高效液相色谱串联质谱法同时测定人血浆中厄贝沙坦和氨氯地平浓度

Determination of irbesartan and amlodipine in human plasma by liquid chromatography tandem mass spectrometry

目的建立快速测定人血浆中厄贝沙坦和氨氯地平浓度的液相色谱-质谱联用法。方法蛋白沉淀法,色谱柱为ACQUITY UPLC BEH C18(2.1 mm×50 mm,1.7μm),柱温为40℃,梯度洗脱,流动相为乙腈(B)-0.1%甲酸水溶液(A),流速为0.40 mL·min^(-1),进样量为5μL,分析时间为3.00 min。ESI离子源,多反应监测正离子模式,定量离子对分别为m/z 429.2→m/z 195.1(厄贝沙坦)、m/z 409.3→m/z 238.1(氨氯地平)、m/z 433.2→m/z 195.1(厄贝沙坦-d4)和m/z 413.3→m/z 238.1(氨氯地平-d4)。考察方法的专属性、标准曲线和定量下限、精密度、基质效应、提取回收率和稳定性。结果人血浆中厄贝沙坦的标曲线性范围为10~5000 ng·mL^(-1),线性回归方程为Y=7.03×10^(-4)x+7.35×10^(-4)(r=0.998),批内、批间精密度在1.2%~9.4%,相对偏差在-6.1%~4.0%,低、中、高质量浓度内标归一化基质效应因子的精密度为0.4%~6.6%,提取回收率为102.0%~105.2%。人血浆中氨氯地平的标曲线性范围为0.05~10 ng·mL^(-1),线性回归方程为Y=4.86×10-2x+6.66×10^(-4)(r=0.999),批内、批间精密度在1.1%~10.1%,相对偏差在-10.0%~8.3%,低、中、高质量浓度内标归一化基质效应因子的精密度为0.7%~11.6%,提取回收率为98.4%~106.6%。稳定性均良好。结论方法简单高效且灵敏度高,适用于人血浆中厄贝沙坦和氨氯地平浓度的检测及药代动力学和生物等效性等研究。

Objective To develop a rapid LC-MS/MS method to determinate the concentration of irbesartan and amlodipine in human plasma.MethodsSamples were pre-treated using a protein precipitation method and separated by ACQUITY UPLC BEH C18(2.1mm×50 mm,1.7μm)column in gradient elution model with a mobile phase of acetonitrile(B)-0.1%formic acid in water(A)during 3 min.The column temperature was 40℃and the flow rate was 0.40m L·min^(-1).Detection of those two analytes was achived in positive ion ESI in MRM mode.The MS/MS ion transitions monitored were m/z429.2→m/z 195.1(irbesartan),m/z 409.3→m/z 238.1(amlodipine),m/z 433.2→m/z 195.1(irbesartan-d4)and m/z 413.3→m/z 238.1(amlodipine-d4).The specificity,standard curves and lower limit of quantitation(LLOQ),precision,matrix effect,recovery and stability were investigated.Results For irbesartan,the linearity range was 10-5000 ng·m L-1and the standard curve was Y=7.03×10^(-4)x+7.35×10^(-4)(r=0.998).The intra-and inter-precision were 1.2%-9.4%,while the relative deviation was-6.1%-4.0%.The matrix effect normalized by internal standard for low middle high QCs was 0.4%-6.6%,and the recovery was 102.0%-105.2%.For amlodipine,the linearity range was0.05-10 ng·m L-1and the standard curve was Y=4.86×10-2x+6.66×10^(-4)(r=0.999).The intra-and inter-precision were 1.1%-10.1%,while the relative deviation was-10.0%-8.3%.The matrix effect normalized by internal standard for low middle high QCs was 0.7%-11.6%,and the recovery was 98.4%-106.6%.Both of those two analytes’stability were good.Conclusion This LC-MS/MS method was simple as well as sensitive,could be used for pharmacokinetics and bioequivalence studies.