雜環胺化合物是指在其化學結構中含有兩種以上不同元素和至少一個胺基（-NRR'）的環狀結構化合物。許多文獻和臨床研究已指出雜環胺化合物具有高度致癌性和致突變性。分子模印技術是基於“抗原－抗體結合理論”的延伸性應用，該技術提供對目標分析物極佳的特異性和識別能力，而利用該技術合成的高分子材料則稱為分子模印聚合物（MIPs）。本篇研究目的為開發具有高度選擇性的分子模印聚合物，並使用液相層析－串聯式質譜技術定量複雜基質中的雜環胺化合物。本研究選擇IQ作為模板分子，並使用兩種官能基單體AMPS和MAA，以及交聯劑EGDMA分別透過總體聚合和沉澱聚合的方式合成分子模印聚合物，並搭配使用掃描式電子顯微鏡觀察兩種分子模印聚合物的表面特徵。之後透過對模板分子IQ的吸附能力來評估與優化合成比例，結果顯示當模板分子：官能基單體：交聯劑的比例為1：8：40時具有最好的吸附效果。此外，我們將兩種分子模印聚合物與固相萃取法結合並優化其萃取條件。實驗結果為使用MAA作為官能基單體所製備的分子模印聚合物不僅對 IQ具有高度選擇性，且MeIQ和8-MeIQx兩個雜環胺也可以被有效地偵測。最後，本實驗開發的方法可以成功應用於真實樣品的分析。

Heterocyclic amines (HCAs) refer to compounds that have ring structure which contains more than two different elements and at least one amine group (-NRR') in their chemical structures. Many studies and literature have pointed out that HCAs are highly carcinogenic and mutagenic. Molecularly imprinted technology is an extending application based on the "antigen-antibody binding theory". This technology provides excellent specificity and recognition ability, and the material synthesized by this method called molecularly imprinted polymers (MIPs).  The aim of this study was to develop the highly specific molecularly imprinted polymers for the purification and quantification of heterocyclic amine compounds in complex matrices using liquid chromatography-tandem mass spectrometry. In this study, IQ was selected as a template molecule, and two functional monomers, AMPS and MAA were used with cross-linking agent EGDMA for the synthesis of molecularly imprinted polymers by means of bulk polymerization and precipitation polymerization. The characteristics and surface of the MIPs were investigated through the scanning electron microscope. Afterwards, the synthesis ratios were evaluated and optimized by their adsorption capacity to the template molecules and the selectivity to other HCAs. As a result, template/functional monomer/cross-linking agent at a ratio of 1:8:40 gave better adsorption effect. Furthermore, two kinds of molecularly imprinted polymers were incorporated with solid-phase extraction and optimized the extraction conditions. The results indicated that excellent selectivity for IQ was obtained using MAA as functional monomer, while MeIQ and 8-MeIQx could also be effectively detected. Finally, our method was successfully applied to the real samples.

1. Nagao, M., et al., Mutagenicities of smoke condensates and the charred surface of fish and meat. Cancer Letters, 1977. 2(4): p. 221-226.
2. Anastassiades, M., et al., Fast and easy multiresidue method employing acetonitrile extraction/partitioning and “dispersive solid-phase extraction” for the determination of pesticide residues in produce. Journal of AOAC international, 2003. 86(2): p. 412-431.
3. Oz, F., Quantitation of heterocyclic aromatic amines in ready to eat meatballs by ultra fast liquid chromatography. Food chemistry, 2011. 126(4): p. 2010-2016.
4. Puangsombat, K., W. Jirapakkul, and J.S. Smith, Inhibitory activity of Asian spices on heterocyclic amines formation in cooked beef patties. Journal of food science, 2011. 76(8).
5. Ristic, A., M. Cichna, and G. Sontag, Determination of less polar heterocyclic aromatic amines in standardised beef extracts and cooked meat consumed in Austria by liquid chromatography and fluorescence detection. Journal of Chromatography B, 2004. 802(1): p. 87-94.
6. Turesky, R.J., et al., Quantitation of carcinogenic heterocyclic aromatic amines and detection of novel heterocyclic aromatic amines in cooked meats and grill scrapings by HPLC/ESI-MS. Journal of agricultural and food chemistry, 2005. 53(8): p. 3248-3258.
7. Toribio, F., et al., Heterocyclic amines in griddled beef steak analysed using a single extract clean-up procedure. Food and Chemical Toxicology, 2007. 45(4): p. 667-675.
8. Gross, G., A. Grüter, and S. Heyland, Optimization of the sensitivity of high-performance liquid chromatography in the detection of heterocyclic aromatic amine mutagens. Food and Chemical Toxicology, 1992. 30(6): p. 491-498.
