Isolation and Characterization of Two New Antimicrobial...
6
Research Article Isolation and Characterization of Two New Antimicrobial Acids from Quercus incana (Bluejack Oak) Rizwana Sarwar , 1 Umar Farooq , 1 Sadia Naz, 1 Nadia Riaz , 2 Syed Majid Bukhari, 1 Abdur Rauf , 3 Yahia N. Mabkhot , 4 and Salim S. Al-Showiman 4 1 Department of Chemistry, COMSATS Institute of Information Technology, Abbotabad 22060, Pakistan 2 Department of Environmental Sciences, COMSATS Institute of Information Technology, Abbotabad 22060, Pakistan 3 Department of Chemistry, University of Swabi, Anbar, Khyber Pakhtunkhwa 23430, Pakistan 4 Department of Chemistry, College of Science, King Saud University, P.O. Box 2455, Riyadh 1451, Saudi Arabia Correspondence should be addressed to Umar Farooq; [email protected], Abdur Rauf; [email protected], and Yahia N. Mabkhot; [email protected] Received 24 August 2017; Accepted 27 November 2017; Published 28 January 2018 Academic Editor: Nikos Chorianopoulos Copyright © 2018 Rizwana Sarwar et al. is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Two new compounds [1-2] were purified from ethyl acetate fraction of Quercus incana. e structure of these compounds is mainly established by using advanced spectroscopic technique such as UV, IR, one-dimensional (ID) and two-dimensional (2D) NMR techniques, and EI mass. e structural formula was deduced to be 4-hydroxydecanoic acid [1] and 4-hydroxy-3-(hydroxymethyl) pentanoic acid [2]. Both isolated compounds were tested for their antimicrobial potential and showed promising antifungal activity against Aspergillus niger and Aspergillus flavus. 1. Introduction e family Fagaceae is large family comprising 8 genera and about 800–1100 species. Quercus is the largest genus of family Fagaceae having huge medicinal importance and mostly found in dry conditions [1]. e genus Quercus have long been considered among the clades of woody angiosperms in terms of species diversity, horticultural merit, ecological dominance, and industrial and economic values [2]. e Quercus robur is the only cultivated species while other 600 known species are found in temperate regions of the Northern Hemisphere, Southward through Central America to Colombia and through Turkey to Pakistan [3]. e wood is durable, is attractively grained, and is mostly utilized for timber purposes; it is particularly important in shipbuilding, construction for flooring, furniture, railroad ties, and veneers. e bark of Quercus spp. has been used for medicinal purposes and is an important source of phenolic compounds like tannins which are used for tanning leather and wine production [4]. e fruit (acorn) of Quercus has husk coating which is edible and highly nutritious and rich in carbohydrates and protein. Quercus (oak) species are utilized in conventional pharmaceutical, as astringent, antiseptic, and hemostatic and in addition to the treatment of acute diarrhea, hemorrhoid, and oral, genital, and anal mucosa inflammation. Moreover, the decoction plants from this genus can be used against burns and added to ointments for the healing of cuts [5]. Oak seeds are a major source of sugar, amino acids, lipids, and different sterols [6]. Quercus species have been utilized against problems of skin, wounds and gastrointestinal illnesses [7], astringent, mellow germ- free, small cuts [8], and mouth washes [9] all suggesting their antimicrobial potential. Genus Quercus is characterized by six species found mostly in Northern areas of Pakistan. e most promising timber specie is Quercus incana Roxb. (Blue jack oak or cinnamon oak) locally called Ban shindar, Kharpata serci (Punjabi), Rein (Hindko), and Serie (Pushto) [10]. e Q. incana has huge medicinal usage; it may be used as astringent [11], diuretic, and antidiarrheal agent and for treatment of asthma. Bark and leaves of Q. incana may be used as antipyretic, antirheumatism, antidiabetic, and antiarthritic Hindawi BioMed Research International Volume 2018, Article ID 3798105, 5 pages https://doi.org/10.1155/2018/3798105
Transcript of Isolation and Characterization of Two New Antimicrobial...
Research ArticleIsolation and Characterization of Two New Antimicrobial Acidsfrom Quercus incana (Bluejack Oak)
Rizwana Sarwar 1 Umar Farooq 1 Sadia Naz1 Nadia Riaz 2 Syed Majid Bukhari1
Abdur Rauf 3 Yahia N Mabkhot 4 and Salim S Al-Showiman4
1Department of Chemistry COMSATS Institute of Information Technology Abbotabad 22060 Pakistan2Department of Environmental Sciences COMSATS Institute of Information Technology Abbotabad 22060 Pakistan3Department of Chemistry University of Swabi Anbar Khyber Pakhtunkhwa 23430 Pakistan4Department of Chemistry College of Science King Saud University PO Box 2455 Riyadh 1451 Saudi Arabia
Correspondence should be addressed to Umar Farooq umarfciitnetpk Abdur Rauf mashaljcsyahoocomand Yahia N Mabkhot yahiaksuedusa
Received 24 August 2017 Accepted 27 November 2017 Published 28 January 2018
Academic Editor Nikos Chorianopoulos
Copyright copy 2018 Rizwana Sarwar et alThis 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
Two new compounds [1-2]were purified from ethyl acetate fraction ofQuercus incanaThe structure of these compounds is mainlyestablished by using advanced spectroscopic technique such as UV IR one-dimensional (ID) and two-dimensional (2D) NMRtechniques and EI massThe structural formula was deduced to be 4-hydroxydecanoic acid [1] and 4-hydroxy-3-(hydroxymethyl)pentanoic acid [2] Both isolated compounds were tested for their antimicrobial potential and showed promising antifungal activityagainst Aspergillus niger and Aspergillus flavus
1 Introduction
The family Fagaceae is large family comprising 8 genera andabout 800ndash1100 speciesQuercus is the largest genus of familyFagaceae having huge medicinal importance and mostlyfound in dry conditions [1] The genus Quercus have longbeen considered among the clades of woody angiospermsin terms of species diversity horticultural merit ecologicaldominance and industrial and economic values [2] TheQuercus robur is the only cultivated species while other600 known species are found in temperate regions of theNorthern Hemisphere Southward through Central Americato Colombia and through Turkey to Pakistan [3]
The wood is durable is attractively grained and is mostlyutilized for timber purposes it is particularly important inshipbuilding construction for flooring furniture railroadties and veneers The bark of Quercus spp has been used formedicinal purposes and is an important source of phenoliccompounds like tannins which are used for tanning leatherand wine production [4] The fruit (acorn) of Quercus hashusk coating which is edible and highly nutritious and
