60 g of the pulverized samples wereextracted with 50 ethanol by cold maceration for 48 hoursat room temperature on a shaker (Rocking LaboratoryShaker) (e mixture was separated by centrifugation andeach supernatant was evaporated under reduced pressureusing a rotary evaporator (Buchi R-205 Switzerland) (eextracts were dried using a vacuum freeze dryer (HetoPowerDry LL3000 UK) to obtain the respective crude ex-tracts which included D erecta leaves and unripe and ripefruits 15 g of each crude extract was sequentially extractedwith solvents of increasing polarity starting with petroleumether followed by ethyl acetate and methanol (e residualportion was designated as hydro fraction (e variousfractions namely petroleum ether ethyl acetate methanoland hydro fractions were all air-dried at room temperatureand used for the tests

24 Phytochemical Screening (e powdered samples of Derecta leaves unripe and ripe fruits and respective crudeextracts were subjected to qualitative analysis for identifi-cation of tannins (Gelatin test) glycosides (Molischrsquos test)alkaloids (Dragendorffrsquos reagents) coumarins (FeCl3 test)saponins (foam test) flavonoids (alkaline reagent) tri-terpenoids (Salkowskirsquos test) and sterols (LiebermannndashBurchard test) according to the standard methods describedby Harborne [11] and Sazada et al [12]

25 Heavy Metal Analysis Prior to sample analysis theglassware used for the analysis was soaked overnight in 10(vv) nitric acid followed by washing with 10 (vv)hydrochloric acid then rinsed with double-distilled waterand dried to eliminate interference Accurately 1 g of each ofthe extracts and the raw samples were weighed into a 50mLdigestion tube Each sample was wet-digested using amixture of 1mL of H2O 2mL of HCl 5mL of 1 1 HNO3 HClO4 and 2mL of H2SO4 for 20 minutes (emixture washeated in a digestion block at 150degC(e digested sample wasallowed to cool and diluted with 50mL of distilled water(edigests were analysed to determine the levels of heavy metalsin each extract Heavy metals including lead copper nickelzinc and iron were analysed using an atomic absorptionspectrometer (Varian AA 240FS) with a long path airacetylene burner and cathode lamp for respective metals

26 UV and FTIR Spectroscopic Analysis (e fractions ofleaves and fruits were diluted to 1 10 with respective sol-vents (e fractions were scanned in the wavelength ranging

2 Biochemistry Research International

from 200 to 700 nm using a double-beam ultraviolet spec-trophotometer (PerkinElmer USA) to detect the charac-teristic peaks 10mg of the dried extracts was encapsulated in100mg of KBr pellet to prepare translucent sample discs(epowdered sample of each extract was loaded in a FTIRspectroscope (UATR Spectrum 2 PerkinElmer) with a scanrange from 400 to 4000 cmminus 1 and a resolution of 4 cmminus 1 (epeak values of the UV and FTIR were recorded

27 Determination of Total Phenolic Content (e totalphenolic content of the fractions was assessed by FolinndashCiocalteu (FC) procedure [13] with some modificationsGallic acid was used as the standard Approximately 50 μL ofthe extract was mixed with 3mL of distilled water and 250 μLof FC reagent (e mixture was allowed to stand for5minutes and then 750 μL of 20 Na2CO3 was added (eresulting mixture was vigorously vortexed for 2min and thenincubated for 30min at room temperature (e absorbanceof the solution was measured at 760 nm using a UV-VISspectrophotometer ( UV1201 Shimadzu Japan) All de-terminations were performed in triplicate Gallic acid(02mgmiddotmLminus 1 04mgmiddotmLminus 1 06mgmiddotmLminus 1 08mgmiddotmLminus 1 and1mgmiddotmLminus 1) was used as standard to prepare a calibrationcurve from which polyphenolic content in terms of the gallicacid equivalent in one gram of each extract was determined(e total phenolic content was calculated from the calibrationcurve and final results expressed as mg GAE100 g DM

28 Determination of Total Flavonoid Content (e alu-minium chloride colorimetric assay method [14] wasemployed to evaluate total flavonoid content (TFC) usingquercetin as standard An aliquot of 500 μL of the extract andfractions were mixed with 15mL of 999 ethanol (EtOH)100 μL of 1M potassium acetate 100 μL of 10 aluminiumchloride and 3mL of distilled water (e mixture wasshaken vigorously and left to stand in dark at room tem-perature (e resulting mixtures were incubated for 30minat room temperature and corresponding absorbance mea-sured at 415 nm All determinations were carried out intriplicate A standard calibration curve was constructedusing quercetin standard solutions of 20 μgmiddotmLminus 140 μgmiddotmLminus 1 60 μgmiddotmLminus 1 80 μgmiddotmLminus 1 100 μgmiddotmLminus 1 and120 μgmiddotmLminus 1 500 μL of each standard was treated in the samemanner as the samples above to generate calibration curve(e total flavonoid content of each extract was determinedfrom the curve and the results recalculated and expressed asμg QE100 g DM

29 Determination of Total Tannins (e amount of tanninsin plant extract and fractions was determined by theFolinndashCiocalteu method with slight modifications [13]100 μL of the sample extract and fractions was added to 5mLof distilled water 500 μL of FolinndashCiocalteu reagent 1mLand of 35 Na2CO3 solution (e mixture was shaken welland kept at room temperature for 30min A set of referencestandard solutions of gallic acid (02 04 06 08 and1mgmiddotmLminus 1) were prepared in the same manner as described

earlier Absorbance for test and standard solutions weremeasured against the blank at 725 nm with an UVVisiblespectrophotometer (e total tannin content was de-termined from the calibration curve and the resultsexpressed tannin content was in terms of mg of GAE100 gDM

