急性瘦肉精中毒检材的QuEChERS净化-液质联用快速确证方法
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摘要
目前液相色谱串联质谱法测定动物源性食品中瘦肉精普遍采用酶水解提取、固相萃取净化的方式处理样品,该方法耗时长、分析成本高。本研究采用酸水解和酶水解两种提取方式结合QuEChERS(Quick,Easy,Cheap,Effective,Rugged,Safe)净化和高效液相色谱串联质谱建立了动物性食品中特布他林、沙丁胺醇、莱克多巴胺和克伦特罗4种β2受体激动剂的快速定性确证和定量检测方法。采用酶水解提取-QuEChERS净化处理样品,在电喷雾离子源正离子扫描(ESI+)和多反应监测(MRM)模式下,4种瘦肉精在1. 0～20. 0μg/L浓度之间呈线性,线性相关系数均大于0. 999,检出限和定量限分别为0. 1和0. 3μg/kg,猪肉和牛肉样品中4种化合物的回收率为78. 5%～112%,相对标准偏差RSD为2. 8%～8. 9%。采用酸水解法代替酶水解提取时,尽管对于样品中莱克多巴胺测定结果稍偏低,但样品前处理时间大大缩短。两种提取方法结合使用对于提高瘦肉精引起的食物中毒应急处置效率具有重要意义。
The pretreatment procedure of enzymatic hydrolysis and extraction coupled with solid phase extraction( SPE) purification is usually employed for the determination of β-agonists by high performance liquid chromatography( HPLC) with tandam mass spetroscopy( MS) in food originated from animals. In this study,a HPLC-MS/MS with QuEChERS( Quick,Easy,Cheap,Effective,Rugged,Safe) purification procedures was developed for the determination of terbutaline,salbutamol,ractopamine and clenbuterol in food of animal origin using both acid hydrolysis extraction method and enzymatic hydrolysis extraction method in food poisoning event caused by clenbuterol and analogs. Using enzymatic hydrolysis extraction with QuEChERS purification and electrospray ionization mass spectrometry in positive mode adopting multiple reaction monitoring( MRM) mode,four analytes were analyzed in the range of 1. 0 ～ 20. 0 μg/L with correlation coefficients( R) higher than 0. 999. The limit of detection( LOD) and the limit of quantification( LOQ) was 0. 1 μg/kg and 0. 3 μg/kg,respectively,and recoveries for 4 β2-agonists in pork and beef samples ranged from 78. 5% to 112% with relative standard deviation( RSD) of 2. 8% ～ 8. 9%. When acid hydrolysis extraction was used alternatively,the pretreatment time was shortened greatly,but the result for ractopamine was slightly lower than that using the enzymatic hydrolysis extraction procedure. The combined use of two pretreatment procedures will promote the emergency response in real food poisoning case caused by clenbuterol and analogs.
The pretreatment procedure of enzymatic hydrolysis and extraction coupled with solid phase extraction( SPE) purification is usually employed for the determination of β-agonists by high performance liquid chromatography( HPLC) with tandam mass spetroscopy( MS) in food originated from animals. In this study,a HPLC-MS/MS with QuEChERS( Quick,Easy,Cheap,Effective,Rugged,Safe) purification procedures was developed for the determination of terbutaline,salbutamol,ractopamine and clenbuterol in food of animal origin using both acid hydrolysis extraction method and enzymatic hydrolysis extraction method in food poisoning event caused by clenbuterol and analogs. Using enzymatic hydrolysis extraction with QuEChERS purification and electrospray ionization mass spectrometry in positive mode adopting multiple reaction monitoring( MRM) mode,four analytes were analyzed in the range of 1. 0 ～ 20. 0 μg/L with correlation coefficients( R) higher than 0. 999. The limit of detection( LOD) and the limit of quantification( LOQ) was 0. 1 μg/kg and 0. 3 μg/kg,respectively,and recoveries for 4 β2-agonists in pork and beef samples ranged from 78. 5% to 112% with relative standard deviation( RSD) of 2. 8% ～ 8. 9%. When acid hydrolysis extraction was used alternatively,the pretreatment time was shortened greatly,but the result for ractopamine was slightly lower than that using the enzymatic hydrolysis extraction procedure. The combined use of two pretreatment procedures will promote the emergency response in real food poisoning case caused by clenbuterol and analogs.
引文
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[8]WANG Quanshuai,SUN Weiming,LIU Jianrong.Determination of Four Clenbuterol Residues in Pork by Solid Phase Extraction and Gas Chromatography-Mass Spectrometry[J].Occup Health,2015,31(2):181-183(in Chinese).王权帅,孙卫明,刘建荣.猪肉中特步他林等4种瘦肉精残留量的固相萃取-气相色谱质谱联用测定法[J].职业与健康,2015,31(2):181-183.
[9]CAI Yuzhi,LUO Xiaoyan,SU Wenzhou.Clenbuterol Hydrochloride Residue Determination by High Performance Liquid Chromatography in Pork[J].Chinese J Lab Technol,2002,12(2):193-194(in Chinese).蔡玉枝,罗晓燕,苏文周.高效液相色谱法测定猪组织中盐酸克伦特罗残留量[J].中国卫生检验杂志,2002,12(2):193-194.
