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1 August 30, 2016 Archives 2016 vol EVALUATION OF THE ANTIOXIDANT CAPACITY AND CONTENT OF POLYPHENOLS OBTAINED FROM TEA (Cmelli Sinensis) OF FOUR BRANDS SOLD IN COLOMBIA BY EXTRACTION AT ROOM TEMPERATURE Rmiréz-Aristizl, L. S. 1* ; Ortiz, A. 2 ; Restrepo-Aristizl, M.F. 1 ; Slins-Villd, J.F. 1 1 Universidd Tecnológic de Pereir, Crrer 27 #10-02 Brrio Almos Risrld, Colomi 2 Centro Ncionl de Investigciones de Cfé Cenicfé Sede Plnlto, km. 4 ví Chinchiná-Mnizles. Mnizles (Clds) - Colomi * lurmire@utp.edu.co Astrct Herl medicines hve long een used to tret chronic diseses such s cncer, neurodegenertion nd dietes, usully in the form of herl tes, lso clled tisnes [1]. Te, second only to wter, is the most highly consumed everge worldwide nd its medicinl nd helth properties, long known to erly Chinese civiliztions, hve een widely explored [2]. The consumption of green te in Colomi is recent trend nd the mrket is continuously growing, then the most common commercilly ville types of green te were tested in this study; Orientl, Lipton, Hindú nd Jiel. The min ojective of this study ws to determine nd compre the mount of polyphenols present in te smples considering extrction in wter t room temperture nd stirring every 30 seconds for 5 minutes. The ntioxidnt cpcity ws determined y the 1,1- diphenyl-2-picrylhydrzyl (DPPH) free-rdicl scvenging ssy nd the oxygen rdicl sornce cpcity (ORAC) ssy. Concentrtions rnging from ± 0.98 to ± 0.86 mg Trolox equivlents / g dry smple for DPPH ssy nd ± 1.31 to ± 2.05 mg Trolox equivlent / g dry simple were determined for ORAC ssy. It ws lso found tht the mount of totl phenols in te smples rnging from 2.53 ± 0.25 to ± 0.53 mg gllic cid equivlent (GAE) / g dry smple nd totl flvonoid content concentrtion ws otined from 2.67 ± 0.20 to 7.08 ± 0.38 mg ctechin equivlent / g dry simple. The ntioxidnt ctivity nd totl flvonoid content were highly correlted for oth DPPH (r 2 = ) s ORAC (r 2 = ). Te extrcts from the Orientl rnd hd the highest polyphenol content showing greter iologicl ctivity, contrry to Jiel te rnd which recorded the lowest concentrtions in ll nlyzes. Key words: Antioxidnt ctivity, cmelli sinensis, green te, room temperture

2 PhOL Rmiréz-Aristizl et l 2 (1-9) Introduction Green te (Cmelli sinensis) is well known for vrious helth enefits ssocited with risk reduction of wide rnge of chronic diseses, such s cncer, dietes, nd crdiovsculr diseses [3]. In the pst few yers, specil ttention hs een pid to the ntioxidnt ctivities of the polyphenolic compounds present in green te due to their phrmceuticl properties. The performed studies report tht green te extrct (GTE) shows mny helth eneficil properties, prticulrly ginst the dmge cused y pollution, stress, cigrette smoke nd other toxins, prevention of crdiovsculr diseses, cncer, dietes [4]. Te is rich in polyphenolic compounds, nmely the ctechins, (+)-ctechin (C), ( )-epictechin (EC), ( )- epictechin gllte (ECG), ( )-epiglloctechin (EGC), nd ( )-epiglloctechin gllte (EGCG) re thought responsile for such eneficil effects [5]. The methods of prepring the everge lso vry worldwide: in Chin, te leves re steeped in hot wter (70 80 C for green te) for s, nd the sme te leves re usully repetedly