210 Determination of Antioxidant Activity (e free radicalscavenging ability of the extracts against DPPH free radicalwas evaluated as described by Oliveira et al [15] with somemodifications Briefly 200 μL of each extract was added to38mL of 0004 DPPH methanolic solution After 60minof incubation at room temperature in dark the absorbancewas measured at 517 nm A blank sample containing onlymethanol was used to zero the spectrophotometer Eachexperiment was performed in triplicate

Inhibition (I ) was calculated as follows

I Abs0 minus Abs1

Abs01113890 1113891lowast 100

Abs0 absorbance of 0004DPPHwithout analyte

Abs1 absorbance of 0004DPPHplus the test compound

(1)

211 Preparation of CultureMedia and Inoculum (emediaused in the assay comprised nutrient agar (Oxoid CM0003England) bacteriological peptone (Sigma-Aldrich P0556Germany) and Mueller-Hinton agar (Oxoid CM0337Oxoid Ltd England) (e media were prepared according tothe manufacturerrsquos instructions

2111 Preparation of Inoculum (e assay factored ninebacteria and yeast isolates (e stock cultures of Pseudo-monas aeroginosa (ATCC 27853) Escherichia coli (ATCC25922) Salmonella typhi (ATCC 19430) Staphylococcusaureus (ATCC 25923) Proteus mirabilis (ATCC 49565)Candida albicans Klebsiella pneumonia (ATCC 33495)Paratyphi B and Staphylococcus saprophyticus (ATCC15305) were subcultured onto fresh nutrient agar (OxoidCM0003 England) plates to obtain pure colonies Singlecolonies of each organism were suspended in test tubescontaining 5mL of sterilized bacteriological peptone(Sigma-Aldrich P0556 Germany) and incubated at 37degC for10ndash18 hours to attain the turbidity of 05 McFarland stan-dards [16] (e turbidity of the actively growing brothcultures which were not in conformity to the standard(exceeded the standard) were adjusted with sterile bacteri-ological peptone to obtain turbidity optically comparablewith that of 05 McFarland standard (approximately 1-2times108CFUmL for E coli ATCC 25922) [17]

2112 Preparation of Plant Fractions for the BioassaysConsidering the preparation of stock solutions for thebioassays 100mgmiddotmLminus 1 200mgmiddotmLminus 1 and 300mgmiddotmLminus 1 of

Biochemistry Research International 3

the fractionated plant extracts were reconstituted using 20vv DMSO (Sigma D5879 Germany) resulting in per-centage concentrations of 10 20 and 30 for the de-termination of antimicrobial activity (e crude extract wasdissolved in sterile distilled water for the aqueous extractpreparation of same concentrations (e stock solutionswere stored in a refrigerator until needed

2113 Antimicrobial Activity Assay (Agar DiffusionMethod) (e antimicrobial activity was carried out usingthe agar diffusion method [17] (e inoculums were in-oculated by swabbing onto sterile Mueller-Hinton agar(Oxoid CM0337 Oxoid Ltd England) A sterilized cockborer of an internal diameter of about 4mm was used topunch six holes in the medium (e plant extracts weredispensed into the holes to a volume of 100 μL (e positivecontrol was an antibiotic disc of 30 μg chloramphenicol(Oxoid CT00143 Oxoid Ltd England) and 20 vv DMSO(Sigma D5879 Germany) was used as negative control in atriplicate fashion (e plates were refrigerated for about 4 hto allow for absolute diffusion of the extract and incubated at37degC for 24 h after which the diameter of each zone ofinhibition was measured in millimetres (mm) with a ster-ilized ruler

212 Data Analysis (e results were expressed asmean plusmn standard error mean (SEM) and compared byone-way analysis of variance followed by Tukeyrsquos multiplecomparison test at the 95 significant level usingGraphPad Prism version 60

3 Results and Discussion

31 Results

311 Phytochemical Constituents Qualitative phytochemi-cal screening of raw leaves and fruits of D erecta indicatedpresence of tannins glycosides saponins flavonoids andtriterpenoids Alkaloids were present in unripe fruits onlywhile coumarins were not detected in any of the parts of Derecta (Table 1)

312 Heavy Metal Content in D erecta (e concentrationlevels of Ni and Pb were below detectable limits of1times 10minus 5mgmiddotLminus 1 for the leaves and fruits (e highest meanconcentration for Fe (217plusmn 005mgmiddotLminus 1) was found in leaves(raw) and the lowest (033plusmn 001mgmiddotLminus 1) in the ripe fruitextracts (Table 2) (e permissible limit of Fe in medicinalplants is 20mgmiddotmLminus 1 [17]

313 UV-Vis Spectroscopic Analysis (e UV-Vis spec-troscopy of extractfractions of D erecta was done at a rangeof 200 to 700 nm (e number of peaks and maximumabsorption range is shown in Table 3

314 FTIR Spectra (e FTIR spectra are shown inFigures 1ndash3 and these enabled the identification of possible

functional groups of the active components present in thefractions of D erecta

315 Total Phenolic Content (e total polyphenolic con-tents varied among the extracts and fractions (e meanhighest total phenolic level was observed in the hydrofraction of unripe fruit with a corresponding value of1454plusmn 029mg GAE100 g whilst the lowest value of220plusmn 015mg GAE100 g was recorded in petroleum etherfraction of the leaves (is is presented in Figure 4

316 Total Flavonoid Content (e highest total flavonoidcontent was observed in ethyl acetate fraction of the leaves(34349plusmn 345 μg QE100 g) (is is followed by hydro frac-tion of leaves (33870plusmn 418 μg QE100 g) hydro fraction ofunripe fruit (33008plusmn 192 μg QE100 g) ethyl acetate fractionof unripe fruit (28008plusmn 345 μg QE100 g) crude extract ofunripe seeds (24860plusmn 225 μg QE100 g) crude extract of ripeseeds (21800plusmn 345 μg QE100 g) methanolic fraction of ripeseeds (14617plusmn 096 μg QE100 g) and hydro fraction ofunripe seeds (4176plusmn 096 μg QE100 g) (Figure 5)

