Palermo Amelia, Alessi Beatrice, Botrè Francesco, de la Torre Xavier, Fiacco Ilaria, Mazzarino Monica
Laboratorio Antidoping, Federazione Medico Sportiva Italiana, Largo Giulio Onesti, 1, 00197, Rome, Italy.
Dipartimento di Chimica e Tecnologia del Farmaco, 'Sapienza' Università di Roma, Piazzale Aldo Moro 5, 00185, Rome, Italy.
Drug Test Anal. 2016 Sep;8(9):930-9. doi: 10.1002/dta.1897. Epub 2015 Oct 20.
We have studied whether the phase II metabolism of 19-norandrosterone, the most representative metabolite of 19-nortestosterone (nandrolone), can be altered in the presence of other drugs that are not presently included on the Prohibited List of the World Anti-Doping Agency. In detail, we have evaluated the effect of non-prohibited drugs belonging to the classes of anti-fungals, benzodiazepines, and non-steroidal anti-inflammatory drugs on the glucuronidation of 19-norandrosterone. In vitro assays based on the use of either pooled human liver microsomes or specific recombinant isoforms of uridine diphosphoglucuronosyl-transferase were designed and performed to monitor the formation of 19-norandrosterone glucuronide from 19-norandrosterone. Determination of 19-norandrosterone (free and conjugated fraction) was performed by gas chromatography - mass spectrometry after sample pretreatment consisting of an enzymatic hydrolysis (performed only for the conjugated fraction), liquid/liquid extraction with tert-butylmethyl ether, and derivatization to form the trimethylsilyl derivative. In parallel, a method based on reversed-phase liquid chromatography coupled to tandem mass spectrometry in positive electrospray ionization with acquisition in selected reaction monitoring mode was also developed to identify the non-prohibited drugs considered in this study. Incubation experiments have preliminarily shown that the glucuronidation of 19-norandrosterone is principally carried out by UGT2B7 (39%) and UGT2B17 (31%). Inhibition studies have shown that the yield of the glucuronidation reaction is reduced in the presence of the anti-fungals itraconazole, ketoconazole, and miconazole, of the benzodiazepine triazolam and of the non-steroidal anti-inflammatory drugs diclofenac and ibuprofen, while no alteration was recorded in the presence of all other compounds considered in this study. Copyright © 2015 John Wiley & Sons, Ltd.
我们研究了19-去甲睾酮（诺龙）最具代表性的代谢物19-去甲雄酮的II相代谢在存在世界反兴奋剂机构禁用清单中目前未包含的其他药物时是否会发生改变。具体而言，我们评估了抗真菌药、苯二氮䓬类药物和非甾体抗炎药这几类非禁用药物对19-去甲雄酮葡萄糖醛酸化的影响。设计并进行了基于使用人肝微粒体池或尿苷二磷酸葡萄糖醛酸基转移酶的特定重组同工型的体外试验，以监测19-去甲雄酮葡萄糖醛酸苷从19-去甲雄酮的形成。19-去甲雄酮（游离和结合部分）的测定是在样品预处理后通过气相色谱-质谱法进行的，样品预处理包括酶促水解（仅对结合部分进行）、用叔丁基甲基醚进行液-液萃取以及衍生化以形成三甲基硅烷基衍生物。同时，还开发了一种基于反相液相色谱-串联质谱法的方法，在正电喷雾电离下以选择反应监测模式进行采集，以鉴定本研究中考虑的非禁用药物。孵育实验初步表明，19-去甲雄酮的葡萄糖醛酸化主要由UGT2B7（39%）和UGT2B17（31%）进行。抑制研究表明，在抗真菌药伊曲康唑、酮康唑和咪康唑、苯二氮䓬类药物三唑仑以及非甾体抗炎药双氯芬酸和布洛芬存在的情况下，葡萄糖醛酸化反应的产率降低，而在本研究中考虑的所有其他化合物存在的情况下未记录到改变。版权所有© 2015约翰威立父子有限公司。