steeped (seven times). The Jpnese usully prepre green te y steeping leves in hot wter for out 2 min nd using them for 2 3 infusions. In the United Kingdom, Irelnd nd in Cnd, lck te is mostly prepred using oiling wter nd consumed with milk nd often sugr. Americns re lrge consumers of iced te which is mde from hot te cooled with ice. In recent times in Tiwn, especilly in summer, cold wter (4 or 25 C) steeping is new populr wy for mking te [6]. Severl studies hve een done on the vrious rewing conditions to mximise the extrction of these components from green te. The results hve defined optiml wter rewing conditions including the temperture eing mintined t 80 8C for 30 min nd rtio of te to wter of 1:20 g/ml [7]. However, these optimistion studies focused on loose-lef green te nd were crried out under lortory conditions. These conditions re very different from household rewing hits, where te is simply rewed in oiled wter nd left t room temperture for short time (3 min) efore eing consumed [8]. This study is imed to investigte whether the ntioxidnt cpcity of different types of te could e ffected y steeping te gs in cold wter during short time of steeping nd tking into ccount household hits. Methods Te Smples A set of four commercil rnds of green te (Orientl, Lipton, Hindú) nd comintions of green te with certin fruits nd romtic flowers (Jiel with Hiiscus sdriff nd lemon). They were used in the experiment for comprison of cold te infusions. These te smples were purchsed t the locl supermrket nd nlyzed in triplicte ccording with the temperture of infusion. Smple preprtion In order to simulte everge rewing, ech rnd of te were prepred using n queous extrction procedure, to study the effect of different extrction condition on the totl phenols, flvonoids nd ntioxidnt ctivity. Cold te infusions were prepred y dding 150 ml of potle wter t room temperture to teg tht ws weighed (tle 1) nd leving the infusions to stnd t room temperture (27 28 C) for 5 minutes, gitting mnully every minute nd controlling it s temperture every 30 seconds. See figure 1. Three extrctions of ech te rnds (Orientl, Lipton, Hindú nd Jiel) were performed for three consecutive dys for cold extrct. Just three extrctions for ech of the four rnds were retined y type of infusion. Smples were diluted t ppm in distilled wter nd stored t 4 C for susequent nlysis. Totl phenol content (TPC) Totl phenol content of the different te smples ws determined ccording to the Folin-Cioclteu method descried y V. L. Sigleton, R. Orthofer nd R.M. Lmuel-Rventós [9]. The smples were diluted 1:10 in distilled wter, 160 μl of ech smple, nd stndr were dded to polystyrene cells of 3000 μl. Then 1420 μl Folin regent (1:10) nd 1420 μl of N2CO3 7.5% were dded nd homogenized, After mixing, the smples were left for 120 minutes t room temperture in the drk, the sornce ws red t 758 nm ccording with the spectrl scnning (SHIMADZU UV-1700 spectrophotometer). All lnks contined distilled wter nd te, the phenolic compounds were quntified using clirtion curves of Gllic cid nd expressed s mg gllic cid equivlent / g smple (mg GAE/g smple). Kinetics of the rection ws conducted for period of two hours with te smples nd stndrds to determine the stility of the complex formed ccording with time.