rich in carbohydrates and protein Quercus (oak) speciesare utilized in conventional pharmaceutical as astringentantiseptic and hemostatic and in addition to the treatmentof acute diarrhea hemorrhoid and oral genital and analmucosa inflammation Moreover the decoction plants fromthis genus can be used against burns and added to ointmentsfor the healing of cuts [5] Oak seeds are a major source ofsugar amino acids lipids and different sterols [6] Quercusspecies have been utilized against problems of skin woundsand gastrointestinal illnesses [7] astringent mellow germ-free small cuts [8] and mouth washes [9] all suggesting theirantimicrobial potential
Genus Quercus is characterized by six species foundmostly in Northern areas of Pakistan The most promisingtimber specie is Quercus incana Roxb (Blue jack oak orcinnamon oak) locally called Ban shindar Kharpata serci(Punjabi) Rein (Hindko) and Serie (Pushto) [10] The Qincana has hugemedicinal usage it may be used as astringent[11] diuretic and antidiarrheal agent and for treatmentof asthma Bark and leaves of Q incana may be used asantipyretic antirheumatism antidiabetic and antiarthritic
HindawiBioMed Research InternationalVolume 2018 Article ID 3798105 5 pageshttpsdoiorg10115520183798105
2 BioMed Research International
HO
OH
1 2 3 4 5 67
8
9
10
O
HO
OH
OH
HH
12
34 5
O
12
CHCH
1
Figure 1 Structure of compounds [1-2]
purposes [12] These medical applications and therapeuticpotential of Quercus incana prompted us to carry out thephytochemical investigation to explore biologically activecompounds
2 Material and Methods
21 Experimental Procedures The ethyl acetate soluble frac-tion was selected for isolation of bioactive compounds usingcolumn chromatographic analysis having column silica andflash silica gel as an adsorbent material The column waseluted by using n-hexane and ethyl acetate with increas-ing polarity which yield two new compounds 4-hydroxydecanoic acid [1] and 4-hydroxy-3-(hydroxymethyl) pen-tanoic acid [2] by increasing polarity The purity of com-pounds [1-2] was checked by using precoated TLC platesThe IR spectrum was recorded by using spectrophotometerJASCO-320A The EI mass was recoded on double focus-ing Varian MAT-312 Spectrometer 1H-NMR and 13C-NMRspectra were measured by using advance Bruker AMX-300 spectrometer machine The chemical shifts in partsper million (120575) relative to tetramethylsilane as an internalstandard and scalar (J) were described in Hz
22 Plant Extraction and Fractionation Extraction and frac-tionation of Quercus incana were reported in our previousstudy [13] Ethyl acetate soluble fraction was subjected torepeated column chromatography which yielded two purecompounds [1-2]
23 Antibacterial Activity of Compounds [1-2] Antibacterialactivity was performed by agar well diffusion method withsome modifications [14] Three Gram-positive (Staphylococ-cus aureus Micrococcus luteus and Bacillus subtilis) andGram-negative (Escherichia coli Pseudomonas pickettii andShigella flexneri) pathogens were used in study 10120583g of eachcompounds [1-2] was dissolved in 1mL DMSO Standarddrug and each sample (20 120583L) were poured in 6mmwellTheassay plates were incubated at 37∘C for 24 hrs The zone ofinhibition was dignified in mm and DMSO was used as anegative control in the experiment
24 Antifungal Assay Disc diffusion methods were used fordetermination of antifungal effects by using two selected fun-gal strains such asAspergillus niger andAspergillus flavus [15]
DMSO was used as a solvent before applying compounds onpetri plates DMSO was completely evaporated
Characterization of Compound 1 Colorless oil IR (KBr) 120592max3622 br (OH) 1714 (C=O) cmminus1 [120572]D
25 + 3480∘ (119888 = 078CHCl
3) EI-MS119898119911 (rel int) 188 [M]+ (15) 176 (9) 157 (35)
128 (5) 115 (9) HR-EI-MS 119898119911 188 1420 (calcd for 1881412for C10H20O3) 1H-NMR (CDCl
3 300MHz) 120575 362 (1H m
H-4) 088 (3H t J = 77Hz H-10) 208 (2H m H-2) 149162 (2H m H-3) 145 (2H m H-5) 147 133 (2H m H-6)130 (2H m H-7) 128 (2H m H-8) 129 (2H m H-9) 13C-NMR (CDCl
3 75MHz) 120575 1796 (C-1) 721 (C-4) 139 (C-10)
346 (C-2) 349 (C-3) 379 (C-5) 261 (C-6) 298 (C-7) 320(C-8) 221 (C-9)
Characterization of Compound 2 Colorless oil IR (KBr) 120592max3495 (OH) 1708 (C=O) cmminus1 [120572]D
25 + 5360∘ (119888 = 097CHCl
3) EI-MS119898119911 (rel int) 131 [M-OH]+ 131 (10) 115 (8)
86 (100) 71 (31) HR-EI-MS 119898119911 [M-OH]+ 1310744 (calcd1310736 for C
6H11O3-OH) 1H-NMR (CDCl
3 300MHz) 120575
235 (1H m H-2) 204 (1H m H-2) 258 (1H m H-3) 392(1H m H-4) 123 (3H d J = 69Hz H-5) 431 (2H m H-11015840)13C-NMR (CDCl
3 75MHz) 120575 1775 (C-1) 273 (C-2) 459
(C-3) 681 (C-4) 209 (C-5) 655 (C-11015840)
3 Result and Discussion
Ethyl acetate soluble fraction was subjected to repeatedcolumn chromatography on silica gel using n-hexane andethyl acetate as a solvent with gradual increasing in polarityup to 100 ethyl acetate which resulted in four subfractions(Fractions AndashD) The fractions obtained based on TLC pro-file were resubjected to pencil column chromatography andeluted with n-hexane EtOAc 25 75 and n-hexane EtOAc30 70 to purify compound 1 (105mg) and compound 2(98mg) (Figure 1)
Compound 1 was isolated as a colorless oil and hasmolecular formula of C
10H20O3as suggested by molecular
ion peak at 119898119911 188 [M]+ in HR-EIMS The other fragmentpeaks were obtained at 119898119911 176 157 128 and 115 The HR-EIMS gave exact mass of compound 1 which was at 1198981199111881420 (calcd 119898119911 1881412) The IR spectrum displayedabsorption bands for hydroxyl and carbonyl groups at 3622and 1714 cmminus1 respectively The 1H-NMR spectrum of com-pound 1 exhibited typical signal for aliphatic acid skeletonwhich was strongly supported by DEPT experiment The
BioMed Research International 3
Table 1 1H-NMR (CDCl3 300MHz) data of compounds [1-2] in
ppm J in Hz
Position 1 21 - -
2 208 m 204 m235 m
3 162 m149 m 258 m
4 362 m 392 m5 145 m 123 (1H d J = 69Hz)
6 147 m133 m -
7 130 m -8 128 m -9 129 m -10 088 (t J = 77) -11015840 - 431 m
13C-NMR spectrum revealed the presence of one methylone methine seven methylene and one quaternary carbonsignals The methine signal appearing at 120575H 362 (1H m) wasassigned to H-4 while methyl group resonated at 120575H 088(3H t J = 77Hz) Typical methylene signals were resonatedas multiplet for seven methylene carbons at 120575H 208 (2Hm) 120575H 149 (1H m) 120575H 145 (2H m) 120575H 133 (1H m) 120575H147 (1H m) 120575H 130 (2H m) 120575H 128 (2H m) and 120575H129 (2H m) assigned to H-2 H-3 H-5 H-6 H-7 H-8and H-9 respectively (Table 1) The 13C-NMR spectrum (BBand DEPT) corroborated the presence of seven methylenecarbons one methine carbon one terminal methyl carbonand one quaternary carbon The carbonyl carbon showedsignals at 120575C 1796 whereas methine signal centered at 120575C 721The 13C-NMR chemical shift of CH
3-C10 was observed at
