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estudios fitoquimicos
   Original Article   PHYTOCHEMICALS, ANTIOXIDANT ACTIVITY AND PHENOLIC PROFILING OF DIPLOCYCLOS PALMATUS   (L! C EFFERY #SMANGANI A ATTAR, SAVALIRAM G GHANE$ De%art&ent ' )'tan*, S+ia-i #ni*, /'l+a%0r 123441, Ma+ara.+tra, In5ia E&ail6 .gg7'tan*80ni.+ia-iacin Received: 02 Jan 2017 Revised and Accepted: 14 e! 2017    A)STRACT   O7-ectie6 The aim of this study was to analyze phytochemicals, antioxidant potential and phenolic profiling of leaf and fruit extracts of Diplocyclos  palmatus .   Met+'5.6 The leaves and fruits were subjected for sequential extraction with hexane, chloroform, methanol and water. All extracts were subjected to biochemical studies such as phenols, tannins, flavonoids, terpenoids and antioxidant assays such as 1,1diphenyl1picryl hydrazyl ! ##$%, &,&' Azinobis !(ethylbenzothiozoline)sulfonic acid !A*T+%, ferric reducing antioxidant property !-A#%, metal chelating and phospho molybdenum reduction assay. urther methanolic extract was used for phenolics characterization by reversed phasehigh performance liquid chromatography !-#$#/%. Re.0lt.6 0t was observed that methanol fruit extract showed significantly higher phenolics !.&23.31 mg tannic acid equivalent !TA4%5g extract%, flavonoids !16.3&23.) mg catechin equivalent !/4%5g extract% and terpenoids !&7).7(23.7) mg ursolic acid equivalent !8A4%5g extract%. $owever, chloroform extracts of leaf and fruit exhibited a high amount of tannins !&&.3723.3), ).23.13 mg /45g extract% respectively. The extracts were subjected to assess their antioxidant potential using various in vitro  systems such as ##$, A*T+, -A#, metal chelating and phospho molybdenum reduction. Among the various extracts, methanol fruit extract had highest ##$ radical scavenging activity !&).7(23.19 mg ascorbic acid equivalent !AA4%5g extract%, metal chelating activity !3.:323.31 mg 4 TA equivalent !44%5g extract% and phospho molybdenum activity !&1.&92&.1 mg AA45g extract%. 0n A*T+ radical scavenging assay, aqueous leaf extract !1&.1123.37 mg trolox equivalent !T4%5g extract% showed the best response. The effective ferric reducing antioxidant property !191.69213.1& mg e !00%5g extract% was exhibited by aqueous fruit extract. ;verall, methanol and water were found to be the best solvents for the extraction of antioxidant compounds from fruit and leaf. 0n the -#$#/ analysis, the major bioactive phenolic compounds such as catechin !/A% and hydroxybenzoic acid !$*A% were recorded in leaf as compared to fruit. 0n leaf, /A and chlorogenic acid !/A% were principal compounds in leaf and fruit respectively. $owever, gallic acid !<A%, $*A, /A and vanillic acid !=A% were widespread in leaf and fruit.  C'ncl0.i'n6 ;n the basis of the results, it was found that D. palmatus  may serve as a novel and rich source of natural antioxidants and it can be further explored for pharmaceutical purposes. /e*9'r5.6 Diplocyclos palmatus , Antioxidant, -eactive oxygen species, -#$#/ > &317 The Authors. #ublished by 0nnovare Academic +ciences #vt td. This is an open access article under the // *? license !http@55creativecommons.org5licenses5by59.35% ;0@ http@55dx.doi.org513.&&165ijpps.&317vi9.1):1 INTROD#CTION #lants have been used as a source of food and medicine from the beginning of civilization. They are a diverse source of nutrients and bioactive compounds that are valuable for human health. any ethanobotanical surveys and traditional Bnowledge of medicinal plant has been proved their significance in curing various diseases C1D. 0n recent year, interest has been focused on plantbased antioxidant compounds due to their safer and nontoxic nature. #lants contain various phytochemicals that showed antioxidant properties liBe phenolics, flavonoids, carotenoids, vitamins, terpenoids and nitrogenous compounds C&D. Antioxidants are those compounds which have the ability to neutralize the harmful effects of reactive oxygen species !