3 PhOL Rmiréz-Aristizl et l 3 (1-9) Totl flvonoid content (TFC) The TFC ws determined ccording to D.O Kim, K.W. Lee, H.J. Lee nd C.Y. Lee [10]. The solution is mixture of sodium nitrite solution followed y luminum thrichloride with sodium hydroxide nd distilled wter, which is mixed well nd the sornce is mesured ginst freshly prepred regent lnk. The extrct ws rected with luminum chloride for determintion of flvonoid content. A pink color ppers fter few minutes which indicte the presence of flvonoids ccording with J. Zhishen, T. Mengcheng, nd W. Jinming [11]. Briefly, 600μL of distilled wter ws dded to 150 μl of ech smple of hot nd cold extrction te nd stock solution of ctechin intro polystyrene cells of 3000 μl, 10 μl of 5% sodium nitrte (NNO3) ws dded. The mixture ws incuted in the drk for 5 minutes. A 45 μl volume of luminum chloride (AlCl3, 10%) ws dded to the mixture nd incuted for further 6 minutes in the drk. A 300 μl of NOH, 1M, ws dded to the resulting mixture followed y ddition of 360 μl of distilled wter. The sornce ws red t 505 nm ccording with the spectrl scnning (SHIMADZU UV-1700 spectrophotometer). The smples were diluted 2:10 in distilled wter, ech lnk contins te extrct nd distilled wter. The flvonoids contents were quntified using clirtion curves of Ctechin nd expressed s mg Ctechin equivlent/g smple (mg CE/g smple). Kinetics of the rection ws conducted for period of one hour with te smples nd stndrds to determine the stility of the complex formed ccording with time. Antioxidnt ctivity DPPH ssy DPPH ssy ws crried out in photometric cells of 3mL tking into ccount chnges in the method of O. P. Shrm [12]. A volume of 300 µl te cold extrct (ll rnds were 10 diluted times) nd/ or trolox stndrd with their respective controls; positive control ws hydroquinone 1000 ppm; negtive control ws methnol 96%, were dded to photometric cells followed y 2700 ml of methnol solution of DPPH t 30 ppm except the lnk smple. The smples were incuted in the drk t room temperture for 30 minutes (ccording to the kinetics of the rection ws determined for the methnolic solution of DPPH nd ech of the te smples nlyzed: Orientl, Hindú, Jiel nd Lipton) nd red t wvelength of 517 nm y spectrophotometer (SHIMADZU UV spectrophotometer). The scvening cpcity ws clculted s: % A.A= [A c (-) A te /Ac (-) ]*100, where Ac (-) is the sornce of the control nd A te is the sornce of the tested smple. Trolox ws used s stndrd. Free rdicl scvenging cpcities of te were expressed s mg Trolox Equivlent/ g smple. (mg TE/ g smple). Kinetics of the rection ws conducted for period of one hour with te smples nd stndrds to determine the stility of the complex formed ccording with time. ORAC ssy The ntioxidnt ctivity of te smples were lso nlyzed y ORAC (Oxigen Rdicl Asornce Cpcity) ccording to the method of K. M. Gillespie [13]. Dilutions of te extrcts rnging from 80 to 50 diluted times in PBS t 75 mm hving te cold extrcts (Orientl nd Lipton: 80 diluted times; Hindú nd Jiel: 50 diluted times). The diluted smples were dded to 96 wells with solid white plte followed y 187 ul of 80 nm fluorescein diluted in PBS (phosphte uffered sline) t 75 mm. After 15 minutes incution in the drk t 37 C solution of 140 mm AAPH diluted in 75 mm PBS is prepred nd dded to ech well of the plte; AAPH is responsile for strting the decy of fluorescein so it is importnt to hve fst nd strict control of the ddition, recording dt every 120 seconds until decy of fluorescein, using n emission wvelength of 493 nm nd filter excittion 515 nm using fluorescence spectrophotometer (Vrin, Cry Eclipse, version 1.1 (135)). The net AUC (re under the fluorescence decy curve) for ech smple/stndrd ws otined y sutrcting the re of the lnk smple (PBS). Antioxidnt ctivity ws expressed s mg Trolox equivlent/ g smple (mg TE/g smple) using the liner regression vlue otined from the trolox clirtion curve. Results Totl phenol content (TPC) The totl phenolic content of the 4 rnds green te re shown in figure 2 nd tle 2, were quntified using clirtion curves of gllic cid (1-30 mg/l) performed every dy of the ssy. The totl phenolic compounds were found etween mg gllic cid Equivlent /g smple. The highest levels were mesured in Orientl rnd (14.63 ± 0.53 mg equivlent gllic cid / g smple), similr mounts were lso otined in Lipton (10.38 ± 0.55 mg gllic cid equivlent / g smple) nd Hindú (9.89 ± 0.66 mg gllic cid equivlent / g smple), while Jiel contined the lowest mount (2.53 mg gllic cid equivlent / g smple).

