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1 KOREAN J. FOOD SCI. TECHNOL. Vl. 41, N. 6, pp. 673~680 (2009) The Krean Sciety f Fd Science and Technlgy ü ³ e w Á Á½Ÿ * w tœw / l Effects f Schizandra chinensis Extract n the Grwth f Intestinal Bacteria Related with Obesity Eun-Ji Jeng, Wn-Jng Lee, and Kwang-Yup Kim* Department f Fd Science and Technlgy, the Research Center fr Biresurce and Health, Chungbuk Natinal University Abstract This study was cnducted t screen fr plant fd materials that imprve human intestinal micrflra, especially micrrganisms assciated with besity. Amng 30 tested plant fd materials, the extract f Schizandra chinensis inactivated Eubacterium limsum, Bacterides fragilis and Clstridium spp. Additinally, S. chinensis extract was als fund t have a grwth-prmting effect n Bifidbacterium spp.. The antimicrbial activity and antixidant activity f the water extract did nt decrease in respnd t heating. Additinally, the water extract f S. chinensis did nt shw a txic effect n the grwth f Cac-2 cells. In viv feeding tests were perfrmed t investigate the influence f extract n the intestinal micrflra in rats. Althugh the extract did nt reduce besity induced by a high fat diet, it led t significant increase in the ppulatin f Bifidbacterium spp. and a decrease in the ppulatin f Clstridium spp. in rats. Taken tgether, these results indicate that S. chinensis culd be useful as a functinal fd cmpnent t cntrl intestinal micrbial flra. Key wrds: intestinal micrflra, antimicrbial activity, besity, Schizandra chinensis s ü w ù wš» w ³ ü q j ù, w y w ww w ³ ƒ w (1). ü³ ³ w³ ù ³x w kƒ (2). w w Bifidbacterium ³ w Clstridium perfringens w ³ k ü ³ g w (3). w ³ ü jš w³ w x wù (4). w l y» š š w w³ w š. Kim Shin(5) ƒ w Clstridium perfringens wš Bifidbacterium bifidum, Lactbacillus acidphilus, L. plantarum w š šw. ü³ k, w w w ü³ ³x j y *Crrespnding authr: Kwang-Yup Kim, Department f Fd Science and Technlgy, Chungbuk Natinal University, Chengju, Chungbuk , Krea Tel: Fax: kimky@chungbuk.ac.kr Received April 22, 2009; revised September 18, 2009; accepted September 19, 2009 w š. x y ƒ w. w y w ³x w. ù w w, p ü w» w mw w ü x. Turnbaugh (6) wùƒ w ü, ü ƒ w ƒ š t w. 12 y 1, k y w w ü w Firmicutes Bacteridetes w, Bacteridetes ƒ ƒ ³ w (7). y w 90% Bacillus, Clstridium, Lactbacillus, Streptcccus, Staphyllcccus w Fermicutes š Bacterides fragilis, B. thetaitamicrn, B. acidifaciens, B. distasnis w Bacteridetes 5% w ù, 1, k y w y w Bacteridetes ù 20% w ùkù Bacteridetes s w ú w ü ³ w w (6). Turnbaugh (6) mw, x mw ü y ƒw y w. Fermicutes Bacteridetes³ x û (6). ù (Schizandraceae) w ù 673

