临床干血斑样本中12种抗凝血鼠药的液相色谱-串联质谱法同时定量检测

摘要

To address the fast detection need towards poisoned patients in clinic,a sensitive,accurate liquid chromatographic-tandem mass spectrometric method was developed here for the qualitative and quantitative determinations of twelve anticoagulant rodenticides in dried blood spots.For the thoroughly punched dried blood spots,conditions influencing the extraction efficiency,including the kind of filter paper card,the water-soaking step and the kind of extraction solvent were investigated.A C1s column was employed as separation phase while ammonium acetate in water (5 mmol/L)-ammonium acetate in methanol (5 mmol/L) was used as mobile phase by gradient elution.Then the sample was detected using negative electrospray ionization under multiple reaction monitoring mode.The results showed that,with the 903 type filter card as supporting matrix and a water-soaking step,followed by an extraction with methanol solvent containing internal standard for 5 min,the extraction efficiencies of the twelve rodenticides ranged from 66％ to 115％ with their intra-day relative standard deviations less than 15％.Good method validation results were obtained for all the anticoagulant rodenticides.The linearity for pindone was from 20 to 500 ng/mL with a correlation coefficient(r2) of 0.998 7,and that for the other eleven analytes were from 5 to 500 ng/mL with their r2 of 0.998 8-0.999 6.The recoveries for all the analytes ranged from 61％ to 105％,with the matrix effects of 71％-193％ and the intra-day relative standard deviations less than 15％.The established method was accurate,sensitive,rapid and convenient,and was successfully applied in the rapid detection of rodenticides in three clinic blood samples.It provides a new approach for clinical diagnosis and forensic toxicant analysis on anticoagulant rodenticide intoxication accidents.%为应对临床中毒快检的需求,建立了干血斑样本中12种抗凝血鼠药的液相色谱-串联质谱定性定量方法.针对全打孔的干血斑样本,考察了滤纸卡种类、润湿步骤、提取溶剂种类对提取效果的影响.采用C18色谱柱进行分离,以乙酸铵(5 mmol/L)水溶液-乙酸铵(5 mmol/L)甲醇溶液为流动相进行梯度洗脱,在电喷雾负离子电离方式下使用三重四极多反应监测模式检测.结果表明,采用Whatman903滤纸卡为基底,以水为溶剂充分润湿,再以含内标的甲醇溶液提取5 min,各鼠药的提取率为66％ ～ 115％,且结果稳定(日内RSD小于15％).除杀鼠酮的线性范围为20 ～500 ng/mL(r2 =0.998 7)外,其它11种鼠药的线性范围为5～500 ng/mL(r2 =0.998 8 ～0.999 6);12种鼠药的回收率为61％～ 105％,基质效应为71％ ～193％(日内RSD小于15％).该方法准确、灵敏、快速且操作简便,成功应用于3例临床鼠药中毒病人样本的快速检测,为临床毒物检测和法医毒物分析的快速筛查和准确定量提供了新的技术方法.