Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
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Thermal Stable Soluble Proteins in Pork Fat and Meat, and Their Antigenicity

돈지육 및 돈육 중 열안정성 단백질의 존재 확인과 이의 항원성 확인

  • Kim, Jeong-Sook (Division of Applied Life Science, Graduate School, Gyeongsang National University) ;
  • Lee, Jeong-Eun (Division of Applied Life Science, Graduate School, Gyeongsang National University) ;
  • Shim, Won-Bo (Department of Agricultural Chemistry and Food Science & Technology, Gyeongsang National University)
  • 김정숙 (경상대학교 응용생명과학부) ;
  • 이정은 (경상대학교 응용생명과학부) ;
  • 심원보 (경상대학교 농화학식품공학과)
  • Received : 2016.08.30
  • Accepted : 2016.10.14
  • Published : 2016.12.30
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Abstract

Thermal-stable soluble proteins (TSSP) in livestock products has been recently reported. Therefore, the development of antibodies and immunoassay using a TSSP is useful because the presence of TSSP can be measured on processed food. In this study, the existence of TSSPs in pork fat and meat was confirmed and their antigenicity was investigated. The extracts from pork fat and meat by heating method were analyzed by SDS-PAGE with 5% stacking and 12% separating gels. The protein profiles from the raw pork fat and meat extracts (major band ranged 25 to 100 kDa) without cooking and heating treatments were significantly different compared to those from cooked and heated pork fat and meat extracts (several major bands > 100 kDa and < 30 kDa). This meant that non thermal-stable soluble proteins ranged from 25 to 100 kDa may be denaturated to insoluble proteins by cooking and heating treatments, and TSSPs were in pork fat and meat at kept their properties. The confirmed TSSPs were used as an immunogen to investigate their antigenicity. Eight mice (5 mice for pork fat and 3 mice for pork meat) were separately immunized with the TSSPs of pork fat and meat, and the anti-sera obtained from the immunized mice showed high titer values. Polyclonal antibodies against each target protein showed the specific reaction to pork fat and meat, individually. These indicated that TSSP could be used as an immunogen to produce antibodies such as monoclonal and polyclonal antibodies. In addition, antibodies specific to TSSP from pork fat and meat may be used as a bio-receptor in immunoassays for the identification of fraudulent adulteration with pork fat and meat in livestock products.

본 연구에서는 돈지육 및 돈육 조직 내에 열안정성 수용성 단백질의 존재 여부를 확인하고 항체 생산에 있어 항원으로의 사용 가능 여부를 확인하고자 하였다. 이를 위해 돈지육 및 돈육을 생(raw) 시료와 조리된(cooked) 시료로 구분하여 비열처리 및 열처리법으로 단백질을 추출한 후 단백질 존재여부를 단백질 정량과 SDS-PAGE로 확인하였다. 그 결과 돈지육과 돈육 모두 생 시료를 비열처리법으로 추출한 시료의 경우 25~100 kDa 사이의 다양한 단백질이 확인된 반면 시료를 가열하거나 추출 시 열처리를 한 경우 돈지육에는 100 kDa 이상의 단백질과 30 kDa 및 15 kDa 이하의 일부 단백질이, 돈육에는 100 kDa 이상과 30 kDa 이하의 단백질이 확인되어 돈지육과 돈육에 열안정성 수용성 단백질이 존재하는 것으로 확인되었다. 이들 열안정성 수용성 단백질을 마우스에 면역 후 항혈청 역가를 측정한 결과 면역한 모든 마우스에서 높은 역가를 나타내었고, 생산된 혈청은 돈지육과 돈육에 각각 특이적인 반응성을 보인 반면 다른 축육과 지방육에 대해서는 반응성이 상대적으로 낮았다. 이러한 연구결과를 볼 때 돈지육 및 돈육에 존재하는 열안정성 수용성 단백질이 돈지육과 돈육에 특이적으로 반응하는 항체를 개발하는데 유용한 마커로서 활용이 가능하며, 열안정성 수용성 단백질에 대한 항체개발은 열처리된 축육 가공품 중 돈지육 및 돈육의 분석에도 매우 유용하게 활용할 수 있을 것으로 판단된다.

