Evaluation of Dry Rehydratable Film Method for Enumeration of Microorganisms in Korean Traditional Foods

Transcription

1 J. Fd Hyg. Safety 19(4), (2004) w m t w v sƒ ½ Á Áw 1 Á 2 Á ³ 2 Áy t 3 Á½ v 3 Á w œw w tœw w t œw w z Evaluation of Dry Rehydratable Film Method for Enumeration of Microorganisms in Korean Traditional Foods Kwan-Sik Kim, Eun-Kyung Bae, Sang Do Ha 1, Young Seo Park 2, Chul Kyoon Mok 2, Kwan Pyo Hong 3, Sang Phil Kim 3, and Jiyong Park %FQBSUNFOUPG#JPUFDIOPMPHZ:POTFJ6OJWFSTJUZ %FQBSUNFOUPG'PPE4DJFODFBOE5FDIOPMPHZ$IVOH"OH6OJWFSTJUZ %FQBSUNFOUPG'PPEBOE#JPFOHJOFFSJOH,ZVOHXPO6OJWFSTJUZ.,PSFB$PNQBOZ ABSTRACT Dry rehydratable film methods were compared to conventional methods for the enumeration of microorganisms in Korean traditional foods. Kimchi, doenjang, kochujang, kanjang, takju, sujeongkwa and sikhe were used as Korean traditional foods. Petrifilm TM aerobic count plate, Petrifilm TM coliform count plate, Petrifilm TM E. coli/coliform count plate, Petrifilm TM yeast and mold count plate and Petrifilm TM staph express count plate were compared to plate count agar, most probable number (MPN) for coliform, MPN for E. coli, potato dextrose agar and coagulase test, respectively. Regression analysis indicated that correlation coefficient values were , , , and for total aerobic bacteria, yeast and mold, coliform, E. coli and S. aureus, respectively. There were no significant differences between two methods, suggesting that Petrifilm TM plates can be used as an alternative to conventional method for the determination of microorganisms in Korean traditional foods. Key words: microbial isolation, dry rehydratable film method, conventional method, 3M Petrifilm TM t ³, ³ ³, y s ³, z q w y x t sƒw w. t 1) ³ y w ³ w t sq m š ù ƒ š. w wš x z Petrifilm TM aerobic count plate (PAC, 3M Center, St. Paul, MN, USA), Simplate TM TPC (Biocontrol Systems, Inc., Bellevue, WA, USA). Petrifilm TM w, w š t w ³ d w. ³ ³ d w x MPN, Violet Red Bile Agar (VRBA), 2) Anderson- Baird-Parker MPN ƒ 3) Author to whom correspondence should be addressed. š. ù MPN w ³ d 4,5) 6» w w z û. ³ w Petrifilm TM Association of Official Analytical Council (AOAC) w œ x t 48 ü ³ ³ y w. z q w ³, w» w m x ƒ. ù ü ³ w š, w z. w w w» w 3,6-8) dichloran rose bengal chlortetracycline agar (DRBC), 9) oxytetracycline glucose yeast extract agar (OGYE), 10) molybdate agar ƒ 11). ù œm q w j ƒw» t w z q w š. 3M Petrifilm TM yeast and mold count plate (PYM) z q wù d w. 209

