Antimicrobial Effect of Lithospermum erythrorhizon Extracts on the Food-borne Pathogens

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(1)KOREAN J. FOOD SCI. TECHNOL. Vol. 36, No. 5, pp. 823~827 (2004). ©The Korean Society of Food Science and Technology. æ~ºÂb~ 7ë9 bö & Î 8 æ ** ê«L ®'·. Antimicrobial Effect of Lithospermum erythrorhizon Extracts on the Food-borne Pathogens Ji-Hyun Bae* Department of Food Science and Nutrition, Keimyung University Antimicrobial effect of Lithospermum erythrorhizon extracts against food-borne pathogens was investigated. L. erythrorhizon was extracted with methanol at room temperature, and the extraction was sequentially fractionated using petroleum ether, chloroform, ethyl acetate, and methanol. Antimicrobial activity of L. erythrorhizon extracts was determined using paper disc method against food-borne pathogens and food spoilage bacteria. Ethyl acetate extracts of L. erythrorhizon showed the highest activity against Staphylococcus aureus and Shigella dysenteriae. Synergistic effect was found in combined extracts of L. erythrorhizon and Sophora subprostrata as compared with each extract alone. Growth inhibition curve was determined using ethyl acetate extracts of L. erythrorhizon, against S. aureus and S. dysenteriae. Ethyl acetate extract of L. erythrorhizon, showed strong antimicrobial activity against S. aureus at 4,000 ppm, retarding growth of S. aureus more than 48 hr and S. dysenteriae up to 12 hr. Key words: Lithospermum erythrorhizon, antimicrobial activity, food-borne pathogens. *. . ¶ ~ ªbê ®ÒÚæ , ¢ 5 7 æö " ¢ (3). æ~~ «« erythrorhizonö ¾æ¾ ® erythroº ¯f ïj, rhizonf òÒ¢ ~~º, æ~ òÒ~ bö F>Ú ®º ¯f ï²º AÆ¦V _&& ò º N~¢º Ç" ;;ö ¯f ïj b V *~ ÒÏ ~ zb, êê s"~ ãÖê ï²¢ Ï &' F . æ~ òÒ~ z[ö F>Ú ®º 1,4-dinaphthoquinonê bî zò(shikonin)f &¦V j~ ¯f " ïB Ï>Ú^ z, "öê Â ï² Ï~¶ ~ º & Úæ ® (4). ~~ "ïö & º *Û¶"ö & (5)¢ j~ ¶."~ "Î"ö & þ(6), ¶"ï²ö ~ ÒRF~ "ïö & (7), 5 ¶.ï²~ Wª" ßWj 7b ¶. ï²~ " ïWö & (8)& Úæ ® . zò~ Wö & ö ~~ ï²zb 20-50 µg/mL ³êöB Wj ¾æÚº ©b > ® (9). $ æ~º , " 5 çÒ~ £ÒÎ"& ®º ©b rJ^ .¦V '« b¦÷, zç, .ç, ÿç 5 ~î~ ~òö Ï>Ú z b, ¢öBº ¶Ú Ï£®~ öò ÒÏ>Vê . (10). f ? bÒò 5 £Bö & æ~~ Ïö & · & Ú^ zb¾, ® " &NB 7 ëW bö & Î"ö &Bº Ú' & ¦ ;¢ öBº æ~ºÂb '« 7ë FB ^ö ~º Wj ¦Ã , £Òò þ ¾O> ®º Öv""~ çßÎ"ê ¦Ã ¶ ~& .. " ^z~ Bö V Û&~ æzö ~ ¶" B >º 7ëf Salmonella³ " Vibrio parahaemolyticus, Listeria monocytogenes, GB® 5 &.~öB ¦ÂB E. coli O157:H7 ö > ®b, 7ëf ® * >ö ö 7º*~² ª>Ú ® . ®~ ¦Nf æîf " b~ ·Ïö ~ ¢Ú¾º, ¢ Oæ~V * &¾Ò, ïË, ïÿ, B Î&, OÒF Ò · O » j ÒÏ~ &ËV*~ Ëj ê~ ® . &¾Ò ¾ ïË 5 ïÿ O»f B®~ î~ &~¾ &ËjÏ~ Ã& ¢ &^R > ®b, WB~ ãÖº n*Wö & ÖJ¢ Ù ®Ú "¾öº ²j¶ ~ ; » Ã&ö V ¢ 6N ÒÏïj B~Jº º^ . ç j ~V * O»~ ~¾ Â W bîj ¦ï~ © j ®ö Î&~ B ÒÏb F" n*Wj ÿö òÊ¶ ~º ô ê¯> ® (1,2). æ~(Lithospermum erythrorhizon. Sieb. et Zucc.)º æ~" (Boraginaceae)ö ³~º j .bB æ., ¶., ¶", *Corresponding author: Ji-Hyun Bae, Department of Food Science and Nutrition, Keimyung University, Shindang-dong, Dalseo-gu, Taegu 704-701, Korea Tel: 82-53-580-5875 Fax: 82-53-580-5875 E-mail: [email protected] 823.

