Rapid quantification of HIV protease inhibitors in human plasma by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry
Rapid quantification of HIV protease inhibitors in human plasma by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry
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DOI：
10.1002/jms.425
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发表时间：
2003-02-01
影响因子：
2.3
通讯作者：
Beijnen, JH
中科院分区：
文献类型：
--
作者：
Crommentuyn, KML;Rosing, H;Beijnen, JH
HIV蛋白酶抑制剂是重要的抗逆转录病毒药物，可大大降低与HIV-1感染相关的发病率和死亡率。最近的数据显示了蛋白酶抑制剂的血浆浓度与临床反应之间的关系，这使得治疗药物监测变得有价值。我们开发并验证了一种检测方法，使用液相色谱结合电喷雾串联质谱 (LC/MS/MS)，对血浆中六种许可的蛋白酶抑制剂（安普那韦、茚地那韦、洛匹那韦、奈非那韦、利托那韦和沙奎那韦）以及药理活性奈非那韦代谢物 M8 进行常规定量。样品预处理包括仅使用甲醇和乙腈混合物进行蛋白质沉淀。 100μl血浆。在 Inertsil ODS3 柱（50 x 2.0 nun i.d.，粒径 5 μm）上进行色谱分离，使用乙酸盐缓冲液（pH 5）和甲醇进行快速逐步梯度，流速为 0.5 ml min(-1)。分析运行时间为 5.5 分钟。使用 96 孔板自动进样器可以批量处理多达 150 个患者样本。三重四极杆质谱仪在正离子模式下操作，并使用多反应监测进行药物定量。该方法的验证浓度范围为：茚地那韦和沙奎那韦为 0.01-10 mug ml(-1)，安普那韦为 0.1-10 mug ml(-1)，奈非那韦和利托那韦为 0.05-10 mug ml(-1)，洛匹那韦为 0.1-20 mug ml(-1)，洛匹那韦为 0.01-5 mug ml(-1)。 M8。沙奎那韦-d(5) 和茚地那韦-d(6) 用作内标。变异系数始终为
HIV protease inhibitors are important antiretroviral drugs which have substantially reduced the morbidity and mortality associated with HIV-1 infection. Recent data have shown relationships between plasma concentrations of the protease inhibitors and clinical response, which makes therapeutic drug monitoring valuable. We have developed and validated an assay, using liquid chromatography coupled with electrospray tandem mass spectrometry (LC/MS/MS), for the routine quantification of the six licensed protease inhibitors (amprenavir, indinavir, lopinavir, nelfinavir, ritonavir and saquinavir) and the pharmacologically active nelfinavir metabolite M8 in plasma.The sample pretreatment consisted of protein precipitation with a mixture of methanol and acetronitrile using only 100 mul of plasma. Chromatographic separation was performed on an Inertsil ODS3 column (50 x 2.0 nun i.d., particle size 5 mum), with a quick stepwise gradient using an acetate buffer (pH 5) and methanol, at a flow rate of 0.5 ml min(-1). The analytical run time was 5.5 min. The use of a 96-well plate autosampler allowed batch sizes up to 150 patient samples. The triple-quadrupole mass spectrometer was operated in the positive ion mode and multiple reaction monitoring was used for drug quantification. The method was validated over the concentration ranges 0.01-10 mug ml(-1) for indinavir and saquinavir, 0.1-10 mug ml(-1) for amprenavir, 0.05-10 mug ml(-1) for nelfinavir and ritonavir, 0.1-20 mug ml(-1) for lopinavir and 0.01-5 mug ml(-1) for M8. Saquinavir-d(5) and indinavir-d(6) were used as internal standards. The coefficients of variation were always