120575C 139 and seven methylene carbons appeared at 120575C 346349 379 261 298 320 and 221 for C-2 C-3 C-5 C-6 C-7C-8 and C-9 respectively (Table 2) The HMBC and COSYspectra were quite helpful for accurate placement of varioussubstituents in the molecule The HMBC spectrum showedstrong correlation of methine proton at 120575H 362 (H-4) withC-2 C-3 C-5 and C-6 [13] The methyl proton at 120575H 088showed strong HMBC correlation with C-9 (120575C 221) C-8(120575C 320) which was quite supportive in the establishmentof structure Finally all spectral data confirmed compound1 as an aliphatic acid having straight chain of -(CH
2)7-CH3-
moiety [16] and was proposed to be 4-hydroxy decanoic acidCompound 2 was isolated as colorless oil Its structure
was mainly established by 1H-NMR and high resolutionmass spectroscopy and supported by 13C-NMR spectrumIts molecular formula C
6H12O4was concluded from the
accurate mass measurement of peak at 119898119911 [M-OH]+ 131corresponding to molecular composition C
6H11O3-OH In
addition to its molecular ion peak it showed some charac-teristic fragments at 119898119911 115 86 and 71 The HR-EIMS gaveexactmass of compound 2 at119898119911 1310744 (calcd 1310736 forC6H11O3-OH)The IR spectrum showed absorption bands at
Table 2 13C-NMR (CDCl3 75MHz) of compounds [1-2] in ppm
Position 1 21 1796 17752 346 2733 349 4594 721 6815 379 2096 261 -7 298 -8 320 -9 221 -10 139 -11015840 - 655
3495 cmminus1 and 1708 cmminus1 indicating presence of the hydroxylgroup and carbonyl carbon respectively Similarly broadabsorption centered at 2935 cmminus1 suggested the presenceof carboxylic acid The 1H-NMR showed a signal for asecondary methyl group at 120575 123 (3H d J = 69Hz H-5)connected to amethine group resonated at 120575H 392 (1HmH-4) possessing hydroxyl group while the other methine signalappeared at 120575H 258 (1H m H-3) The downfield methylenebearing hydroxyl group appeared at 120575H 431 (2HmH-11015840) andthe other methylene centered at 120575H 204 (1H m H-2) and 120575H235 (1H m H-2) was directly connected to carboxylic acid(Table 1) The 13C-NMR spectrum confirmed the presenceof one methyl carbon two methylene groups two methinecarbons and one quaternary carbon in the structure
In 13C-NMR spectrum signal for secondary methylappeared at 120575C 209 whereas the methine signal bearinghydroxyl group was observed at 120575C 681 for C-4 The signalfor another methine appeared at 120575C 459 for C-3 The sidechain methylene having free hydroxyl group resonated at120575C 655 while the second methylene group at position C-2appeared at 120575C 273 Similarly the quaternary carbon in theform of carboxylic acid showed signal at 120575C 1775 (Table 2)Based on the HMBC and H-H COSY correlation (Figure 2)the connectivity of the C-1 to C-5 chain was found inagreement with literature [17] The HMBC spectrum showedH-C correlation of CH
3-5 with that of C-4 and C-3 Similarly
the position of hydroxyl group at C-11015840 was confirmed bystrong HMBC correlation of CH
2-11015840 with C-3 C-2 and C-
4 and weak interaction with C-1 The structure of compound2 was mainly established by 1H-NMR high resolution massspectrometry and supported by 13C-NMRspectrum Fromallspectral data it was evident that compound 2was 4-hydroxy-3-(hydroxymethyl) pentanoic acid
31 Antibacterial Activity The antibacterial activity of iso-lated compounds [1-2] was determined by agar well diffusionmethod (Table 3) Compound 1 was significantly activeagainst Bacillus subtilis Staphylococcus aureus andMicrococ-cus luteus (Gram-positive) Both compounds [1-2] showedpromising antibacterial activity against Staphylococcus aureuswith 16mm and 13mm zone of inhibition Compound 2 was
4 BioMed Research International
HO
OH
1 2 3 4 5 67
8
9
10
Important HMBC correlation of 1
O
HO
OH
OH
HH
12
3
45
Important HMBC correlation of 2
O
CH
CH
1
Figure 2 HMBC correlation of compounds [1-2]
Table 3 Antibacterial activity of isolated compounds [1-2]
S number Culture Zone of inhibition (mm)1 2 Ciprofloxacin
1 Bacillus subtilis 8 5 82 Staphylococcus aureus 16 13 163 Micrococcus luteus 11 9 184 Pseudomonas pickettii 0 0 05 Escherichia coli 0 0 06 Shigella flexneri 6 9 14
Table 4 Antifungal activity of isolated compounds [1-2]
Extract Pathogenic fungiAspergillus flavus Aspergillus niger
1 12mm plusmn 050 15mm plusmn 0702 17mm plusmn 028 22mm plusmn 057Nystatin (standard) 16mm plusmn 092 21mm plusmn 028Note Each value in the table was obtained by calculating the average of threeexperiments
moderately active against Bacillus subtilis and Micrococcusluteus with 5mm and 9mm zone of inhibition Both com-pounds were inactive against Escherichia coli and Shigellaflexneri
32 Antifungal Activity Antifungal activity of both com-pounds [1-2] was done against Aspergillus flavus andAspergillus niger Both compounds [1-2] showed immenseactivity against Aspergillus niger with 15mm plusmn 070 and22mm plusmn 057 zone of inhibition (Table 4) Moderate activitywas observed by compound 1 againstAspergillus flavushaving12mm plusmn 050 zone of inhibition
4 Conclusion
The current study describes the isolation characterizationand antimicrobial activity of isolated compounds from ethylacetate fraction of Quercus incana Both compounds dis-played promising antimicrobial activity against human bac-terial and fungal strainsTherefore these isolated compounds
may be considered as the lead compounds as an antimicrobialagents
Conflicts of Interest
There are no conflicts of interest regarding this paper
Acknowledgments
The authors extend their appreciation to the Deanship ofScientific Research at King Saud University for its fundingthis prolific research group no RGP-007
References
[1] K C Nixon and W L Crepet ldquoTrigonobalanus (Fagaceae)Taxonomic Status and Phylogenetic Relationshipsrdquo AmericanJournal of Botany vol 76 no 6 p 828 1989
[2] M Kappelle ldquoNeotropical montane oak forests overview andoutlookrdquo in Ecology and Conservation of Neotropical MontaneOak Forests pp 449ndash467 Springer 2006
[3] E Nisar and S I Ali Flora of West Pakistan Botany Depart-ment Gordon College Rawalpindi Pakistan 1976
[4] S T Shah H Ahmad and R Zamir ldquoPollen morphologyof three species of Quercus (Family Fagaceaerdquo Journal ofAgriculture Forestry and the Social Sciences vol 1 no 4 pp 359-360 2005
[5] L BremnessHerbs (Handbooks) DorlingKindersley PublishersLTD London UK 1994
[6] S Rakic S Petrovic J Kukic et al ldquoInfluence of thermaltreatment on phenolic compounds and antioxidant properties
BioMed Research International 5
of oak acorns from Serbiardquo Food Chemistry vol 104 no 2 pp830ndash834 2007
[7] L Viegi A Pieroni P M Guarrera and R Vangelisti ldquoA reviewof plants used in folk veterinary medicine in Italy as basis fora databankrdquo Journal of Ethnopharmacology vol 89 no 2-3 pp221ndash244 2003
