Determination of Heterocyclic Amines in Roasted Fish and Shellfish by Liquid Chromatography-Electrospray Ionization/Mass Spectrometry

Liquid chromatography-mass spectrometry를 이용한 가열 조리된 어패류에서의 heterocyclic amines 함량 분석

  • Lee, Jae-Hwan (Department of Food Science and Technology and Institute of Lotus Functional Food Ingredient, Dongguk University) ;
  • Back, Yoo-Mi (Department of Food Science and Technology and Institute of Lotus Functional Food Ingredient, Dongguk University) ;
  • Lee, Kwang-Geun (Department of Food Science and Technology and Institute of Lotus Functional Food Ingredient, Dongguk University) ;
  • Shin, Han-Seung (Department of Food Science and Technology and Institute of Lotus Functional Food Ingredient, Dongguk University)
  • 이재환 (동국대학교 식품공학과 및 Lotus기능성식품소재연구소) ;
  • 백유미 (동국대학교 식품공학과 및 Lotus기능성식품소재연구소) ;
  • 이광근 (동국대학교 식품공학과 및 Lotus기능성식품소재연구소) ;
  • 신한승 (동국대학교 식품공학과 및 Lotus기능성식품소재연구소)
  • Published : 2009.06.30

Abstract

Heterocyclic aromatic amines (HCAs) are mutagenic and carcinogenic substances that are formed during the heating of protein-rich foods. HCAs are generally found at low amounts in a complex matrix, which requires sophisticated analysis. In this study, HCAs were extracted from lyophilized fish and shellfish samples using solid-phase extraction (SPE) and determined by liquid chromatography-electrospray ionization/mass spectrometry (LC-ESI-MS). The HCA recoveries in the fish and shellfish ranged from 15.7 to 74.7% with standard deviations from 0.2 to 7.63%. And HCA concentrations ranged from 0.8 to 1,117.7 $ng/g^{-1}$ in cooked food samples. 1-methyl-9H-pyrido[3,4-b]indole (Harman), 9H-pyrido[3,4-b]indole (Norharman), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) were the most abundant HCAs formed in the muscle of fried mackerel, at levels of 1,117.7, 926.6, and 133.7 ng/g, respectively. 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), 2-aminodipiryrido[1,2-a:3,2-d]imidazole(Glu-P-2), 2-amino-9H-pyrido[2,3-b]indole(A${\alpha}$C), 2-amino-3methyl-9H-pyrido [1,2-a:3,2-d]imidazole(MeA${\alpha}$C), 2-amino-3,4,7,8-tetramethylimidazo[4,5-f]quinoxaline (TriMeIQx), 2-amino-3,7,8-trimethylimidazo [4,5-f]quinoxaline(7,8-DiMeIQx), and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) were only detected by small quantities ranged from 1.5 to 98.6 ng/g. Overall, this study provides useful information on HCA levels in fish and shellfish products consumed in Korea.

국내에서 주로 섭취되는 어패류(고등어, 삼치, 조기, 대합, 바지락, 꼬막, 백합)를 가열조리 하였을 때 형성되는 HCAs를 분리, 동정하여 그 함량을 모니터링 하였다. 추출 및 정제는 고체상 추출 방법을 사용하였고 정성, 정량을 위한 분석장비는 LC/MS를 사용하였다. 모니터링 결과 생선류에서는 고등어(2.2-1,1117.7 ng/g), 삼치(9.1-443.6 ng/g), 조기(17.5-179.5 ng/g)순으로 HCAs가 많이 검출되었으며 특히 고등어 muscle에서 많은 양(1,1117.7 ng/g, Harman)이 검출되었다. 조개류 중에서는 대합(12.2-196.4 ng/g), 꼬막(2.2-76.2 ng/g), 바지락(1.7-25.8 ng/g), 백합(1.5-2.7 ng/g)순으로 많이 검출되었으며 대합의 Norharman(196.4 ng/g)이 가장 많이 검출되었다. 15종의 HCAs 중 ${beta}$-carbolines에 속하는 Norharman과 Harman이주로 검출되었으며 그 밖에 PhIP, Trp-P-1. Trp-P-2 등의 발암가능성이 있는 물질들이 검출되었다. 조리방법 측면에서 보면 간장을 이용한 조림방법보다는 직접적인 가열을 통한 구이방법이 많은 양의 HCAs를 형성시켰으며 구이 중에서도 muscle만 있는 부위에서의 검출함량보다는 muscle과 skin이 함께 있는 부위에서 검출함량이 더 많았다. 가열 조리한 어패류 분석방법에 대한 유효성을 확인해 본 결과 정량한계는 0.8-23.9 ng/mL, 검출한계는 0.2-7.2 ng/mL이었으며 회수율은 15.7-74.7%이었다.

