利用高效液相色谱-飞行时间质谱和高效液相色谱-三重四级杆质谱检测南美白对虾的过敏原
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摘要
为检测南美白对虾的过敏原,并寻找表征过敏原的特征肽段,采用测序级胰蛋白酶对南美白对虾的蛋白进行酶解,经高效液相色谱-飞行时间质谱的分离检测和数据库检索,并建立了三重四级杆多反应监测(MRM)快速检测表征过敏原肽段的方法。结果表明,南美白对虾中共检测出7种过敏原,分别为原肌球蛋白、精氨酸激酶、肌球蛋白轻链、血蓝蛋白亚基L1、血蓝蛋白亚基L2、血蓝蛋白亚基L3、肌钙蛋白C1,对应的肽段条数分别为214、16、84、145、225、163和93条。MRM结果显示,该方法灵敏度高、选择性好,为分析南美白对虾过敏原提供了参考。
To detect allergens of Litopenaeus vannamei and find peptides for representation of allergens,high-performance liquid chromatography coupled to quadrupole time of flight mass spectrometry( HPLC-QTOF) with database search was used to separate and detect all sequencing grade modified trypsin-digested peptides.And multiple reaction monitoring( MRM) method on triple quadrupole mass spectrometer was established for rapid detection of the peptides of allergens.The results showed that there were seven kinds of allergens in Litopenaeus vannamei,which were Lit v 1 tropomyosin,arginine kinase,Lit v 3 allergen myosin light chain,hemocyanin subunit L1,hemocyanin subunit L2,hemocyanin subunit L3 and troponin C1,and the numbers of identified peptides of allergens were 214,16,84,145,225,163 and 93 respectively. The MRM result showed that the established method was of high sensitivity and good selectivity.This paper would providea reference for the analysis of allergens of Litopenaeus vannamei.
To detect allergens of Litopenaeus vannamei and find peptides for representation of allergens,high-performance liquid chromatography coupled to quadrupole time of flight mass spectrometry( HPLC-QTOF) with database search was used to separate and detect all sequencing grade modified trypsin-digested peptides.And multiple reaction monitoring( MRM) method on triple quadrupole mass spectrometer was established for rapid detection of the peptides of allergens.The results showed that there were seven kinds of allergens in Litopenaeus vannamei,which were Lit v 1 tropomyosin,arginine kinase,Lit v 3 allergen myosin light chain,hemocyanin subunit L1,hemocyanin subunit L2,hemocyanin subunit L3 and troponin C1,and the numbers of identified peptides of allergens were 214,16,84,145,225,163 and 93 respectively. The MRM result showed that the established method was of high sensitivity and good selectivity.This paper would providea reference for the analysis of allergens of Litopenaeus vannamei.
引文
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[19]贺建华,邵纯君,鹿麟,等.福林酚法测定甘露聚糖肽口服溶液中蛋白质的含量[J].中国药物评价,2017,34(2)92-94.
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[21]Mi R,Sun Y,Li J,et al.Immune-related proteins detected through iTRAQ-based proteomics analysis of intestines from Apostichopus japonicus in response to tussah immunoreactive substances[J].Fish&Shellfish Immunology,2018,74:436-443.
[2]Michael K,Renate B,Uta T,et al.Long-lasting sensitization to food during the first two years precedes allergic airway disease[J].Pediatric Allergy&Immunology Official Publication of the European Society of Pediatric Allergy and Immunology,1998,9(2):61-67.
[3]罗奕铭.虾过敏原酶联免疫快速检测方法的研究[D].广州:华南农业大学,2011.
[4]冯汉利,胡小钟,郭明星,等.甲壳类食品过敏原研究及其在出入境检验检疫中的地位[J].湖北农业科学,2010,49(1):207-210.
[5]Schfer T,B9hler E,Ruhdorfer S,et al.Epidemiology of food allergy/food intolerance in adults:Associations with other manifestations of atopy[J].Allergy,2015,56(12):1172-1179.
[6]吕相征,刘秀梅,杨晓光.健康人群食物过敏状况的初步调查[J].中国食品卫生杂志,2005,17(2):119-121.
[7]李振兴,林洪.不同种类甲壳类动物过敏原及其过敏原性的研究[C]//中国水产学会青年学术年会,2004.
[8]李振兴,林洪,李明华,等.不同虾类的过敏原及其过敏原性[J].水产学报,2006,30(2):281-284.
[9]Altman D R,Chiaramonte L T.Public perception of food allergy[J].Journal of Allergy&Clinical Immunology,1996,97(6):1247-1251.
[10]刘光明,沈苑,曹敏杰,等.虾类过敏原的识别、纯化和检测技术研究[J].中国食品学报,2008,8(6):142-148.
[11]张轶群.海产品过敏原免疫芯片检测方法的研究[D].青岛:中国海洋大学,2009.
[12]宫瑞.南美白对虾常见病害及防治方法[J].中国水产,2009(2):58-58.
[13]陈东亮,刘金明,杜宗强,等.关于南美白对虾的养殖技术之三:南美白对虾养殖常见疾病及防治[J].中国水产,2001(11):62-63.
[14]García-Orozco K D,Aispuro-Hernández E,YepizPlascencia G,et al.Molecular characterization of arginine kinase,an allergen from the shrimp Litopenaeus vannamei[J].International Archives of Allergy&Immunology,2007,144(1):23-28.
[15]吴海明.南美白对虾主要过敏原鉴定及酶法消减技术的研究[D].天津:天津商业大学,2011.
[16]Shim Y S,Kim K J,Seo D,et al.Rapid method for the determination of vitamins A and E in foods using ultra-highperformance liquid chromatography[J].Journal of Aoac International,2012,95(2):517-522.
[17]肖遂,薛飞群.UPLC/MS在兽药残留分析中的应用[C].中国畜牧科技论坛,2011.
[18]徐清,林向阳,王征.同位素稀释三重串联四级杆质谱法测定饮用水中4种增塑剂[J].食品安全质量检测学报,2015(1):239-244.
[19]贺建华,邵纯君,鹿麟,等.福林酚法测定甘露聚糖肽口服溶液中蛋白质的含量[J].中国药物评价,2017,34(2)92-94.
[20]Wis'niewski J R,Zougman A,Nagaraj N,et al.Universal sample preparation method for proteome analysis[J].Nature Methods,2009,6(5):359-362.
[21]Mi R,Sun Y,Li J,et al.Immune-related proteins detected through iTRAQ-based proteomics analysis of intestines from Apostichopus japonicus in response to tussah immunoreactive substances[J].Fish&Shellfish Immunology,2018,74:436-443.