[8] M L Leporatti and S Ivancheva ldquoPreliminary comparativeanalysis of medicinal plants used in the traditional medicine ofBulgaria and Italyrdquo Journal of Ethnopharmacology vol 87 no2-3 pp 123ndash142 2003
[9] T Aburjai M Hudaib R Tayyem M Yousef and M QishawildquoEthnopharmacological survey of medicinal herbs in Jordanthe Ajloun Heights regionrdquo Journal of Ethnopharmacology vol110 no 2 pp 294ndash304 2007
[10] M Hussain and A Ghani ldquoHerbal remidies used for gastroin-testinal disorders in Kaghan valleyrdquo Pakistan Journal of WeedScience Research in SearchWorks vol 14 no 3-4 pp 169ndash2002008
[11] F Haq H Ahmad andM Alam ldquoTraditional uses of medicinalplants of Nandiar Khuwarr catchment (District Battagram)Pakistanrdquo Journal of Medicinal Plants Research vol 5 no 1 pp39ndash48 2011
[12] M I Shinwari and M A Khan ldquoFolk use of medicinalherbs of margalla hills National Park Islamabad rdquo Journal ofEthnopharmacology vol 69 no 1 pp 45ndash56 2000
[13] R Sarwar U Farooq A Khan et al ldquoEvaluation of antioxidantfree radical scavenging and antimicrobial activity of Quercusincana Roxbrdquo Frontiers in Pharmacology vol 6 article no 2772015
[14] J Harit A Barapatre M Prajapati K R Aadil and S SenapatildquoAntimicrobial activity of rhizome of selected Curcuma varietyrdquoInternational Journal of Life Sciences Biotechnology and PharmaResearch vol 2 pp 183ndash189 2013
[15] V Bobbarala V R Vadlapudi and C Naidu ldquoAntimicrobialpotentialities of mangrove plant Avicennia marinardquo Journal ofPharmacy Research vol 2 no 6 2009
[16] T N Misra R S Singh J Upadhyay and R SrivastavaldquoIsolation of a natural sterol and an aliphatic acid fromVernoniacinereardquo Phytochemistry vol 23 no 2 pp 415ndash417 1984
[17] K Krohn U Farooq U Florke et al ldquoSecondary metabolitesisolated froman endophytic Phoma sp -Absolute configurationof tetrahydropyrenophorol using the solid-state TDDFT CDmethodologyrdquo European Journal of Organic Chemistry no 19pp 3206ndash3211 2007
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Submit your manuscripts atwwwhindawicom
2 BioMed Research International
HO
OH
1 2 3 4 5 67
8
9
10
O
HO
OH
OH
HH
12
34 5
O
12
CHCH
1
Figure 1 Structure of compounds [1-2]
purposes [12] These medical applications and therapeuticpotential of Quercus incana prompted us to carry out thephytochemical investigation to explore biologically activecompounds
2 Material and Methods
21 Experimental Procedures The ethyl acetate soluble frac-tion was selected for isolation of bioactive compounds usingcolumn chromatographic analysis having column silica andflash silica gel as an adsorbent material The column waseluted by using n-hexane and ethyl acetate with increas-ing polarity which yield two new compounds 4-hydroxydecanoic acid [1] and 4-hydroxy-3-(hydroxymethyl) pen-tanoic acid [2] by increasing polarity The purity of com-pounds [1-2] was checked by using precoated TLC platesThe IR spectrum was recorded by using spectrophotometerJASCO-320A The EI mass was recoded on double focus-ing Varian MAT-312 Spectrometer 1H-NMR and 13C-NMRspectra were measured by using advance Bruker AMX-300 spectrometer machine The chemical shifts in partsper million (120575) relative to tetramethylsilane as an internalstandard and scalar (J) were described in Hz
22 Plant Extraction and Fractionation Extraction and frac-tionation of Quercus incana were reported in our previousstudy [13] Ethyl acetate soluble fraction was subjected torepeated column chromatography which yielded two purecompounds [1-2]
23 Antibacterial Activity of Compounds [1-2] Antibacterialactivity was performed by agar well diffusion method withsome modifications [14] Three Gram-positive (Staphylococ-cus aureus Micrococcus luteus and Bacillus subtilis) andGram-negative (Escherichia coli Pseudomonas pickettii andShigella flexneri) pathogens were used in study 10120583g of eachcompounds [1-2] was dissolved in 1mL DMSO Standarddrug and each sample (20 120583L) were poured in 6mmwellTheassay plates were incubated at 37∘C for 24 hrs The zone ofinhibition was dignified in mm and DMSO was used as anegative control in the experiment
24 Antifungal Assay Disc diffusion methods were used fordetermination of antifungal effects by using two selected fun-gal strains such asAspergillus niger andAspergillus flavus [15]
DMSO was used as a solvent before applying compounds onpetri plates DMSO was completely evaporated
Characterization of Compound 1 Colorless oil IR (KBr) 120592max3622 br (OH) 1714 (C=O) cmminus1 [120572]D
25 + 3480∘ (119888 = 078CHCl
3) EI-MS119898119911 (rel int) 188 [M]+ (15) 176 (9) 157 (35)
128 (5) 115 (9) HR-EI-MS 119898119911 188 1420 (calcd for 1881412for C10H20O3) 1H-NMR (CDCl
3 300MHz) 120575 362 (1H m
H-4) 088 (3H t J = 77Hz H-10) 208 (2H m H-2) 149162 (2H m H-3) 145 (2H m H-5) 147 133 (2H m H-6)130 (2H m H-7) 128 (2H m H-8) 129 (2H m H-9) 13C-NMR (CDCl
3 75MHz) 120575 1796 (C-1) 721 (C-4) 139 (C-10)
346 (C-2) 349 (C-3) 379 (C-5) 261 (C-6) 298 (C-7) 320(C-8) 221 (C-9)
Characterization of Compound 2 Colorless oil IR (KBr) 120592max3495 (OH) 1708 (C=O) cmminus1 [120572]D
25 + 5360∘ (119888 = 097CHCl
3) EI-MS119898119911 (rel int) 131 [M-OH]+ 131 (10) 115 (8)
86 (100) 71 (31) HR-EI-MS 119898119911 [M-OH]+ 1310744 (calcd1310736 for C
6H11O3-OH) 1H-NMR (CDCl
3 300MHz) 120575
235 (1H m H-2) 204 (1H m H-2) 258 (1H m H-3) 392(1H m H-4) 123 (3H d J = 69Hz H-5) 431 (2H m H-11015840)13C-NMR (CDCl
3 75MHz) 120575 1775 (C-1) 273 (C-2) 459
(C-3) 681 (C-4) 209 (C-5) 655 (C-11015840)
3 Result and Discussion
Ethyl acetate soluble fraction was subjected to repeatedcolumn chromatography on silica gel using n-hexane andethyl acetate as a solvent with gradual increasing in polarityup to 100 ethyl acetate which resulted in four subfractions(Fractions AndashD) The fractions obtained based on TLC pro-file were resubjected to pencil column chromatography andeluted with n-hexane EtOAc 25 75 and n-hexane EtOAc30 70 to purify compound 1 (105mg) and compound 2(98mg) (Figure 1)
Compound 1 was isolated as a colorless oil and hasmolecular formula of C
10H20O3as suggested by molecular
ion peak at 119898119911 188 [M]+ in HR-EIMS The other fragmentpeaks were obtained at 119898119911 176 157 128 and 115 The HR-EIMS gave exact mass of compound 1 which was at 1198981199111881420 (calcd 119898119911 1881412) The IR spectrum displayedabsorption bands for hydroxyl and carbonyl groups at 3622and 1714 cmminus1 respectively The 1H-NMR spectrum of com-pound 1 exhibited typical signal for aliphatic acid skeletonwhich was strongly supported by DEPT experiment The
BioMed Research International 3
Table 1 1H-NMR (CDCl3 300MHz) data of compounds [1-2] in
ppm J in Hz
Position 1 21 - -
2 208 m 204 m235 m
3 162 m149 m 258 m
4 362 m 392 m5 145 m 123 (1H d J = 69Hz)
6 147 m133 m -
7 130 m -8 128 m -9 129 m -10 088 (t J = 77) -11015840 - 431 m