Keywords

References

  1. Matsukura N, Kawachi T, Morino K, Ohgaki H, Sugimura T, Takayama S. Carcinogenicity in mice of mutagenic compounds from a tryptophan pyrolysate. Science 213: 346-347 (1981) https://doi.org/10.1126/science.7244619
  2. Shin HS, Lee YS. Influence of commercial marinades on heterocyclic aromatic amine formation and overall mutagenicity in fried beef steaks. Food Sci. Biotechnol. 14: 323-327 (2005)
  3. Sugimura T. Food and cancer. Toxicology 182: 17-21 (2002) https://doi.org/10.1016/S0300-483X(02)00250-0
  4. Ohgaki H, Takayama S, Sugimura T. Carcinogenicities of heterocyclic amines. Mutat. Res. 259: 399-410 (1991) https://doi.org/10.1016/0165-1218(91)90130-E
  5. Jagerstad M, Reutersward AL, Olsson R, Grivas S, Nyhammar T, Olsson K, Dahlqvist A. Creatin(in)e and maillard reaction products as precursors of mutagenic compounds: Effects of various amino acids. Food Chem. 12: 239-44 (1983)
  6. Sugimura T, Kawachi T, Nagao M, Yahagi T, Seino Y, Okamato T, Shudo K, Kosuge T, Tsuji K, Wakabayashi K, Iitaka Y, Itai A. Mutagenic principle(s) in tryptophan and phenylalanine pyrolysis products. P. Jpn. Acad. 53: 58-61 (1977)
  7. Becher G, Knize MG, Nes IF, Felton JS. Isolation and identification of mutagens from a fried Norwegian meat product. Carcinogenesis 9: 247-1253 (1988) https://doi.org/10.1093/carcin/9.2.247
  8. Hatch FT, Felton JS, Knize MG. Mutagens Formed in Foods During Cooking. ICI Atlas of Science: Pharmacology. ISI Press, Philadelphia, PA, USA. pp. 222-228 (1988)
  9. Knize MG, Hopmans E, Happe JA. The identification of a new heterocyclic amine mutagen from a heated mixture of creatine, glutamic acid, and glucose. Mutat. Res. 260: 313-319 (1991) https://doi.org/10.1016/0165-1218(91)90016-F
  10. Sugimura T, Wakabayashi K. Mutagens and carcinogens in food. Prog. Clin. Biol. Res. 347: 1-18 (1990)
  11. Augustsson K, Skog K, Jagerstad M, Steineck G. Assessment of the human exposure to heterocyclic amines. Carcinogenesis 18: 1931-1935 (1997) https://doi.org/10.1093/carcin/18.10.1931
  12. Rohrmann S, Becker N. Development of a short questionnaire to assess the dietary intake of heterocyclic aromatic amines. Public Health Nutr. 5: 699-705 (2002)
  13. Felton JS, Knize MG. New mutagens from cooked food. Prog. Clin. Biol. Res. 347: 19-38 (1990)
  14. Yamashita K, Umemoto A, Grivas S, Kato S, Sato S, Sugimura T. Heterocyclic amines -DNA adducts analysed by 32 P-postlabeling method. Nucleic Acids Res. Symp. Ser. 14: 111-114 (1988)
  15. Skog K. Cooking procedures and food mutagens: A literature review. Food Chem. Toxicol. 31: 655-675 (1993) https://doi.org/10.1016/0278-6915(93)90049-5
  16. Aoyama S, Yamamoto Y. Antioxidant activity and flavonoid content of Welsh onion (Allium fistulosum) and the effect of thermal treatment. Food Sci. Technol. Res. 13: 67-72 (2007) https://doi.org/10.3136/fstr.13.67
  17. Arimoto S, Hayatsu H. Role of hemin in the inhibition of mutagenic activity of 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) and other aminoazaarenes. Mutat. Res. 213: 217-226 (1989) https://doi.org/10.1016/0027-5107(89)90153-X
  18. Balogh Z, Gray JI, Gomaa EA, Booren AM. Formation and inhibition of heterocyclic aromatic amines in fried ground beef patties. Food Chem. Toxicol. 38: 395-401 (2000) https://doi.org/10.1016/S0278-6915(00)00010-7
  19. Gibis M. Effect of oil marinades with garlic, onion, and lemon juice on the Formation of heterocyclic aromatic amines in fried beef patties. J. Agr. Food Chem. 55: 10240–10247 (2007) https://doi.org/10.1021/jf071720t