4 PhOL Rmiréz-Aristizl et l 4 (1-9) Totl flvonoid content. (TFC) The totl flvonoid content of the te extrct were low compred with TPC, nd rnged from 2.67 ± ± 0.38mg ctechin equivlent / g smple, (see figure 3 nd tle 2) nd quntified using clirtion curve of ctechin (1-30 mg/l) performed every dy of the ssy. Orientl nd Hindú hd the highest levels compred to other rnds (Orientl: 7.08 ± 0.38 mg ctechin equivlent / g smple, Hindú: 6.71 ± 0.36 mg ctechin equivlent / g smple), for Lipton rnd ws otined content of 4.87 ± 0.26 mg ctechin equivlent/ g smple, while Jiel contined the lowest mount (2.67 ± 0.20 mg ctechin equivlent / g smple). Antioxidnt Activity The ntioxidnt ctivity of the te infusions ws evluted using two independent ssys, DPPH nd ORAC. A clirtion curve of trolox ( um) llowed to compre ntioxidnt ctivity in different rnds of te expressed s μmol Trolox equivlent / g dry smple. The results otined from DPPH ssy reported in the figure 4 nd tle 2, shown vlues rnged from ± ± 0.86 mg Trolox equivlent / g dry smple, similr to the vlues otined y ORAC ssy reported in the figure 5, which hve rnge from ± ± 2.05 mg Trolox equivlent / g dry smple. The Orientl nd Hindú rnd te hd the highest DPPH nd ORAC vlues while the Jiel rnd showed the lowest. Discussion The degree of oxidtion of the leves defines the type of te: white, yellow, green, oolong, pu-erh nd lck te. Green te is the lest processed, resulting from quick drying of the fresh leves, with miniml oxidtion, which mke it richer in ioctive polyphenols comprtively to more processed tes, where these compounds re degrded during the process. The consumption of te, especilly green te, hs severl wellestlished helth ene- fits, nmely the reduction of the incidence of oxidtive stress relted diseses nd crdiovsculr disorders, for exmple [14]. The helth enefits of green te re minly ttriuted to their high phenolic content, which mke these everges one of the mjor sources of helth promoting polyphenols in our diet [1]. Extrction is the initil nd the most importnt step in the recovery nd purifiction of ioctive compounds from plnt mterils. In generl, the conventionl techniques for green te extrction re heting, oiling, Soxhlet extrction nd cold extrction, which re ll limited y long extrction periods nd low extrction efficiency [15]. The results otined from the nlysis of the ntioxidnt ctivity of te infusions y E. Dmini nd T. Bcchetti [16] showed tht green te exhiited greter ctivity when steeped for 2 h in wter t room temperture.the different ffinities of the extrction solvents for totl te lef constituents in terms of their different extrction conditions, such s polrity of extrcting solvents, nd temperture ply n importnt role while investigting the phytochemicl profile nd ntioxidnt functions of te [17]. In this study the long extrction times t room temperture is not used, since the min ojective ws to simulte household te extrction in wy tht would llow to otin the polyphenolic compounds in green te using short time period. Totl phenolic content (TPC) of green te re presented en tle 2, the highest content ws Orientl (14.63 ± 0.53 mg gllid cid equivlent/g smple) nd the lowest ws Jiel (2.53 ± 0.25 mg gllid cid equivlent/g smple). In generl the four rnds showed decresing ehviour Orientl > Lipton Hindú > Jiel. An nlysis of vrince