13C-NMR spectrum revealed the presence of one methylone methine seven methylene and one quaternary carbonsignals The methine signal appearing at 120575H 362 (1H m) wasassigned to H-4 while methyl group resonated at 120575H 088(3H t J = 77Hz) Typical methylene signals were resonatedas multiplet for seven methylene carbons at 120575H 208 (2Hm) 120575H 149 (1H m) 120575H 145 (2H m) 120575H 133 (1H m) 120575H147 (1H m) 120575H 130 (2H m) 120575H 128 (2H m) and 120575H129 (2H m) assigned to H-2 H-3 H-5 H-6 H-7 H-8and H-9 respectively (Table 1) The 13C-NMR spectrum (BBand DEPT) corroborated the presence of seven methylenecarbons one methine carbon one terminal methyl carbonand one quaternary carbon The carbonyl carbon showedsignals at 120575C 1796 whereas methine signal centered at 120575C 721The 13C-NMR chemical shift of CH
3-C10 was observed at
120575C 139 and seven methylene carbons appeared at 120575C 346349 379 261 298 320 and 221 for C-2 C-3 C-5 C-6 C-7C-8 and C-9 respectively (Table 2) The HMBC and COSYspectra were quite helpful for accurate placement of varioussubstituents in the molecule The HMBC spectrum showedstrong correlation of methine proton at 120575H 362 (H-4) withC-2 C-3 C-5 and C-6 [13] The methyl proton at 120575H 088showed strong HMBC correlation with C-9 (120575C 221) C-8(120575C 320) which was quite supportive in the establishmentof structure Finally all spectral data confirmed compound1 as an aliphatic acid having straight chain of -(CH
2)7-CH3-
moiety [16] and was proposed to be 4-hydroxy decanoic acidCompound 2 was isolated as colorless oil Its structure
was mainly established by 1H-NMR and high resolutionmass spectroscopy and supported by 13C-NMR spectrumIts molecular formula C
6H12O4was concluded from the
accurate mass measurement of peak at 119898119911 [M-OH]+ 131corresponding to molecular composition C
6H11O3-OH In
addition to its molecular ion peak it showed some charac-teristic fragments at 119898119911 115 86 and 71 The HR-EIMS gaveexactmass of compound 2 at119898119911 1310744 (calcd 1310736 forC6H11O3-OH)The IR spectrum showed absorption bands at
Table 2 13C-NMR (CDCl3 75MHz) of compounds [1-2] in ppm
Position 1 21 1796 17752 346 2733 349 4594 721 6815 379 2096 261 -7 298 -8 320 -9 221 -10 139 -11015840 - 655
3495 cmminus1 and 1708 cmminus1 indicating presence of the hydroxylgroup and carbonyl carbon respectively Similarly broadabsorption centered at 2935 cmminus1 suggested the presenceof carboxylic acid The 1H-NMR showed a signal for asecondary methyl group at 120575 123 (3H d J = 69Hz H-5)connected to amethine group resonated at 120575H 392 (1HmH-4) possessing hydroxyl group while the other methine signalappeared at 120575H 258 (1H m H-3) The downfield methylenebearing hydroxyl group appeared at 120575H 431 (2HmH-11015840) andthe other methylene centered at 120575H 204 (1H m H-2) and 120575H235 (1H m H-2) was directly connected to carboxylic acid(Table 1) The 13C-NMR spectrum confirmed the presenceof one methyl carbon two methylene groups two methinecarbons and one quaternary carbon in the structure
In 13C-NMR spectrum signal for secondary methylappeared at 120575C 209 whereas the methine signal bearinghydroxyl group was observed at 120575C 681 for C-4 The signalfor another methine appeared at 120575C 459 for C-3 The sidechain methylene having free hydroxyl group resonated at120575C 655 while the second methylene group at position C-2appeared at 120575C 273 Similarly the quaternary carbon in theform of carboxylic acid showed signal at 120575C 1775 (Table 2)Based on the HMBC and H-H COSY correlation (Figure 2)the connectivity of the C-1 to C-5 chain was found inagreement with literature [17] The HMBC spectrum showedH-C correlation of CH
3-5 with that of C-4 and C-3 Similarly
the position of hydroxyl group at C-11015840 was confirmed bystrong HMBC correlation of CH
2-11015840 with C-3 C-2 and C-
4 and weak interaction with C-1 The structure of compound2 was mainly established by 1H-NMR high resolution massspectrometry and supported by 13C-NMRspectrum Fromallspectral data it was evident that compound 2was 4-hydroxy-3-(hydroxymethyl) pentanoic acid
31 Antibacterial Activity The antibacterial activity of iso-lated compounds [1-2] was determined by agar well diffusionmethod (Table 3) Compound 1 was significantly activeagainst Bacillus subtilis Staphylococcus aureus andMicrococ-cus luteus (Gram-positive) Both compounds [1-2] showedpromising antibacterial activity against Staphylococcus aureuswith 16mm and 13mm zone of inhibition Compound 2 was
4 BioMed Research International
HO
OH
1 2 3 4 5 67
8
9
10
Important HMBC correlation of 1
O
HO
OH
OH
HH
12
3
45
Important HMBC correlation of 2
O
CH
CH
1
Figure 2 HMBC correlation of compounds [1-2]
Table 3 Antibacterial activity of isolated compounds [1-2]
S number Culture Zone of inhibition (mm)1 2 Ciprofloxacin
1 Bacillus subtilis 8 5 82 Staphylococcus aureus 16 13 163 Micrococcus luteus 11 9 184 Pseudomonas pickettii 0 0 05 Escherichia coli 0 0 06 Shigella flexneri 6 9 14
Table 4 Antifungal activity of isolated compounds [1-2]
Extract Pathogenic fungiAspergillus flavus Aspergillus niger
1 12mm plusmn 050 15mm plusmn 0702 17mm plusmn 028 22mm plusmn 057Nystatin (standard) 16mm plusmn 092 21mm plusmn 028Note Each value in the table was obtained by calculating the average of threeexperiments
moderately active against Bacillus subtilis and Micrococcusluteus with 5mm and 9mm zone of inhibition Both com-pounds were inactive against Escherichia coli and Shigellaflexneri
32 Antifungal Activity Antifungal activity of both com-pounds [1-2] was done against Aspergillus flavus andAspergillus niger Both compounds [1-2] showed immenseactivity against Aspergillus niger with 15mm plusmn 070 and22mm plusmn 057 zone of inhibition (Table 4) Moderate activitywas observed by compound 1 againstAspergillus flavushaving12mm plusmn 050 zone of inhibition
4 Conclusion
The current study describes the isolation characterizationand antimicrobial activity of isolated compounds from ethylacetate fraction of Quercus incana Both compounds dis-played promising antimicrobial activity against human bac-terial and fungal strainsTherefore these isolated compounds
may be considered as the lead compounds as an antimicrobialagents
Conflicts of Interest
There are no conflicts of interest regarding this paper
Acknowledgments
The authors extend their appreciation to the Deanship ofScientific Research at King Saud University for its fundingthis prolific research group no RGP-007
References
[1] K C Nixon and W L Crepet ldquoTrigonobalanus (Fagaceae)Taxonomic Status and Phylogenetic Relationshipsrdquo AmericanJournal of Botany vol 76 no 6 p 828 1989
[2] M Kappelle ldquoNeotropical montane oak forests overview andoutlookrdquo in Ecology and Conservation of Neotropical MontaneOak Forests pp 449ndash467 Springer 2006