-;+% and protect the biological system from possible deterioration C(D. <enerally, -;+ are generated in both plant and animal as a part of normal cell metabolism and play an important role in various cell signaling pathway and gene expression. ;verproduction of -;+ leads to oxidative stress and subsequent damages of biological molecules such as lipids, proteins and nucleic acids C9D. urthermore, they also play a significant role in development and progression of many chronic diseases liBe ageing, arthritis, carcinogenesis, diabetes, mellitus, cardiovascular, respiratory, Bidney, malignant tumors, cataracts, hypertension, atherosclerosis, #arBinson's and Alzheimer's diseases C6D. Among the natural antioxidants, phenolics are widely distributed a group of compounds in the plant Bingdom. +everal studies have proved that phenolic compounds are directly associated with the antioxidant activities and have diverse health benefits in the prevention of various oxidative stress related diseases C)D. Diplocyclos palmatus !amily /ucurbitaceae% is neglected medicinal plant commonly called as +hivalingi and ollipop climber. eaves and fruits are eaten as a vegetable in Eenya and +outh4ast Asia. The different parts of the plant have been used by the traditional practitioner for curing various ailments. The fruits are laxative, expectorant and mainly used in reproductive medicines to treat impotency, female infertility and leucorrhoea. +eeds are aphrodisiac, febrifuge, antiinflammatory and used for promoting conception in women C7, :D. 0t is also used in the treatment of a cough, flatulence, rheumatic pain, asthma, sBin diseases and snaBes bite. #lant possess gynaecological, anticonvulsant, antivenom, antidote, antimicrobial, antidermatophytic, analgesic and antiarthritic properties C1&D. 0n earlier studies, worB has been done mostly on pharmacological activities, but no reports are available on its phytochemicals and antioxidant properties. 0n this respect, the aim of the present investigation was to analyze the chemical constituents !phenolics, tannins, flavonoids, terpenoids% and antioxidant activities such as ##$, A*T+, -A#, metal chelating and phospho molybdenum reduction using leaf and fruit extracts . urthermore, characterization of phenolics was also carried out using reversed phasehigh performance liquid chromatography !-#$#/% from methanolic extracts of leaf and fruit.   International Journal of Pharmacy and Pharmaceutical Sciences ISSN- 0975-1491 Vol 9, Issue 4, 2017  G+ane et a #Int J P$a%& P$a%& Sci' () *' Iss+e 4' 101,10-   102 MATERIALS AND METHODS C+e&ical. an5 reagent. <A, /A, $*A, /A, =A, 8A and trolox were purchased from +igmaAldrich !+t. ouis, o, 8+A%. ##$, A*T+, ethylenediamine tetraacetic acid !4 TA%, &,9,)tripyridyl&triazine !T#TF%, ascorbic acid, ferrous chloride, ferrozine, ferric chloride, ferrous sulphate, sodium phosphate, ammonium molybdate, hydrochloric acid, sulfuric acid and solvents such as hexane, chloroform, ethanol, methanol were purchased from +isco research laboratory #vt. td., umbai. All the reagents used were of analytical grade. Pre%arati'n ' .'lent e:tract. eaves and fruits were collected freshly during the month Govember &319 from Eolhapur, aharashtra, 0ndia. The location lies between G1)H93'61.I atitude and 479H16'1(.