配合饲料中64种药物超高效液相色谱-三重四极杆串联质谱检测方法的研究

doi:10.12160/j.issn.1672-5190.2024.03.002

摘要/Abstract

摘要： [目的]建立同时检测配合饲料中64种药物的超高效液相色谱-三重四极杆串联质谱（ultra-high performance liquid chromatography-triple quadrupole tandem mass spectrometry,UHPLC-MS/MS）法,提高非法添加物的检测效率。[方法]采用Waters HSS T3型色谱柱（2.1 mm×100 mm,1.8 μm）进行分离,流动相A为0.1%甲酸水溶液,流动相B为含0.1%甲酸的乙腈溶液,梯度洗脱,流速为0.40 mL/min,进样量为2 μL;采用电喷雾离子源正离子扫描模式进行检测,多反应监测模式进行信号采集。比较4种样品提取溶剂以及2种固相萃取柱处理对目标药物的回收率,确定样品前处理的最佳方法。利用建立的UHPLC-MS/MS法对宁夏回族自治区不同来源的100批次配合饲料样品进行64种药物检测。[结果]配合饲料样品均质后,用含0.2%甲酸的乙腈水溶液（乙腈∶水=8∶2,V/V）提取,利用Oasis PRiME HLB型固相萃取柱对样品净化,多数目标药物的回收率在60%以上。64种药物在浓度为5.0~200.0 μg/L的范围内线性关系良好,相关系数（R）均大于0.99;不同药物的定量限在5.0~10.0 μg/kg;阳性添加5.0、20.0、50.0 μg/kg 3个浓度的平均回收率在41.00%~120.49%,批内相对标准偏差（RSD）在0.54%~15.94%,批间RSD在1.25%~13.64%。在100个批次的配合饲料样品中均未检出目标药物。[结论]建立的UHPLC-MS/MS法线性关系良好、回收率高、精密度好,具有较高的重现性和较好的可操作性,可用于配合饲料中非法添加64种药物的筛查。

关键词: 配合饲料, 超高效液相色谱-三重四极杆串联质谱法, 非法添加, 兽药

Abstract: [Objective] The aims of the present study were to establish an ultra-high performance liquid chromatography-triple quadrupole tandem mass spectrometry （UHPLC-MS/MS） method for simultaneous detection of 64 drugs in compound feed, and to improve the detection efficiency of illegal additives. [Method] Waters HSS T3 column （2.1 mm×100 mm, 1.8 μm） was used for separation. Gradient elution was performed using 0.1% formic acid aqueous solution as mobile phase A and acetonitrile containing 0.1% formic acid as mobile phase B, respectively, with a flow rate of 0.40 mL/min and sample size of 2 μL. Positive-ion electrospray ionization （ESI） source scanning mode was used for detection, and multiple reaction monitoring （MRM） mode was used for signal acquisition. The recovery rates of the target drugs obtained with 4 kinds of sample extraction solvents and 2 types of solid-phase extraction column were compared to determine the optimal method of sample pretreatment. Detection of the 64 drugs in 100 batches of compound feed samples of different sources collected in Ningxia Hui Autonomous Region was carried out with the established UHPLC-MS/MS method. [Result] The compound feed samples were extracted with acetonitrile aqueous solution （acetonitrile∶water=8∶2, V/V） containing 0.2% formic acid after homogenization. Oasis PRiME HLB solid-phase extraction column was used to purify the samples. The recovery rates of most target drugs was above 60%. In the concentration range of 5.0-200.0 μg/L, a good linear relationship of the 64 drugs was observed, and their correlation coefficients （R） were all greater than 0.99. The limit of quantitation of different drugs was 5.0-10.0 μg/kg. The average recovery rates of 3 spiking levels at 5.0, 20.0 and 50.0 μg/kg ranged from 41.00% to 120.49%. The intra- and inter-assay relative standard deviations （RSDs） were 0.54%-15.94% and 1.25%-13.64%, respectively. No target drug was detected in the 100 batches of compound feed samples. [Conclusion] The established UHPLC-MS/MS method has good linear relationship, high recovery rate, good precision, high reproducibility and good operability, and can be used for screening the 64 drugs illegally added in compound feed.

Key words: compound feed, ultra-high performance liquid chromatography-triple quadrupole tandem mass spectrometry method, illegal addition, veterinary drug

中图分类号:

陈海燕, 陈娟, 李永琴, 马春芳, 张慧宁, 杨俊华, 杨奇. 配合饲料中64种药物超高效液相色谱-三重四极杆串联质谱检测方法的研究[J]. 畜牧与饲料科学, 2024, 45(3): 11-19.

CHEN Haiyan, CHEN Juan, LI Yongqin, MA Chunfang, ZHANG Huining, YANG Junhua, YANG Qi. Establishment of an Ultra-high Performance Liquid Chromatography-Triple Quadrupole Tandem Mass Spectrometry（UHPLC-MS/MS）Method for Simultaneous Detection of 64 Drugs in Compound Feed[J]. Animal Husbandry and Feed Science, 2024, 45(3): 11-19.

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随着生活水平的不断提高,人们对畜禽产品的质量越来越关注.饲料安全是关乎畜禽产品质量的重要因素之一^[1].畜禽养殖业中抗生素的不合理使用甚至滥用对畜禽产品质量造成了严重危害,也给公共卫生安全带来了巨大挑战.新型高效生物饲料的开发作为饲料行业的新业态,通过在饲料中应用生物技术和生物饲料添加剂代替抗生素的使用,具有降低饲料原料中抗营养因子含量、提高饲料消化率、改善动物肠道健康、提高动物免疫力、提升动物生产性能^[2-4]等作用.本文介绍了生物饲料的定义和分类,以发酵饲料和菌酶协同发酵饲料为例,综述生物饲料在牛、羊、猪生产中的应用研究进展,分析当前生物饲料应用存在的问题并提出相关建议,以期为生物饲料的开发及应用提供参考.