[3] E Nisar and S I Ali Flora of West Pakistan Botany Depart-ment Gordon College Rawalpindi Pakistan 1976
[4] S T Shah H Ahmad and R Zamir ldquoPollen morphologyof three species of Quercus (Family Fagaceaerdquo Journal ofAgriculture Forestry and the Social Sciences vol 1 no 4 pp 359-360 2005
[5] L BremnessHerbs (Handbooks) DorlingKindersley PublishersLTD London UK 1994
[6] S Rakic S Petrovic J Kukic et al ldquoInfluence of thermaltreatment on phenolic compounds and antioxidant properties
BioMed Research International 5
of oak acorns from Serbiardquo Food Chemistry vol 104 no 2 pp830ndash834 2007
[7] L Viegi A Pieroni P M Guarrera and R Vangelisti ldquoA reviewof plants used in folk veterinary medicine in Italy as basis fora databankrdquo Journal of Ethnopharmacology vol 89 no 2-3 pp221ndash244 2003
[8] M L Leporatti and S Ivancheva ldquoPreliminary comparativeanalysis of medicinal plants used in the traditional medicine ofBulgaria and Italyrdquo Journal of Ethnopharmacology vol 87 no2-3 pp 123ndash142 2003
[9] T Aburjai M Hudaib R Tayyem M Yousef and M QishawildquoEthnopharmacological survey of medicinal herbs in Jordanthe Ajloun Heights regionrdquo Journal of Ethnopharmacology vol110 no 2 pp 294ndash304 2007
[10] M Hussain and A Ghani ldquoHerbal remidies used for gastroin-testinal disorders in Kaghan valleyrdquo Pakistan Journal of WeedScience Research in SearchWorks vol 14 no 3-4 pp 169ndash2002008
[11] F Haq H Ahmad andM Alam ldquoTraditional uses of medicinalplants of Nandiar Khuwarr catchment (District Battagram)Pakistanrdquo Journal of Medicinal Plants Research vol 5 no 1 pp39ndash48 2011
[12] M I Shinwari and M A Khan ldquoFolk use of medicinalherbs of margalla hills National Park Islamabad rdquo Journal ofEthnopharmacology vol 69 no 1 pp 45ndash56 2000
[13] R Sarwar U Farooq A Khan et al ldquoEvaluation of antioxidantfree radical scavenging and antimicrobial activity of Quercusincana Roxbrdquo Frontiers in Pharmacology vol 6 article no 2772015
[14] J Harit A Barapatre M Prajapati K R Aadil and S SenapatildquoAntimicrobial activity of rhizome of selected Curcuma varietyrdquoInternational Journal of Life Sciences Biotechnology and PharmaResearch vol 2 pp 183ndash189 2013
[15] V Bobbarala V R Vadlapudi and C Naidu ldquoAntimicrobialpotentialities of mangrove plant Avicennia marinardquo Journal ofPharmacy Research vol 2 no 6 2009
[16] T N Misra R S Singh J Upadhyay and R SrivastavaldquoIsolation of a natural sterol and an aliphatic acid fromVernoniacinereardquo Phytochemistry vol 23 no 2 pp 415ndash417 1984
[17] K Krohn U Farooq U Florke et al ldquoSecondary metabolitesisolated froman endophytic Phoma sp -Absolute configurationof tetrahydropyrenophorol using the solid-state TDDFT CDmethodologyrdquo European Journal of Organic Chemistry no 19pp 3206ndash3211 2007
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
BioMed Research International 3
Table 1 1H-NMR (CDCl3 300MHz) data of compounds [1-2] in
ppm J in Hz
Position 1 21 - -
2 208 m 204 m235 m
3 162 m149 m 258 m
4 362 m 392 m5 145 m 123 (1H d J = 69Hz)
6 147 m133 m -
7 130 m -8 128 m -9 129 m -10 088 (t J = 77) -11015840 - 431 m
13C-NMR spectrum revealed the presence of one methylone methine seven methylene and one quaternary carbonsignals The methine signal appearing at 120575H 362 (1H m) wasassigned to H-4 while methyl group resonated at 120575H 088(3H t J = 77Hz) Typical methylene signals were resonatedas multiplet for seven methylene carbons at 120575H 208 (2Hm) 120575H 149 (1H m) 120575H 145 (2H m) 120575H 133 (1H m) 120575H147 (1H m) 120575H 130 (2H m) 120575H 128 (2H m) and 120575H129 (2H m) assigned to H-2 H-3 H-5 H-6 H-7 H-8and H-9 respectively (Table 1) The 13C-NMR spectrum (BBand DEPT) corroborated the presence of seven methylenecarbons one methine carbon one terminal methyl carbonand one quaternary carbon The carbonyl carbon showedsignals at 120575C 1796 whereas methine signal centered at 120575C 721The 13C-NMR chemical shift of CH
3-C10 was observed at
120575C 139 and seven methylene carbons appeared at 120575C 346349 379 261 298 320 and 221 for C-2 C-3 C-5 C-6 C-7C-8 and C-9 respectively (Table 2) The HMBC and COSYspectra were quite helpful for accurate placement of varioussubstituents in the molecule The HMBC spectrum showedstrong correlation of methine proton at 120575H 362 (H-4) withC-2 C-3 C-5 and C-6 [13] The methyl proton at 120575H 088showed strong HMBC correlation with C-9 (120575C 221) C-8(120575C 320) which was quite supportive in the establishmentof structure Finally all spectral data confirmed compound1 as an aliphatic acid having straight chain of -(CH
2)7-CH3-
moiety [16] and was proposed to be 4-hydroxy decanoic acidCompound 2 was isolated as colorless oil Its structure
was mainly established by 1H-NMR and high resolutionmass spectroscopy and supported by 13C-NMR spectrumIts molecular formula C
6H12O4was concluded from the
accurate mass measurement of peak at 119898119911 [M-OH]+ 131corresponding to molecular composition C
6H11O3-OH In
addition to its molecular ion peak it showed some charac-teristic fragments at 119898119911 115 86 and 71 The HR-EIMS gaveexactmass of compound 2 at119898119911 1310744 (calcd 1310736 forC6H11O3-OH)The IR spectrum showed absorption bands at
Table 2 13C-NMR (CDCl3 75MHz) of compounds [1-2] in ppm
Position 1 21 1796 17752 346 2733 349 4594 721 6815 379 2096 261 -7 298 -8 320 -9 221 -10 139 -11015840 - 655
3495 cmminus1 and 1708 cmminus1 indicating presence of the hydroxylgroup and carbonyl carbon respectively Similarly broadabsorption centered at 2935 cmminus1 suggested the presenceof carboxylic acid The 1H-NMR showed a signal for asecondary methyl group at 120575 123 (3H d J = 69Hz H-5)connected to amethine group resonated at 120575H 392 (1HmH-4) possessing hydroxyl group while the other methine signalappeared at 120575H 258 (1H m H-3) The downfield methylenebearing hydroxyl group appeared at 120575H 431 (2HmH-11015840) andthe other methylene centered at 120575H 204 (1H m H-2) and 120575H235 (1H m H-2) was directly connected to carboxylic acid(Table 1) The 13C-NMR spectrum confirmed the presenceof one methyl carbon two methylene groups two methinecarbons and one quaternary carbon in the structure
In 13C-NMR spectrum signal for secondary methylappeared at 120575C 209 whereas the methine signal bearinghydroxyl group was observed at 120575C 681 for C-4 The signalfor another methine appeared at 120575C 459 for C-3 The sidechain methylene having free hydroxyl group resonated at120575C 655 while the second methylene group at position C-2appeared at 120575C 273 Similarly the quaternary carbon in theform of carboxylic acid showed signal at 120575C 1775 (Table 2)Based on the HMBC and H-H COSY correlation (Figure 2)the connectivity of the C-1 to C-5 chain was found inagreement with literature [17] The HMBC spectrum showedH-C correlation of CH