2 674 w t wz 41 «6y (2009) t w (8). schizandrl, schizandrin A, B, C, angelylgmisin O, P, epigmisin, pregmisin, dexyschizandrin lignan yw,»,, w w š, w» ƒ š t š (9). w w Hikin y (10) w šw, Lee Lee(11) g w, She (12) w y w šw. w Mn(9) x w»,, ƒ w schizandrin w šw. K (13) z α-glucamylase» w j z ƒ 2x w z z ƒ šw. m v» t w l w,,, w y, g w, x y, g l w, š x y,, w w w y ùkü (14,15). m, y,, ƒw, sd j, p, ½e, š» d š (15). Eubacterium spp. wš Bacterides spp. j k w š, mw w ü w w z x mw ü ³ y y wš w. ù ƒ ƒw ü ³» wš w. 30 p w w, x w 30 Table 1 ùkü. Blender(HALLDE MASK- INER, Kista, Sweden) w Standard Testing Sieve(APER- TURE 1.70 mm, Chung Gye Sang Gng Sa, Seul, Krea) w. 15 g 20 (, 80% ethanl) ƒ w 60 C 200 rpm 3 w (», w mdel Unin55R, 3000 rpm, 10 )w z œ»(eyela CCA-1110, Tky, Japan) 40 C w w» w š w 100 mg/ml w w. Table 1. Flra material used in study Scientific name 1 Cix lachrymajbi var. mayuen 2 Angelicae Gigantis Radix 3 Eucmmia ulmides Oliver 4 Sasa brealis var. gracilis 5 Hrdeum vulgare Linne 6 Crnus ffcinalis 7 Schizandra chinensis 8 Lycpersicn esculentum Mill. 9 Perilla frutescens 10 Oenanthe javanicadc. 11 Brassica leracea var. italica 12 zingiber fficinale 13 Cucurbita spp. 14 Hvenia dulcis 15 Zizyphus jujuba var. 16 Acanthpanax senticsus 17 Cassia btusiflia 18 Glycyrrhiza uralensis Fisch 19 Suaeda japnica 20 Chrysanthemum indicum 21 Atractyldes japnica 22 Carthamus tinctrius 23 Cinnammumcassia 24 Curcuma armatica 25 Brassica leraceae L. 26 Massa Medicata Fermentata 27 Lycium chinense Mill. 28 Slanum melngena L. 29 Sinmenium acutum 30 Rehmannia glutinsa ³ x w ³ Eubacterium limsum KCTC 3266, Clstridium perfringens KCTC 5101, C. difficile KCTC 5009, Bacterides fragilis KCTC 3688, Bifidbacterium bifidum KCTC 3418 w l(krean Cllectin fr Type Cultures, KCTC) w. RCM media(reinfrced Clstridial Medium, Difc, Detrit, MI, USA), MRS Brth (DeMan, Rgsa, Sharpe brth, Difc), MRSA (DeMan, Rgsa, Sharpe Agar, Difc) wš 37 C 24 x» w. w³ k 30% DMSO 100 mg/ml (Micr High speed centrifuge, Visin VS CFN, 10,000 rpm, 5 min)w w. x ³ w l ³ y yw 650 nm O.D. 0.5ƒ w w 10 2 cfu/ml³ w. 80% k w 96 well plate w w d 96 well plate wš 15, 7.5, 3.75 µl wš 10 2 ³ 100 µl w 0, 24 hr z 650 nm O.D. d wš w (%) y w. Inhibitry effect (%) ={1 (24 hr sample O.D. 0 hr sample blank O.D.)/ (cntrl O.D. cntrl blank O.D.)} 100

