亲水作用色谱串联质谱法测定人血浆中的拉米夫定

Determination of lamivudine in human plasma by hydrophilic interaction chromatography (HILIC) tandem mass spectrometry

目的：建立简便、快速的亲水作用色谱串联质谱法（HILIC-MS/MS）测定人血浆中的拉米夫定。方法：以13C1,15N2（拉米夫定为内标,血浆样品经乙腈沉淀蛋白后,采用Luna HILIC（100 mm×3.0 mm,3μm）柱分离。流动相为乙腈-5 mmol·L-1醋酸铵-甲酸（95∶5∶0.01,v/v/v）,进样体积2μL,样品分析时间3 min。采用ESI源正离子模式、多反应监测（MRM）,用于定量分析的离子反应分别为m/z230（112（拉米夫定）和m/z233（115（内标）。结果：测定人血浆中拉米夫定的线性范围为8.0～2000 ng·mL-1,定量下限为8.0 ng·mL-1,日内、日间精密度（RSD）均小于9.0%,准确度（RE）在-7.1%~2.7%之间。本法成功应用于健康受试者口服2种拉米夫定片后的生物等效性研究。结论：采用稳定同位素内标的HILIC-MS/MS法更为简便、快捷和准确,适用于人血浆样品中拉米夫定的测定。

Objective：To develop and validate a hydrophilic interaction chromatography-tandem mass spectrometric（HILIC-MS/MS） method for the determination of lamivudine in human plasma.Method：The plasma sample was protein precipitated with acetonitrile using the stable labelled 13C1,15N2-lamivudine as the internal standard.After sample preparation,the analyte and the IS were separated on a Luna HILIC（100 mm×3.0 mm,3 μm）column using 2 μL injection volume with a run time of 3.0 min.The mobile phase consisited of acetonitrile-5 mmol·L-1 ammonium acetate-formic acid（95∶ 5∶ 0.01,v/v/v）.Detection was performed by tandem mass spectrometry using an electrosprasy source（ESI） in the positive ion mode,operating in the multiple reaction monitoring（MRM） of the transitions of m/z 230→112 and m/z 233→115 for lamivudine and the IS,respectively.Results：The linear calibration curves were obtained in the concentration range of 8.0-2000 ng·mL-1.The lower limit of quantification was 8.0 ng·mL-1.The intra-day and inter-day relative standard deviation over the entire concentration range was less than 9.0%.The accuracy was in the range of-7.1% to 2.7% in terms of relative error.The method was applied to evaluate the bioequivalence of two lamivudine tablet formulations in healthy volunteers.Conclusion：This method is simple,selective,rapid and suitable for the determination of lamivudine in human plasma.