(2) ®"²æ B 36 ² B 5 ^ (2004). 824. Òò 5 O». Table 1. List of microorganisms used for antimicrobial activity test. ò þöB ÒÏ W þ &ç b æ~º Öb, & 7 ÎW £* ÏöB 2003j 10ú, ç~ ©j «~& . ®Bbj B~V * &ó² 2® >^~ Î ê, ^~² îê~ ºÂÏ ò ÒÏ ~& .. Strains. ÒÏ " 5 Væ æ~ ºÂb~ þö ÒÏ "º Gram(+) ^ 2« " Gram(-) ^ 7«b C 9«j "VFö « ²öB ª· Aj ÒÏ~& (Table 1). Î "~ V· ö tryptic soy broth(Difco, USA)¢ Væ ÒÏ~&b, 37oC, incubatoröB 18-24* V·~ Gj &V~& . W. þö ÒÏ ÚVæº tryptic soy agar(Difco, USA)& . 9 bî~ ºÂ Î æ~ 500 gö & æ~ 7ï~ 2V ªï petroleum ether, chloroform, ethyl acetate, methanolj ÒÏ~ W bîj ºÂ~& . ºÂ&ö îêÎ æ~¢ I 1 L~ methanolj If ê NöB 6* O~ ê, Whatman No. 2(Whatman international Ltd., England)ö "~ ®Bbj B ~& . "B Ïf 6{³»V(EYELA, N-N. Series, Japan)¢ ÒÏ~ 45oCöB 6{Á³»~&b ³» methanol ºÂbf petroleum ether, chloroform, ethyl acetate 5 methanolj '' ÒÏ~ Ï êÛ ª³~& . r methanol º Âb" '« FVÏ ¢ ªêz&Vö I 5ª* >·ëb z Ú b ê, 15ª* Nö O~Î ê ªÒ~& . æ ~~ >ºÂbj FVÏ ºÂ~ Îf ºÒö 1N Ã ~>¢ I 100oCöB 30ª* y ê ÿ¢ O»b " ~& . "B Ïj 6{³»V(EYELA, N-N. Series, Japan) ¢ ÒÏ~ 45oCöB 6{Á³»~&b ' ³ê C~ þö ÒÏ~& . þ~ çßÎ" ªCö ÒÏ Öv" ºÂbê æ~ ºÂb" ÿ¢ O»b &j~& . æ~ ºÂb~ 9 G; W bîj ¦ï~V * þöBº paper discO» j ÒÏ~& (11). Tryptic soy broth(TSB)Væö V· ^ j Spectrophotometer(Nontron instruments. Italy)¢ ÒÏ~ optical density8 0.4(620 nm) 7ê¢ .~ pour-plate methodö V¢ tryptic soy agar(TSA)Væ& ª" B V· 7 ö ¢~² Df ê NöB : . Væ *ö B paper disc¢ ò>ö ² RÒ &OÎ ê æ~~ petroleum ether, chloroform, ethyl acetate, methanol, >ºÂbj ' ' 100 ppm, 250 ppm, 500 ppm 5 1,000 ppmb C~ 20 µLO ÂÂ® >V . Control æ~ ºÂb Ú®æ pf 70% ethanolj þ" ÿ¢ O»b 6' ~& . &jB Î plate¢ 37oCöB 24* V· ê disc "æö WB clear zone(mm)~ V¢ G;~ ' ª³b~ W ;ê¢ G;~& . K~ çßÎ G; æ~ ºÂbj W b ºÂb" b®j K~ çß ¦¢ {~¶ Öv" ºÂb"~ bj ê. Gram positive bacteria. Gram negative bacteria. Staphylococcus aureus ATCC 25923 Bacillus cereus ATCC 27348 Escherichia coli ATCC 25922 Pseudomonas aeruginosa ATCC 27853 Salmonella typhimurium ATCC 14028 Salmonella enteritidis ATCC 13076 Shigella sonnei ATCC 25931 Shigella dysenteriae ATCC 9199 Shigella flexneri ATCC 12022. Table 2. Yield of organic solvents and water extracts from Lithospermum erythrorhizon Fraction. Dried weight (g). Yield (%). Petroleum ether extract Chloroform extract Ethyl acetate extract Methanol extract Aqueous extract. 2.0 30.2 2.2 58.5 114.3. 0.4 6.0 0.4 11.7 22.3. ~& . þ~ .j þöB K ®r «ÃB æ~ ~ ethyl acetate ºÂb" Öv"~ ethyl acetate ºÂbj '' 1,000 ppmO D, æ~~ ethyl acetate ºÂb 2,000 ppm 5 Ö v"~ ethyl acetate ºÂb 2,000 ppm" Kj jv~& . &ç "º Shigella dysenteriae¢ ÒÏ~ &b 70% ethanolj ' òf ÿ¢ · 20 µLO ª"~ ¦Ã~& . b~ G . G; æ~~ ethyl acetate ºÂbj membrane filter(0.2 µm, pore size. Toyoroshi kaisha Ltd., Japan) BÊ, ÚVæö ' ºÂbj 1,000 ppm, 2,000 ppm, 5 4,000 ppm ³êê Î&~ & . Vö O.D.8j 0.4 ¾ ^ V·j 109V C ~ Z'b 7« ê 37oCöB 72* V·~, 12* î ^ V·~ Ã;ê¢ 620 nm2Ë~ spectrophotometer öB G;~& (12).. Ö 5 8 æ~~ '« F8Ï 5 >ºÂb~ >N æ~~ methanol ºÂbj petroleum ether, chloroform, ethyl acetate 5 methanol '' ªÒ Ö", ' ª³b~ ºÂ > Nf Table 2" ? ¾æÒ . æ~~ petroleum ether, chloroform, ethyl acetate, methanol ª³b 5 >ºÂbf '' 0.4, 6.0, 0.4, 11.7 5 22.3% ¾æ¾, petroleum ether~ >N & Ë Ô~ >ºÂb~ >N &Ë ¸~ . æ~~ F8Ï 5 >ºÂb~ 9 ¦ï Paper disc O»b æ~~ '« FVÏ ª³b 5 > ºÂbj '« ®¦N 5 7ëö 'ÏB W j þ : r" ?f Ö"¢ áj > ®î . Gram ·Wö & æ~~ petroleum ether, chloroform, ethyl acetate, methanol ºÂb 5 >ºÂb~ Wf Table 3".