317 Total Tannins Content (TTC) (e TTC was expressedin terms of gallic acid equivalent per 100 g of the extract (eTTC varied from 1382plusmn 004mg GAE100 g to355plusmn 007mg GAE100 g (Figure 6)

318 DPPH Free Radical Scavenging Activity (e DPPHfree radical scavenging activity was expressed as percentage() (e highest scavenging activity of 900 was registeredfor methanolic fraction of the leaves whilst the lowestmethanolic fraction of 54was seen in unripe fruit (Figure 7)

319 Antimicrobial Activity (e antimicrobial activityassay of the ripe fruit unripe fruit and leaves of D erectaproved negative for all the crude hydro and methanolfractions with no recorded antimicrobial activity against thetest organisms (e positive control (standard antibiotic)however recorded positive responses to the test organisms

32 Discussion In the present study we carried out phy-tochemical investigation on Duranta erecta with the view ofdetermining its usefulness in traditional medicine (e

Table 1 Phytochemical constituents of D erecta

Test Leaves Unripe fruits Ripe fruitsTannins + + +Glycosides + + +Alkaloids minus + minus

Coumarins minus minus minus

Saponins + + +Flavonoids + + +Triterpenoids + + +Sterols + + +Note + present minus not detected


leaves and ripened fruits contained tannins glycosidessaponins flavonoids and triterpenoids whilst the unripefruits contained all the aforementioned in addition to al-kaloids (Table 1)(e absence of alkaloids in the leaves is notconsistent with earlier reports [9 18] that detected an ap-preciable level of alkaloids in D erecta leaves Phyto-chemicals play important role by combining with nutrientsand dietary fibre to us against diseases though some havebitter taste Alkaloids have been shown to have antioxidantproperties via alleviating H2O2-induced oxidative damage[19] Alkaloids have also been implicated in antimalarialactivity of many plants by blocking protein synthesis inPlasmodium falciparum [20] Tannins are known potentantioxidants and this activity is achieved by chelation ofmetal ions such as Fe (II) and interfering with one of thereaction steps in the Fenton reaction thereby retardingoxidation [21] Saponins possess antioxidant potentialthrough hydrogen peroxide scavenging capability [21]Flavonoids have the ability to inhibit lipid peroxidation bysugar substitutions in the phenolic C ring [19] Triterpenoidsand steroid saponins have been found detrimental to Pfalciparum [19] (is goes to buttress the use of D erecta inmalaria in folk medicine (e presence of these phyto-chemicals account for their multipurpose in traditionalmedicine

(e concentration levels of Ni and Pb were below de-tectable limits of 1times 10ndash5mgmiddotLminus 1 for the leaves and fruits(e highest mean concentration for Fe (217plusmn 005mgmiddotLminus 1)was found in leaves (raw) and the lowest (033plusmn 001mgmiddotLminus 1)in the ripened fruit extracts (Table 2) (e permissible limitof Fe in medicinal plants is 20mgmiddotLminus 1 [22] (e concen-tration of Zn in the analysed samples ranged from135plusmn 004mgmiddotLminus 1 to 040plusmn 002mgmiddotLminus 1 (e highest con-centration level of Zn was recorded in the raw leaf samples(ese values were within FAOWHO permissible limits of274mgmiddotLminus 1 in medicinal plants and were regarded as safe(e concentration of Cu in the samples varied (e highestCu level of 011plusmn 001mgmiddotLminus 1 was observed in ripened fruitfollowed by unripe 004plusmn 001mgmiddotLminus 1 and leaves002plusmn 001mgmiddotLminus 1 (e permissible limit of Cu in plantsrecommended by WHO is 10mgmiddotkgminus 1 In all the analysedsamples concentration of Cu recorded was below thepermissible limit

(e use of medicinal plants for healthcare presents a wayto humans to get exposed to trace elements and heavymetals Major and trace elements have significant roles incombating a variety of human ailments and diseases [23]Also beyond certain limits these may present serious healthhazards to humans (e knowledge on minerals and heavymetal content is essential for safe use of medicinal plants Allthe elements evaluated were within the WHOFAO per-missible limits (e presence of these elements and in theappropriate amounts in the plant confers useful values onthis plant Fe is an important trace element and iron-proteinmixtures play a vital role in metabolism It is essential in thebiosynthesis of haemoglobin and transport of oxygen andelectrons throughout the body [24] Zn is a component ofmany metalloenzymes especially enzymes which play acentral role in nucleic acid metabolism It is also essential inbone formation wound healing brain development andnormal growth processes [25] Cu is an essential componentof many enzymes and it plays a significant role in a widerange of physiological processes including Fe utilization freeradical elimination bone and connective tissues develop-ment and melanin production [24]

(e presence of phytochemicals was confirmed byspectroscopic studies showing the characteristic spectraobtained in the ultraviolet and visible spectrum(e UV-Visspectra of the extracts of the various parts had absorptionmaxima in the region of 2030ndash3851 nm 202ndash4230 nm and2030ndash3790 nm for the leaves unripe fruits and ripe fruitsrespectively (ese absorption bands are characteristic for