3 t w³y w y (disc plate methd) w w³y : 30 w k w y ( ) w þ w w. k w z w ƒƒ 100 mg/ml w w. ³ sq ³ y w ƒ 3% ƒw ml plate w. Paper disc(advantec 8 mm, Ty Rshi Kaisha, Ltd, Japan) 100 mg/ml v 30 µl w w ³ sq k z, x» e(mart Micrbilgy, ANOXOMAT WS80, Lichtenvrde, The Netherlands) 37 C, 24 hr w z paper disc w y y wš Vernier caliper(mituty, , N10, Kawasaki, Japan) y j» d w. w (MIC, minimum inhibitry cncentratin) d : w brth micrdilutin (17) w d w. ƒƒ ³ w z ³ 650 nm O.D. 0.4(10 CFU/mL)ƒ 6 w z 96 well plate ƒ x³ 100 µl wš, ƒ 0, 0.10, 0.25, 0.50, 1.00, 1.50, 2.00 mg/ml 100 µl w 24 w z ELISA reader (Shimadzu UV-160, Kyt, Japan) mw 650 nm d w w. t» w³y d :» š ƒœ y y» w w³y w w ³ (18) 96 C 15» w š ³» w 60, 70, 80, 90, 100 C 10 w. ³ ³ w³y x w w. w y w³y w z v ƒƒ 30 µl w z w y clear zne j» d wš w. s r w d : s r w 2g 40 ml ƒw 60 C 3 w w. s r w Dewant (19) Flin-Cicalteu reagent ƒ s r yw w y w w. ƒ 100 µl 2% Na 2 CO 3 2mL ƒw z 3 ew 50% Flin- Cicalteu reagent 100 µl ƒw. 30 z, Ÿ 750 nm d w š, t tannic acid(sigma Chemical C., St. Luis, MO, USA) w. z s r w 100 g mg tannic acid ùkü. DPPH w œ d : DPPH(1,1-diphenyl-2-picryl hydrazyl) w œ (Electrn dnating ability, EDA) Chi(20) d w., 0.2 mm DPPH(Sigma-Aldrich C., St. Luis, MO, USA) g 99.9% ethanl ww 100 ml w ü ³ e w w w z w. DPPH 0.8 ml 0.2 ml ƒw z 30 ew 520 nm Ÿ e d w. œ ƒ ƒ Ÿ ùkü. MTT assay w s w. MEM w , 0.125, 0.25, 0.5, 1, 2, 3, 4, 5 mg/ml w. x s v s s Cac-2 cell MEM(Mdified Eagle s medium, GIBCO, USA) 10% FBS (Fetal bvine serum, Hyclne, USA), 1% 5 mm nn-essential amin acid(neaa, GIBCO, NY, USA), 1% Penicillin-streptmycin(GIBCO, USA) sw g 37 C, 5% CO 2 Incubatr w s y y k living cell mitchndria succinyl dehydrgenase w yellw-mtt purple frmazan y k z, frmazan ELISA reader d w. MTT(3-(4,5-dimethylthiazl-2- yl)2,5-diphenyl tetrazlium brmide, Sigma-Aldrich C. USA) PBS 5 mg/mlƒ þ w w. Cac-2 cell 2 10 cells/ml w 5 96-multi well plate ƒ well 100 µl wš, 48 w s k z ywš w š 24 s w. ƒ well MTT (5 mg/ml in PBS) 20 µl wš 4 wš, š 100% DMSO 150 µl w z ELISA reader w 550 nm d w. Antiprliferative activity (%)=[1 [A sample /A cntrl ]] 100 x Rdent Diet with 45%Kcal Fat w. x 5 Sprague-Dawley (Charles River Labratry Inc., USA) 24 w k z, cntrl w ù š (High-fat-diet) w š, š w In vitr test mw t w 0.5 g/kg n wš 2 y d w. 6 z 12 w. d w. ü³ y w z 0.85% NaCl ³ yw š Mitsuka (2) xw š ƒ ³ p w. ùkù w Mitsuka (2) ³ xk w (genus) w š ƒƒ ³ d w ü³ y y w.» ù (Betula platyphylla Sap) ƒ þ 4 C w k z w. ù xylitl yww membrane filter kit(pre size 0.22, Crning, NY, USA) w w. ƒ v Table 2 ùkü. p w z t w 16 w w. sƒ (clr), (taste), w(flavr) š»y (verall acceptance) w 9»y w w, w w sƒw.