[10]Li L B,Du H W,Yu H,et al.Application of Ionic Liquid as Additive in Determination of ThreeβAgonists by Capillary Electrophoresis with Amperometric Detection[J].Electrophoresis,2013,34:277-283.
[11]Wang G M,Zhao J,Peng T,et al.Matrix Effects in the Determination ofβReceptor Agonists in Animal-Derived Foodstuffs by Ultra-Performance Liquid Chromatography Tadem Mass Spectrometry with Immunoaffinity Solid-Phase Extraction[J].JSep Sci,2013,36:796-802.
[12]Suo D C,Zhao G L,Wang P L,et al.Simultaneous Determination ofβAgonists and Psychiatric Drugs in Feeds by LC-MS-MS[J].J Chromatogr Sci,2014,52(7):604-608.
[13]LIU Guihua,MAO Lisha,ZHU Zhou,et al.Determination of Residues of NineβAgonists in Edible Meat and Liver by Liquid Chromatography Tandem Mass Spectrometry[J].South China J Prev Med,2014,40(5):425-430(in Chinese).刘桂华,毛丽莎,朱舟,等.液相色谱串联质谱法测定食用肉类及肝脏中9种β受体激动剂[J].华南预防医学,2014,40(5):425-430.
[14]SHI Na,HAO Jie,JIANG Jie,et al.Simultaneous Determination of 35β-agonist and 11β-Blocker Residues in AnimalOrigin Foods by Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry[J].Anal Instrum,2018,1:20-31(in Chinese).史娜,郝杰,姜洁,等.超高效液相色谱-串联质谱法同时检测动物源性食品中残留的35种β-受体激动剂和11种β-受体阻断剂[J].分析仪器,2018,1:20-31.
[15]QU Bin,GENG Shiwei,LU Guiping,et al.Rapid Determination ofβAgonists Residuals in Pig Liver by Liquid Chromatography Tandem Mass Spectrometry Combined with Novel Qu ECh ERS Method[J].J Food Saf Food Qual,2015,6(12):4747-4754(in Chinese).曲斌,耿士伟,陆桂萍,等.新型Qu ECh ERS方法结合液相色谱串联质谱法快速测定猪肝中β受体激动剂残留[J].食品安全质量检测学报,2015,6(12):4747-4754.
[16]LI Lei,LI Haichang,GAO Jing,et al.Determination of 8βAgonists by Qu ECh ERS EMR-Lipid-LC/MS/MS[J].Food Res Dev,2016,37(9):178-182(in Chinese).李磊,李海畅,高婧,等.Qu ECh ERS EMR-Lipid-LC/MS/MS测定8种β-受体激动剂[J].食品研究与开发,2016,37(9):178-182.
[17]ZHU Weixia,YANG Yizhou,GUO Huiling,et al.Quick Determination of 16β-Agonist Residues in Pig Urine by Turboflow Cation Online Solid Phase Extraction and Liquid Chromatography-Tandem Mass Spectrometry[J].J Instrum Anal,2015,34(7):802-806(in Chinese).祝伟霞,杨翼州,郭慧领,等.阳离子涡流在线固相萃取/液相色谱-串联质谱快速分析猪尿液中16种β受体激动剂[J].分析测试学报,2015,34(7):802-806.
[2]YE Ni,SUN Lei,YIN Hui,et al.Determination ofβAgonists in Animal Derived Food by UPLC-MS/MS[J].Chinese JVeterin Drug,2015,49(9):51-59(in Chinese).叶妮,孙雷,尹晖,等.UPLC-MS/MS法检测动物性食品中19种β受体激动剂残留[J].中国兽药杂志,2015,49(9):51-59.
[3]XIONG Lin,LI Weihong,GAO Yaqin,et al.A Review of Inspection Technology ofβAgonist Drug Residues in Meat[J].JFood Saf Food Qual,2015,6(2):528-533(in Chinese).熊琳,李维红,高雅琴,等.肉制品中β受体激动剂类药物残留检测技术研究进展[J].食品安全质量检测学报,2015,6(2):528-533.
[4]China's Ministry of Agriculture Bulletin No.1025-18-2008,the Catalogue of Forbidden Drug in Feed and Drinking Water of Animal[Z].2002(in Chinese).农业部公告第176号,禁止在磁疗和动物饮水中使用的药物品种目录[Z].2002.
[5]DONG Fengguang,WANG Guiqiang,GONG Chunbo,et al.Pollution Status and Risk Assessment of Heavy Metals andβ-Receptor Agonist in Commercially Available Livestock and Poultry Meat in Yantai City from 2012-2017[J].Occup Health,2018,34(6):764-768(in Chinese).董峰光,王桂强,宫春波,等.2012-2017年烟台市市售畜禽肉中重金属和β-受体激动剂污染状况及危险性评估[J].职业与健康,2018,34(6):764-768.