Table 2 Heavy metal content of raw plant materials and extracts

MaterialMetals

Fe (mgmiddotLminus 1) Zn (mgmiddotLminus 1) Ni (mgmiddotLminus 1) Cu (mgmiddotLminus 1) Pb (mgmiddotLminus 1)Unripe fruit raw 197plusmn 006 070plusmn 001 BDL 004plusmn 001 BDLUnripe fruit extract 064plusmn 004 040plusmn 002 BDL BDL BDLRipe fruit raw 203plusmn 006 064plusmn 001 BDL 011plusmn 001 BDLRipe fruit extract 033plusmn 001 043plusmn 001 BDL BDL BDLLeaves raw 217plusmn 005 135plusmn 004 BDL 002plusmn 001 BDLLeaves extract 096plusmn 003 126plusmn 0045 BDL BDL BDLNote BDL below detection limit detection limit 000001mgmiddotLminus 1

Table 3 UV-Vis peaks and absorption range

Extractfractions Number ofpeaks

Wavelengthrange (nm)

LeavesPetroleum ether fraction 5 2030ndash2681Ethyl acetate fraction 5 2040ndash2260Methanol fraction 5 2042ndash2159Hydro fraction 3 2852ndash3851Crude extract 2 2840ndash3829

UnripePetroleum ether fraction 5 2059ndash2181Ethyl acetate fraction 5 2040ndash2260Methanol fraction 3 2037ndash3223Hydro fraction 5 2871ndash3815Crude extract 5 2862ndash4231

RipePetroleum ether fraction 5 2030ndash2173Ethyl acetate fraction 5 2070ndash2230Methanol fraction 5 2015ndash3841Hydro fraction 4 2869ndash3769Crude extract 3 2859ndash3790


flavonoid and its derivatives (e flavonoid spectra mainlyconsist of two absorption maxima in the ranges of 230ndash285 nm and 300ndash350 nm [26] Similar results were con-firmed in an earlier study [27] that reported UV-Visspectrum of leaf extract in the absorption range of 270 and340 nm

(e FTIR spectrum was used to establish identity of thefunctional groups present in extract based on the peak valuesin the region of infra-red radiation (e results of FTIR

analysis revealed the presence of alcohols phenols alkanesaldehydes ketones aromatics aliphatic amines aromaticamines amides carboxylic acids esters nitro compoundsalkynes primary and secondary amines and alkyl halides(Figures 1ndash3) (ese were confirmed by FTIR spectroscopicanalysis that predicted the presence of the following groupsC-Br O-H C-H CC CO CequivC N-H C-H C-N C0(e presence of the various characteristic functional groupsmay be responsible medicinal properties of D erecta

101

86400

335604cmndash1

285447cmndash1

292542cmndash1

171086cmndash1

137785cmndash1

117330cmndash1 102502cmndash1

72226cmndash1

146173cmndash1

86889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

92400

9293949596979899

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

101

42400

4550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

101

400

333904cmndash1

210684cmndash1

163439cmndash143092cmndash1

41890cmndash1

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)

Figure 1 FTIR spectrum of petroleum ether (a) ethyl acetate (b) methanolic fraction (c) and hydro fraction (d) of D erecta leaf extract


100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

Figure 2 FTIR spectrum of ethyl acetate (a) methanolic fraction (b) and hydro fraction (c) of D erecta unripe fruit extract

100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued


(e hydro fraction of the unripe fruit recorded thehighest total phenolic content (1454plusmn 029mg GAE100 g)whilst the petroleum ether fraction of the leaves exhibitedthe lowest value (220plusmn 015mg GAE100 g Figure 4) (epolyphenolic content in the fraction depends on the polarityof the extraction solvent used High solubility of phenols inpolar solvents provides high concentration of these com-pounds when polar solvent is used in the extraction process[28] Polyphenolic compounds are important plant con-stituents due to their scavenging ability (e scavengingability is by virtue of the hydroxyl groups which has highaffinity for hydrogen atom thereby conferring antioxidativeproperty on the compounds (e mode of action of phenoliccompounds in free radical moping activity is by inactivatinglipid free radicals or by preventing the decomposition ofhydroperoxides into free radicals [29] (e antioxidantproperty makes such compounds useful in the managementof oxidative stress in humans

From Figure 5 the total flavonoid content varied from34349 plusmn 345 μg QE100 g to 47 plusmn 096 μg QE100 g (ehighest total flavonoid content was recorded in the ethylacetate fraction of the leaves and the lowest in the hydrofraction of unripe fruits (e concentration of flavonoids inextract depends on the polarity of solvents and the part ofplant material used for extractions [30] (e highestconcentration of flavonoids was recorded in solvent ofmoderate polarity (e antioxidative properties of flavo-noids can be linked to several different mechanisms suchas scavenging of free radicals chelation of metal ions suchas iron and copper and inhibition of enzymes responsiblefor free radical generation [29] Flavonoids also possessantibacterial activity (e antibacterial mechanisms ofaction of selected flavonoids are attributed to inhibition ofDNA gyrase cytoplasmic membrane function and com-plexing with bacterial cell wall and licochalcones A and Cenergy metabolism [31]

100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)

Figure 3 FTIR spectrum of petroleum ether (a) ethyl acetate (b) methanolic (c) and hydro fraction (d) of D erecta ripe fruit extract


(e crude extract of the leaves recorded the highestconcentration of tannins level 1382plusmn 004mg GAE100 gwhile the least (355plusmn 007mg GAE100 g) was observed inpetroleum ether fraction of the leaves Comparing our datawith a similar work [22 32] done elsewhere in Africa thecurrent data showed higher tannin levels (e differencescould be partly due to climatic conditions and the part of theplant used in the extraction Tannins inhibit lipid perox-idation by downregulating activity of cyclo-oxygenase en-zyme through inflammation pathway However tanninsinteract with protein molecules forming large cross-linkedcomplexes thereby rendering them indigestible