4 676 w t wz 41 «6y (2009) Table 2. Prductin f Betula platyphylla sap cntaining sugars and Schizandra chinensis water extracts Sample A B C D Betula platyphylla sap 100% 100% 100% 100% S. chinensis water extract 5% 10% 5% 10% xylitl 3% 3% 5% 5% m x w m SAS prgram(sas Institute, Cary, NC, USA) w ƒ x s³ ùkþ neway ANOVA w z Duncan s multiple range test w x wš, p 0.05 w q w. š w³ k» w³ š š t w 96 well plate 80% ethanl w³y Fig. 1, 2 80% ethanl Eubacterium limsum 46% jš, Bacterides fragilis -89% g. š E. limsum 97% jš, B. fragilis 100% g. w E. limsum jš B. fragilis j (Schizandra chinensis)ƒ. w³» w 96 well plate mw Clstridium perfringens Bifidbacterium bifidum w w³y., ü³ w³ C. perfringens wš ³ ³ B. bifidum g. t w³y w y (disc plate methd) w w³y 96 well plate w screeningw ƒ š, paper disc w E. limsum B. fragilis, C. perfringens, C. difficile, B. bifidum w w³y xw, 80% etanl, E. limsum B. fragilis, C. perfringens, C. difficile w w³y clear zne š, B. bifidum w w³y ùkù. Fig. 3, 4 ùkü š clear zne j» Table 3 ùk ü. w w³ w³ r, k Bacillus subtilis, Staphylcccus aureus, Escherichia cli, Klebsiella pneumniae, Pseudmnas aeruginsa, Salmnella Typhimurium(21) Vibri parahaemlyticus, Helicbacter pylri(22) w w³y š š. Fig. 1. Antibacterial activities f flra material extract. - -, Antibacterial activities f flra material water extract against Bacterides fragilis; -þ-, Antibacterial activities f flra material 80% ethanl extract against Bacterides fragilis; - ü-, Antibacterial activities f flra material water extract against Eubacterium limsum ;-Ü-, Antibacterial activities f flra material 80% ethanl extract against Eubacterium limsum. Fig. 2. Antibacterial activities f flra material extract. - -, Antibacterial activities f flra material water extract against Bacterides fragilis; -þ-, Antibacterial activities f flra material 80% ethanl extract against Bacterides fragilis; - ü-, Antibacterial activities f flra material water extract against Eubacterium limsum ;-Ü-, Antibacterial activities f Flra material 80% ethanl extract against Eubacterium limsum.

5 ü ³ e w 677 Fig. 3. Antimicrbial activities f Schizandra chinensis extracts against Eubacterium limsum. A, S. chinensis 80% ethanl extract; B, S. chinensis water extract; C, 80% ethanl Cntrl; D, water cntrl. Fig. 5. Heat stability f the Schizandra chinensis water extracts against Eubacterium limsum. A, 60 C; B, 70 C; C, 80 C; D, 90 C; E, 100 C. Fig. 4. Antimicrbial activities f Schizandra chinensis water extract against Clstridium perfringens. A, Water cntrl. Table 3. Antimicrbial activity f the Schizandra chinensis extracts Micrrganisms 80% ethanl extract Clean zne (mm) Water extract Cntrl 1) Eubacterium limsum ±0.07 3) 25.55±2.40 ND Bacterides fragilis 27.23± ±0.85 ND Clstridium perfingens 13.65± ±1.59 ND Clstridium difficile 15.85± ±2.26 ND Bifidbacterium bifidum ND 2) ND ND Cncentratin f sample was 100 µg/ml. 1) 80% ethanl, and water cntrl 2) Nt detected. 