  20. Ha YL, Grimm NK, Pariza MW. Anticarcinogens from fried ground beef: heat-altered derivatives of linoleic acid. Carcinogenesis 8: 1881-1887 (1987) https://doi.org/10.1093/carcin/8.12.1881
  21. Lee JH, Shin HS. Influence of genotoxic heterocyclic aromatic amine formation and overall mutagenicity in ground beef patties using korean bramble (Rubus coreanum Miquel). Food Sci. Biotechnol. 16: 576-579 (2007)
  22. Holder CL, Preece SW, Conway SC, Pu YM, Doerge DR. Quantification of heterocyclic amine carcinogens in cooked meats using isotope dilution liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry. Rapid Commun. Mass Sp. 11: 1667-1672 (1997) https://doi.org/10.1002/(SICI)1097-0231(19971015)11:15<1667::AID-RCM58>3.0.CO;2-N
  23. Pais P, Knize MG. Chromatographic and related techniques for the determination of aromatic heterocyclic amines in foods. J. Chromatogr. B 747: 139-169 (2000) https://doi.org/10.1016/S0378-4347(00)00118-3
  24. Richling E, Decker C, Haring D, Herderich M, Schreier P. Analysis of heterocyclic aromatic amines in wine by high-performance liquid chromatography-electrospray tandem mass spectrometry. J. Chromatogr. A 791: 71-77 (1997)
  25. Gross GA, Gruter A. Quantitation of mutagenic/carcinogenic heterocyclic amines in food products. J. Chromatogr. 592: 271-278 (1992) https://doi.org/10.1016/0021-9673(92)85095-B
  26. Tai CY, Lee KH, Chen BH. Effects of various additives on the formation of heterocyclic amines in fried fish fibre. Food Chem. 75: 309-316 (2001) https://doi.org/10.1016/S0308-8146(01)00200-X
  27. Totsuka T, Ushiyama H, Ishihara J, Sinha R, Goto S, Sugimura T, Wakabayashi K. Quantification of the co-mutagenic ${\beta}-carbolines$, norharman and harman, in cigarette smoke condensates and cooked foods. Cancer Lett. 143: 139-143 (1999) https://doi.org/10.1016/S0304-3835(99)00143-3
  28. Sugimura T, Wakabayashi K, Nakagama H, Nagao M. Heterocyclic amines: Mutagens/carcinogens produced during cooking of meat and fish. Cancer Sci. 95: 290-299 (2004) https://doi.org/10.1111/j.1349-7006.2004.tb03205.x
  29. Lan CM, Chen BH. Effects of soy sauce and sugar on the formation of heterocyclic amines in marinated foods. Food Chem. Toxicol. 40: 989-1000 (2002) https://doi.org/10.1016/S0278-6915(02)00013-3
  30. Harris DC. Quantitative Chemical Analysis. 7th ed. W. H. Freeman and Company, New York, NY, USA. pp. 78-95 (2006)
  31. Sentellas S, Moyano E, Puignou L, Galceran MT. Optimization of a clean-up procedure for the determination of heterocyclic aromatic amines in urine by field-amplified sample injection-capillary electrophoresis-mass spectrometry. J. Chromatogr. A 1032: 193-201 (2004) https://doi.org/10.1016/j.chroma.2003.11.011
  32. Vollenbroker M, Eichner K. A new quick solid-phase extraction method for the quantification of heterocyclic aromatic amines. Eur. Food Res. Technol. 212: 122-125 (2000) https://doi.org/10.1007/s002170000188
  33. Abdulkarim BG, Smith JS. Heterocyclic amines in fresh and processed meat products. J. Agr. Food Chem. 46: 4680-4687 (1998) https://doi.org/10.1021/jf980175g
  34. Knize MG, Salmon CP, Hopmans EC, Felton JS. Analysis of foods for heterocyclic aromatic amine carcinogens by solid-phase extraction and high-performance liquid chromatography. J. Chromatogr. A 763: 179-185 (1997) https://doi.org/10.1016/S0021-9673(96)00720-0