LC-MS/MS法测定人血浆中硝苯地平和去氢硝苯地平

Rapid and simultaneous determination of nifedipine and dehydronifedipine in human plasma

硝苯地平是一种二氢吡啶类钙拮抗剂,通过CYP3A4代谢生成主要代谢物去氢硝苯地平。本文建立了液相色谱-串联质谱(LC-MS/MS)法同时测定人血浆中的硝苯地平及其代谢物去氢硝苯地平。采用d_6-硝苯地平/d_6-去氢硝苯地平作为内标,血浆样品经乙腈沉淀蛋白后,通过Hypersil Gold C_(18)(50 mm×2.1 mm, 1.9μm)色谱柱分离,以甲醇-0.1%甲酸的5 mmol·L^(-1)醋酸铵水溶液作为流动相。采用电喷雾离子源(ESI源),以多反应监测正离子模式检测,用于定量分析的硝苯地平和去氢硝苯地平的离子对分别为m/z 347.3→254.1和m/z 345.2→283.9,内标的离子对分别为m/z 353.3→257.1和m/z 351.2→286.9。线性范围硝苯地平为0.10～80.0 ng·mL^(-1),去氢硝苯地平为0.050～40.0 ng·mL^(-1)。与文献报道方法相比,色谱分离得到改善,检测灵敏度显著提高。本方法经验证后,成功应用于人血浆中硝苯地平(CYP3A4底物)与阿帕替尼(CYP3A4抑制剂)的相互作用研究。本文临床试验经伦理委员会批准,并在中国医科大学第一附属医院进行。

Nifedipine, a calcium channel antagonist, is metabolized mainly by CYP3A4 to dehydronifedipine.A rapid and sensitive liquid chromatography-tandem mass spectrometry(LC-MS/MS) method was developed to simultaneously determine nifedipine and dehydronifedipine in human plasma using d6-nifedipine/d6-dehydronife‐dipine as internal standards. After extraction from the plasma by protein precipitation, the analytes and internal standard were separated on a Hypersil Gold C18(50 mm×2.1 mm, 1.9 μm). The mobile phase consisted of methanol and 5 mmol · L-1ammonium acetate aqueous solution(0.1% formic acid). Positive electrospray ionization was performed using multiple reaction monitoring(MRM) with transitions of m/z 347.3→254.1 for nifedipine, m/z345.2→283.9 for dehydronifedipine, m/z 353.3→257.1 for d6-nifedipine, m/z 351.2→286.9 for d6-dehydronife‐dipine. The method had a linear calibration curves over the concentrations of 0.10-80.0 ng · mL-1for nifedipine and0.050-40.0 ng · mL-1for dehydronifedipine. The validated LC-MS/MS method has been successfully used study pharmacokinetic interactions of apatinib(CYP3A4 inhibitor) and nifedipine(CYP3A4 substrate) in human. This clinical trial was approved by the society of ethics and conducted in the first hospital of China medical university.