9. Toribio, F., et al., Ion-trap tandem mass spectrometry for the determination of heterocyclic amines in food. Journal of Chromatography A, 2002. 948(1-2): p. 267-281.
10. Frandsen, H., H. Frederiksen, and J. Alexander, 2-Amino-1-methyl-6-(5-hydroxy-) phenylimidazo [4, 5-b] pyridine (5-OH-PhIP), a biomarker for the genotoxic dose of the heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo [4, 5-b] pyridine (PhIP). Food and chemical toxicology, 2002. 40(8): p. 1125-1130.
11. Oz, F. and M. Kızıl, Determination of heterocyclic aromatic amines in cooked commercial frozen meat products by ultrafast liquid chromatography. Food Analytical Methods, 2013. 6(5): p. 1370-1378.
12. Hodge, J.E., Dehydrated foods, chemistry of browning reactions in model systems. Journal of agricultural and food chemistry, 1953. 1(15): p. 928-943.
13. Martins, S.I., W.M. Jongen, and M.A. Van Boekel, A review of Maillard reaction in food and implications to kinetic modelling. Trends in food science & technology, 2000. 11(9-10): p. 364-373.
14. Skog, K., M. Johansson, and M. Jägerstad, Carcinogenic heterocyclic amines in model systems and cooked foods: a review on formation, occurrence and intake. Food and Chemical Toxicology, 1998. 36(9-10): p. 879-896.
15. Jägerstad, M., et al., Formation of heterocyclic amines using model systems. Mutation Research/Genetic Toxicology, 1991. 259(3-4): p. 219-233.
16. Nagao, M., S. Sato, and T. Sugimura, Mutagens produced by heating foods. 1983, ACS Publications.
17. Ferguson, L.R., M. Philpott, and N. Karunasinghe, Dietary cancer and prevention using antimutagens. Toxicology, 2004. 198(1-3): p. 147-159.
18. Sugimura, T., Mutagen-carcinogens in foods with special reference to highly mutagenic pyrolytic products in broiled foods. Origins of human cancer, 1977.
19. Ohgaki, H., S. Takayama, and T. Sugimura, Carcinogenicities of heterocyclic amines in cooked food. Mutation Research/Genetic Toxicology, 1991. 259(3-4): p. 399-410.
20. Adamson, R.H., et al., Carcinogenicity of 2‐Amino‐3‐methylimidazo [4, 5‐f] quinoline in Nonhuman Primates: Induction of Tumors in Three Macaques. Cancer Science, 1990. 81(1): p. 10-14.
21. Jinap, S., et al., Heterocyclic aromatic amines in deep fried lamb meat: The influence of spices marination and sensory quality. Journal of food science and technology, 2016. 53(3): p. 1411-1417.
22. Bouvard, V., et al., Carcinogenicity of consumption of red and processed meat. The Lancet Oncology, 2015. 16(16): p. 1599-1600.
23. Cancer, I.A.f.R.o., Consumption of red meat and processed meat. IARC Working Group: Lyon, France, 2015.
24. Fahrer, J. and B. Kaina, Impact of DNA repair on the dose-response of colorectal cancer formation induced by dietary carcinogens. Food and Chemical Toxicology, 2017. 106: p. 583-594.
25. Cormack, P.A. and K. Mosbach, Molecular imprinting: recent developments and the road ahead. Reactive and Functional Polymers, 1999. 41(1-3): p. 115-124.
26. Matějíček, D., et al., Online molecularly imprinted solid‐phase extraction coupled to liquid chromatography‐tandem mass spectrometry for the determination of hormones in water and sediment samples. Journal of separation science, 2013. 36(9-10): p. 1509-1515.
27. Sun, X., et al., Determination of tetracyclines in food samples by molecularly imprinted monolithic column coupling with high performance liquid chromatography. Talanta, 2009. 79(3): p. 926-934.
28. Svenson, J. and I.A. Nicholls, On the thermal and chemical stability of molecularly imprinted polymers. Analytica Chimica Acta, 2001. 435(1): p. 19-24.
29. Chen, L., S. Xu, and J. Li, Recent advances in molecular imprinting technology: current status, challenges and highlighted applications. Chemical Society Reviews, 2011. 40(5): p. 2922-2942.
30. Pauling, L., A theory of the structure and process of formation of antibodies. Journal of the American Chemical Society, 1940. 62(10): p. 2643-2657.
31. Wulff, G. and A. Sarhan, Über die Anwendung von enzymanalog gebauten Polymeren zur Racemattrennung. Angewandte Chemie, 1972. 84(8): p. 364-364.
32. Arshady, R. and K. Mosbach, Synthesis of substrate‐selective polymers by host‐guest polymerization. Macromolecular Chemistry and Physics, 1981. 182(2): p. 687-692.