DPPH is a stable radical relative to other in vitro-gen-erated free radicals such as hydroxyl radical and superoxideanion DPPH has an added advantage of being not affectedby certain side reactions such as metal-ion chelation and

enzyme inhibition It is used to evaluate the free radicalmopping capacity of compounds In this study the DPPHscavenging activity ranged from 900 to 535 with the leafshowing the highest activity (e high scavenging activityassociated with the leaves explain their relevance in tradi-tional medicine Free radical scavenging is one of the knownmechanisms by which antioxidants inhibit lipid perox-idation (is is associated with the activity of antioxidantenzymes such as superoxide dismutase catalase and glu-tathione peroxidase

From this study (Tables 4ndash6) the extractfractions did notshow antimicrobial activity against the test organism (is isnot consistent with the other previous studies that recordedantimicrobial activity of Duranta erecta against Pseudomonasaeruginosa Proteus mirabilis Salmonella typhi Escherichiacoli and Klebsiella pneumonia Phytochemicals such as

Total phenols

0

5

10

15

20

mg

GA

E

Unripe fruits LeavesRipe fruitsSamples

CrudePetEtherEthylAc

MetOHHydro

Figure 4 Total phenolic content of extract and fractions of Derecta

mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro

SamplesRipe fruits Unripe fruits Leaves

Figure 5 Total flavonoid content of extract and fractions of Derecta

mg

GA

E

Total tannis15

10

5

0


MetOHHydro


Figure 6 Total tannins content of extract and fractions ofD erecta

in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro

Ripe fruits Unripe fruits Leaves

Figure 7 (e DPPH free radical scavenging activity of D erecta


flavonoids triterpenoids and alkaloids are widely known toexhibit some level of antimicrobial activity against somebacteria [33] Biological activity of the extract is moreeffective at higher concentrations and antimicrobial activitydecreases considerably with reduction in the concentration ofthe extract [22]

4 Conclusion

(e leaves and fruits of Duranta erecta contained importantphytochemicals including flavonoids phenols saponinssterols tannins alkaloids and triterpenoids and mineralssuch as Fe Zn and Cu which were within FAOWHOpermissible limits (e various parts demonstrated impor-tant antioxidant and beneficial properties which are nec-essary for its usefulness in traditional medicine (e resultsof FTIR analysis confirmed the presence of alcohols phe-nols alkanes aldehydes ketones aromatics aliphaticamines aromatic amines amides carboxylic acids estersnitro compounds alkynes primary and secondary aminesand alkyl halides

Data Availability

(e data used to support the findings of this study areavailable from the corresponding author upon request

Conflicts of Interest

(e authors declare that they have no conflicts of interest

References

[1] M F Mahomoodally ldquoTraditional medicines in Africa anappraisal of ten potent African medicinal plantsrdquo Evidence-Based Complementary and Alternative Medicine vol 2013Article ID 617459 14 pages 2013

[2] T A Abere P E Okoto and F O Agoreyo ldquoAntidiarrhoeaand toxicological evaluation of the leaf extract of Dissotisrotundifolia Triana (Melastomataceae)rdquo BMC Complemen-tary and Alternative Medicine vol 10 no 1 p 71 2010

[3] C Larbie C Owusu Nyarkoh and C Owusu Adjei ldquoPhy-tochemical and safety evaluation of hydroethanolic leaf ex-tract of Tecoma stans (L) juss ex kunthrdquo Evidence-BasedComplementary and Alternative Medicine vol 2019 ArticleID 7417624 12 pages 2019

[4] G A C Aymard and J R A Grande ldquoDuranta neblinensis(Verbenaceae Duranteae) a new species from Sierra de laNeblina Amazonas state Venezuelardquo Brittonia vol 64 no 3pp 246ndash251 2012

[5] S Hiradate H Yada T Ishii et al ldquo(ree plant growthinhibiting saponins from Duranta repensrdquo Phytochemistryvol 52 no 7 pp 1223ndash1228 1999

[6] F Ijaz N Ahmad I Ahmad A ul Haq and F Wang ldquoTwonew anti-plasmodial flavonoid glycosides from

Table 4 Antimicrobial activity of fractions of ripe fruits against test organisms

Extract TO 1 TO 2 TO 3 TO 4 TO 5 TO 6 TO 7 TO 8 TO 9Ex 1 mdash mdash mdash mdash mdash mdash mdash mdash mdashEx 2 mdash mdash mdash mdash mdash mdash mdash mdash mdashEx 3 mdash mdash mdash mdash mdash mdash mdash mdash mdashCtrl N mdash mdash mdash mdash mdash mdash mdash mdash mdashCtrl P 25plusmn 15 20plusmn 23 23plusmn 17 10plusmn 06 17plusmn 33 26plusmn 10 21plusmn 15 19plusmn 33 22plusmn 41Note TO test organism Ex extract fractions Ctrl N negative control Ctrl P positive control TO 1 Pseudomonas aeroginosa TO 2 Proteus mirabilis TO 3Salmonella typhi TO 4 Staphylococcus saprohyticus TO 5 Candida albicans TO 6 Staphylococcus aureus TO 7 Escherichia coli TO 8Klebsiella pneumoniaTO 9 Paratyphi B Ex 1 crude extract Ex 2 hydro fraction Ex 3 methanol fraction

Table 5 Antimicrobial activity of fractions of leaves against test organisms

Extract TO 1 TO 2 TO 3 TO 4 TO 5 TO 6 TO 7 TO 8 TO 9Ex 1 mdash mdash mdash mdash mdash mdash mdash mdash mdashEx 2 mdash mdash mdash mdash mdash mdash mdash mdash mdashEx 3 mdash mdash mdash mdash mdash mdash mdash mdash mdashCtrl N mdash mdash mdash mdash mdash mdash mdash mdash mdashCtrl P 25plusmn 15 20plusmn 23 23plusmn 17 10plusmn 06 17plusmn 33 26plusmn 10 21plusmn 15 19plusmn 33 22plusmn 41Note TO test organism Ex extract fractions Ctrl N negative control Ctrl P positive control TO 1 Pseudomonas aeroginosa TO 2 Proteus mirabilis TO 3Salmonella typhi TO 4 Staphylococcus saprophyticus TO 5 Candida albicans TO 6 Staphylococcus aureus TO 7 Escherichia coli TO 8 Klebsiellapneumonia TO 9 Paratyphi B Ex 1 crude extract Ex 2 hydro fraction Ex 3 methanol fraction

