UPLC-MS/MS法测定人血浆中伏立诺他的浓度及其药代动力学研究

目的：建立超高效液相色谱-串联质谱法(UPLC-MS/MS)测定人血浆中伏立诺他的浓度，评价伏立诺他胶囊在皮肤T细胞淋巴瘤患者体内的药代动力学特征。方法：血浆经乙腈蛋白沉淀后，采用Waters Acquity-UPLC BEH^® C₁₈色谱柱(2.1 mm×50 mm，1.7 μm)作为分析柱，以0.5%乙酸水(A)-0.5%乙酸乙腈(B)为流动相，梯度洗脱(0~0.2 min，20%B;0.2~1.7 min，20%B→90%B，1.7~2.2 min，90%B;2.3~3.0 min，90%B→20%B)，流速0.3 mL·min^-1，进样器内温度4 ℃，柱温40 ℃；电喷雾离子化(ESI+)串联质谱检测，喷雾电压5 kV，温度450 ℃，去簇电压、入口电压及碰撞室出口电压值分别为90、10和13 V；伏立诺他和苯海拉明(内标)的多反应监测(MRM)扫描离子对分别为m/z 265.3→232.0和m/z 256.1→167.0。受试者给予伏立诺他口服治疗(400 mg·d^-1)，分别采集第1d及第22d静脉血以进行单次给药和多次给药的药代动力学分析。结果：伏立诺他浓度在1~1000 ng·mL^-1范围内线性关系良好(r=0.9996)，最小检出浓度为0.1 ng·mL^-1；日内和日间精密度(RSD)均小于14%，准确度为87.6%~109.2%；提取回收率和基质效应分别为93.6%~105.7%和92.3%~93.5%。共完成4例受试者的单次给药(400 mg·d^-1)药代动力学分析，达峰浓度(C_max)为(402.5±50.2)ng·mL^-1，半衰期(T_1/2)为(2.0±2.3)h；多次给药(400 mg·d^-1，连续22d)的C_max为(800±247.2)ng·mL^-1，T_1/2为(2.04±1.4)h，蓄积指数为1.00，未见明显蓄积。结论：该方法的样本前处理简单，样本检测时间为3 min，适用于伏立诺他临床药代动力学研究中高通量检测的要求。

Objective: To develop an ultra high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)method to determine the concentration of vorinostat in human plasma,and to evaluate the pharmacokinetics of oral vorinostat in patients with cutaneous T cell lymphoma. Methods: Vorinostat and diphenhydramine(internal standard)were determined by UPLC-MS/MS using a Waters Acquity-UPLC BEH⁽] declustering potential,90 V;entrance potential,10 V;collision cell potential, 13 V.The precursor ion→product ion transitions m/z 265.3→232.0 for vorinostat and m/z 256.1→167.0 for the internal standard were monitored.Blood samples for pharmacokinetic measurements were taken on day 1(equivalent to a single-dose evaluation)and on day 22 for multiple-dosing evaluation. Results: The calibration curve was linear in the range of 1 to 1000 ng·mL^-1 with a correlation coefficient of 0.9996,and the limit of detection(LOD)of vorinostat in plasma was 0.1 ng·mL^-1.The intra- and inter-day precisions(RSD)were all less than 14%,and the accuracy was 87.6%-109.2%.The extracted recovery and the matrix effect were 93.6%-104.5% and 92.3%-93.5%,respectively.The single dose pharmacokinetics of four patients were evaluated,and the maximal concentration(C_max)and the mean apparent half life(t_1/2)were(402.5±50.2)ng·mL^-1 and(2.0±2.3)h,respectively.The C_max and t_1/2 for multiple-dosing were(800±247.2)ng·mL^-1 and(2.04±1.4)h,and the accumulation index was 1.00. Conclusion :The UPLC-MS/MS method was quick and sensitive with a total run time of 3 min using a simple protein precipitation,which can be used for clinical pharmacokinetic studies of vorinostat in human plasma.