3-5 with that of C-4 and C-3 Similarly
the position of hydroxyl group at C-11015840 was confirmed bystrong HMBC correlation of CH
2-11015840 with C-3 C-2 and C-
4 and weak interaction with C-1 The structure of compound2 was mainly established by 1H-NMR high resolution massspectrometry and supported by 13C-NMRspectrum Fromallspectral data it was evident that compound 2was 4-hydroxy-3-(hydroxymethyl) pentanoic acid
31 Antibacterial Activity The antibacterial activity of iso-lated compounds [1-2] was determined by agar well diffusionmethod (Table 3) Compound 1 was significantly activeagainst Bacillus subtilis Staphylococcus aureus andMicrococ-cus luteus (Gram-positive) Both compounds [1-2] showedpromising antibacterial activity against Staphylococcus aureuswith 16mm and 13mm zone of inhibition Compound 2 was
4 BioMed Research International
HO
OH
1 2 3 4 5 67
8
9
10
Important HMBC correlation of 1
O
HO
OH
OH
HH
12
3
45
Important HMBC correlation of 2
O
CH
CH
1
Figure 2 HMBC correlation of compounds [1-2]
Table 3 Antibacterial activity of isolated compounds [1-2]
S number Culture Zone of inhibition (mm)1 2 Ciprofloxacin
1 Bacillus subtilis 8 5 82 Staphylococcus aureus 16 13 163 Micrococcus luteus 11 9 184 Pseudomonas pickettii 0 0 05 Escherichia coli 0 0 06 Shigella flexneri 6 9 14
Table 4 Antifungal activity of isolated compounds [1-2]
Extract Pathogenic fungiAspergillus flavus Aspergillus niger
1 12mm plusmn 050 15mm plusmn 0702 17mm plusmn 028 22mm plusmn 057Nystatin (standard) 16mm plusmn 092 21mm plusmn 028Note Each value in the table was obtained by calculating the average of threeexperiments
moderately active against Bacillus subtilis and Micrococcusluteus with 5mm and 9mm zone of inhibition Both com-pounds were inactive against Escherichia coli and Shigellaflexneri
32 Antifungal Activity Antifungal activity of both com-pounds [1-2] was done against Aspergillus flavus andAspergillus niger Both compounds [1-2] showed immenseactivity against Aspergillus niger with 15mm plusmn 070 and22mm plusmn 057 zone of inhibition (Table 4) Moderate activitywas observed by compound 1 againstAspergillus flavushaving12mm plusmn 050 zone of inhibition
4 Conclusion
The current study describes the isolation characterizationand antimicrobial activity of isolated compounds from ethylacetate fraction of Quercus incana Both compounds dis-played promising antimicrobial activity against human bac-terial and fungal strainsTherefore these isolated compounds
may be considered as the lead compounds as an antimicrobialagents
Conflicts of Interest
There are no conflicts of interest regarding this paper
Acknowledgments
The authors extend their appreciation to the Deanship ofScientific Research at King Saud University for its fundingthis prolific research group no RGP-007
References
[1] K C Nixon and W L Crepet ldquoTrigonobalanus (Fagaceae)Taxonomic Status and Phylogenetic Relationshipsrdquo AmericanJournal of Botany vol 76 no 6 p 828 1989
[2] M Kappelle ldquoNeotropical montane oak forests overview andoutlookrdquo in Ecology and Conservation of Neotropical MontaneOak Forests pp 449ndash467 Springer 2006
[3] E Nisar and S I Ali Flora of West Pakistan Botany Depart-ment Gordon College Rawalpindi Pakistan 1976
[4] S T Shah H Ahmad and R Zamir ldquoPollen morphologyof three species of Quercus (Family Fagaceaerdquo Journal ofAgriculture Forestry and the Social Sciences vol 1 no 4 pp 359-360 2005
[5] L BremnessHerbs (Handbooks) DorlingKindersley PublishersLTD London UK 1994
[6] S Rakic S Petrovic J Kukic et al ldquoInfluence of thermaltreatment on phenolic compounds and antioxidant properties
BioMed Research International 5
of oak acorns from Serbiardquo Food Chemistry vol 104 no 2 pp830ndash834 2007
[7] L Viegi A Pieroni P M Guarrera and R Vangelisti ldquoA reviewof plants used in folk veterinary medicine in Italy as basis fora databankrdquo Journal of Ethnopharmacology vol 89 no 2-3 pp221ndash244 2003
[8] M L Leporatti and S Ivancheva ldquoPreliminary comparativeanalysis of medicinal plants used in the traditional medicine ofBulgaria and Italyrdquo Journal of Ethnopharmacology vol 87 no2-3 pp 123ndash142 2003
[9] T Aburjai M Hudaib R Tayyem M Yousef and M QishawildquoEthnopharmacological survey of medicinal herbs in Jordanthe Ajloun Heights regionrdquo Journal of Ethnopharmacology vol110 no 2 pp 294ndash304 2007
[10] M Hussain and A Ghani ldquoHerbal remidies used for gastroin-testinal disorders in Kaghan valleyrdquo Pakistan Journal of WeedScience Research in SearchWorks vol 14 no 3-4 pp 169ndash2002008
[11] F Haq H Ahmad andM Alam ldquoTraditional uses of medicinalplants of Nandiar Khuwarr catchment (District Battagram)Pakistanrdquo Journal of Medicinal Plants Research vol 5 no 1 pp39ndash48 2011
[12] M I Shinwari and M A Khan ldquoFolk use of medicinalherbs of margalla hills National Park Islamabad rdquo Journal ofEthnopharmacology vol 69 no 1 pp 45ndash56 2000
[13] R Sarwar U Farooq A Khan et al ldquoEvaluation of antioxidantfree radical scavenging and antimicrobial activity of Quercusincana Roxbrdquo Frontiers in Pharmacology vol 6 article no 2772015
[14] J Harit A Barapatre M Prajapati K R Aadil and S SenapatildquoAntimicrobial activity of rhizome of selected Curcuma varietyrdquoInternational Journal of Life Sciences Biotechnology and PharmaResearch vol 2 pp 183ndash189 2013
[15] V Bobbarala V R Vadlapudi and C Naidu ldquoAntimicrobialpotentialities of mangrove plant Avicennia marinardquo Journal ofPharmacy Research vol 2 no 6 2009
[16] T N Misra R S Singh J Upadhyay and R SrivastavaldquoIsolation of a natural sterol and an aliphatic acid fromVernoniacinereardquo Phytochemistry vol 23 no 2 pp 415ndash417 1984
[17] K Krohn U Farooq U Florke et al ldquoSecondary metabolitesisolated froman endophytic Phoma sp -Absolute configurationof tetrahydropyrenophorol using the solid-state TDDFT CDmethodologyrdquo European Journal of Organic Chemistry no 19pp 3206ndash3211 2007
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
4 BioMed Research International
HO
OH
1 2 3 4 5 67
8
9
10
Important HMBC correlation of 1
O
HO
OH
OH
HH
12
3
45
Important HMBC correlation of 2
O
CH
CH
1
Figure 2 HMBC correlation of compounds [1-2]
Table 3 Antibacterial activity of isolated compounds [1-2]
S number Culture Zone of inhibition (mm)1 2 Ciprofloxacin
1 Bacillus subtilis 8 5 82 Staphylococcus aureus 16 13 163 Micrococcus luteus 11 9 184 Pseudomonas pickettii 0 0 05 Escherichia coli 0 0 06 Shigella flexneri 6 9 14
Table 4 Antifungal activity of isolated compounds [1-2]
Extract Pathogenic fungiAspergillus flavus Aspergillus niger
1 12mm plusmn 050 15mm plusmn 0702 17mm plusmn 028 22mm plusmn 057Nystatin (standard) 16mm plusmn 092 21mm plusmn 028Note Each value in the table was obtained by calculating the average of threeexperiments
