目的:建立LC-MS/MS法同时测定血浆中霉酚酸(MPA)及其代谢物MPAG与AcMPAG浓度,用于治疗药物监测与药动学研究。方法:100μl血浆标本加入300μl含内标吲哚美辛的乙腈沉淀,离心后取上清液进样。色谱柱为Agilent Eclipse XDB-C₁₈柱(3.5μm, 2.1 mm×100 mm),流动相为2 mmol·L^-1甲酸铵水溶液-甲醇,采用梯度洗脱的方法,流速0.3 ml·min^-1。用多反应监测进行定量,ESI负离子方式进行检测,用于定量分析的检测离子分别为m/z 319.1→274.9(MPA)、m/z 495.3→174.4(MPAG与AcMPAG)与m/z 356.1→312.0(内标吲哚美辛)。24例肝移植患者,采用包括霉酚酸钠肠溶片(EC-MPS)的三联免疫抑制方案,测定服药后MPA及其代谢物浓度并计算药动学参数。结果:MPA在0.10~50.5μg·ml^-1,MPAG在1.13~450μg·ml^-1,AcMPAG在0.11~22.4μg·ml^-1范围内线性良好(r²>0.99)。MPA、MPAG、AcMPAG提取回收率为77.3%~92.6%;基质效应为76.0%~86.7%;回收率为94.2%~116.4%;日内及日间变异均小于15%。24例肝移植患者用药1周后MPA、MPAG、AcMPAG的主要药动学参数分别如下:峰浓度C_max为(18.8±10.9),(154±118),(3.07±2.85)μg·ml^-1;达峰时间T_max为(3.82±2.66),(4.74±2.51),(4.51±2.72)h;曲线下面积AUC_0-12为(45.2±20.3),(1456±1195),(18.3±16.2)μg·h·ml^-1;消除半衰期t_1/2为(3.21±2.56),(9.26±4.33),(5.57±5.76)h。结论:本研究所建立的方法快速准确、灵敏、专属性强,适用于MPA及其代谢物的血药浓度监测和人体药动学研究。

OBJECTIVE To establish an LC-MS/MS method for the determination of mycophenolic acid(MPA) and its metabolites MPAG and AcMPAG in human plasma and investigate the pharmacokinetic characteristics in Chinese liver-transplanted patients. METHODS 300μl of acetontrile containing indomethacin(internal standard) was added in 100μl plasma to precipitate protein. After centrifugation, the supernatant was eluted through Agilent Eclipse XDB-C₁₈(3.5μm, 2.1 mm×100 mm) column by water(2 mmol·L^-1 ammonium acetate)-methanol through a gradient assay, the flow rate was set as 0.3 ml·min^-1. Electrospray ionization(ESI) source was applied and operated in the negative ion mode. Multiple reaction monitoring(MRM) mode with the transition of m/z 319.1→274.9(MPA), 495.3→174.4(MPAG and AcMPAG) and 356.1→312.0(IS) was used for quantification. Plasma concentration of MPA, MPAG, AcMPAG of 24 liver-transplanted patients received immunosuppressive therapy including enteric-coated mycophenolate sodium were determined and pharmacokinetic parameters were calculated. RESULTS The method was proved to be linear in the range of 0.10-50.5μg·ml^-1 for MPA, 1.13-450μg·ml^-1 for MPAG, 0.11-22.4μg·ml^-1 for AcMPAG(r²>0.99). The extract recovery, matrix effect, recovery of MPA and its metabolites was 77.3%-92.6%, 76.0%-86.7% and 94.2%-116.4%, respectively. The within-day and between-day variation were both lower than 15%. The pharmacokinetic parameters of MPA and its metabolites of 24 liver transplant patients were as follows:C_max:(18.8±10.9),(154±118),(3.07±2.85)μg·ml^-1; t_max:(3.82±2.66),(4.74±2.51),(4.51±2.72) h; AUC_0-12:(45.2±20.3),(1456±1195),(18.3±16.2)μg·h·ml^-1;t_1/2:(3.21±2.56),(9.26±4.33),(5.57±5.76)h, respectively. CONCLUSION The method established is rapid, accurate, sensitive and specific, which is suitable for the therapeutic drug monitoring(TDM) and pharmacokinetic study of MPA and its metabolites.