((I ongitude. /ollected plant samples were washed under running tap water and dried in hot air oven at 63 J/ for 7& h. ;ven dried plant samples were ground into fine powder using spice mill. #owdered samples were used for sequential extraction with hexane, chloroform, methanol and water using maceration method. About &3 g of powder was added in a Bnown volume of respective solvent and allowed for shaBing !1& h, 133 rpm, (3 J/% on orbital shaBer incubator !Allied +cientific #roducts, 0ndia%. After filtering through filter paper, centrifuged at 6,333 rpm for 16 min to remove debris. The supernatant was Bept at room temperature for complete evaporation of the solvent. inally, dried extracts were weighed, dissolved in a Bnown volume of respective solvents and Bept at 9 J/ until used for analysis. The extract recovery from various solvent was expressed in percent !K%. T'tal %+en'lic. Total phenolics content were determined by the method described by +ingleton and -ossi C1(D. A Bnown volume of plant extract !133 Ll% was mixed with & ml of prediluted !1@13% olin and /iocalteu reagent. After 6 min incubation at room temperature, 3.: ml of !7.6 K w5v% sodium carbonate !7.6K w5v% was added. The reaction mixture was shaBen thoroughly and allowed to stand for )3 min at room temperature. The absorbance was taBen at 7)6 nm using 8=visible spectrophotometer !+himadzu 8=1:33, Mapan%. A calibration curve was plotted using a standard solution of tannic acid !&31&3 Lg5ml%. Total phenolics were expressed as mg TA45g extract !- &  N 3.:%. T'tal tannin. Total tannins were estimated by using the vanillin$/l method with minor modification C19D. #lant extract !133 Ll% were mixed separately with & ml of reagent consist of 9 K !w5v% vanillin in methanol and : K !v5v% $/l in methanol !1@1 ratio%. The mixture was allowed to stand for &3 min incubation, and the absorbance of samples was measured at 633 nm. /atechin solution !63(33 Lg5ml% was used for the preparation of calibration curve. The amount of total tannins content was expressed as mg /45g extract !- &  N 3.3%. T'tal la'n'i5. Total flavonoids were estimated by the following method of +aBanaBa et al.  C16D. #lant extract !63 Ll% was taBen into a test tube, and volume was made to !3.6 ml% with distilled water followed by addition of 76 Ll of 6 K !w5v% sodium nitrite solution. After ) min incubation, 163 Ll of 13 K !w5v% aluminium trichloride solution was added and incubated at room temperature for 6 min. urther, 3.6 ml of 1  sodium hydroxide was added and final volume made up to & ml with distilled water. The absorbance was read immediately at 613 nm. ifferent concentrations of !63(33 Lg5ml% of catechin were used to prepared standard curve. The amounts of total flavonoids were expressed as mg /45g extract !- &  N 3.3%. T'tal ter%en'i5. Total terpenoids content was determined by the spectroscopic method described by /hang and in C1)D. #lant material !133 Ll% from each solvent extracts was mixed with 163 Ll freshly prepared 6 K !w5v% vanillin in glacial acetic acid and 633 Ll perchloric acid. All the reaction mixture was heated for 96 min at )3 J/ then cooled in ice bath. urther, &.&6 ml glacial acetic acid was added to the reaction mixture and absorbance was measured at 69: nm. 8A !&6633 Lg5ml% was used for the preparation of calibration curve. The amount of total terpenoids content was expressed as mg 8A45g extract !- &  N 3.:7%.  Anti':i5ant a..a*. DPPH; ra5ical .caenging actiit* ##$ radical scavenging activity was measured according to the method of *randOilliams et al. C17D. A methanolic solution of ##$ !3.3&6 g5l% was prepared freshly. Aliquot of plant extract !1 ml% of different concentrations !