(3) æ~ºÂb~ 7ëW bö & Î" Table 3. Antimicrobial activities of each solvent fraction from Lithospermum erythrorhizon against Gram positive bacteria. 825. Table 4. Antimicrobial activities of each solvent fraction from Lithospermum erythrorhizon against Gram negative bacteria. Clear zone on plate (mm)1) Strains. Clear zone on plate (mm)1). Fraction conc. (ppm). PE. C. EA. M. W. Staphylococcus aureus. 100 250 500 1,000. -2) 6 7. -2) 7. -2) 11 13 16. -2) 7 8 11. -2) 7 8. Bacillus cereus. 100 250 500 1,000. 7. 7 9. 9 10 13. 6 7 9. 9. Strains. Fraction conc. (ppm). PE. C. EA. M. W. Escherichia coli. 100 250 500 1,000. -2) -. -2) -. -2) 7 9 12. -2) 7 9. -2) 9. Pseudomonas aeruginosa. 100 250 500 1000. -. -. 7 8 9. 6 7 11. 7 10 11. Salmonella typhimurium. 100 250 500 1,000. -. -. 7 9 13. 7 8. 8. Salmonella enteritidis. 100 250 500 1,000. 8. 7. 7 10 12. 6 7. 7 8 10. Shigella sonnei. 100 250 500 1,000. -. 7. 9 10 13. 6 7. 8 8 9. Shigella dysenteriae. 100 250 500 1,000. 8 9. 8 9. 9 11 14. 7 10 12. 9 10 12. Shigella flexneri. 100 250 500 1,000. 8. -. 7 9 11. 7 8. 9. 1). Diameter, 2)No inhibitory zone was formed. PE: Petroleum ether extract of Lithospermum erythrorhizon, C: Chloroform extract of Lithospermum erythrorhizon, EA: Ethyl acetate extract of Lithospermum erythrorhizon, M: Methanol extract of Lithospermum erythrorhizon, W: Water extract of Lithospermum erythrorhizon.. ? ¾æ¾ discö 6' æ~~ '« ºÂb~ ³ê& Ã& > W ² ¾æÒ . ¯ ³ê& Ã&> Wj ¾æÚº inhibition zone~ V& Ã&~ ethyl acetate ºÂb~ ãÖ S. aureusö & 1,000 ppm ³êöB 16 mm &Ë Wê¢ ¾æÚî (Fig. 1). æ~ ºÂb~ «~ 5 ³êö V¢ ' " ö & Wj ¾æÚ ~ «~ö V¢ 'V Wj ¾æÚ, æ~~ petroleum ether ºÂbf S. aureusö & "B Wj ¾æÚî, æ~~ chloroform ºÂbf B. cereusöB &Ë Wj ¾ æÚî . æ~~ ethyl acetate ºÂbf þö ÒÏ Î "ö & Wj ¾æÚî 250 ppm ³êöBê Î"& ¾æÒ . 9~ öêR ºÂb 7 ethyl acetate ª ³bf S. aureusö &B &Ë ¸f W ®r B : ®º(13), þöBê æ~~ ethyl acetate ºÂb S. aureusö & &Ë ; Î"¢ ¾æÚî . þö ÒÏ '« æ~ ºÂb~ Gram rWö & K ¦ï Ö"º Table 4f ? ¾æ¾, æ~~ methanol ºÂb" >ºÂbf S. dysenteriaeö & &Ë Î"' Wj &, æ~~ ethyl acetate ºÂb Î "ö & ; Kj ¾æÚî . 7 S. dysenteriae& &Ë "6~² >w æ~~ ethyl acetate ºÂb 1,000 ppmöB 14 mm~ clear zonej ¾æÚî . ¾" æ~~ ethyl acetate ºÂbf Gram ·W" Gram rWö & 9f Kj æî ®rj r > ®îº Kim (14)f Ö.