UPLC-MS/MS法测定动物源性食品中氯苯胍及其代谢物残留

doi:10.16431/j.cnki.1671-7236.2020.06.036

摘要/Abstract

摘要： 为快速有效地检测动物源性食品中氯苯胍及其代谢物残留，研究建立一种在鸡蛋、鸡肉、牛肉、鱼肉和猪肉5种动物源性食品中，同时检测氯苯胍及其代谢物（对氯苯甲酸、对氯苯甲酰氨基乙酸）残留的超高效液相色谱-串联质谱（UPLC-MS/MS）法。样品经过2%（V/V）甲酸乙腈溶液提取，无水硫酸钠去除水分，氮吹浓缩后甲醇复溶，正己烷除脂，高速冷冻离心，得到净化后的样品进行上机测定。选用Waters ACQUITY UPLC BEH C18色谱柱（2.1 mm×100 mm，1.7 μm），将甲醇-0.1%甲酸水溶液作为流动相进行梯度洗脱。通过多反应检测（MRM），在正/负离子模式下，采用基质匹配外标法，同时对3种化合物进行定性和定量分析。结果显示，氯苯胍、对氯苯甲酸和对氯苯甲酰氨基乙酸在各自浓度范围内线性关系良好，相关系数R²>0.999。氯苯胍的检出限（LOD）和定量限（LOQ）分别为0.5和1.0 μg/kg，对氯苯甲酸的LOD和LOQ分别为2.5和5.0 μg/kg，对氯苯甲酰氨基乙酸的LOD和LOQ分别为1.0和2.5 μg/kg。不同基质中，3种化合物在4个添加水平（氯苯胍：1.0、25、50、100 μg/kg；对氯苯甲酸：5.0、25、50、100 μg/kg；对氯苯甲酰氨基乙酸：2.5、25、50、100 μg/kg）的平均回收率为76.0%~95.9%，相对标准偏差（RSDs，n=6）为2.6%~10.6%。基质效应|ME|为0.2%~26.2%，其中氯苯胍在鸡蛋中，对氯苯甲酰氨基乙酸在鸡肉、牛肉和鱼肉中存在较强的基质效应（|ME|>20%），空白猪肉可作为代表基质用于3种化合物的定量分析。本方法前处理简单，灵敏度较高，重现性好，可用于动物源食品中氯苯胍及其代谢物残留的测定。

关键词: 超高效液相色谱－串联质谱(UPLC-MS/MS); 动物源性食品; 氯苯胍; 代谢物

Abstract: In order to detect robenidine and its metabolites residues in animal derived food rapidly and effectively,an ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed for the simultaneous determination of robenidine and its metabolites (4-chlorobenzoic acid,4-chlorobenzoylamino-acetic acid) residues in five kinds of animal derived food,including eggs,chicken,beef,fish and pork.Samples were extracted with 2% (V/V) formic acid-acetonitrile solution,removed water with anhydrous sodium sulfate,fixed volume with methanol,then excepted fat with n-hexane.The purified samples were analyzed by UPLC-MS/MS after freezed and high speed centrifuged.UPLC was achieved using a Waters ACQUITY UPLC BEH C18 column (2.1 mm×100 mm,1.7 μm);the mobile phase consisted of methanol and 0.1% formic acid-water solution.The qualitative analysis was performed with MS/MS under multiple reaction monitoring (MRM) mode with positive and negative electrospray ionization;the matrix matched external standard method was used for quantitation.The results showed that there were good linearities of the three compounds in their respective concentration range (R²>0.999).The limits of detection (LODs) and the limits of quantity (LOQs) for robenidine were 0.5 and 1.0 μg/kg,for 4-chlorobenzoic acid were 2.5 and 5.0 μg/kg,for 4-chlorobenzoylamino-acetic acid were 1.0 and 2.5 μg/kg,respectively.While blank samples spiked four levels (robenidine:1.0,25,50,100 μg/kg;4-chlorobenzoic acid:5.0,25,50,100 μg/kg;4-chlorobenzoylamino-acetic acid:2.5,25,50,100 μg/kg),the recoveries and RSDs (n=6) of the three compounds were 76.0%-95.9% and 2.6%-10.6%,respectively.The matrix effects (|ME|) of the developed method were 0.2%-26.2%.There was a strong matrix effect (|ME|>20%) of robenidine in eggs and 4-Chlorobenzoylamino-acetic acid in chicken,beef and fish;blank pork could be used as representative matrix for quantitative analysis of three compounds.Therefore,the method was simple in pretreatment,high in sensitivity and good in reproducibility,and could be used for the determination of robenidine and its metabolites in five kinds of animal derived food.

Key words: ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS); animal derived food; robenidine; metabolite

中图分类号:

S859.84

郑幸果, 郭灵安, 赵珊, 李曦, 雷欣宇, 仲伶俐. UPLC-MS/MS法测定动物源性食品中氯苯胍及其代谢物残留[J]. 中国畜牧兽医, 2020, 47(6): 1961-1970.

ZHENG Xingguo, GUO Lingan, ZHAO Shan, LI Xi, LEI Xinyu, ZHONG Lingli. Determination of Robenidine and Its Metabolites in Animal Derived Food by UPLC-MS/MS[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(6): 1961-1970.

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