&6(33 Lg5ml% were added to ( ml of ##$ solution. -eaction mixtures were Bept in the darB for (3 min and a decrease in absorbance was measured at 616 nm. Activities were measured using a standard curve obtained from various concentrations of ascorbic acid !&3133 Lg5ml% and it expressed in mg AA45g extract !- &  N 3.3%.    A)TS;<ra5ical .caenging actiit* A*T+ ;< radical scavenging activity was carried out using the method of -e et al.  C1:D. A*T+ radical cation produced by reacting 7 mmol aqueous solutions of A*T+ with an aqueous solution of &.96 mmol potassium per sulfate. urther reagent mixture was allowed to stand for 1&1) h at room temperature. The reagent solution was diluted in ethanol !1@: v5v% and equilibrated at (3 J/ to give absorbance at 7(9 nm of 3.723.3&. #lant extract of different concentrations !&3)3 Lg5ml% were combined with ( ml of A*T+ ; Pdiluted solution. Absorbance was recorded after (3 min at 7(9 nm. The calibration curve was obtained for trolox standard solution !&31&3 Lg5ml% and results were expressed as mg T45g extract !- &  N 3.)%. Metal c+elating actiit* The chelating of ferrous ion was estimated by the method of inis et al.  C1D. #lant extract !93 Ll% was mixed with 63 Ll of & mmol e/l &. The reaction was initiated by adding 133 Ll of 6 mmol ferrozine. The final volume of the reaction was made to 1633 Ll by adding distilled water and mixture was shaBen vigorously. The absorbance was measured at 6)& nm after &3 min incubation at room temperature. The chelating activity of the extract was estimated by using 4 TA as standard !&31&3 Lg5ml%. The results are expressed as mg 445g extract !- &  N 3.6%.  FRAP a..a* The antioxidant properties in terms of -A# were estimated according to the protocol of *enzie and +train C&3D. -A# reagent prepared freshly and incubated at (7 J/ prior to use. -A# reagent contained &.6 ml of 13 mmol T#TF in 93 mmol $/l, &.6 ml of e/l ( .)$ & ; and &6 ml of (33 mmol acetate buffer !p$ (.)%. #lant extract !163 Ll% was allowed to react with &:63 Ll -A# reagent and incubated at (7 J/ for (3 min. The increase in absorbance was measured at 6( nm. A standard calibration curve was prepared using different concentrations !&31&3 Lg5ml% of e+; 9 .7$ & ;. -esults were expressed in mg e !00% equivalent5g extract !- &  N 3.:(%. P+'.%+'&'l*75en0& a..a* The phospho molybdenum assay was performed by the method of #rieto et al.  C&1D. A Bnown volume of plant extract !133 Lg5ml% was mixed with 1 ml reagent !3.)  sulfuric acid, &: mmol sodium phosphate and 9 mmol ammonium molybdate%. The vials were capped and incubated in water bath at 6 J/ for 3 min. After cooling the samples, absorbance was measured at )6 nm against blanB. ifferent concentrations !&31&3 Lg5ml% of ascorbic acid in distilled water were used to plot calibration curve and antioxidant activity was expressed as mg AA45g extract !- &  N 3.&%. =0alitatie an5 >0antitatie anal*.i. ' %+en'lic c'&%'0n5. 7* RP?HPLC Pre%arati'n ' .a&%le an5 .tan5ar5 .'l0ti'n The dried powder of leaf and fruit !633 mg% with methanol !13 ml% were Bept on orbital shaBer incubator at 1132& rpm !&6 H/, 9: h%. The further mixture was filtered through Ohatman filter Go. 1 and centrifuged at 13,333 rpm for &3 min at 9 H/. +upernatant was  G+ane et a #Int J P$a%& P$a%& Sci' () *' Iss+e 4' 101,10-   103 collected and stored at 9 H/ until $#/ analysis. #henolic compounds such as <A, /A, $*A, /A, =A and coumaric acid !/;A% !