[6]ZHOU Yuerong,LI Danni,GU Xin,et al.Determination of 11β2-Agonists Residues in Swine and Chicken Manure by Enzyme Linked Immunosorbent Assay[J].Chinese J Veterin Drug,2015,49(8):39-45(in Chinese).周悦榕,李丹妮,顾欣,等.酶联免疫吸附分析法测定猪、鸡粪便中11种β2-受体激动剂残留[J].中国兽药杂志,2015,49(8):39-45.
[7]YUE Hanxiao,LEI Wen,DU Xiaoning,et al.Multi-residue Analysis of 4β-Agonists in Pork Using Isotope Dilution Gas Chromatography-Tandem Mass Spectrometry[J].J Chinese Mass Spectrom Soc,2018,39(1):61-68(in Chinese).岳韩笑,雷雯,杜晓宁,等.同位素稀释-气相色谱-串联质谱法测定猪肉中残留的4种β-受体激动剂[J].质谱学报,2018,39(1):61-68.
[8]WANG Quanshuai,SUN Weiming,LIU Jianrong.Determination of Four Clenbuterol Residues in Pork by Solid Phase Extraction and Gas Chromatography-Mass Spectrometry[J].Occup Health,2015,31(2):181-183(in Chinese).王权帅,孙卫明,刘建荣.猪肉中特步他林等4种瘦肉精残留量的固相萃取-气相色谱质谱联用测定法[J].职业与健康,2015,31(2):181-183.
[9]CAI Yuzhi,LUO Xiaoyan,SU Wenzhou.Clenbuterol Hydrochloride Residue Determination by High Performance Liquid Chromatography in Pork[J].Chinese J Lab Technol,2002,12(2):193-194(in Chinese).蔡玉枝,罗晓燕,苏文周.高效液相色谱法测定猪组织中盐酸克伦特罗残留量[J].中国卫生检验杂志,2002,12(2):193-194.
[10]Li L B,Du H W,Yu H,et al.Application of Ionic Liquid as Additive in Determination of ThreeβAgonists by Capillary Electrophoresis with Amperometric Detection[J].Electrophoresis,2013,34:277-283.
[11]Wang G M,Zhao J,Peng T,et al.Matrix Effects in the Determination ofβReceptor Agonists in Animal-Derived Foodstuffs by Ultra-Performance Liquid Chromatography Tadem Mass Spectrometry with Immunoaffinity Solid-Phase Extraction[J].JSep Sci,2013,36:796-802.
[12]Suo D C,Zhao G L,Wang P L,et al.Simultaneous Determination ofβAgonists and Psychiatric Drugs in Feeds by LC-MS-MS[J].J Chromatogr Sci,2014,52(7):604-608.
[13]LIU Guihua,MAO Lisha,ZHU Zhou,et al.Determination of Residues of NineβAgonists in Edible Meat and Liver by Liquid Chromatography Tandem Mass Spectrometry[J].South China J Prev Med,2014,40(5):425-430(in Chinese).刘桂华,毛丽莎,朱舟,等.液相色谱串联质谱法测定食用肉类及肝脏中9种β受体激动剂[J].华南预防医学,2014,40(5):425-430.
[14]SHI Na,HAO Jie,JIANG Jie,et al.Simultaneous Determination of 35β-agonist and 11β-Blocker Residues in AnimalOrigin Foods by Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry[J].Anal Instrum,2018,1:20-31(in Chinese).史娜,郝杰,姜洁,等.超高效液相色谱-串联质谱法同时检测动物源性食品中残留的35种β-受体激动剂和11种β-受体阻断剂[J].分析仪器,2018,1:20-31.
[15]QU Bin,GENG Shiwei,LU Guiping,et al.Rapid Determination ofβAgonists Residuals in Pig Liver by Liquid Chromatography Tandem Mass Spectrometry Combined with Novel Qu ECh ERS Method[J].J Food Saf Food Qual,2015,6(12):4747-4754(in Chinese).曲斌,耿士伟,陆桂萍,等.新型Qu ECh ERS方法结合液相色谱串联质谱法快速测定猪肝中β受体激动剂残留[J].食品安全质量检测学报,2015,6(12):4747-4754.
[16]LI Lei,LI Haichang,GAO Jing,et al.Determination of 8βAgonists by Qu ECh ERS EMR-Lipid-LC/MS/MS[J].Food Res Dev,2016,37(9):178-182(in Chinese).李磊,李海畅,高婧,等.Qu ECh ERS EMR-Lipid-LC/MS/MS测定8种β-受体激动剂[J].食品研究与开发,2016,37(9):178-182.
[17]ZHU Weixia,YANG Yizhou,GUO Huiling,et al.Quick Determination of 16β-Agonist Residues in Pig Urine by Turboflow Cation Online Solid Phase Extraction and Liquid Chromatography-Tandem Mass Spectrometry[J].J Instrum Anal,2015,34(7):802-806(in Chinese).祝伟霞,杨翼州,郭慧领,等.阳离子涡流在线固相萃取/液相色谱-串联质谱快速分析猪尿液中16种β受体激动剂[J].分析测试学报,2015,34(7):802-806.