Table 6 Antimicrobial activity of fractions of unripe fruits against test organisms



Durantarepensrdquo Journal of Enzyme Inhibition and MedicinalChemistry vol 25 no 6 pp 773ndash778 2010

[7] L M Abou-Setta N M Nazif and A A Shahat ldquoPhyto-chemical investigation and antiviral activity of Durantarepensrdquo Journal of Applied Sciences Research vol 3pp 1426ndash1433 2007

[8] K Bhar L K Kantha S Manna P Nagalaxmi K Eswar andC Satya ldquoIn-vitro cytotoxic activity and anthelmintic activityof chloroform extract of D erecta L ripe fruitsrdquo ResearchJournal of Pharmaceutical Biological and Chemical Sciencesvol 7 no 6 pp 860ndash865 2016

[9] A J Ekenma O D Ejikene F Ekeh M Uwagbae G Ngwuand C Ehilegbu ldquoBioefficacy of Duranta erecta extract onyellow fever and dengue vector Aedes aegypti Linn inNigeriardquo Journal of Medicinal Plants Research vol 12 no 11pp 124ndash132 2018

[10] P Sharma S Khandelwal T Singh and R VijayvergialdquoPhytochemical analysis and antifungal potential of Durantaerecta against some phytopathogenic fungirdquo InternationalJournal of Pharmaceutical Sciences and Research vol 3pp 2686ndash2689 2012

[11] J B Harborne Phytochemical Methods A Guide to ModernTechniques of Plant Analysis Chapman amp Hall London UK3rd edition 1998

[12] S Sazada A Verma A A Rather F Jabeen andM K Meghvansi ldquoPreliminary phytochemicals analysis ofsome important medicinal and aromatic plantsrdquo Advances inBiological Research vol 3 pp 188ndash195 2009

[13] V L Singleton R Orthofer R M Lamuela-Raventos andP Lester Methods in Enzymology Academic PressCambridge MA USA 1999

[14] J Zhishen T Mengcheng and W Jianming ldquo(e de-termination of flavonoid contents in mulberry and theirscavenging effects on superoxide radicalsrdquo Food Chemistryvol 64 no 4 pp 555ndash559 1999

[15] I Oliveira A Sousa I C F R Ferreira A BentoL Estevinho and J A Pereira ldquoTotal phenols antioxidantpotential and antimicrobial activity of walnut (Juglans regiaL) green husksrdquo Food and Chemical Toxicology vol 46 no 7pp 2326ndash2331 2008

[16] F C Mills-Robertson A F Aboakye G Duker-EshunS Kaminta and S Agbeve ldquoIn vitro antimicrobial activity ofCryptolepis sanguinolenta (Perilocaceae)rdquo African Journal ofPharmacy and Pharmacology vol 3 no 9 pp 476ndash480 2009

[17] National Centre for Infectious Disease Centre for DiseaseControl and Prevention WHO Laboratory Methods for theDiagnosis of Epidemic Dysentery and Cholera Centre forDisease Control and Prevention Atlanta Georgia USA 199

[18] A S Ogbuagu K Okoro M U Akpuaka J O OgbuaguU E Ekpunobi and E C Ohaekenyem ldquoQuantitative de-termination of some secondary metabolites and the anti-bacterial effects of the leave extracts of Duranta eractardquoAmerican Journal of Biomedical Science and Engineeringvol 1 no 5 pp 82ndash87 2015

[19] G W Plumb K R Price and G Williamson ldquoAntioxidantproperties of flavonol glycosides from green beansrdquo RedoxReport vol 4 no 3 pp 123ndash127 1999

[20] A L Souto J F Tavares M S da Silva M d F F M DinizP F de Athayde-Filho and J M Barbosa Filho ldquoAnti-in-flammatory activity of alkaloids an update from 2000 to2010rdquo Molecules vol 16 no 10 pp 8515ndash8534 2011

[21] Y Chen Y Miao L Huang et al ldquoAntioxidant activities ofsaponins extracted from Radix trichosanthis an in vivo and in

vitro evaluationrdquo BMC Complementary and AlternativeMedicine vol 14 no 1 p 86 2014

[22] FAOWHO ldquoContaminantsrdquo in Codex Alimentariusvol Volume XVII 1st edition 1984

[23] K Shirin S Imad S Shafiq and K Fatima ldquoDetermination ofmajor and trace elements in the indigenous medicinal plantWithania somnifera and their possible correlation withtherapeutic activityrdquo Journal of Saudi Chemical Societyvol 14 no 1 pp 97ndash100 2010

[24] R Ullah J A Khader I Hussain N M AdElsalam M Talhaand N Khan ldquoInvestigation of macro and micro-nutrients inselected medicinal plantsrdquo African Journal of Pharmacy andPharmacology vol 6 no 25 pp 1829ndash1832 2012

[25] S Jabeen M T Shah S Khan and M Hayat ldquoDeterminationof major and trace elements in ten important folk therapeuticplants of Haripur basin Pakistanrdquo Journal of Medicinal PlantsResearch vol 4 no 7 pp 559ndash566 2010

[26] S M Dhivya and K Kalaichelvi ldquoUV-Vis spectroscopic andFTIR analysis of Sarcostemma brevistigma wight and arnrdquoInternational Journal of Herbal Medicine vol 9 no 3pp 46ndash49 2017

