Dong Ying, Yan Kuan, Ma Yanhua, Yang Zhiyong, Zhao Jun, Ding Jinglin
National Anti-Doping Laboratory, China Anti-Doping Agency, China.
National Anti-Doping Laboratory, China Anti-Doping Agency, China.
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Jun 1;1022:153-158. doi: 10.1016/j.jchromb.2016.04.023. Epub 2016 Apr 13.
Glycerol and mannitol have the potential to act as plasma volume expanders and have been prohibited as masking agents by the World Anti-Doping Agency (WADA) accordingly. In this study, an improved strategy was developed and validated for the determination of urinary glycerol and mannitol levels simultaneously using a liquid chromatography/tandem mass spectrometry technique within 7min in an initial testing procedure. For confirmation, mannitol and all possible hexitols (allitol, altritol, galactitol, iditol and sorbitol) that can occur in human urine were baseline separated. This method made use of the derivatization of glycerol and mannitol by benzoyl chloride followed by analysis via LC-ESI-MS/MS with limited sample preparation. The limit of detection (LOD) for glycerol and mannitol was lower than 50ng/mL. The limit of quantitation (LOQ) for both substances was below 150ng/mL. The assay was linear from 0.15 to 1000μg/mL for glycerol and mannitol in human urine. The coefficients of variation of all inter- and intra-assay determinations at three concentration levels (0.5, 500, 900μg/mL) were better than 13% for glycerol and under 15% for mannitol. The method also afforded satisfactory results in terms of accuracy, derivatization yield, extraction recovery, matrix effect and specificity for both substances.
甘油和甘露醇有作为血浆容量扩充剂的潜力，因此被世界反兴奋剂机构（WADA）禁止用作掩蔽剂。在本研究中，开发并验证了一种改进策略，可在初始测试程序中使用液相色谱/串联质谱技术在7分钟内同时测定尿中甘油和甘露醇水平。为进行确证，将甘露醇以及人尿中可能出现的所有己糖醇（阿洛糖醇、阿卓糖醇、半乳糖醇、艾杜糖醇和山梨醇）进行基线分离。该方法利用苯甲酰氯对甘油和甘露醇进行衍生化，随后通过液相色谱-电喷雾串联质谱仪进行分析，样品前处理有限。甘油和甘露醇的检测限低于50ng/mL。两种物质的定量限均低于150ng/mL。人尿中甘油和甘露醇的测定在0.15至1000μg/mL范围内呈线性。在三个浓度水平（0.5、500、900μg/mL）下，甘油的所有批间和批内测定变异系数均优于13%，甘露醇的变异系数低于15%。该方法在两种物质的准确性、衍生化产率、提取回收率、基质效应和特异性方面也都给出了令人满意的结果。