ANOVA (tukey s multiple comprison Test, significnt p< 0.05), identified significnt differences etween Orientl, Lipton nd Jiel. (,, c) nd no significnt differences were found etween Lipton nd Hindú (). E. Venditti nd T. Bcchetti compred the totl phenol content levels in hot nd cold tes using Folin Cioclteu s regent. They found tht TPC is lwys higher in hot tes thn in cold tes for green te extrcts. The exception is with white te, where TPC is significntly higher in the cold infusion thn in the hot one. In ddition, TPC, in white te prepred with cold wter steeping, is significntly higher thn in ll other tes prepred in the sme wy [6]. This indictes tht these extrcts nlysed for te is likely to e getting much smller quntities due to the extrction method. Although this only pplies to green te. As one possile reson why in our study lower content ws otined due to temperture conditions nd extrction time. The differences found etween rnds my e due to mnufcturing process used y ech industry, prticle size nd other compounds s Jiel who contins hiiscus nd lemon peel. Totl Flvonoid content (TFC) of green te re presented en tle 2, the highest content ws Orientl (7.08 ± 0.38 mg ctechin equivlent/g smple) nd Hindú (6.71 ± 0.36 mg ctechin equivlent/g smple), while the lowest ws Jiel (2.67 ± 0.20 mg ctechin equivlent/g smple). In generl the four rnds showed decresing ehviour Orientl Hindú >

5 PhOL Rmiréz-Aristizl et l 5 (1-9) Lipton > Jiel. An nlysis of vrince ANOVA (tukey s Multiple Comprison Test, significnt p< 0.05), identified significnt differences etween Orientl, Lipton nd Jiel. (,, c) nd no significnt differences were found etween Orientl nd Hindú(). (Fig. 2) Luximon-Rmm [18] showed contents from 15 to 26 mg/g for totl flvonoids in nine commercilly lck te, considering extrctions were performed t oiling temperture. With these dt reported nd compred to ours extrction continues to oserve lower content of flvonoids for green te extrcts t room temperture. The ntioxidnt ctivity of the te infusions ws ssessed using two independent ssys, DPPH nd ORAC. The DPPH method is sed on the reduction of the reltively stle rdicl 1,1-diphenyl-2-picrylhydrzyl, to the formtion of non rdicl form in the presence of hydrogen donting ntioxidnt. The te smples showed ntioxidnt ctivity y the reduction of purple colored DPPH to the yellow colored diphenylpicrylhydrzine derivtives t wvelength of 517 nm [19]. The ORAC ssy originlly developed y Glzer nd Ghiselli [20]. The ORAC ssy offers severl dvntges over DPPH ssy. It uses peroxyl rdicls tht re etter models of ntioxidnt rections with oxidizing lipids nd rective oxygen species (ROS) in foods nd in vivo, nd it provides continuous genertion of rdicls on relistic time scle (more like ctul rections in situ).the completely hydrogen tom trnsfer rdicl quenching mechnism presents contrst nd comprison to electron trnsfer in DPPH ssys. The ssy cn e dpted to detect oth hydrophilic nd hydrophoic ntioxidnts generlly or specificlly y ltering the rdicl source, solvent, nd trget molecule, nd hs een routinely utomted [21]. The ntioxidnt ctivity of green te re presented in tle 2, the highest content ws Orientl (41.29 ± 0.86 mg Trolox equivlent/g smple) nd Hindú (38.64 ± 1.72 mg Trolox equivlent/g smple) determined y DPPH ssy, while the lowest ws Jiel (22.36± 