+igmaAldrich, +t. ouis, o, 8+A% were weighed accurately and dissolved separately in methanol to obtain a standard stocB solution !1 mg5ml%. urther, worBing stocB solution was prepared by diluting the stocB solution with methanol to get five different concentrations for calibration curves. All the stocB and worBing stocB solutions were stored at 9 J/ until further use. RP?HPLC anal*.i. -#$#/ analysis of phenolic compounds was performed on Masco chromatographic system !odel no. /&333 #lus% equipped with a quaternary pump, autosampler, and 8= detector !8= &373%. The separation was performed using $iber /1: column !6 Qm, &639, ) mm%. The built in /hromGA= software system was used for data processing. The mobile phase consisted of water@ acetonitrile@ glacial acetic acid !3@6@6% was used with 3. ml5min flow rate and &3 Ql injection volume. The peaBs were monitored at &:3 nm with 63 min as run time. The identification of phenolic compounds in extracts was performed by comparing their retention times with those of standards. 0dentified phenolics were confirmed by spiBing with Bnown concentration of the respective standard. A standard curve of selected phenolics with four different concentrations !&6R&33 Lg5ml% was prepared and expressed as micrograms per gram of dry weight !mg5g O%. The system suitability was assessed by triplicate injection of standard solutions and extracts. The peaB areas of three replicate injections of standard solutions and extracts were considered to evaluate the repeatability of the method. All solutions !mixed standards, samples and spiBed solutions% were filtered through 3.&& Lm nylon syringe filter. Stati.tical anal*.i. All the analyses were done in triplicate and values are represented as mean2standard error !+4%. The data were subjected to oneway analysis of variance !AG;=A% and significant differences between mean values were determined by uncan's multiple range test ! -T% !  p S3.36% using +#++ version 1).3. RES#LTS AND DISC#SSION T'tal %+en'lic. #henolics are a natural group of secondary metabolites that effectively scavenge free radicals from a biological system. 0t was found that high content of phenolics linearly correlated with high antioxidant activity C&&D. #henolics have been receiving much more attention among researcher and consumer for its diverse health benefits. Apart from these, it possesses anticarcinogenic, antiatherogenic, antiinflammatory, antimicrobial properties C), &(D. 0n the present investigation, we found that methanol extract of leaf and fruit showed higher phenolic content !7.6123.3: and .&23.31 mg TA45g extract% !table 1%. Aqueous extract of leaf and fruit exhibited moderate and hexane and chloroform extract showed least phenolics content. The results are agreed with Eumari C&9D who found that methanol extract of Pithecellobium dulce  leaves recorded relatively higher level of total phenols !::.& mg <A45g dry weight%. T'tal tannin. Tannins are high molecular weight phenolics with multiple biological activities liBe metal ion chelation, protein precipitation and considered as natural antioxidants. 0t has been reported that tannins are more effective antioxidant than low molecular weight phenolics C&6D. Ta7le 26 E:tract *iel5, t'tal %+en'lic., t'tal tannin., t'tal la'n'i5. an5 t'tal ter%en'i5. c'ntent ' lea an5 r0it e:tract. ' D# pa &at+ P+*t'c+e&ical. Lea e:tract Fr0it e:tract He:ane C+l'r''r& Met+an'l A>0e'0. He:ane C+l'r''r& Met+an'l A>0e'0. 4xtract yield 1  1.7) 3. 1.63 .(7 &.17 3.:6 1.&6 :.6 Total phenolics &  3.))23.31 e  3.:623.3& d  7.6123.3: b  1.7923.33 c  3.()23.33 f   3.)723.33 e  .&23.31 a  1.:&23.33 c  Total tannins (  9.:(23.39 d  &&.3723.3) a  7.6)23.&1 b  9.6)23.13 d  3.7323.36 f   ).23.1( c  &.723.36 e  3.923.39 f   Total flavonoids (  3.:23.1& f   &.9923.&7 e  7.623.&1 c  .6623.)6 b  1.:)23.3: e  &.6123.