moderately active against Bacillus subtilis and Micrococcusluteus with 5mm and 9mm zone of inhibition Both com-pounds were inactive against Escherichia coli and Shigellaflexneri
32 Antifungal Activity Antifungal activity of both com-pounds [1-2] was done against Aspergillus flavus andAspergillus niger Both compounds [1-2] showed immenseactivity against Aspergillus niger with 15mm plusmn 070 and22mm plusmn 057 zone of inhibition (Table 4) Moderate activitywas observed by compound 1 againstAspergillus flavushaving12mm plusmn 050 zone of inhibition
4 Conclusion
The current study describes the isolation characterizationand antimicrobial activity of isolated compounds from ethylacetate fraction of Quercus incana Both compounds dis-played promising antimicrobial activity against human bac-terial and fungal strainsTherefore these isolated compounds
may be considered as the lead compounds as an antimicrobialagents
Conflicts of Interest
There are no conflicts of interest regarding this paper
Acknowledgments
The authors extend their appreciation to the Deanship ofScientific Research at King Saud University for its fundingthis prolific research group no RGP-007
References
[1] K C Nixon and W L Crepet ldquoTrigonobalanus (Fagaceae)Taxonomic Status and Phylogenetic Relationshipsrdquo AmericanJournal of Botany vol 76 no 6 p 828 1989
[2] M Kappelle ldquoNeotropical montane oak forests overview andoutlookrdquo in Ecology and Conservation of Neotropical MontaneOak Forests pp 449ndash467 Springer 2006
[3] E Nisar and S I Ali Flora of West Pakistan Botany Depart-ment Gordon College Rawalpindi Pakistan 1976
[4] S T Shah H Ahmad and R Zamir ldquoPollen morphologyof three species of Quercus (Family Fagaceaerdquo Journal ofAgriculture Forestry and the Social Sciences vol 1 no 4 pp 359-360 2005
[5] L BremnessHerbs (Handbooks) DorlingKindersley PublishersLTD London UK 1994
[6] S Rakic S Petrovic J Kukic et al ldquoInfluence of thermaltreatment on phenolic compounds and antioxidant properties
BioMed Research International 5
of oak acorns from Serbiardquo Food Chemistry vol 104 no 2 pp830ndash834 2007
[7] L Viegi A Pieroni P M Guarrera and R Vangelisti ldquoA reviewof plants used in folk veterinary medicine in Italy as basis fora databankrdquo Journal of Ethnopharmacology vol 89 no 2-3 pp221ndash244 2003
[8] M L Leporatti and S Ivancheva ldquoPreliminary comparativeanalysis of medicinal plants used in the traditional medicine ofBulgaria and Italyrdquo Journal of Ethnopharmacology vol 87 no2-3 pp 123ndash142 2003
[9] T Aburjai M Hudaib R Tayyem M Yousef and M QishawildquoEthnopharmacological survey of medicinal herbs in Jordanthe Ajloun Heights regionrdquo Journal of Ethnopharmacology vol110 no 2 pp 294ndash304 2007
[10] M Hussain and A Ghani ldquoHerbal remidies used for gastroin-testinal disorders in Kaghan valleyrdquo Pakistan Journal of WeedScience Research in SearchWorks vol 14 no 3-4 pp 169ndash2002008
[11] F Haq H Ahmad andM Alam ldquoTraditional uses of medicinalplants of Nandiar Khuwarr catchment (District Battagram)Pakistanrdquo Journal of Medicinal Plants Research vol 5 no 1 pp39ndash48 2011
[12] M I Shinwari and M A Khan ldquoFolk use of medicinalherbs of margalla hills National Park Islamabad rdquo Journal ofEthnopharmacology vol 69 no 1 pp 45ndash56 2000
[13] R Sarwar U Farooq A Khan et al ldquoEvaluation of antioxidantfree radical scavenging and antimicrobial activity of Quercusincana Roxbrdquo Frontiers in Pharmacology vol 6 article no 2772015
[14] J Harit A Barapatre M Prajapati K R Aadil and S SenapatildquoAntimicrobial activity of rhizome of selected Curcuma varietyrdquoInternational Journal of Life Sciences Biotechnology and PharmaResearch vol 2 pp 183ndash189 2013
[15] V Bobbarala V R Vadlapudi and C Naidu ldquoAntimicrobialpotentialities of mangrove plant Avicennia marinardquo Journal ofPharmacy Research vol 2 no 6 2009
[16] T N Misra R S Singh J Upadhyay and R SrivastavaldquoIsolation of a natural sterol and an aliphatic acid fromVernoniacinereardquo Phytochemistry vol 23 no 2 pp 415ndash417 1984
[17] K Krohn U Farooq U Florke et al ldquoSecondary metabolitesisolated froman endophytic Phoma sp -Absolute configurationof tetrahydropyrenophorol using the solid-state TDDFT CDmethodologyrdquo European Journal of Organic Chemistry no 19pp 3206ndash3211 2007
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
BioMed Research International 5
of oak acorns from Serbiardquo Food Chemistry vol 104 no 2 pp830ndash834 2007
[7] L Viegi A Pieroni P M Guarrera and R Vangelisti ldquoA reviewof plants used in folk veterinary medicine in Italy as basis fora databankrdquo Journal of Ethnopharmacology vol 89 no 2-3 pp221ndash244 2003
[8] M L Leporatti and S Ivancheva ldquoPreliminary comparativeanalysis of medicinal plants used in the traditional medicine ofBulgaria and Italyrdquo Journal of Ethnopharmacology vol 87 no2-3 pp 123ndash142 2003
[9] T Aburjai M Hudaib R Tayyem M Yousef and M QishawildquoEthnopharmacological survey of medicinal herbs in Jordanthe Ajloun Heights regionrdquo Journal of Ethnopharmacology vol110 no 2 pp 294ndash304 2007
[10] M Hussain and A Ghani ldquoHerbal remidies used for gastroin-testinal disorders in Kaghan valleyrdquo Pakistan Journal of WeedScience Research in SearchWorks vol 14 no 3-4 pp 169ndash2002008
[11] F Haq H Ahmad andM Alam ldquoTraditional uses of medicinalplants of Nandiar Khuwarr catchment (District Battagram)Pakistanrdquo Journal of Medicinal Plants Research vol 5 no 1 pp39ndash48 2011
[12] M I Shinwari and M A Khan ldquoFolk use of medicinalherbs of margalla hills National Park Islamabad rdquo Journal ofEthnopharmacology vol 69 no 1 pp 45ndash56 2000
[13] R Sarwar U Farooq A Khan et al ldquoEvaluation of antioxidantfree radical scavenging and antimicrobial activity of Quercusincana Roxbrdquo Frontiers in Pharmacology vol 6 article no 2772015
[14] J Harit A Barapatre M Prajapati K R Aadil and S SenapatildquoAntimicrobial activity of rhizome of selected Curcuma varietyrdquoInternational Journal of Life Sciences Biotechnology and PharmaResearch vol 2 pp 183ndash189 2013
[15] V Bobbarala V R Vadlapudi and C Naidu ldquoAntimicrobialpotentialities of mangrove plant Avicennia marinardquo Journal ofPharmacy Research vol 2 no 6 2009
[16] T N Misra R S Singh J Upadhyay and R SrivastavaldquoIsolation of a natural sterol and an aliphatic acid fromVernoniacinereardquo Phytochemistry vol 23 no 2 pp 415ndash417 1984
[17] K Krohn U Farooq U Florke et al ldquoSecondary metabolitesisolated froman endophytic Phoma sp -Absolute configurationof tetrahydropyrenophorol using the solid-state TDDFT CDmethodologyrdquo European Journal of Organic Chemistry no 19pp 3206ndash3211 2007
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