0.98 mg Trolox equivlent/g smple). In generl the four rnds showed decresing ehviour Orientl Hindú > Lipton > Jiel. An nlysis of vrince ANOVA (tukey s Multiple Comprison Test, significnt p< 0.05), identified significnt differences etween Orientl, Lipton nd Jiel. (,, c) nd no significnt differences were found etween Orientl nd Hindú (). (Fig. 3). The results otined y ORAC llowed to see tht the highest ntioxidnt ctivity ws to Orientl te (46.25 ± 2.05 mg Trolox equivlent/g smple) nd the lowest ws Jiel (22.95 ± 1.31 mg Trolox equivlent/g smple), nd their ehviour of ll smples ws Orientl > Hindú Lipton > Jiel. Antioxidnt ctivity of ll smples showed lmost the sme ehviour ssessed y two methods. According to the results otined from the nlysis performed, Jiel te rnd showed the sme results for oth ssys (TPC nd TFC). Perhps t this extrction conditions ll the phenols presented in the solution re flvonoids. See tle 2. Jiel showed the lowest levels of phenolic compounds, flvonoids nd ntioxidnt ctivity, possily due to the ddition of hiiscus nd lemon peel tht could msk other compounds [22]. Therefore from the ove results, one cn deduce tht ntioxidnt ctivity in the vrious te rnds tested is correlted with their totl flvonoid content. For DPPH ssy the correltion is given y r 2 = nd for ORAC ssy the correltion is given y r 2 = In ccordnce with the liner correltions is possile to hve some fctors tht could ese the form to get results out ntioxidnt ctivity hving previously nlyzed the totl flvonoid content. Correltion etween DPPH nd TFC hs the eqution elow: Correltion etween ORAC nd TFC hs the eqution elow: The coefficients TFC re relted to the totl flvonoid content expressed in mg gllic cid equivlent/g smple nd A.A to ntioxidnt ctivity expressed in mg Trolox equivlent/g smple. To dte, there pper to e no thorough studies on how ntioxidnt ctivity of tes my e ffected y hot or cold wter steeping nd how this my e relted to their polyphenol content. The results otined contriute to gining further knowledge on how the potentil helth enefits of this populr everge my e mximized y the different methods of preprtion. Acknowledgments The uthors would like to thnk the Universidd Tecnológic de Pereir for finncil support. References 1. Büyüklci A, El SN. Determintion of In Vitro Antidietic Effects, Antioxidnt Activities nd Phenol Contents of Some Herl Tes. Plnt Foods Hum Nutr. 2008;63(1):27-33.

6 PhOL Rmiréz-Aristizl et l 6 (1-9) 2. McKy DL, Blumerg JB. The Role of Te in Humn Helth: An Updte. J Am Coll Nutr. 2002;21(1): Hollmn PC., Ktn M. Dietry Flvonoids: Intke, Helth Effects nd Biovilility. Food Chem Toxicol. 1999;37(9-10): doi: /s (99) Mne A-M, Vsile BS, Meghe A. Antioxidnt nd ntimicroil ctivities of green te extrct loded into nnostructured lipid crriers. Comptes Rendus Chim. 2014;17(4): Rshidinejd A, Birch EJ, Everett DW. Antioxidnt ctivity nd recovery of green te ctechins in full-ft cheese following gstrointestinl simulted digestion. J Food Compos Anl. 2016;48: Venditti E, Bcchetti T, Tino L, Crloni P, Greci L, Dmini E. Hot vs. cold wter steeping of different tes: Do they ffect ntioxidnt ctivity? Food Chem. 2010;119(4): Komes D, Horžić D, Belščk A, Gnić KK, Vulić I. Green te preprtion nd its influence on the content of ioctive compounds. Food Res Int. 2010;43(1): Astill C, Birch