2 210,XBO4JL,JNFUBM y s ³ w FDA Baird- Parker (B-P) agar wš ù, 12) B-P agar p û» ³ w y s ³ g w»ƒ š. w ƒ šƒ š 13) y x v w y x w ³ ƒ w ƒ w. w 14) B-P agar w Petrifilm TM Staph express count plate (STX, 3M Center, St. Paul, MN, USA), y s ³ w staphylococcal w ƒ wz w w p š x» k.» m x ƒ š w wš x y w, w t Petrifilm» x w ü TM w š, ù m t 15-17) w w. x ù m t š š š yw y x w v ƒ. ƒ» x w yw x» w» x sƒ ƒ. ù m t Petrifilm w v» x TM mw tü y x wwš w v z sƒw. x t z t m w, z t ½e(kimchi), š (kochujang), (doenjang), (kanjang) w š, m x(sikhe), (sujeongkwa), k (takju) w. t w. ³ E. coli (ATCC 25922) S. aureus (ATCC 12600) w w, ü ³ w. t w ³ z, q w t 37 o C» w 12 w. ½e, k, x, t Whatman No.2 filter membrane w w z šx w. r p, š t t 9 ƒ ³ stomacher bag š stomacher w 120 ³ yw z w x w. ³ ³, y s ³ ½e, k, x, t Whatman No.2 filter membrane w w z, šx wš, r p t šx t» stomacher w 120 ³ yw w ƒ 10 2, 10 3, 10 4, 10 5, 10 6, 10 CFU/mL ³ 7 ( ³ ³ ) y s ³(y s ³ ) w z 1 37 C w o w x w. m w t w tœ 18) w w. ³ PCA w 37 o C 24 w z ³ d w. Coliform d lactose broth (LB, Difco, MI, USA) w š, 3-tube series MPN w. LB broth tube wš, 35 o C 48 w z durham tube»sƒ x q w. q x ƒ š tœ 18) y t w coliform d w. E. coli d AOAC œ EC broth (EC, Difco, MI, USA) w MPN w. EC broth durham tube swwš EC broth tube w z 44.5 o C 24 w ƒ ykw wš durham tube»sƒ s E. coli presumptive positive q w. z q d ³ d w xw., ³ 10% ³ ƒw ph 3.5 w PDA w 25 o C 5-7 w z w w. y s ³ d 10% NaCl ƒw tryptic soy broth 37 o C 16 ³ w z, ùy ƒ m w w 37 o C w. ùy ƒ m w y n ( m w) ùkùš, ykw y(ùy ) y x w. v w ³ PAC, ³ ³ ƒƒ PCC, PEC w š, z q

3 &WBMVBUJPOPG%SZ3FIZESBUBCMF'JMN.FUIPEGPS&OVNFSBUJPOPG.JDSPPSHBOJTNTJO,PSFBO5SBEJUJPOBM'PPET 211 PYM, y s ³ STX ƒƒ w. v w 3M w. ³ d PAC v v š ü w q r z 35 o C z colony w. Coliform d filmü w z 35 o C 24 z ƒ g w. E. coli d w z, 35 o C 24 w ƒ g w. z q d PYM w z 35 o C 5-7 w z vj š ƒ y z ³ w š, w j œ wš ƒ y q ³ w. y s ³ STX w z 35 o C 24 w z Petrifilm TM staph express disk wš 35 o C 3 w z vj y ùkù g w. m ƒ w x 3 w s³ wš CFU/mL log10 yw m w. tœ w Petrifilm TM w xz w w. z Excel 2000 software package (Microsoft Corp., Redmond, USA) w wš l (r),»»(slope), r(intercept) l x w. š ³ ³ PCA w t w v PAC w w. PCA w t w k t w ü w ³ w, PAC 2 film w film k š, film y 2,3,5-triphenyltetrazolium chloride (TTC), film w z 35 o C w z colony ³ w. ³ d w t w. PAC w ³ log x wš PCA w ³ log y w w d ³ plottingw w z»», r Table 1 ùkü, t d ƒ ùkù. t ü w xƒ ƒ û ƒ ƒ. t z»» w ùkü. t ³ log s³ PAC 6.09, PCA 6.07 ƒ. PAC ³ w š ½e k 80% ƒ PAC ³ w. MacAllister 15) Ginn 16) PCA w PAC t l ³ w Table 1. Comparison of Petrifilm TM aerobic count plate (PAC) and plate count agar (PCA) methods for the enumeration of total aerobic bacteria in Korean traditional foods Mean log (CFU/mL)±S.D. PAC=PCA Sample N 1) r 2) a 3) b 4) P 5) PAC PCA PAC>PCA PAC<PCA Kimchi ± ± (0.0) 37(80.4) 9(19.6) Doenjang ± ±0.74 1(2.5) 16(41.0) 22(56.5) Kochujang ± ± (0.0) 11(36.7) 19(63.3) Kanjang ± ± (0.0) 27 (100.0) 0 (0.0) Takju ± ± (0.0) 23(79.3) 6(20.7) Sujeongkwa ± ±0.57 1(3.4) 12(41.4) 16(55.2) Sikhe ± ±0.74 1(3.6) 14(50.0) 13(46.4)