33. Muldoon, M.T. and L.H. Stanker, Molecularly imprinted solid phase extraction of atrazine from beef liver extracts. Analytical Chemistry, 1997. 69(5): p. 803-808.
34. Hennion, M.-C., Solid-phase extraction: method development, sorbents, and coupling with liquid chromatography. Journal of chromatography A, 1999. 856(1-2): p. 3-54.
35. Yang, Y., et al., Molecularly imprinted solid-phase extraction for selective extraction of bisphenol analogues in beverages and canned food. Journal of agricultural and food chemistry, 2014. 62(46): p. 11130-11137.
36. Hu, X., et al., Molecularly imprinted polymer coated solid-phase microextraction fibers for determination of Sudan I–IV dyes in hot chili powder and poultry feed samples. Journal of Chromatography A, 2012. 1219: p. 39-46.
37. Gao, D., et al., In Vivo Selective Capture and Rapid Identification of Luteolin and Its Metabolites in Rat Livers by Molecularly Imprinted Solid-Phase Microextraction. Journal of agricultural and food chemistry, 2017. 65(6): p. 1158-1166.
38. Sarafraz-Yazdi, A. and N. Razavi, Application of molecularly-imprinted polymers in solid-phase microextraction techniques. TrAC Trends in Analytical Chemistry, 2015. 73: p. 81-90.
39. Wei, Z.-H., et al., Imprinted monoliths: recent significant progress in analysis field. TrAC Trends in Analytical Chemistry, 2017. 86: p. 84-92.
40. Xie, C., et al., Electrochemical sensor for 2, 4-dichlorophenoxy acetic acid using molecularly imprinted polypyrrole membrane as recognition element. Microchimica Acta, 2010. 169(1-2): p. 145-152.
41. Ye, L. and K. Haupt, Molecularly imprinted polymers as antibody and receptor mimics for assays, sensors and drug discovery. Analytical and bioanalytical chemistry, 2004. 378(8): p. 1887-1897.
42. Van Royen, G., P. Dubruel, and E. Daeseleire, Development and evaluation of a molecularly imprinted polymer for the detection and cleanup of benzylpenicillin in milk. Journal of agricultural and food chemistry, 2014. 62(35): p. 8814-8821.
43. Liu, L., et al., Efficient molecular imprinting strategy for quantitative targeted proteomics of human transferrin receptor in depleted human serum. Analytical chemistry, 2015. 87(21): p. 10910-10919.
44. Whitcombe, M.J., et al., The rational development of molecularly imprinted polymer-based sensors for protein detection. Chemical Society Reviews, 2011. 40(3): p. 1547-1571.
45. Jiang, W., L. Liu, and Y. Chen, Simultaneous Detection of Human C-Terminal p53 Isoforms by Single Template Molecularly Imprinted Polymers (MIPs) Coupled with Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS)-Based Targeted Proteomics. Analytical chemistry, 2018. 90(5): p. 3058-3066.
46. Sellergren, B., Polymer-and template-related factors influencing the efficiency in molecularly imprinted solid-phase extractions. TrAC Trends in Analytical Chemistry, 1999. 18(3): p. 164-174.
47. Beltran, A., et al., Synthesis by precipitation polymerisation of molecularly imprinted polymer microspheres for the selective extraction of carbamazepine and oxcarbazepine from human urine. Journal of Chromatography A, 2009. 1216(12): p. 2248-2253.
48. Vasapollo, G., et al., Molecularly imprinted polymers: present and future prospective. International journal of molecular sciences, 2011. 12(9): p. 5908-5945.
49. Richter, B.E., et al., Accelerated solvent extraction: a technique for sample preparation. Analytical Chemistry, 1996. 68(6): p. 1033-1039.
50. Snyder, L.R., J.J. Kirkland, and J.W. Dolan, Introduction to modern liquid chromatography. 2011: John Wiley & Sons.
51. Meyer, V.R., Practical high-performance liquid chromatography. 2013: John Wiley & Sons.
52. Xu, P., D.M. Duong, and J. Peng, Systematical optimization of reverse-phase chromatography for shotgun proteomics. Journal of proteome research, 2009. 8(8): p. 3944-3950.
53. Yamashita, Y., et al., Alterations in gastric mucin with malignant transformation: novel pathway for mucin synthesis. J Natl Cancer Inst, 1995. 87(6): p. 441-6.
54. Laylin, J.K., Nobel laureates in chemistry, 1901-1992. 1993: Chemical Heritage Foundation.
55. James, A.T. and u.A. Martin, Gas-liquid partition chromatography: the separation and micro-estimation of volatile fatty acids from formic acid to dodecanoic acid. Biochemical Journal, 1952. 50(5): p. 679.