(3 e  16.3&23.) a  9.(:23.3& d  Total terpenoids 9 11&.)32(.(7 e  1:).392&.9: c  &&7.7721.7: b  (:.&621.6) g  :9.)(21.&) f   1(7.&32&.&: d  &7).7(23.7) a &.923.) h   1 K of dry extract, &  mg TA45g extract, (  mg /45g extract, (  mg 8A45g extract. =alues are the means of three replicates2+4. ean values with different alphabets showed statistically significant differences !p3.36% according to -T. 0n our results, irrespective of solvent used the leaf extract showed higher tannins content than fruit extract except for chloroform !table 1%. Total tannin content of chloroform leaf extract was higher !&&.3723.3) mg /45g extract% than methanol, hexane and aqueous leaf extract. +imilarly, chloroform fruit extract contained significantly higher tannins !).23.13 mg /45g extract% than other fruit extracts. The results are comparable with Cucumis dipsaceus  fruit C&)D. T'tal la'n'i5. lavonoids are a most common group of antioxidants. ue to their high redox potential, they act as singlet oxygen quenchers, reducing agents, hydrogen donors and metal chelator C&7D. 0n our analysis, fruit extract showed higher flavonoids content than leaf except for aqueous leaf extract !table 1%. Aqueous extract of leaf and methanol extract of fruit showed higher flavonoids content !.6623.)6 and 16.3&23.) mg /45g extract% compared to other extract. Archana and =ijayalaBshmi C&:D recorded a considerable amount of flavonoids in ethanol root extract of Glycyrrhiza glabra.   T'tal ter%en'i5. Terpenoids are the largest and diverse group of compounds and served as a growth regulator, pollinator attractants, antifeedants, toxins and antioxidant C&D. any of which are efficient in the treatment of human diseases liBe cancer, malaria, inflammation and a variety of viral and bacterial diseases C(3D. 0n present study, highest terpenoids were recorded in methanol leaf and fruit extract !&&7.7721.7: and &7).7(23.7) mg 8A45g extract% while least were observed in aqueous leaf and fruit !(:.&621.6) and &.923.) mg 8A45g extract respectively% !table 1%.  Anti':i5ant a..a*. DPPH ra5ical .caenging actiit* Fig   26 DPPH; ra5ical .caenging actiit* ' lea an5 r0it e:tract. ' D# pa &at+s  Val0e. 9ere t+e &ean. ' t+ree re%licate.@SE The method is based on the reduction of ##$ radical by antioxidant and then form of more stable ##$ molecule C17D.  G+ane et a #Int J P$a%& P$a%& Sci' () *' Iss+e 4' 101,10-   104 ethanolic solution of ##$ is with deep violet color that showed a strong absorption at 617 nm. The absorption decreases as a result of a change in color from violet to yellow due to ##$ radical scavenged by the plant antioxidant. 0n the present study, among leaf extracts, water !&6.3123.37 mg AA45g extract% showed a higher level of radical scavenging activity. ;n the other hand, in fruit, methanol extract found better !&).7(23.19 mg AA45g extract%. 0n leaf, the order of scavenging capacity in the extract was aqueousUchloroformU hexaneUmethanol. $owever, in fruit extract, the order of scavenging capacity was methanolUaqueousU chloroformUhexane !fig. 1%. ;ur results are also in good agreement with Eumari C&9D who confirmed that methanol extract of Pithecellobium dulce  leaves !0/ 63  79.: Lg5ml% showed significant ##$ radical scavenging activity.  A)TS ra5ical .caenging actiit* Fig   6 A)TS;<ra5ical .caenging actiit* ' lea an5 r0it e:tract. ' D# pa &at+s  Val0e. 9ere t+e &ean. ' t+ree re%licate.