MR, Dcome C, Humphrey PG, Mrtin PT. Fctors Affecting the Cffeine nd Polyphenol Contents of Blck nd Green Te Infusions. J Agric Food Chem. 2001;49(11): Singleton VL, Orthofer R, Lmuel-Rventós RM. Oxidnts nd Antioxidnts Prt A. Elsevier; Kim D-O, Lee KW, Lee HJ, Lee CY. Vitmin C Equivlent Antioxidnt Cpcity (VCEAC) of Phenolic Phytochemicls. J Agric Food Chem. 2002;50(13): Zhishen J, Mengcheng T, Jinming W. The determintion of flvonoid contents in mulerry nd their scvenging effects on superoxide rdicls. Food Chem. 1999;64(4): Shrm OP, Bht TK. DPPH ntioxidnt ssy revisited. Food Chem. 2009;113(4): Gillespie KM, Che JM, Ainsworth EA. Rpid mesurement of totl ntioxidnt cpcity in plnts. Nt Protoc. 2007;2(4): Shhidi F. Antioxidnt fctors in plnt foods nd selected oilseeds. BioFctors. 2000;13(1-4): Pn X, Niu G, Liu H. Microwve-ssisted extrction of te polyphenols nd te cffeine from green te leves. Chem Eng Process. 2003;42(2): Dmini E, Bcchetti T, Pdell L, Tino L, Crloni P. Antioxidnt ctivity of different white tes: Comprison of hot nd cold te infusions. J Food Compos Anl. 2014;33(1): Bhorun T, Neergheen-Bhujun V, Toolsee NA, Somnh J, Luximon-Rmm A, Aruom OI. Bioctive Phytophenolics nd Antioxidnt Functions of Aqueous nd Orgnic Te Extrcts. Elsevier; Luximon-Rmm A, Bhorun T, Crozier A, et l. Chrcteriztion of the ntioxidnt functions of flvonoids nd pronthocynidins in Muritin lck tes. Food Res Int. 2005;38(4): Zhng Y, Li Q, Xing H, et l. Evlution of ntioxidnt ctivity of ten compounds in different te smples y mens of n on-line HPLC-DPPH ssy. Food Res Int. 2013;53(2): Glzer AN. Oxygen Rdicls in Biologicl Systems Prt B: Oxygen Rdicls nd Antioxidnts. Elsevier; Schich KM, Tin X, Xie J. Reprint of Hurdles nd pitflls in mesuring ntioxidnt efficcy: A criticl evlution of ABTS, DPPH, nd ORAC ssys. J Funct Foods. 2015;18: Rmirez LS, Ospin LF, Ortiz A. Evlution of the Antioxidnt Cpcity nd Chrcteriztion of Phenolic Compounds Otined From Te ( Cmelli Sinensis ) for Products of Different Brnds Sold in Colomi. Phrmcologyonline. 2015;3:

7 m g c te c h in E q / g s m p le m g A c id g llic E q / g s m p le T e m p e r tu r e C PhOL Rmiréz-Aristizl et l 7 (1-9) O R IE N T A L 1 O R IE N T A L 2 O R IE N T A L 3 L IP T O N 1 L IP T O N 2 L IP T O N 3 H IN D Ú 1 H IN D Ú 2 H IN D Ú 3 J A IB E L 1 J A IB E L 2 J A IB E L E x tr c tio n d y s Figure 1. Temperture Control c 0 O RIENTAL LIP TO N HINDÚ J AIBEL Figure 2. Totl phenol content (TPC) c 2 0 O R IE N T AL L IP T O N H IN D Ú J AIB E L Figure 3: Totl flvonoid content (TFC).

8 m g T R O L O X E q / g s m p le m g T R O L O X E q / g s m p le PhOL Rmiréz-Aristizl et l 8 (1-9) c O R IE N T AL L IP T O N H IN D Ú J AIB E L Figure 4: DPPH ssy c O R IE N T AL L IP T O N H IN D Ú J AIB E L Figure 5. ORAC ssy. Figure 7. Correltion ORAC Vs TFC.

9 PhOL Rmiréz-Aristizl et l 9 (1-9) Tle 1. Weight smples. SAMPLE DAY WEIGHT (g) ORIENTAL LIPTON HINDÚ JAIBEL Tle 2. Summry, TPC, TFC, DPPH nd ORAC results of green te t room temperture. Totl phenol content (TPC) Totl flvonoid content (TFC) DPPH ssy ORAC ssy Smple mg GAE / g smple Percent mg /g smple eq CAT Percent mg Trolox eq/ g smple Percent mg Trolox eq/ g smple Percent ORIENTAL ± % 7.08 ± % ± % ± % LIPTON ± % 4.87 ± % ± % ± % HINDÚ 9.89 ± % 6.71 ± % ± % ± % JAIBEL 2.53 ± % 2.67 ± % ± % ± %