超高效液相色谱-串联质谱法同时检测人尿中34种小肽类禁用物质

Simultaneous detection of 34 prohibited small peptides in human urine by UHPLC-MS/MS

采用超高效液相色谱-串联质谱技术,建立了人尿中的34种小肽类禁用物质的测定方法。使用Oasis WCX固相萃取小柱(1 cc,30 mg)对尿样进行前处理,洗脱液在38℃氮气流下吹干,复溶后使用Kinetex 2. 6μC18 2. 1 mm×100 mm色谱柱以及Agilent 1290-6470 C进行分析。以含0. 1%甲酸的水和乙腈为流动相进行梯度洗脱,使用电喷雾离子源以正离子动态多反应监测模式进行扫描。该方法对于34种小肽类禁用物质的方法选择性、可靠性和稳健度良好,检出限不高于2 ng/mL(S/N> 3),回收率在0. 5%~200. 2%之间,基质效应分布范围为76. 8%~5293. 8%。该方法简单,特异性、灵敏度均可满足世界反兴奋剂机构(WADA)技术文件要求,目前已应用于北京兴奋剂检测实验室的常规检测中。

An ultra-high performance liquid chromatography-tandem mass spectrometry( UHPLC-MS/MS)method was developed for the simultaneous detection of 34 prohibited small peptides in human urine. Urine samples were pre-treated with Oasis WCX cartridge and the eluants were evaporated under nitrogen flow at 38 ℃and resolved before injection into UHPLC-MS/MS. The mobile phase in the gradient elution of UHPLC was comprised of 0. 1% formic acid in water and acetonitrile. The targeted small peptides were analyzed using dynamic multiple reaction monitoring( MRM) mode under positive electrospray ionization in tandem mass spectrometry. The results showed that the method had good selectivity,reliability and robustness. The limits of detection were no more than 2 ng/m L,and the spiked recoveries and matrix effects ranged from 0. 5% to200. 2% and 76. 8% to 5293. 8%,respectively. The described method is simple with good specificity and sensitivity which meets the demand listed in World Anti-Doping Agency( WADA) technical document,and has been applied for the routine analysis of urine samples in Beijing National Anti-Doping Laboratory.