在线固相萃取-超高效液相色谱-串联质谱法测定尿液中芬太尼及其代谢物的含量  

Determination of Fentanyl and Its Metabolite in Urine by On-line SPE Combined with UHPLC-MS/MS

作　　者：赵清佳 汪蓉[2] 徐智儒 盛振海 梁晨[2] 张玉荣[2] ZHAO Qingjia;WANG Rong;XU Zhiru;SHENG Zhenhai;LIANG Chen;ZHANG Yurong(State Key Laboratory of New Drug and Pharmaceutical Process,China State Institute of Pharmaceutical Industry,Shanghai 200040,China;Shanghai Key Laboratory of Crime Scene Evidence,Shanghai Institute of Forensic Science,Shanghai 200083,China)

机构地区：[1]中国医药工业研究总院创新药物与制药工艺国家重点实验室,上海200040 [2]上海市公安局物证鉴定中心上海市现场物证重点实验室,上海200083

出　　处：《理化检验（化学分册）》2021年第6期511-518,共8页Physical Testing and Chemical Analysis(Part B:Chemical Analysis)

摘　　要：建立了在线固相萃取(on-line SPE)-超高效液相色谱-串联质谱法(UHPLC-MS/MS)测定尿液中芬太尼及其代谢物去甲芬太尼的方法,并将该方法运用于大鼠尿液中芬太尼及其代谢物的检测时限研究。将尿液用pH 9.2的2 mmol·L^(-1)甲酸铵溶液稀释,离心后直接进样。采用Oasis HLB固相萃取小柱通过在线固相萃取富集净化目标物,用pH 9.2的2 mmol·L^(-1)甲酸铵溶液淋洗,以分析泵的初始流动相将目标物洗脱至Hypesil GOLD C_(18)分析柱上,以不同体积比的含有2 mmol·L^(-1)甲酸铵和0.1%(体积分数)甲酸的甲醇溶液、含有2 mmol·L^(-1)甲酸铵和0.1%(体积分数)甲酸的水溶液的混合液为流动相进行梯度洗脱。串联质谱分析中采用电喷雾正离子源(ESI+)和选择离子监测(SRM)模式检测。将芬太尼以20μg·kg^(-1)的给药剂量对大鼠尾静脉注射后进行尿液收集,考察原体和代谢物的检测时限。结果表明:芬太尼和去甲芬太尼的质量浓度在2.0~800 ng·L^(-1)内与其峰面积呈线性关系,检出限均为0.200 ng·L^(-1),测定值的相对标准偏差(n=5)均小于15%。在大鼠尿液中芬太尼原体的检测时限为5~7 d,而其代谢物去甲芬太尼的检测时限长达28 d。A method was established for the determination of fentanyl and its metabolite(norfentanyl)in urine by on-line SPE combined with UHPLC-MS/MS,which was applied to the study of detection window of fentanyl and its metabolite in rat urine.The urine was diluted with 2 mmol·L^(-1) ammonium formate buffer solution at pH 9.2,and the diluted sample was directly injected after centrifugation.The target compounds were enriched and purified by on-line SPE with Oasis HLB solid phase extraction column,and eluted onto the Hypesil GOLD C_(18) analytical column with the initial mobile phase of the analytical pump.A mixture of methanol solution containing 2 mmol·L^(-1) ammonium formate and 0.1%(φ)formic acid,and aqueous solution containing 2 mmol·L^(-1) ammonium formate and 0.1%(φ)formic acid in different ratios was used as the mobile phase for gradient elution.ESI+and the mode of selected ion monitoring(SRM)were adopted in MS/MS.Rat urine samples were collected after the tail vein injection fentanyl of rats at a dose of 20μg·kg^(-1),and the detection window of the protosome and metabolite was studied by this method.As shown by the results,linear relationships between values of peak area and mass concentration of fentanyl and norfentanyl were found in the same range of 2.00-800 ng·L^(-1),with the same detection limit(3 S/N)of 0.200 ng·L^(-1).RSDs(n=5)of the determined values were all less than 15%.The detection window of protosome fentanyl in rat urine was 5-7 d,while the detection window of its metabolite norfentanyl could be up to 28 d.

关 键 词：超高效液相色谱-串联质谱法 在线固相萃取 芬太尼及其代谢物 检测时限 

分 类 号：O657.63[理学—分析化学]