3) Values are mean±sd (n=3). MIC(minimum inhibitry cncentratin) d w w (MIC) d Fig. 6. Heat stability f the Schizandra chinensis water extracts against Clstridium difficile. A, 60 C; B, 70 C; C, 80 C; D, 90 C; E, 100 C. E. limsum, B. fragilis, C. perfringens, C. difficile w w³ ƒ š z» yw» w ƒ š w. 0.10, 0.25, 0.50, 1.00, 2.00 µg/ml xw, E. limsum, B. fragilis, C. perfringens, C. difficile 4³ 1.00 µg/ml ƒ w ùkû. t» w³y d : w³y Fig. 5, 6 Table 4 C. perfingens w clear zne j»ƒ ƒw. C. perfingens w 3³ 70 C clear zne w š 80 C clear zne ƒw. ³ 80 C w w³y. w w³y šw Chung (22) š w C 1

6 678 w t wz 41 «6y (2009) Table 4. Heat stability f the Schizandra chinensis water extract Micrrganisms Clean zne (mm) 60 C 70 C 80 C 90 C 100 C Eubacterium limsum ±0.78 1) 11.78± ± ± ±0.04 Bacterides fragilis 14.20± ± ± ± ±0.04 Clstridium perfingens 11.15± ± ± ± ±0.42 Clstridium difficile 15.93± ± ± ± ±0.78 Cncentratin f sample was 100 µg/ml 1) Values are mean±sd (n=3) Table 5. Ttal plyphenl cmpund cntent f Schizandra chinensis extract Table 6. Antixidant activities f water extract frm Schizandra chinensis Extractin cnditins Cntent (mg/g) Extractin cnditins EDA (%) Temperature ( C) ±0.00 a ±0.00 a ±0.01 b ±0.00 a ±0.01 ab 1) Values are mean±s.d. f triplicate analyses (n=3). 2) Any means in the same clumn fllwed by the same letter are nt significantly (p<0.05) different by Duncan s multiple range test w z w³ d w w³ yƒ. w w y w ùkü š ƒ. s r w d : r yw phenlic hydrxyl» ƒ»»k w w ƒ, w yz y» ƒ (20), w r yw w w C 10 C y x w, 7.50 mg/g s r w ùkü. w Table 5 10 C y s r w š, w w ùkþ s r w, Lee (23) s r w ƒ w š r yw w r yw y ù, w r yw» ƒ. ³ w š. DPPH w œ d : DPPH e w œ (EDA %) d w Table 6 10 mg/ml C w w ù kü (p<0.05). 100 C C û 44.43% w ùkù 100 C š w z q ƒ. MTT assay w s : v s w» w (0.0625, 0.125, 0.25, 0.5, 1, 2, 3, 4, 5 mg/ml) w s w., v s w z ƒ. Temperature ( C) ±0.25 b ±1.02 b ±0.37 b ±2.30 b ±0.33 a 1) Values are mean±sd f triplicate analyses (n=3). 2) Any means in the same clumn fllwed by the same letter are nt significantly (p<0.05) different by Duncan s multiple range test. Table 7. Effects f Schizandra chinensis n bdy weight gain and epididymal adipse in the rats Bdy weight change Grup Initial weight (g) Final weight (g) Final/Initial Nrmal 1) 223.9±8.3 a 377.9±18.3 a 1.69±0.08 Cntrl 2) 227.4±6.9 a 409.8±25.7 b 1.80±0.09 Extract 0.5 g/kg 239.3±9.3 b 408.7±35.9 c 1.83±0.15 Values are mean±se f 8 rats. Any means in the same clumn fllwed by the same letter are nt significantly (p<0.05) different by Duncan s multiple range test 1) Nrmal: cmmercial fd 2) Cntrl: high-fat-diet x x y Table 7 cntrl high-fat-diet rats n w rats ƒ ùkù. n» 6» ƒ ùkù. ü³ Table 8 ùkù Bifidbacterium n cntrl n ³ 9.30% 16.92% y w ƒw. wr ü t w³ Clstridium cntrl n ³ 19.77% 7.38% x w. w Staphylcccus, Streptcccus w ùkûš, n Lactbacillus š Eubacterium ³ w. Bacteridaceae in vitr x z ƒ ù ³ w w³ w q» ƒ v w. in viv x ƒ t w³ Clstridium ³ Bifidbacterium ü³ j z ƒ. w³ Clstridium ³ x