Keywords

References

  1. MFDS, A study for quantitative analysis method of meat and meat products. research report (2013).
  2. Available from http://www.mt.co.kr/view/mtview.php?type=1&no=2011070512384035352&outlink=1, Accessed Jul. 14 (2016).
  3. Available from http://www.foodsafetynews.com/2013/02/ukstepping-up-hunt-for-horse-pig-dna-in-the-beef/#.V4eWXIaweUk, Accessed Jul. 14 (2016).
  4. Available from http://www.foodsafetynews.com/2014/05/recallpork-nda-traces-found-incadbury-chocolate-in-malaysia/#.V4eWvYaweUk, Accessed Jul. 14 (2016).
  5. Heo, E.J., Ko, E.K., Seo, K.H., Kim, Y.J., Park, H.J., Wee, S.H. and Moon, J.S.: Validation of PCR and ELISA test kits for identification of domestic animal species in raw meat and meat products in Korea. J. Fd Hyg. Safety, 29, 158-163 (2014). https://doi.org/10.13103/JFHS.2014.29.2.158
  6. Soares, S., Amaral, J.S., Mafra, I. and Oliveira, M.B.P.: Quantitative detection of poultry meat adulteration with pork by a duplex PCR assay. Meat Sci., 85, 531-536 (2010). https://doi.org/10.1016/j.meatsci.2010.03.001
  7. Aida, A.A., Man, Y.C., Wong, C.M.V.L., Raha, A.R. and Son, R.: Analysis of raw meats and fats of pigs using polymerase chain reaction for Halal authentication. Meat Sci., 69, 47-52 (2005). https://doi.org/10.1016/j.meatsci.2004.06.020
  8. Aristoy, M.C. and Toldra, F.: Histidine dipeptides HPLCbased test for the detection of mammalian origin proteins in feeds for ruminants. Meat Sci., 67, 211-217 (2004). https://doi.org/10.1016/j.meatsci.2003.10.008
  9. Abbas, O., Pierna, J.F., Codony, R., Von Holst, C. and Baeten, V.: Assessment of the discrimination of animal fat by FTRaman spectroscopy. J. Mol. Struct., 924, 294-300 (2009).
  10. Man, Y.C., Gan, H.L., NorAini, I., Nazimah, S.A.H. and Tan, C.P.: Detection of lard adulteration in RBD palm olein using an electronic nose. Food Chem., 90, 829-835 (2005). https://doi.org/10.1016/j.foodchem.2004.05.062
  11. Kim, K.H., Kim, M.R., Park, Y.E., Kim, Y.S., Lee, H.Y., Park, Y.C., Kim, S.Y., Choi, J.D. and Jang, Y.M.: Detection and differentiation of intentional and unintentional mixture in raw meats using real-time PCR. J. Fd Hyg. Safety, 29, 340-346 (2014). https://doi.org/10.13103/JFHS.2014.29.4.340
  12. Chen, F.C. and Hsieh, Y-H.P.: Porcine troponin I: a thermostable species marker protein. Meat Sci., 61, 55-60 (2002). https://doi.org/10.1016/S0309-1740(01)00162-0
  13. Ofori, J.A. and Hsieh, Y-H.P.: Characterization of a 12 kDa thermal-stable antigenic protein in bovine blood. J. Food Sci., 76, C1250-C1256 (2011). https://doi.org/10.1111/j.1750-3841.2011.02393.x
  14. Ofori, J.A. and Hsieh, Y-H.P.: Characterization of a 60-kDa thermally stable antigenic protein as a marker for the immunodetection of bovine plasma-derived food ingredients. J. Food Sci., 80, C1654-C1660 (2015). https://doi.org/10.1111/1750-3841.12963
  15. Hsieh, Y-H.P., Ofori, J.A., Rao, Q. and Bridgman, R.C.: Monoclonal antibodies specific to thermostable proteins in animal blood. J. Agric. Food Chem., 55, 6720-6725 (2007). https://doi.org/10.1021/jf070031e
  16. Hsieh, Y-H.P. and Ofori, J.A.: Detection of horse meat contamination in raw and heat-processed meat products. J. Agric. Food Chem., 62, 12536-12544 (2014). https://doi.org/10.1021/jf504032j
  17. Nhari, R., Hafidz, R.M., Hamid, M., Rasli, N.M., Omar, A.R., Sheikha, E. and Mustafa, S.: Monoclonal antibodies specific to heat-treated porcine blood. J. Sci. Food Agric., 96, 2524-2531 (2015).
  18. Chen, F.C., Hsieh, Y-H.P. and Bridgman, R.C.: Monoclonal antibodies to porcine thermal-stable muscle protein for detection of pork in raw and cooked meats. J. Food Sci., 63, 201-205 (1998).
  19. Laemmli, U. K.: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 227, 680-685 (1970). https://doi.org/10.1038/227680a0