56. Baldwin, M. and F. McLafferty, Liquid chromatography‐mass spectrometry interface–I: The direct introduction of liquid solutions into a chemical ionization mass spectrometer. Journal of Mass Spectrometry, 1973. 7(9): p. 1111-1112.
57. Olivares, J.A., et al., On-line mass spectrometric detection for capillary zone electrophoresis. Analytical Chemistry, 1987. 59(8): p. 1230-1232.
58. Fenn, J., Electrospray ionization mass spectrometry: how it all began. Journal of biomolecular techniques: JBT, 2002. 13(3): p. 101.
59. Trufelli, H., et al., An overview of matrix effects in liquid chromatography–mass spectrometry. Mass spectrometry reviews, 2011. 30(3): p. 491-509.
60. Dempster, A., A new method of positive ray analysis. Physical Review, 1918. 11(4): p. 316.
61. Munson, M.S. and F.-H. Field, Chemical ionization mass spectrometry. I. General introduction. Journal of the American Chemical Society, 1966. 88(12): p. 2621-2630.
62. Dass, C., Fundamentals of contemporary mass spectrometry. Vol. 16. 2007: John Wiley & Sons.
63. El-Aneed, A., A. Cohen, and J. Banoub, Mass spectrometry, review of the basics: electrospray, MALDI, and commonly used mass analyzers. Applied Spectroscopy Reviews, 2009. 44(3): p. 210-230.
64. Fenn, J.B., et al., Electrospray ionization for mass spectrometry of large biomolecules. Science, 1989. 246(4926): p. 64-71.
65. Tanaka, K., et al., Protein and polymer analyses up to m/z 100 000 by laser ionization time‐of‐flight mass spectrometry. Rapid communications in mass spectrometry, 1988. 2(8): p. 151-153.
66. Takats, Z., et al., Mass spectrometry sampling under ambient conditions with desorption electrospray ionization. Science, 2004. 306(5695): p. 471-473.
67. Cody, R.B., J.A. Laramée, and H.D. Durst, Versatile new ion source for the analysis of materials in open air under ambient conditions. Analytical chemistry, 2005. 77(8): p. 2297-2302.
68. Domon, B. and R. Aebersold, Mass spectrometry and protein analysis. science, 2006. 312(5771): p. 212-217.
69. Evans, A.M., et al., Integrated, nontargeted ultrahigh performance liquid chromatography/electrospray ionization tandem mass spectrometry platform for the identification and relative quantification of the small-molecule complement of biological systems. Analytical chemistry, 2009. 81(16): p. 6656-6667.
70. Wilm, M.S. and M. Mann, Electrospray and Taylor-Cone theory, Dole's beam of macromolecules at last? International Journal of Mass Spectrometry and Ion Processes, 1994. 136(2-3): p. 167-180.
71. Bruins, A.P., Mechanistic aspects of electrospray ionization. Journal of Chromatography A, 1998. 794(1-2): p. 345-357.
72. Thomson, J.J., XIX. Further experiments on positive rays. The London, Edinburgh, and Dublin Philosophical Magazine and Journal of Science, 1912. 24(140): p. 209-253.
73. McLafferty, F.W., Tandem mass spectrometry. Science, 1981. 214(4518): p. 280-287.
74. Holčapek, M., R. Jirásko, and M. Lísa, Recent developments in liquid chromatography–mass spectrometry and related techniques. Journal of Chromatography A, 2012. 1259: p. 3-15.
75. Yost, R. and C. Enke, Selected ion fragmentation with a tandem quadrupole mass spectrometer. Journal of the American Chemical Society, 1978. 100(7): p. 2274-2275.
76. Senko, M.W., J.P. Speir, and F.W. McLafferty, Collisional activation of large multiply charged ions using Fourier transform mass spectrometry. Analytical Chemistry, 1994. 66(18): p. 2801-2808.
77. Allen, J.S., The detection of single positive ions, electrons and photons by a secondary electron multiplier. Physical Review, 1939. 55(10): p. 966.
78. Ozmen, M.M., M.V. Dinu, and O. Okay, Preparation of macroporous poly (acrylamide) hydrogels in DMSO/water mixture at subzero temperatures. Polymer Bulletin, 2008. 60(2-3): p. 169-180.
79. Kabiri, K., et al., Solvent-, ion-and pH-specific swelling of poly (2-acrylamido-2-methylpropane sulfonic acid) superabsorbing gels. Journal of polymer research, 2010. 17(2): p. 203-212.
80. Lorenzo, R.A., et al., To remove or not to remove? The challenge of extracting the template to make the cavities available in molecularly imprinted polymers (MIPs). International journal of molecular sciences, 2011. 12(7): p. 4327-4347.
81. Batlokwa, B.S., et al., Optimal template removal from molecularly imprinted polymers by pressurized hot water extraction. Chromatographia, 2011. 73(5-6): p. 589-593.