7 ü ³ e w 679 Table 8. Analysis f intestinal micrflra in the rats fed with Schizandra chinensis extract (CFU/mL) Nrmal 1) Cntrl 2) Extract Enterbacteriaceae (0.68) 2) (9.77) (53.85) Yeast (0.15) (6.74) (2.46) Bifidbacterium (21.58) (9.30) (16.92) Staphylcccus (5.26) (0.49) (0.01) Clstridium (3.58) (19.77) (7.38) Streptcccus (0.26) (23.26) (0.05) Bacillus (5.79) - - Lactbacillus (40.00) (6.74) (3.54) Bacteridaceae (8.37) (5.85) Eubacterium (5.81) (4.62) Gram negative rds (22.11) (18.60) - Ttal (100) 3) (100) (100) 1) Nrmal: cmmercial fd 2) Cntrl: high-fat-diet 3) Percentage abut number f selected stains in test strains Table 9. Sensry evaluatin f Betula platyphylla sap cntaining sugars and Schizandra chinensis water extract (panel: 16) Sample A B C D Clr 4.94±1.84 a 5.25±1.54 a ±1.86 ab 6.81±1.97 b Flavr 4.25±1.61 a 4.69±1.18 a 5.06±1.77 a 4.75±1.5 a 0 Sur 4.19±1.48 a 4.31±1.52 a 5.19±1.82 a 5.06±2.09 a Sweet 4.44±1.41 a 4.31±1.21 a 6.44±1.71 b 4.81±1.87 a verall 4.81±1.33 a 4.63±1.39 a 6.68±1.54 b 5.44±1.67 a Any means in the same clumn fllwed by the same letter are nt significantly (p<0.05) different by Duncan s multiple range test Eubacterium z, Bacteridaceae z ³ Bifidbacterium in vitr x ³ w ü, ƒ ü ³ w ³ j t ƒ š ƒ.» w w w sƒ Table 9. sƒ w sample Dƒ 6.81, w sample Cƒ 5.06, sample Cƒ 5.19, sample Cƒ 6.44 ùkû,»y sample Cƒ 6.68 sample w (p<0.05). 5% xylitl, 5% ƒw sample Cƒ w» ƒ w ƒ w ƒ. wù ü³ y wš in vitr k w š w³y» wš s w x ww. ù ƒ t» ƒ w ƒw ƒ y wš w. 96 well plate w screeningw, ³ B. fragilis B. spp. y jš, w³ E. limsum C. perfringens j w. w³y paper disc methd w xw E. limsum, B. fragilis, C. perfringens, C. difficile w w³ ƒ š B. bifidum w w³ ùkü. k MIC d w, E. limsum, B. fragilis, C. perfringens, C. difficile 1µg/ ml w w.» ƒ w ty w ³ 96 C, 15» C 10 C y 10 w xw, w³y w š j w³ w 80 C ³ w. w s r w œ w w y y d w w. w. w w³y w y ùkü s r w œ w w s r w œ w w³y. v s w y w» w v s Cac-2 w w MTT assay mw y w. š 5µg/ ml s w. mw xw», in vitr s w q w w x w y w. x high-fat-diet rats 0.5 mg/kg/day n w w e. ü ³ y cntrl extract n w, in vitr x Bifidbacterium spp. 7.62% jš Clstridium spp % g. w Eubacterium g. ƒ ³ jš, w³ j ü ³ w ³ j t ƒ w». ƒw sƒ ü w» w ù ü ³ y w» š m ƒw sƒw»y ù 100%, 5%, m 5%

8 680 w t wz 41 «6y (2009) ƒw sample Cƒ 6.68 sample w (p<0.05). 5% xylitl, 5% ƒw sample Cƒ w», t ƒ w ƒ w ƒ. Áw» x l w l t» ( y: ) w. x 1. Gibsn GR, Rberfrid MB. Dietary mdulatin f the human clnic micrbita: intrducing the cncept f prebitics. J. Nutr. 125: (1995) 2. Mistuka T. Recent trends in research n intestinal flra. Bifidbacteria Micrflra 1: 3-24 (1982) 3. Mistuka T. The Intestinal Flra and Bihmertasis. Japan Scientific Scieties Press. Tky, Japan (1989) 4. Shin H-K, Shin O-H, K Y-J. Effects f ptat prtein n the grwth f Clstridium perfringens and ther intestinal micrrganisms. Krean J. Appl. Micrbial. Bitechnl. 