[27] N Rani S Sharma and M Sharma ldquoPhytochemical analysisof Meizotropis pellita by FTIR and UV-VIS spectrophotom-eterrdquo Indian Journal of Science and Technology vol 9 no 31pp 1ndash4 2016

[28] S M Mohsen and A S M Ammar ldquoTotal phenolic contentsand antioxidant activity of corn tassel extractsrdquo FoodChemistry vol 112 no 3 pp 595ndash598 2008

[29] S K Sharma and A P Singh ldquoIn vitro antioxidant and freeradical scavenging activity of Nardostachys jatamansi DCrdquoJournal of Acupuncture and Meridian Studies vol 5 no 3pp 112ndash118 2012

[30] M S Stankovic M D Topuzovic S Solujic and VMihailovildquoAntioxidant activity and concentration of phenols and fla-vonoids in the whole plant and plant parts of Teucriumchamaerdys L var glandiferumHausskrdquo Journal of MedicinalPlants Research vol 4 no 20 pp 2092ndash2098 2010

[31] E dos Santos M Pereira C da Silva et al ldquoAntibacterialactivity of the alkaloid-enriched extract from Prosopis juliflorapods and its influence on in vitro ruminal digestionrdquo In-ternational Journal of Molecular Sciences vol 14 no 4pp 8496ndash8516 2013

[32] M J Bangou N T R Meda A M E (iombiano et alldquoAntioxidant and antibacterial activities of five Verbenaceaespecies from Burkina Fasordquo Current Research Journal ofBiological Sciences vol 4 no 6 pp 665ndash672 2012

[33] C D Romero S F Chopin G Buck E Martinez M Garciaand L Bixby ldquoAntibacterial properties of common herbalremedies of the Southwestrdquo Journal of Ethnopharmacologyvol 99 no 2 pp 253ndash257 2005


Hindawiwwwhindawicom

International Journal of

Volume 2018

Zoology

Hindawiwwwhindawicom Volume 2018

Anatomy Research International

PeptidesInternational Journal of



Journal of Parasitology Research

GenomicsInternational Journal of


Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018


BioinformaticsAdvances in

Marine BiologyJournal of



Neuroscience Journal


BioMed Research International

Cell BiologyInternational Journal of



Biochemistry Research International

ArchaeaHindawiwwwhindawicom Volume 2018


Genetics Research International


Advances in

Virolog y Stem Cells International



Enzyme Research



MicrobiologyHindawiwwwhindawicom

Nucleic AcidsJournal of

Volume 2018

Submit your manuscripts atwwwhindawicom

In folk medicine plants are important source of naturalremedies used to treat many diseases Overdependence ontraditional folk medicine has brought to light the issue ofsustaining our fast-degrading forest In Ghana most of thetraditional medicine practitioners depend on our forests fortheir trade while our backyards are overgrown with orna-mental plants which are equally good source of herbalmedicine (us if we want to conserve our forest for sus-tainable development then there is the need to screen or-namental plants for biological and pharmacological propertiesby researchers(is is part of studies identifying alternative usefor ornamental plant [3]

Durantabelongs to the Verbenaceae family and itcomprises 35 species It is a native plant of Asia Africa andSouth and Central America [4] Duranta erecta (synony-mous to D repens) commonly known as ldquogolden dewdroprdquo is an upright scrambling shrub with a height of 1ndash3m It is grown as a hedge or ornamental plants in varioushomes in Ghana Phytoconstituents including coumar-inolignoids (E)-cinnamic acid (E)-p-methoxycinnamicacid and lamiide have been isolated from the genusDuranta [5 6] A number of bioactive compounds such asβ-sitosterol naringenin acteoside lamiide sucrose andraffinose have been identified and isolated from Durantarepens [7] Different parts of the plant are used to treatvariety of diseases (e fruit and leaves are used in tradi-tional folk medicine for the treatment of malaria intestinalworms and abscess and in some cases serve as vermifugeor diuretic D erecta is said to possess antitumor activityand significant antibacterial activity [8] It has numerousnatural uses including antifungal and insecticidal properties[9] (e ethyl acetate extract of leaves reportedly exhibitedsignificant antiplasmodial activity against the chloroquine-sensitive and chloroquine-resistant strains of Plasmodiumfalciparum [5] Methanolic extracts of different parts such asleaves stem and roots of D erecta exhibited antifungalproperties against Aspergillus flavus Alternaria sp Peni-cillium sp Rhizopus sp and Trichoderma sp [10] (e aimof the current study was to carry out the phytochemical andantioxidant screening ofD erecta grown as hedges in Ghanaas a justification for its use in traditional medicine

2 Materials and Methods

21 Chemicals FolinmdashCiocalteu phenol reagent (FCR)gallic acid 2 2-diphenylminus 1-picrylhydrazyl hydrate (DPPH)quercetin ascorbic acid and DMSO were from SigmaAlCl3 Na2CO3 CH3COOK HClO4 CHCl3 NaOH FeCl3and NaCl were fromMerck (Darmstadt Germany) Ethanolmethanol petroleum ether ethyl acetate nitric acid andsulphuric acid were from Janssen Chimica (Beerse Bel-gium) Nutrient agar Mueller-Hinton agar and chloram-phenicol were procured from Oxoid England All chemicalswere of analytical grade

22 Plant Collection and Authentication Plant materials(leaves ripe and unripe fruits) were collected from KNUSTcampus in October 2017 and authenticated by Dr George

Sam at the Department of Herbal Medicine Faculty ofPharmaceutical Sciences KNUST and the specimen wasdeposited in the facultyrsquos herbarium (voucher numberKNUSTHM12017L011)

