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目的建立LC-MS/MS法测定人血浆中阿托伐他汀及其主要活性代谢物2-羟基阿托伐他汀和4-羟基阿托伐他汀的浓度。方法分别以阿托伐他汀-d5,2-羟基阿托伐他汀-d5和4-羟基阿托伐他汀-d5为内标,加入5%甲酸水溶液,用甲基叔丁基醚进行液液萃取后,进样分析。色谱柱为CAPCELL PAK C_(18)(2 mm×100 mm,TYPE:MGⅡ,5μm),流动相为乙腈-5 mmol·L~(-1)乙酸胺水溶液-甲酸=46:54:0.143,流速为0.40 mL·min~(-1)。电喷雾离子源:正离子多反应监测扫描分析。阿托伐他汀的离子对为m/z 559.3→m/z 440.2,2/4-羟基阿托伐他汀的离子对为m/z 559.3→m/z 440.2。考察其专属性、标准曲线与定量下限、准确度和精密度、提取回收率和基质效应、残留效应和稳定性。结果血浆中阿托伐他汀的标准曲线方程为y=4.35×10~(-2)x+3.43×10~(-4)(r=0.9993),在0.05~100 ng·mL~(-1)线性关系良好,定量下限为0.05ng·mL~(-1);2-羟基阿托伐他汀的标准曲线方程为y=1.03×10~(-1)x+6.93×10~(-4)(r=0.998 7),在0.05~50 ng·mL~(-1)线性关系良好,定量下限为0.05ng·mL~(-1);4-阿托伐他汀的标准曲线方程为y=5.35×10~(-1)x-2.26×10~(-3)(r=0.9984),在0.05~5.0 ng·mL~(-1)线性关系良好,定量下限为0.05ng·mL~(-1)。阿托伐他汀、2-羟基阿托伐他汀和4-羟基阿托伐他汀的日内、日间的相对标准偏差(RSD)均小于15%;提取回收率为61.64%~90.29%;基质效应为88.74%~105.62%。结论 LC-MS/MS法快速、灵敏、准确、选择性强、重复性好,适用于人血浆中阿托伐他汀及其活性代谢产物的浓度测定,可以满足临床药物浓度监测以及药代动力学研究的需要。
Objective To establish a LC-MS / MS method for the determination of atorvastatin and its major active metabolites, 2-hydroxyatoformin and 4-hydroxyatulinism in human plasma. Methods Atorvastatin-d5,2-hydroxy-atorvastatin-d5 and 4-hydroxy-atorvastatin-d5 were respectively used as internal standard, 5% aqueous solution of formic acid and liquid-liquid extraction with methyl tert-butyl ether After injection analysis. The column was CAPCELL PAK C 18 (2 mm × 100 mm, TYPE: MG Ⅱ, 5 μm). The mobile phase was acetonitrile-5 mmol·L -1 aqueous acetic acid solution-formic acid = 46:54: 0.143, 0.40 mL · min -1. Electrospray ionization source: Positive ion multiple reaction monitoring scanning analysis The ion pair of atorvastatin was m / z 559.3 → m / z 440.2, and the ion pair of 2/4-hydroxyatoform was m / z 559.3 → m / z 440.2. Investigate its specificity, standard curve and lower limit of quantification, accuracy and precision, extraction recovery and matrix effects, residual effects and stability. Results The standard curve equation of atorvastatin in plasma was y = 4.35 × 10 -2 x + 3.43 × 10 -4 (r = 0.9993) The calibration curve of 2-hydroxy atorvastatin was y = 1.03 × 10 ~ (-1) x + 6.93 × 10 ~ (-4) ( r = 0.998 7). The linear range was 0.05 ~ 50 ng · mL -1 and the limit of quantification was 0.05 ng · mL -1. The standard curve of 4-atorvastatin was y = 5.35 × The linear range was between 0.05 and 5.0 ng · mL -1 with a lower limit of quantitation of 0.05 ng · mL -1 in the range of 10 -1 to 2.26 × 10 -3 (r = 0.9984) . The relative standard deviations (RSDs) of atorvastatin, 2-hydroxy atorvastatin and 4-hydroxy atorvastatin were both less than 15% and the recoveries were 61.64% ~ 90.29%. The matrix effects were 88.74% ~ 105.62%. Conclusion The LC-MS / MS method is rapid, sensitive, accurate, selective and reproducible. It is suitable for the determination of atorvastatin and its active metabolites in human plasma. It can meet the needs of clinical drug concentration monitoring and pharmacokinetics The need of research