高效液相色谱-串联质谱法测定肉制品和调味料中7种水产品过敏原
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摘要
基于高效液相色谱-串联质谱系统建立了食品中水产品过敏原的快速筛查和定量检测方法。样品经蛋白质提取、纯化、胰蛋白酶解后,利用超高效液相色谱-四极杆/静电场轨道阱高分辨质谱(UPLC-Q/Exactive-HRMS)结合ProteinPilot软件,基于母离子和碎片离子谱分析,实现蛋白质和多肽的鉴定。再通过基本局部比对搜索工具(BLAST)与Uniprot数据库对比分析,筛选出南美白虾、大闸蟹、青蟹、金枪鱼、大西洋鲑鱼的7种过敏原蛋白的30个特征肽。利用高效液相色谱-三重四极杆质谱(UPLC-QqQ-MS)系统对特征肽进行验证和多反应监测(MRM)定量研究。结果表明,该方法在5～250 mg/kg范围内线性关系良好,检出限为2～3.5 mg/kg,平均回收率为88.7%～110.2%。该方法重现性好,通量高,可应用于肉制品和调味料中7种过敏原的快速筛查和定量分析。
A liquid chromatography-tandem mass spectrometry method for the identification of marker peptides of aquatic product allergens and quantitative detection of multiple allergens in meat products and seasonings was developed. The samples were prepared by protein extraction, protein purification, and trypsin hydrolysis. The proteins and peptides were identified using ProteinPilot by the data analysis of the ion spectrum of polypeptide fragments using ultra-performance liquid chromatography-quadrupole/electrostatic orbitrap high-resolution mass spectrometry(UPLC-Q/Exactive-HRMS). The identification of 30 species-specific marker peptides in Penaeus vannamei, Eriocheir, Scylla serrata, Thunnus thynnus, and Atlantic salmon by comparison of the basic local alignment search tool(BLAST) with the UniProt database was achieved. The verification and multiple reaction monitoring(MRM) quantitative studies of these marker peptides were performed using a triple quadrupole mass spectrometry(UPLC-QqQ-MS) system. The proposed method showed a good linear relationship in the range of 5-250 mg/kg. The limits of quantitation and observed recoveries were in the range of 2-3.5 mg/kg and 88.7%-110.2%, respectively. This method presents various advantages such as good repeatability and high throughput, suitability for rapid screening, and quantitative analysis of seven aquatic allergens in meat products and seasonings.
A liquid chromatography-tandem mass spectrometry method for the identification of marker peptides of aquatic product allergens and quantitative detection of multiple allergens in meat products and seasonings was developed. The samples were prepared by protein extraction, protein purification, and trypsin hydrolysis. The proteins and peptides were identified using ProteinPilot by the data analysis of the ion spectrum of polypeptide fragments using ultra-performance liquid chromatography-quadrupole/electrostatic orbitrap high-resolution mass spectrometry(UPLC-Q/Exactive-HRMS). The identification of 30 species-specific marker peptides in Penaeus vannamei, Eriocheir, Scylla serrata, Thunnus thynnus, and Atlantic salmon by comparison of the basic local alignment search tool(BLAST) with the UniProt database was achieved. The verification and multiple reaction monitoring(MRM) quantitative studies of these marker peptides were performed using a triple quadrupole mass spectrometry(UPLC-QqQ-MS) system. The proposed method showed a good linear relationship in the range of 5-250 mg/kg. The limits of quantitation and observed recoveries were in the range of 2-3.5 mg/kg and 88.7%-110.2%, respectively. This method presents various advantages such as good repeatability and high throughput, suitability for rapid screening, and quantitative analysis of seven aquatic allergens in meat products and seasonings.
引文
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[2] Sicherer S H,Sampson H A.J Allergy Clin Immunol,2010,125(2):116
[3] Sicherer S H.J Allergy Clin Immunol,2011,127(3):594
[4] Koeberl M,Clarke D,Lopata A L.J Proteome Res,2014,13(8):3499
[5] Mónica C,CanAs B,José M,et al.J Proteomics,2012,75(11):3211
[6] Watanabe H,Saita K,Akaboshi C,et al.Shokuhin Eiseigaku Zasshi,2014,55:41
[7] Ahsan N,Rao R S,Gruppuso P A,et al.J Proteomics,2016,143:15
[8] Parker C H,Khuda S E,Pereira M,et al.J Agric Food Chem,2015,63:10669
[9] Goto K,Minatani T,Nagai H.J AOAC Int,2015,98(5):1355
[10] Korte R,Monneuse J M,Gemrot E,et al.J Agric Food Chem,2016,64(31):6219
[11] Eischeid A C,Kim B,Kasko S M.J Agric Food Chem,2013,61:5669
[12] Cheng F,Wu J J,Zhang J,et al.Food Chem,2016,199:799
[13] Eischeid A C.Food Control,2016,59:393
[14] Gu S Q,Zhao C M,Cheng J,et al.Chinese Journal of Chromatography,2016,43(7):639 古淑青,赵超敏,程甲,等.色谱,2016,43(7):639
[15] Abdel R A M,Kamath S,Lopata A L,et al.Rapid Commun Mass Spectrom,2010,24(24):3624
[16] Abdel R A M,Lopata A L,Randell E W,et al.Anal Chim Acta,2010,681(1):49
[17] Ortea I,Canas B,Calo-Mata P,et al.J Agric Food Chem,2009,57(13):5665
[18] Siciliano R A.J Agric Food Chem,2008,56(23):11071
[19] Carrera M,Canas B,Vazquez J,et al.J Proteome Res,2010,9(9):4393
[20] Gu S Q,Chen N N,Zhou Y,et al.Food Control,2018,84,89
[21] De C C,Calvano C D,Zambonin C G.J Agric Food Chem,2014,62(39):9401
[22] Zhan L N,Chen Q,Gu S Q,et al.Chinese Journal of Chromatography,2017,35(4):405 詹丽娜,陈沁,古淑青,等.色谱,2017,35(4):405
[23] Gu S Q,Zhan L N,Zhao C M,et al.Chinese Journal of Chromatography,2018,36(12):1269 古淑青,詹丽娜,赵超敏,等.色谱,2018,36(12):1269
[24] Claydon A J,Grundy H H,Charlton A J,et al.Food Addit Contam Part A Chem Anal Control Expo Risk Assess,2015,32(10):1718
[25] Wang L S,Huang J C,Chen Y L,et al.J Agric Food Chem,2015,63:3437
[26] Carrera M,Canas B,Lopez-Ferrer D,et al.Anal Chem,2011,83:5688