23 Preparation and Extraction of PlantMaterial (e leavesand fruits of D erecta were washed under running tap waterto remove dust and other extraneous substances (ey weresubsequently dried under shade for 4 weeks(e shade-driedparts of the plant were coarsely powdered using a grinder(Waring USA) 60 g of the pulverized samples wereextracted with 50 ethanol by cold maceration for 48 hoursat room temperature on a shaker (Rocking LaboratoryShaker) (e mixture was separated by centrifugation andeach supernatant was evaporated under reduced pressureusing a rotary evaporator (Buchi R-205 Switzerland) (eextracts were dried using a vacuum freeze dryer (HetoPowerDry LL3000 UK) to obtain the respective crude ex-tracts which included D erecta leaves and unripe and ripefruits 15 g of each crude extract was sequentially extractedwith solvents of increasing polarity starting with petroleumether followed by ethyl acetate and methanol (e residualportion was designated as hydro fraction (e variousfractions namely petroleum ether ethyl acetate methanoland hydro fractions were all air-dried at room temperatureand used for the tests

24 Phytochemical Screening (e powdered samples of Derecta leaves unripe and ripe fruits and respective crudeextracts were subjected to qualitative analysis for identifi-cation of tannins (Gelatin test) glycosides (Molischrsquos test)alkaloids (Dragendorffrsquos reagents) coumarins (FeCl3 test)saponins (foam test) flavonoids (alkaline reagent) tri-terpenoids (Salkowskirsquos test) and sterols (LiebermannndashBurchard test) according to the standard methods describedby Harborne [11] and Sazada et al [12]

25 Heavy Metal Analysis Prior to sample analysis theglassware used for the analysis was soaked overnight in 10(vv) nitric acid followed by washing with 10 (vv)hydrochloric acid then rinsed with double-distilled waterand dried to eliminate interference Accurately 1 g of each ofthe extracts and the raw samples were weighed into a 50mLdigestion tube Each sample was wet-digested using amixture of 1mL of H2O 2mL of HCl 5mL of 1 1 HNO3 HClO4 and 2mL of H2SO4 for 20 minutes (emixture washeated in a digestion block at 150degC(e digested sample wasallowed to cool and diluted with 50mL of distilled water(edigests were analysed to determine the levels of heavy metalsin each extract Heavy metals including lead copper nickelzinc and iron were analysed using an atomic absorptionspectrometer (Varian AA 240FS) with a long path airacetylene burner and cathode lamp for respective metals

26 UV and FTIR Spectroscopic Analysis (e fractions ofleaves and fruits were diluted to 1 10 with respective sol-vents (e fractions were scanned in the wavelength ranging









I Abs0 minus Abs1

Abs01113890 1113891lowast 100



(1)









31 Results






















MaterialMetals












101

86400

335604cmndash1

285447cmndash1

292542cmndash1

171086cmndash1

137785cmndash1


72226cmndash1

146173cmndash1

86889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

92400

9293949596979899

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

101

42400

4550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

101

400

333904cmndash1

210684cmndash1


41890cmndash1

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)



100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)


100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018









I Abs0 minus Abs1

Abs01113890 1113891lowast 100



(1)









31 Results






















MaterialMetals












101

86400

335604cmndash1

285447cmndash1

292542cmndash1

171086cmndash1

137785cmndash1


72226cmndash1

146173cmndash1

86889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

92400

9293949596979899

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

101

42400

4550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

101

400

333904cmndash1

210684cmndash1


41890cmndash1

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)



100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)


100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018






31 Results






















MaterialMetals












101

86400

335604cmndash1

285447cmndash1

292542cmndash1

171086cmndash1

137785cmndash1


72226cmndash1

146173cmndash1

86889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

92400

9293949596979899

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

101

42400

4550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

101

400

333904cmndash1

210684cmndash1


41890cmndash1

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)



100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)


100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018







MaterialMetals












101

86400

335604cmndash1

285447cmndash1

292542cmndash1

171086cmndash1

137785cmndash1


72226cmndash1

146173cmndash1

86889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

92400

9293949596979899

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

101

42400

4550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

101

400

333904cmndash1

210684cmndash1


41890cmndash1

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)



100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)


100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018





101

86400

335604cmndash1

285447cmndash1

292542cmndash1

171086cmndash1

137785cmndash1


72226cmndash1

146173cmndash1

86889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

92400

9293949596979899

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

101

42400

4550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

101

400

333904cmndash1

210684cmndash1


41890cmndash1

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)



100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)


100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018


100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

102

78400

333964cmndash1

290258cmndash1

294730cmndash1

283290cmndash1

177203cmndash1

151541cmndash1

169475cmndash1

160318cmndash1


128438cmndash1

102313cmndash1

59316cmndash1

40773cmndash1

80828486889092949698

100

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)

400

290000cmndash1

333884cmndash1

282980cmndash1



40804 cmndash1

40290cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)


100

82400

333857cmndash1

283385cmndash1

292394cmndash1

163681cmndash1

144944cmndash1

60718cmndash1

102120cmndash18486889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(a)

Figure 3 Continued




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018




100

85400

86889092949698

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(b)



160228cmndash1


128319cmndash1

63893cmndash1

107196cmndash1

89

99

909192839495969798

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(c)

210445cmndash1

400

333902cmndash1


40373cmndash1

101

2935404550556065707580859095

T

4000 3000 2500 2000 1500 1000 5003500cmndash1

(d)







Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018






Total phenols

0

5

10

15

20

mg

GA

E



MetOHHydro


mg

QE

Total flavanoid400

300

200

100

0

CrudeEthylAc

MetOHHydro



mg

GA

E

Total tannis15

10

5

0


MetOHHydro



in

hibi

tion

Extracts

0

20

40

60

80

100DPPH


MetOHHydro





4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018



4 Conclusion


Data Availability




References














































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018


































Volume 2018

Zoology











Volume 2018

















Advances in




Enzyme Research





Volume 2018


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