4 212,XBO4JL,JNFUBM ü P<0.01 š šw, MacAllister t l ³ 15) w ü PCA plate PAC ƒ w š š w. Ha 17), š», k, ƒ, w ³ PCA plate w Petrifilm TM aerobic count plate w d w ƒ x w ³ d PCA plate w w ykw ³ d Petrifilm TM aerobic count plate TTC w colonyƒ d w» l ³ d w PCA plate w Petrifilm TM aerobic count plateƒ w š w. x x ww PAC w x PCA w x ƒ š ƒ» PCA w š q. ³ ³ MPN PCC w w. t ³ CFU/ ml w 37 C w w z o w ³ d w. ³ d w t w š z»», r Table 2 ùkü. t ƒ ùkù. t ü w z»» w ùkü. ³ log s³ r t ³ log s³ PCC 3.92, MPN w 3.81 PCC. k PCC MPN ³ w. PCC MPN w r ƒ w, MPN PCC ³ w š, š» dryfilm w MPN l ³ šƒ. 19) ù MPN y w ³ s³e m w w» PCC repeatability reproducibility variance MPN š šw. w PCC MPN w š r w. wr Ha 17) š» w VRBA (Difco) PCC 0.91 ù PCC VRBA (Difco) w P<0.05 û ³ w ü, l ³ BGLB (Difco) w MPN PCC w ƒ ù PCC MPN ³ w ü š šw w. Nelson 20) ƒ û ³ PCC ³ š û sƒ ³ ƒ š š w ù t ³ log s³ PCC. wr Park 21) š» š», š» w ³ PCC MPN w ƒ š» 0.83, š» 0.96, š» 0.81 ùkü PCC ƒ œ d m MPN w š šw. x x ww PCC w Table 2. Comparison of Petrifilm TM coliform count plate (PCC) and most probable number (MPN) methods for the enumeration of coliform in Korean traditional foods Mean log (CFU/mL)±S.D. PCC=MPN Sample N 1) r 2) a 3) b 4) P 5) PCC MPN PCC>MPN PCC<MPN Doenjang ± ±1.85 2(11.1) 16(88.9) 0(0.0) Kanjang ± ±1.86 2(4.9) 31(75.6) 8(19.5) Takju ± ± (0.0) 27 (100.0) 0(0.0) Sikhe ± ±1.44 1(4.8) 6(28.6) 14(66.6)

