DoCoLab, UGent, Department of Clinical Chemistry, Microbiology and Immunology, Technologiepark 30, Zwijnaarde, Belgium.
Drug Test Anal. 2009 Nov;1(11-12):554-67. doi: 10.1002/dta.56.
Anabolic steroids are among the most frequently detected compounds in doping analysis. They are extensively metabolized and therefore an in-depth knowledge about steroid metabolism is needed. In this study, a liquid chromatography tandem mass spectometry (LC-MS/MS) method based on a precursor ion scan with a uPA-SCID mouse with humanized liver (a chimeric mouse) was explored for the detection of steroid metabolism. Methandienone was used as a model compound. The application of the precursor ion scan method in positive human samples and chimeric mice samples after methandienone administration allowed the detection of most steroid metabolites without any structural restriction. Three hitherto unreported metabolites were found using this approach. These metabolites were characterized using LC-MS/MS and feasible structures were proposed. The structure of one of them, 6-ene-epimethandienone, was confirmed by the synthesis of the reference compound. A selected reaction monitoring (SRM) method for the specific detection of all these metabolites has been developed. The application of this method to several human and chimeric mouse samples confirmed that more than 80% of the steroid metabolites were found in both samples. Only metabolites that are poorly detectable by LC-MS/MS were not detected in some urine samples. The metabolic nature of the unreported metabolites was also confirmed. A global strategy for the detection of steroid metabolites combining both human and chimeric mouse urine is proposed.
合成代谢类固醇是兴奋剂分析中最常检测到的化合物之一。它们广泛代谢，因此需要深入了解类固醇代谢。在这项研究中，探索了一种基于前体离子扫描的液相色谱串联质谱（LC-MS/MS）方法，该方法使用具有人源化肝脏的 uPA-SCID 小鼠（嵌合小鼠）来检测类固醇代谢。以美雄酮作为模型化合物。在前体离子扫描方法的应用中，在给予美雄酮后，正人类样本和嵌合小鼠样本中可以检测到大多数类固醇代谢物，而没有任何结构限制。使用这种方法发现了三种以前未报道的代谢物。使用 LC-MS/MS 对这些代谢物进行了表征，并提出了可行的结构。其中一种代谢物，6-ene-epimethandienone 的结构通过参考化合物的合成得到了证实。已经开发出一种用于特异性检测所有这些代谢物的选择反应监测（SRM）方法。该方法在几个人类和嵌合小鼠样本中的应用证实，在这两种样本中都发现了超过 80%的类固醇代谢物。只是由于 LC-MS/MS 检测能力有限，在一些尿液样本中未检测到某些代谢物。未报道的代谢物的代谢性质也得到了证实。提出了一种结合人源和嵌合小鼠尿液的检测类固醇代谢物的全局策略。