20: (1992) 5. Kim JD, Shin TS. The grwth prmting effect f Bifidbacterium bifidum by cmbinatin f natural prducts bearing antixidative capacity. Krean J. Micrbil. Bitech. 30: (2002) 6. Turnbaugh PJ, Ley RE, Mahwald MA, Magrini V, Mardis ER, Grdn JI. An besity-assciated gut micrbime with increased capacity fr energy harvest. Nature 444: (2006) 7. Ley RE, Turnbaugh PJ, Klein S, Grdn JI. Human gut micrbes assciated with besity. Nature 444: (2006) 8. Kim KS, Park CG, Ryu SN, Bang JK, Lee BH. Schizandrin, li cmpunds, and their extractin yield in fruits f Schizandra chinensis Bailln. Krean J. Crp Sci. 45: (2000) 9. Mn KS. Ingredients and applicatins f medicinal plants, science encyclpedia. Ilwl, C., Ltd., Seul. pp (1999) 10. Hikin H, Kis Y, Takuchi H, Ikeya Y. Validity f the riental medicines 60. Liver-prtective drugs II, Antihepattxic actin f lignids frm Schizandra chinensis fruits. Planta Med. 50: (1984) 11. Lee JS, Lee SW. Effect f water extract in fruits f mija (Schizandra chinensis Bailln) n alchl metablism. Krean J. Diet. Culture 5: (1990) 12. She HJ, Lee MY, Hwang JS. Effect f Schizandrae fructus extract n bld cnstituents f allxan-induced diabetes rabbits. J. Krean Sc. Fd Sci. Nutr. 16: (1987) 13. K BS, Park Sk, Chi SB, Jun DW, Chi MK, Park SM. A study f hypglycemic effects f crude extracts f Schizandrae fructus. J. Krean Sc. Appl. Bi. Chem. 47: (2004) 14. Mk CK. Quality characteristics f instant tea prepared frm spray-dred mija (Schizandra chinensis Bailln) extract grape juice mixture. Fd Eng. Prg. 9: (2005) 15. Oh SL, Kim SS, Min BY, Chung DH. Cmpsitin f free sugars, free amin acids, nn-vlatile rganic acids, and tannins in the extracts f L. chinensis M, A. acutilba M, S. chinensis B. and A. sessiliflrum S. Krean J. Fd Sci. Technl. 22: (1990) 16. Yn SS. Krean fd (histry and cking). Suhak C., Seul, Krea. p. 372 (1982) 17. Amster D. Susceptibility Testing f Antimicrbial in Liquid Media. 4 th ed. Williams and Wikins, Baltimre, MD, USA. pp (1996) 18. Kseglu SS, Lawhn JT, Lusas EW. Vegetable juices prduced with membrane technlgy. Fd Technl. 45: (1991) 19. Dewant V, Xianzhng W, Liu RH. Prcessed sweet crn has higher antixidant activity. J. Agr. Fd Chem. 50: (2002) 20. Chi Y, Kim MH, Shin JJ, Park JM, Lee J. The antixidant activities f the sme cmmercial teas. Krean J. Sc. Fd Sci. Nutr. 32: (2003) 21. Ch YJ, Ju IS, Kim BC, Lee WS, Kim MJ, Lee BG, An BJ, Kim JH, Kwn OJ. Bilgical activity f mija (Schhizandra chinensis Bailln) extract. J. Krean. Sc. Appl. Bil. Chem. 50: (2007) 22. Chung KH, Lee SH, Lee YC, Kim JT. Antimicrbial activity f mija (Schizandra chinensis) extract. J. Krean Sc. Fd Sci. Nutr. 30: (2001) 23. Lee SC, Jeng SM, Kim SY, Park HR, Nam KC, Ahn DU. Effect f far-infrared radiatin and heat treatment n the antixidant activity f water extracts frm peanut hulls. J. Agr. Fd Chem. 94: (2006)