LC-MS/MS法分析人血浆中西格列汀浓度及其应用研究

摘要：目的 建立一种测定人血浆中西格列汀浓度的液相色谱-串联质谱（LC-MS/MS）分析方法，并将该方法应用于西格列汀在人体内的药代动力学研究。方法 以西格列汀-d4为内标，血浆样品经CleanertPPT沉淀板沉淀后，通过Diamonsil C18色谱柱（100 mm×4.6 mm，5 μm）进行分离，使用甲醇-10 mmol/L甲酸铵水溶液（含10%甲醇，0.1%甲酸）作为流动相，进行梯度洗脱，流速为0.5 mL/min。通过电喷雾电离源（ESI），以多反应监测（MRM）模式进行正离子检测。从选择性、残留、线性范围与定量下限、精密度与准确度、基质效应和回收率、稳定性方面进行方法学验证。同时考察健康人口服西格列汀片100 mg后的主要药代动力学参数。结果 西格列汀、西格列汀-d4的MRM离子对分别为 m/z 408.0→235.2、m/z 412.1→239.0。人血浆中西格列汀在 0.5～1 000 μg/L 浓度范围内线性关系良好（R2>0.99），定量下限为0.5 μg/L；定量下限和质控样品的批内、批间精密度（RSD）在0.83%~12.80%之间，准确度（RE）在±10.0%以内。健康人口服西格列汀片100 mg后主要药代动力学参数：达峰时间（Tmax）、达峰浓度（Cmax）、、生物利用度（AUC）、半衰期（T1/2）分别为（2.44±1.29）h、（375±138）μg/L、（2 915±585）h·μg/L、（11.10±2.41）h。结论 本LC-MS/MS分析方法敏感度高且样品处理方法简单快速，满足生物分析的法规要求，可应用于人体内西格列汀的药代动力学研究。

Analytical methods and application of LC-MS/MS for quantification ofsitagliptin in human plasma

Abstract: Objective To develop a liquid chromatography-tandem mass spectrometry (LC-MS / MS) method forquantification of sitagliptin in human plasma, and to apply it for the pharmacokinetics study of sitagliptin in human.Methods The plasma sample was precipitated by CleanertPPT and sitagliptin-d4 was used as the internal standard (IS).Chromatographic separation was performed on a Diamonsil C18 column (100 mm×4.6 mm, 5 μm). Mobile phase consisted ofmethanol and a mixture of 10 mmol/L ammonium formate-methanol-formic acid (90∶10∶0.1) at a flow rate of 0.5 mL/min.The validation was performed in terms of selectivity, residue, linear range and lower limit of quantitation, precision andaccuracy, matrix effects and recovery, and stability. The main pharmacokinetic parameters (Tmax, Cmax, AUC0-t and T1/2) wereobserved in healthy people after oral administration of sitagliptin tablets. Results Sitagliptin and sitagliptin-d4 (IS) wereionized with an ESI source and operated in positive ion mode. The detected ions were m/z 408.0→235.2 (sitagliptin) and m/z412.1→239.0 (sitagliptin-d4). This validated LC-MS/MS method yielded a good linearity over the range of 0.5-1 000 μg/L(R2=0.99), and the lower limit of quantitation (LLOQ) was 0.5 μg/L. The intra and inter-assay precisions (RSD) were withinthe range of 0.83%~12.80%, and the accuracy (RE) was less than ± 10.0%. The pharmacokinetic parameters, Tmax, Cmax,AUC0-t and T1/2 were (2.44±1.29) h, (375±138) μg/L, (2 915±585) h·μg/L and (11.10±2.41) h. Conclusion This LC-MS /MS method is simple, sensitive, rapid and suitable for pharmacokinetics study of sitagliptin in human being.