~ zêR ºÂb Gram ·W" Gram rW" E. coliö z "6~² >w~ & : ® . Þ þö ÒÏ æ~ ºÂb~ ³ê& 100 ppm ~ ãÖöº Î"¢ ¦Ã > ìî , æ~~ petroleum ether ºÂb" chloroform ºÂb~ ãÖ ê Î "ö & æ Wj ¾æÚæ pj 500 ppm çöBò Kj &V > ®î . b~ ethyl acetate ºÂ[öº Òò Wª, FVÖ~, êò, , VÚ 5 Væ r¢~& " ÏÂ>º ©b rJ^ ®º, . þöB &Ë ¸f Kj æ~~ ethyl acetate ºÂ böê f FÒ Wª F>Ú ®j ©b ÒòB . Hong (15)f FWb~ butanolºÂb Gram ·W S.. 1). Diameter, 2)No inhibitory zone was formed. PE: Petroleum ether extract of Lithospermum erythrorhizon, C: Chloroform extract of Lithospermum erythrorhizon, EA: Ethyl acetate extract of Lithospermum erythrorhizon, M: Methanol extract of Lithospermum erythrorhizon, W: Water extract of Lithospermum erythrorhizon.. aureus, S. faecalis 5 Bacillus sp.ö &~ BGÛB Î"¢ Gram rW E. colif ê Candida albicansö & Bº ÛBÎ"& ì : ®b¾, þö ÒÏ æ~~ ãÖ Gram ·W" rW " *ö ßû' N ¢ æ p~ . æ~ ºÂb ÖL ºÂb~ çß Î æ~~ ethyl acetate ºÂb" Öv"~ ethyl acetate ºÂb j Dîj ãÖ ¾æ¾º Î"º Table 5f ? ¾æÒ. . þöB &Ë "6 Î"¢ &~ S. aureusö & v b ºÂb~ Kf æ~ ºÂb" Öv" ºÂ bj b®j ãÖ z ² ¾æ¾, æ~~ ethyl acetate ºÂ b òj ëb 2,000 ppm & ãÖ(17 mm) æ~~ ethyl acetate ºÂb 1,000 ppmö Öv"~ ethyl acetate ºÂb 1,000 ppmj DÚ & ãÖ& z Kj & (20 mm). S..

(4) ®"²æ B 36 ² B 5 ^ (2004). 826. Table 5. Antimicrobial activity of combinded extracts from Lithospermum erythrorhizon and Sophora subprostrata Clear zone on plate (mm)1) at 1,000 ppm Strains Staphylococcus aureus Shigella dysenteriae. Control. Lithospermum erythrorhizon (2,000 ppm). Sophora subprostrata (2,000 ppm). Both3) (each 1,000 ppm). -2) -. 17 15. 13 17. 20 19. 1). Diameter. No inhibitory zone was formed. 3) Lithospermum erythrorhizon and Sophora subprostrata. 2). Fig. 1. Antimicrobial activities of various extracts of Lithospermum erythrorhizon against Staphylococcus aureus at the concentration of 1,000 ppm. C: control (70% ethanol), I: petroleum ether extract, II: chloroform extract, III: ethyl acetate extract, IV: methanol extract, V: Aqueous extract.. Fig. 2. Antimicrobial activities of ethyl acetate extract of Lithospermum erythrorhizon, ethyl acetate extract of Sophora subprostrata and both extracts against Shigella dysenteriae. C: control (70% ethanol), I: Lithospermum erythrorhizon (2,000 ppm), II: Sophora subprostrata (2,000 ppm), III: Lithospermum erythrorhizon (1,000 ppm) and Sophora subprostrata (1,000 ppm).. dysenteriaeö &Bê v ºÂbj '' 1,000 ppmO DÚ R ãÖ& æ~~ ethyl acetate ºÂb 2,000 ppmj ëb & ãÖ ¸f Kj & (Fig. 5.). æ~~ ethyl acetate ºÂb Gram r9 5 Gram ·9 ~ Ãö ~º 'Ë æ~~ ethyl acetate ºÂbj ³êê(0 ppm, 1,000 ppm, 2,000 ppm 5 4,000 ppm) TSBVæö Î&~, Gram ·W S. aureusf Gram rW S. dysenteriaeö '' 7«B 72 * V·~B ¢; * *Ïb "~ WË ;ê¢ G ; :, Fig. 3 5 Fig. 4f ?f ÃFj áj > ®î. . S. aureus~ ãÖ, æ~~ ethyl acetate ºÂbj Iæ pf control VæöB V·®j 12* ê¦V /Ï Ã&¢ WË ¢ÚÎj &V > ®î . æ~~ ethyl acetate ºÂb 1,000 ppmj Î& VæöBº ~ WË 