盐酸哌甲酯在血浆中的稳定性及其药动学研究

LC-MS/MS assay of methylphenidate: stability and pharmacokinetics in human

研究哌甲酯（MPH）在人血浆中不稳定性，建立HPLC．MS／MS测定方法并运用于人体药代动力学研究。采用HPLC及LC．MS／MS法考察哌甲酯在醇、水、血浆中的稳定性及甲酸水溶液处理对哌甲酯降解的抑制作用。结果表明，于新鲜制备的人血浆样品（200μL）中立即加入2％甲酸水溶液10μL，能够抑制血浆中血浆酯酶的活性。以5mmol·L-1醋酸铵水溶液（含0．1％甲酸）-甲醇（54：46）为流动相；使用SapphireCl8柱进行分离。采用电喷雾离子源（ESI＋）及多反应监测模式（MRM）进行检测，检测离子为m／z234．2—84．1（MPH），m／z260.3—183．1（普萘洛尔，内标）。哌甲酯血浆浓度测定线性范围为0．035-40ng·mL-1；日内、日间精密度（RSD）均小于5％，准确度为±2％之内。应用此法研究了6名健康志愿者单剂量口服哌甲酯36mg后药动学特点。所建方法能准确测定人血浆中哌甲酯的血药浓度，可用于哌甲酯的人体药动学研究。

The study aims to solve the instability problem of methylphenidate （MPH） in plasma, and establish a LC-MS/MS method for simultaneous determining of MPH in human plasma. The stabilities of MPH in different media were studied, and the degradation characteristics of MPH in these media were also investigated by HPLC and LC-MS/MS. To a 200 μL aliquot of freshly collected plasma sample, 10 μL 2% formic acid was added immediately to prevent the hydrolysis of MPH in human plasma samples. Chromatographic separation was performed on a Sapphire C 18 column using the mobile phase of methanol - 5 mmol L-1 ammonium acetate buffer solution containing 0.1% formic acid （46 ： 54）. MPH was quantified by tandem mass spectrometry operating in positive electrospray ionization mode with multiple reaction monitoring. The detection used the transitions of protonated molecules at m/z 234.2→84.1 for MPH and m/z 260.3→183.1 for propranolol （IS）, separately. The intra- and inter-assay precisions were all below 5.0%. The accuracies were all in standard ranges. The linear calibration curve was obtained in the concentration range of 0.035-40 ng.mL-1. The methods fulfilled the demand. The method was used to determine the concentration of MPH in human plasma after a single dose of 36 mg MPH tablet to 6 healthy Chinese volunteers. The method is suitable for the precisely determination of MPH and for pharmacokinetic study of MPH in human plasma.