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目的建立测定人血浆中非布司他及其活性代谢物67M-4质量浓度的LC-MS/MS方法。方法采用乙腈沉淀蛋白法处理血浆样品,采用不同的LC-MS/MS条件分别测定血浆中极性差异较大的非布司他和67M-4的质量浓度。非布司他的测定采用Platisil ODS(150 mm×4.6 mm,5μm)色谱柱和体积分数0.1%甲酸溶液-乙腈(体积比30∶70)为流动相;67M-4的测定采用Zorbax SB-C18(50 mm×4.6 mm,5μm)色谱柱和体积分数0.2%甲酸溶液-乙腈为流动相,梯度洗脱。质谱检测采用多反应离子选择监测(MRM)方式。结果血浆中非布司他和67M-4的质量浓度分别在0.010 00~15.06 mg·L-1和0.505 0~202.0μg·L-1内线性关系良好,提取回收率分别为88.0%~93.5%和88.7%~96.2%,方法的专属性,日内、日间精密度及基质效应均符合测定要求。结论该方法灵敏、可靠,适用于非布司他在人体内的药代动力学研究。
OBJECTIVE To establish a LC-MS / MS method for the determination of febuxostat and its active metabolite 67M-4 in human plasma. Methods Plasma samples were treated with acetonitrile-precipitated protein. The different concentrations of febuxostat and 67M-4 in plasma were determined by different conditions of LC-MS / MS. The febuxostat assay was performed on a Platisil ODS column (150 mm × 4.6 mm, 5 μm) with a 0.1% formic acid solution in acetonitrile (30:70 by volume) as the mobile phase. The determination of 67M-4 was performed on a Zorbax SB-C18 (50 mm × 4.6 mm, 5 μm) column and a 0.2% formic acid solution in acetonitrile as mobile phase with gradient elution. Mass spectrometry detection using multiple reactive ion selective monitoring (MRM) mode. Results The linear correlation between the concentrations of febuxostat and 67M-4 in plasma ranged from 0.010 00 to 15.06 mg · L-1 and from 0.505 0 to 202.0 μg · L-1, respectively. The recovery rates were 88.0% -93.5% And 88.7% ~ 96.2%. The specificity of the method, intra-day and inter-day precision and matrix effects all met the determination requirements. Conclusion The method is sensitive and reliable and suitable for the study of pharmacokinetics of febuxostat in human.