@SE A*T+ radical scavenging assay is an excellent tool for determining the antioxidant activity C(1D. 0n this assay, the green color of A*T+ radical solution gradually vanishes and became colorless due to the reduction of A*T+ radical by antioxidant C1:D. 0n our study, methanol and aqueous extracts of leaf showed higher radical scavenging capacity !:.:123.9 and 1&.1123.37 mg T45g extract%. +imilarly, fruit extracted with methanol and water showed highest radical scavenging activity !11.:123.37 and 13.)23.17 mg T45g of extract% while chloroform and hexane extract registered with least radical scavenging activity !fig. &%. These results are concurred with Givedhini et al.  C&)D wherein methanol extract of Cucumis dipsaceus  fruit showed higher A*T+ radical scavenging activity against chloroform. Metal c+elating actiit* The metal ion is a natural constituent of every biological system, but excess or exposure to certain metal ions induces the generation of -;+. These -;+ causes oxidative deterioration of biological macromolecules and enhances risB of various diseases. $ence, antioxidants protect the biological system either by trapping free radicals or by chelating metal ion C(&D. Fig   B6 Metal c+elating actiit* ' lea an5 r0it e:tract. ' D#  pa &at+s# Val0e. are t+e &ean. ' t+ree re%licate.@SE Mean al0e. 9it+ 5ierent al%+a7et. .+'9e5 .tati.ticall* .igniicant 5ierence. (% 44! acc'r5ing t' DMRT 0n the present investigation, aqueous leaf extract showed higher activity !3.&923.331 mg 445g extract%. Among fruit extracts, methanol exhibited higher activity !3.:323.33: mg 445g extract%. TaBing everything into account, the fruit extracts showed highest metal chelating activity !fig. (%. +imilarly, Givedhini et al.  C&)D reported that methanol extract of Cucumis dipsaceus  fruit exhibited higher chelating activity !1&.9 mg 4 TA equivalent5g extract%. FRAP a..a* -A# assay is based on the ability of antioxidant compounds to reduce the T#TFe!000% complex to the intensely blue colored T#TFe!00% complex in acidic medium with maximum absorption at 6( nm. Fig   16 FRAP acti,it* ' lea an5 r0it e:tract. ' D# pa &at+s  Val0e. are t+e &ean. ' t+ree re%licate.@SE Mean ,al0e. 9it+ 5ierent al%+a7et. .+'9e5 .tati.ticall* .igniicant 5ierence. (%44! acc'r5ing t' DMRT <enerally reducing properties are related with the presence of reductant compounds that inactivate the oxidant by donating electrons C((D. The results indicated that aqueous extract of both leaf and fruit exhibited significantly higher -A# value !11:.32).(7 and 191.69213.1& mg e !00% equivalent5g extract% compared to other extracts. ethanolic extracts of leaf and fruit registered with moderate -A# value !69.923.76 and 6.729.(3 mg e !00% equivalent5g extract%. $owever, lower -A# values were recorded in the chloroform and hexane leaf and fruit extracts !fig. 9%. 0rshad et al.  C(9D reported that aqueous fruit extract of Luffa cylindrica showed significant antioxidant activity than other cucurbits extracts. P+'.%+'&'l*75en0& a..a* The antioxidant capacities of different extracts were evaluated and results are depicted in fig. 6. 0t is based on the reduction of o !=0 to o =% by antioxidants and subsequent formation of a green color phosphateo !=% complex that show maximum absorption at )6 nm C&1D. 0n our study, leaves extracted with water showed significantly higher antioxidant activity !1((.6)2(.33 mg AA45g extract% than another solvent extract. ;n the other hand, in fruit, methanol extract showed highest antioxidant activity !&1.&92&.1 mg AA45g extract%. 0n leaf, the order of antioxidant capacity was aqueousUmethanolUchloroformUhexane. Fig   6 P+'.%+'&'l*75en0& re50cti'n a..a* ' lea an5 r0it e:tract. ' D# pa &at+s  Val0e. are t+e &ean. ' t+ree re%licate.@SE Mean ,al0e. 9it+ 5ierent al%+a7et. .+'9e5 .tati.ticall* .igniicant 5ierence. (%44! acc'r5ing t' DMRT
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