平衡透析并液质联用法同时测定度洛西汀、氟西汀和阿戈美拉汀的人血浆蛋白结合率

摘要

Objective To establish an equilibrium dialysis combined with high performance liquid chromatography-tandem mass spectrometric method(HPLC-MS/MS) for the determination of human plasma protein binding of duloxetine,fluoxetine and agomelatine.Methods The plasma samples were prepared by equilibrium dialysis methods.Diazepam was used as internal standard.The separation was achieved on a WATERS Xterr RP18 column(4.6 mm×100 mm,3.5 μm)with gradient elution by a mobile phase consisting of methanol and 0.3% formic acid containing 5 mmol·L-1 ammonium acetate.Electrospray ionization source was applied and operated in the positive multiple reaction monitoring mode.The specificity,standard curve and lower limit of quantitation,precision and recovery,the matrix effect as well as stability were investigated.Results In plasma and PBS buffer,the good linear relationship of duloxetine,fluoxetine and agomelatine were obtained.The inter-batch and intra-batch precision were all less than 15%,respectively.The average recovery rate was ranged from 85.89% to 106.86%.The matrix factor normalized by internal standard was ranged from 86.55% to 104.94%.Conclusion The method is accurate,sensitive,simple and suitable for the study of plasma protein binding of duloxetine,fluoxetine and agomelatine in hunman plasma.%目的 建立平衡透析并液质联用法(HPLC-MS/MS),同时测定度洛西汀、氟西汀和阿戈美拉汀在人血浆中的蛋白结合率.方法 用平衡透析法处理血浆得透析内液和透析外液.以地西泮作为内标,色谱柱为WATERS Xterra(R) RP18(4.6 mm×100 mm,3.5 μm),以甲醇-水(5 mmol·L-1醋酸铵-0.3%甲酸)为流动相,梯度洗脱,用电喷雾离子源,正离子多反应监测.考察该方法的特异性、标准曲线与最低定量限、精密度与回收率、基质效应以及稳定性. 结果 在血浆和磷酸盐缓冲液(PBS)中,度洛西汀、氟西汀和阿戈美拉汀的标准曲线线性良好,批内、批间精密度均<15%,提取回收率85.89%~106.86%,内标归一化基质效应在86.55%~104.94%,且稳定性均较好. 结论 该方法准确、灵敏、简便,可用于度洛西汀、氟西汀和阿戈美拉汀的人血浆蛋白结合率的研究.