36*r æ ÛBNj &V > ®î, 4,000 ppm~ ³êöBº 48 * çræ ~ Ã æ>º ©j { > ®î (Fig. 3). Chung(16)f S. aureusö & ¶: FË ethanol ºÂ b 3.0 mg/mL çöB Ã æ>î : ® b, Park (17)f 9~ >ÏW ºÂb 1,000-1,200 ppm º *öB ~ Ã ÛB& ·B ~& . $ Jeon (18)f îã~ methanol ºÂb S. aureus~ WËj ÛB : ®º, þöBê æ~~ ethyl acetate ºÂb S. aureus~ Ãö ÛB Î"¢ & . $ Shin . Fig. 3. Effect of ethyl acetate extracts of Lithospermum erythrorhizon against the growth of Staphylococcus aureus. -ü -, control; -þ -, 1000 ppm; - -, 2000 ppm; -ù-, 4000 ppm.. (19)f ¶²´~ ethanol ºÂb S. typhimurium~ G ÛB ¢ 36*ræ æ³Î ~&, Chung (20)ê 'æ º Âb ß'b S. typhimuriumö & Wj <º. : ®Ú, ÂböB ªÒ>º '« W bîj DÚ Ï~ 7ë~ WËj ÎN'b ÛB >ê ® j ©b ÒòB . æ~~ ethyl acetateºÂb S. dysenteriae ö & ~º G & ;ê¢ ÿ¢ O»b 72* ÿ.

(5) æ~ºÂb~ 7ëW bö & Î". Fig. 4. Effect of ethyl acetate extract of Lithospermum erythrorhizon against the growth of Shigella dysenteriae. -ü -: Control, -þ -: 1,000 ppm, - -: 2,000 ppm, -ù-: 4,000 ppm.. n ÚÚ : Fig. 4f ?f Ö"¢ áj > ®î . æ~~ ethyl acetate ºÂbj Î&~æ pf control~ ãÖ V· ê 12*¦V /³ ~ Ãj " > ®î, 4,000 ppm ³ê ¢ Î&~&j ãÖ ~ Ã jò~² Ú^ æ~~ ethyl acetate ºÂb S. aureusf S. dysenteriae~ WËj Î N'b ÛBÒ > ®º ©b 6>î .. º. £. öBº 7ë FB^ö & W Ö> Â W bîj ¦ï~V * .¦V "*" OöB 6Ò Ï>Ú N æ~¢ '« FVÏ ºÂ~ 7ë F B^ö & Wj Ò ~ . æ~¢ methanol ºÂ ê, petroleum ether, chloroform, ethly acetate¢ Ï~ NöB '' Ï ê êÛ ª³~, >ºÂbj áf ê, 9«~ 7ë FB^(S. aureus, B. cereus, Sal. enteritidis, S. flexneri, E. coli, Sal. typhimurium, S. dysenteriae, P. aeruginosa, S. sonnei)ö &~ Î"¢ Ò~& . æ~ ºÂ b~ ³êê W ¦ïöBº æ~~ ethyl acetate ºÂ b &Ë Î"¢ &b S. aureusf S. dysenteriae & &Ë "6~² >w~º "& . æ~~ ethyl acetate º Âb" Öv"~ ethyl acetate ºÂbj b~ Kj G ; Ö" v ºÂbj DÚ Î&®j ãÖ& ëb Ò Ï®j çßÎ"¢ ¾æÚî . $ æ~~ ethyl acetate ºÂb 7ë FB^~ WËö ~º Î"¢ ¦ ;~V * S. aureus 5 S. dysenteriae~ V·ö æ~~ ethyl acetate ºÂbj '' 4,000 ppm ³ê Î&®j , S. aureus~ G 48* çræ ÛBNj &V > ®î, S. dysenteriae~ Gê 12*ræ æÒ > ®î .. ^. ò. 1. Moon BS. Food Sanitation. Shinkwang Publishing Co., Seoul,. 827. Korea (2003) 2. McSwane D, Rue N, Linton R. Essential of Food Safety and Sanitation. Prentice Hall, Upper Saddle River, NJ, USA (2000) 3. Yook CS. Coloured Medicinal Plants of Korea, Academic Publishing Co., Seoul, Korea (1989) 4. Yoon KJ, Park SJ. Lee HW, Yook CS. Studies on the constitutents and their antibacterial effect of the roof of Lithospermum erythrorhizon Sieb. et Zucc. Bul1. K. H. Pharm. Sci. 16: 155-161 (1988) 5. Park YH, Nam YJ. The antibacterial activity and deodorization of fabrics dyed with Lithospermi Radix extract. J. Korean