5 &WBMVBUJPOPG%SZ3FIZESBUBCMF'JMN.FUIPEGPS&OVNFSBUJPOPG.JDSPPSHBOJTNTJO,PSFBO5SBEJUJPOBM'PPET 213 x MPN w x ƒ ùk ù MPN w ùkü y. PCC d y w š q. ³ ³ d w t w š z»», r Table 3 ùkü. t MPN PEC ƒ ùkù. t ü w z»» w ùkü. w t ³ log s³ PEC 4.06, MPN w 4.05, ³ ƒ PEC MPN w. Suwansonthichai 22) w ³ ³ MPN ƒ, chromocult coliform agar (CCA), Fluorocult LMX broth (LMX), PEC w m MPN ƒ w. PEC w d ³ MPN ³ 95.7% ùkû. MPN w ³ 4% yw y x mw ³ y PEC w ³ 1.7% ³ y. w LMX CCA w ³ ƒƒ 8.0% 2.9%ƒ ³ y. x r PEC>CCA=LMX>MPN š w, MPN>LMX=PEC=CCA ¼š, MPN>LMX=CCA>PEC š šw MPN w PEC. x w PEC MPN ùkü z û MPN w q w. z q z q w t w š, z»», r Table 4 ùkü t ƒ ùkù, t ü w š, z»» w ùkü. ƒ t PYM PDA z ƒ, z log s³ PYM 4.32, PDA w 4.41 PDA sƒ. PDA PYM k û ³ ³ w» q. Knight 23) w,, f,, p, f j 6ƒ t 4 q 2 z w w t w z q PYM Food and Drug Administration Bacteriological Analytical Manual œ w d w m ƒ w š šw. l t l z q w» w PYM PDA ƒ, ƒ» m tü r wš k PYM PDA w š q. y s ³ y s ³ w t 30- Table 3. Comparison of Petrifilm TM E. coli count plate (PEC) and most probable number (MPN) methods for the enumeration of E. coli in Korean traditional foods Mean log (CFU/mL)±S.D. PEC=MPN Sample N 1) r 2) a 3) b 4) P 5) PEC MPN PEC>MPN PEC<MPN Doenjang ± ± (0.0) 0 (0.0) 17(100.0) Kanjang ± ±1.54 2(5.3) 28(73.7) 8(21.0) Sujeongkwa ± ± (0.0) 9(25.0) 27(75.0)

6 214,XBO4JL,JNFUBM 60 w š z»», r Table 5 ù kü ƒ ùkù. t ü w š z»» w ùkü. ƒ t 58.3% coagulase x STX y s ³, y s ³ log s³ STX 3.32, coagulase x w 3.29 STX. Baird-Parker (B-P) t (FDA) kwš t w t w S. aureus ³ B-P n y w» ƒ y x v w, y x w w ³ 24) w ƒ w š. w 14) De Buyser 13) B-P ƒ k û ³ w» S. aureus ³ w» š šw. Schoeller š» e 24) B-P STX w S. aureus w w B-P w 84³ 73% S. aureusƒ y w e S. aureus w z w ³ y w w STX B- P w š w. ww t l y s ³ w» w STX tœ coagulase Table 4. Comparison of Petrifilm TM yeast and mold count plate (PYM) and potato dextrose agar (PDA) methods for the enumeration of yeast and mold in Korean traditional foods Sample N 1) r 2) a 3) b 4) P 5) ±S.D. Mean log (CFU/mL) PYM PDA PYM=PDA PYM>PDA PYM<PDA Kimchi ± ± (0.0) 4(14.8) 23(85.2) Doenjang ± ± (0.0) 2(7.4) 25(92.6) Kochujang ± ± (0.0) 2(7.4) 25(92.6) Kanjang ± ± (0.0) 2(7.4) 25(92.6) Takju ± ±0.90 1(3.7) 5(18.5) 21(77.8) Sujeongkwa ± ± (0.0) 5(21.7) 18(78.3) Sikhe ± ± (0.0) 5(18.5) 22(81.5) Table 5. Comparison of Petrifilm TM staph express count plate (STX) and coagulase methods for the enumeration of Staphylococcus aureus in Korean traditional foods Sample N 1) r 2) a 3) b 4) P 5) =Coagulase Mean log (CFU/mL)±S.D. STX STX Coagulase STX >Coagulase STX <Coagulase Kimchi ± ±1.76 0(0.0) 41(68.3) 19(31.7) Doenjang ± ±1.74 3(10.0) 17(56.7) 10(33.3) Kochujang ± ±1.77 2(6.7) 12(40.0) 16(53.3) Kanjang ± ±1.77 0(0.0) 19(63.3) 11(36.7) Takju ± ±1.76 4(13.3) 14(46.7) 12(40.0) Sujeongkwa ± ±1.83 0(0.0) 19(63.3) 11(36.7) Sikhe ± ±1.83 0(0.0) 27(90.0) 3(10.0)

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