近年来,由于生产丰富和生活水平提高,禽蛋的消费量有所增加。因此,禽蛋的安全要求逐渐提高。目前,关于确定禽蛋产卵期间禁用兽药的分析方法的报道很少。因此,建立高通量和高效的筛选方法来监测产蛋期禁用的兽药势在必行。在这项研究中,开发了一种基于超高效液相色谱-串联质谱(UPLC-MS/MS)结合基于QuEChERS技术的分析方法,用于同时测定31种禁用的兽药,包括9种药物类别(大环内酯类,解热和镇痛药物,磺胺类药物,抗菌增效剂,抗球虫药,腐殖质,喹诺酮类药物,四环素,氨酚)在不同类型的禽蛋中。影响反应的主要因素,recovery,和方法的灵敏度,如提取溶剂,净化吸附剂,LC分离条件,和MS/MS参数,在样品预处理和仪器分析过程中进行了优化。用2mL0.1mol/L乙二胺四乙酸二钠溶液和8mL3%乙酸乙腈溶液提取2.00g鸡蛋中的31种兽药残留,并用2克氯化钠盐析。离心后,使用QuEChERS方法用100mg十八烷基硅烷键合的硅胶(C18)净化5mL上清液,50毫克的N-丙基乙二胺(PSA),和50mg的基于NH2的吸附剂。吹氮和再溶解后,在WatersCORTECSUPLCC18分析色谱柱(150mm×2.1mm,1.8μm)在流速下,柱温,注射量为0.4mL/min,30℃,和5μL,分别。在这些分析物中,在正电喷雾电离(ESI+)条件下,使用(A)5mmol/L乙酸铵(pH4.5)和(B)乙腈作为流动相,以动态多反应监测(MRM)模式获得26种分析物。梯度洗脱程序如下:0-2.0分钟,12%B-30%B;2.0-7.5分钟,30%B-50%B;7.5-10.0分钟,50%B;10.0-10.1分钟,50%B-100%B;10.1-12.0分钟,100%B;12.0-12.1分钟,100%B-12%B;使用(A)H2O和(B)乙腈作为流动相,在负电喷雾电离(ESI-)条件下以MRM模式获得其它五种目标分析物。梯度洗脱程序如下:0-2.0分钟,12%B-40%B;2.0-6.0分钟,40%B-80%B;6.0-6.1分钟,80%B-100%B;6.1-8.0分钟,100%B;8.0-8.1分钟,100%B-12%B使用基质匹配的外标校准进行定量。结果表明,各化合物在各自范围内具有良好的线性关系,相关系数>0.99。检测限(LODs)和定量限(LOQs)分别为0.3-3.0μg/kg和1.0-10.0μg/kg,分别。31种禁用兽药的平均回收率达到三个水平(LOQ,最大残留限值(MRL),禽蛋中的2MRL)介于61.2%至105.7%之间,相对标准偏差(RSD)为1.8%~17.6%。该方法用于30份市售禽蛋样品中禁用兽药的检测和分析。包括20个鸡蛋,5个鸭蛋,5个鹅蛋在一个鸡蛋中检测到恩诺沙星,含量为12.3μg/kg。该方法简单,经济,实用,并能够同时测定禽蛋中的多种禁用兽药。
The consumption of poultry eggs has increased in recent years owing to the abundance of production and improvements in living standards. Thus, the safety requirements of poultry eggs have gradually increased. At present, few reports on analytical methods to determine banned veterinary drugs during egg-laying period in poultry eggs have been published. Therefore, establishing high-throughput and efficient screening methods to monitor banned veterinary drugs during egg-laying period is imperative. In this study, an analytical method based on ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) combined with QuEChERS-based techniques was developed for the simultaneous determination of 31 banned veterinary drugs encompassing nine drug classes (macrolides, antipyretic and analgesic drugs, sulfonamides, antibacterial synergists, anticoccidials, antinematodes, quinolones, tetracyclines, amphenicols) in different types of poultry eggs. The main factors affecting the response, recovery, and sensitivity of the method, such as the extraction solvent, purification adsorbent, LC separation conditions, and MS/MS parameters, were optimized during sample pretreatment and instrumental analysis. The 31 veterinary drug residues in 2.00 g eggs were extracted with 2 mL of 0.1 mol/L ethylene diamine tetraacetic acid disodium solution and 8 mL 3% acetic acid acetonitrile solution, and salted out with 2 g of sodium chloride. After centrifugation, 5 mL of the supernatant was cleaned-up using the QuEChERS method with 100 mg of octadecylsilane-bonded silica gel (C18), 50 mg of N-propylethylenediamine (PSA), and 50 mg of NH2-based sorbents. After nitrogen blowing and redissolution, the 31 target analytes were separated on a Waters CORTECS UPLC C18 analytical chromatographic column (150 mm×2.1 mm, 1.8 μm) at a flow rate, column temperature, and injection volume of 0.4 mL/min, 30 ℃, and 5 μL, respectively. Among these analytes, 26 analytes were acquired in dynamic multiple reaction monitoring (MRM) mode under positive electrospray ionization (ESI+) conditions using (A) 5 mmol/L ammonium acetate (pH 4.5) and (B) acetonitrile as mobile phases. The gradient elution program was as follows: 0-2.0 min, 12%B-30%B; 2.0-7.5 min, 30%B-50%B; 7.5-10.0 min, 50%B; 10.0-10.1 min, 50%B-100%B; 10.1-12.0 min, 100%B; 12.0-12.1 min, 100%B-12%B; The five other target analytes were acquired in MRM mode under negative electrospray ionization (ESI-) conditions using (A) H2O and (B) acetonitrile as mobile phases. The gradient elution program was as follows: 0-2.0 min, 12%B-40%B; 2.0-6.0 min, 40%B-80%B; 6.0-6.1 min, 80%B-100%B; 6.1-8.0 min, 100%B; 8.0-8.1 min, 100%B-12%B. Matrix-matched external standard calibration was used for quantification. The results showed that all the compounds had good linear relationships within their respective ranges, with correlation coefficients of >0.99. The limits of detection (LODs) and quantitation (LOQs) were 0.3-3.0 μg/kg and 1.0-10.0 μg/kg, respectively. The average recoveries of the 31 banned veterinary drugs spiked at three levels (LOQ, maximum residue limit (MRL), and 2MRL) in poultry eggs ranged from 61.2% to 105.7%, and the relative standard deviations (RSDs) ranged from 1.8% to 17.6%. The developed method was used to detect and analyze banned veterinary drugs in 30 commercial poultry egg samples, including 20 eggs, 5 duck eggs, and 5 goose eggs. Enrofloxacin was detected in one egg with a content of 12.3 μg/kg. The proposed method is simple, economical, practical, and capable of the simultaneous determination of multiple classes of banned veterinary drugs in poultry eggs.