Soc. Cloth. Text. 27: 60-66 (2003) 6. So HO, Joo YJ. The effect of color fastness on agent in the gromwell dyeing. J. Korean Soc. Cloth. Text. 14: 133-144 (1990) 7. Choi H. Shin YS. Analysis of charactristics and dyeing properties of gromwell colornts (Part 2)-Dyeing properties of silk on gromwel colorants. J. Korean Soc. Cloth. Text. 26: 124-132 (2002) 8. Cho KR. Studies on the natural dyes (II)-dyeing of silk fibers by gromwell color matter, J. Korean Soc. Cloth. Text. 13: 370-379 (1989) 9. Park UY. Jang DS. Cho HR. Antimicrobial effect of Lithospermi radix (Lithospermum erythrorhizon) extract. J. Korean Soc. Food Nutr. 21: 97-100 (1992) 10. Lee HB. Oh MH. Lee JH. Isolation and identification of red color pigments from the Korean Lithospermum erythrorhizon. Korean J. Food Nutr. 13: 379-382 (2000) 11. James GC. Sherman J. Chemotheraperapeutic Agent in Microbiology, a Laboratory Manual Chemical Agents of Control. Eagan Press, St. Paul, MN, USA. pp. 247-254 (1987) 12. Karapinar M. Inhibitiory effects of anethole and eugenol on the growth and toxin production of Aspergillus parasiticus. Int. J. Food Microbiol. 10: 193-200 (1990) 13. Kang SK. Isolation and antimicrobial activity of antimicrobial substance obtained from leaf Mustard (Brassica juncea). Korean J. Food Sci. Technol. 24: 697-698 (1995) 14. Kim SY. Effect of wildlife plants addition on the preservation of bread and rice cake. PhD thesis, Pukyong National University, Busan, Korea (1997) 15. Hong ND, Noh YS, Kim NJ, Kim JS. A study on efficacy of Ulmi Cortex, Korean J. Pharmacog. 21: 217-223 (1990) 16. Chung HJ. Antioxidative and antimicrobial activities of Opuntia ficus indica var. saboten. Korean J. Soc. Food Sci. 16: 164-169 (2000) 17. Park SK, Park JR, Lee SW, Seo KI., Kang SK, Shim KH. Antimicrabial activity and heat stability of water-pretreated extract of leaf mustard dolsan (Brassica juncea). J. Korean Soc. Food Nutr. 24: 710-715 (1995) 18. Jeon YO, Kim KH, Kim SI, Han YS. Screening of antimicrobial activity of the plantain (Plantago asiatica L.) extract. Korean J. Soc. Food Sci. 14: 39-45 (1998) 19. Shin DH, Kim MS, Han JS. Antimicrobial effect of ethanol extracts from some medicinal herbs and their fractionated against food borne bacteria. Korean J Food Sci. Technol. 29: 808-816 (1997) 20. Chung DO, Jung JH. Studies on antimicrobial substances of Ganoderma lucidum. Korean J. Food Sci. Technol. 24: 552-557 (1992) (2004j 4ú 24¢ %>; 2004j 8ú 17¢ j).

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