大鼠体内外细胞色素P450酶活性检测方法的建立及应用

Establishment and application of detection methods for cytochrome P450 enzyme activities in vivo and in vitro of rats

目的建立快速、简便、灵敏可以同时测定大鼠体内外“鸡尾酒”探针药物和代谢产物的LC-MS/MS方法,并用于大鼠肝微粒体外代谢和大鼠体内的药代动力学研究。方法所有样品均加入乙腈进行沉淀处理,然后在UPLC CORTECS C18(2.1 mm×50 mm,1.6μm)色谱柱分离,质谱方法采用多反应监测模式(MRM)。液相方法采用梯度洗脱流动相乙腈-0.1%甲酸水。结果6种探针药和代谢产物的体内外样品方法学验证符合生物样品检测要求。该方法成功应用于大鼠体内外“鸡尾酒”实验中。结论本实验建立的“鸡尾酒”方法可用于体内外快速评价药物或中草药对相应的CYP450亚型酶活性的影响,可以用于潜在的药物相互作用的预测。

Objective To establish a rapid,simple and sensitive LC-MS/MS method for simultaneous determination of"cocktail"probe drugs and metabolites in vivo and in vitro of rats,and to apply it to studies of in vitro metabolism of rats′liver microsomes and in vivo pharmacokinetics of rats.Methods Both in vivo and in vitro samples were treated by acetonitrile precipitation method.Probe drugs and metabolites were detected by UPLC CORTECS C18(2.1 mm×50 mm,1.6μm)chromatographic column combined with UPLC-MS/MS method of multiple reaction monitoring mode(MRM),in which acetonitrile-0.1%formic acid water was used as gradient elution method of mobile phase.Results Methodological verification of samples in vitro and in vivo of 6 probe drugs and metabolites met the requirements of biological sample detection.The method had been successfully applied to"cocktail"experiments in vivo and in vitro of rats.Conclusion The"cocktail"method established in this experiment could be used to rapidly evaluate the efficacies of drugs or Chinese herbal medicines on the activities of the corresponding CYP450 subtypes enzymes in vivo and in vitro and could be used to predict potential drug interactions.