LC-MS/MS同时测定人血浆中洛匹那韦和利托那韦浓度

Simultaneous Determination of Lopinavir and Ritonavir in Human Plasma by LC-MS/MS

关键词:  洛匹那韦  利托那韦  血药浓度监测  液相色谱-质谱联用

Key words:  lopinavir  ritonavir  therapeutic drug monitoring  LC-MS/MS

摘要:

目的 建立一种快速、灵敏并同时测定人体血浆中洛匹那韦(LPV)和利托那韦(RTV)浓度的液相色谱-质谱联用检测方法。方法 采用Agilent Eclipse Plus C₁₈柱(4.6 mm×150 mm，3.5 μm)，流动相为乙腈-水(含0.005 mol·L^-1 甲酸铵，0.1%甲酸)(90∶10)；流速：0.5 mL·min^-1，柱温：40 ℃，以乙酸乙酯为提取剂。采用选择反应监测(SRM)LPV(m/z 629.5→155.1)、RTV(m/z 721.4→296.1)和内标茚地那韦(IDV)(m/z 614.5→465.4)进行测定。结果 LPV高(10 000 μg·L^-1)、中(1 000 μg·L^-1)、低(40 μg·L^-1)3个浓度的平均方法回收率RSD均<15%；线性范围为：20~20 000 μg·L^-1，回归方程为Y=1.669 9X-0.001 3，r=0.998 4(n=7)，定量下限为20 μg·L^-1。RTV高(2 500 μg·L^-1)、中(250 μg·L^-1)、低(10 μg·L^-1)3个浓度的平均方法回收率RSD均<15%；线性范围为5~5 000 μg·L^-1，回归方程为Y=1.723 7X-3.274 8×10^-4，r=0.998 7(n=7)，分析方法的定量下限为5 μg·L^-1。结论 该方法灵敏、准确、简单、快速，可用于LPV和RTV同时应用时两者的临床血药浓度监测和药动学研究。

Abstract:

OBJECTIVE To set up a rapid and sensitive LC-MS/MS method for simultaneous quantitative determination of lopinavir(LPV) and ritonavir(RTV) in human plasma. METHODS LPV and RTV were extracted with ethyl acetate. The residues were analyzed with an Agilent Eclipse Plus CM₁₈ column(4.6 mm×150 mm, 3.5 μm) with the mobile phase consisted of acetonitrile with 0.005 mol·L^-1 ammonium formate(0.1% formic acid)=90∶10, with a flow rate of 0.5 mL·min^-1, temperature of 40 ℃. Selected reaction monitoring(SRM) using the precursor to product ion combinations of m/z 629.5→155.1, 721.4→296.1 and m/z 614.5→465.4 was performed to detect LPV, RTV and the internal standard(indinavir, IDV), respectively. RESULTS The average recovery rate RSD for LPV of three levels of concentration of high (10 000 μg·L^-1), medium (1 000 μg·L^-1) and low (40 μg·L^-1), were less than 15%. The calibration curves for LPV was Y=1.669 9X-0.001 3, r=0.998 4, over the range of 20-20 000 μg·L^-1. The limits of quantitation was 20 μg·L^-1. Meanwhile, the average recovery rate RSD for RTV of three levels of concentration of high (2 500 μg·L^-1), medium (250 μg·L^-1) and low (10 μg·L^-1), were also less than 15%. The calibration curves for RTV was Y=1.723 7X-3.274 8×10^-4，r=0.998 7(n=7), over the range of 5-5 000 μg·L^-1. The limits of quantitation was 5 μg·L^-1. CONCLUSION The method provides a sensitive, accurate, precise and reliable analytical procedure for the therapeutic drug monitoring of LPV and RTV simultaneously in clinic and phamacokinetic studies.