LC-MS/MS法测定人血浆中维格列汀浓度及其在药动学研究中的应用

Determination of vildagliptin in human plasma by LC-MS/MS and its application to a pharmacokinetic study

目的建立测定人血浆中维格列汀质量浓度的液相色谱-串联质谱(liquid chromatography tandem mass spectrometry,LC-MS/MS)联用方法,考察空腹条件下维格列汀片的人体药动学特征。方法血浆样品经蛋白沉淀法处理,采用稳定同位素标记^(13)C_(5)-^(15)N-维格列汀作为内标,采用Hypurity C_(18)(150 mm×2.1 mm,5μm)色谱柱分离,进行梯度洗脱,流动相为5 mmol·L^(-1)甲酸铵水溶液(A)-乙腈(B),流速为0.5 mL·min^(-1)。在正离子条件下选择多反应离子监测模式进行定量分析。将8名健康受试者采用随机双周期双交叉方式进行空腹试验,并通过WinNonlin 8.1软件计算药动学参数。结果人血浆中维格列汀在质量浓度1.11~534.00μg·L^(-1)内线性关系良好。批内和批间准确度偏差均在±15%以内,精密度相对标准偏差均小于15%。各基质内标归一化的基质因子相对标准偏差均小于5%,选择性、提取回收率、残留效应、稀释可靠性和稳定性均符合要求。结论所建立的方法符合生物样本分析要求,可用于人血浆中维格列汀质量浓度检测及其药动学研究。

Objective To investigate the pharmacokinetics of vildagliptin tablet under fasting conditions,a liquid chromatography tandem mass spectrometry(LC-MS/MS)for the determination of vildagliptin in human plasma was developed.Methods Plasma samples were pre-treated using protein precipitation with^(13)C_(5)-^(15)N-vildagliptin as an internal standard.The chromatographic separation was performed on a Hypurity C_(18)(150 mm×2.1 mm,5μm)column using the mobile phase containing 5 mmol·L^(-1)ammonium formate solution(A)and acetonitrile(B)at a constant flow rate of 0.5 mL·min^(-1)by gradient elution mode.The multiple reaction monitoring(MRM)mode was selected for quantitative analysis in positive ion mode.The test was performed in 8 healthy volunteers by a 2-period randomized crossover design.The plasma concentration of vildagliptin was determined by the validated analytical method.And the pharmacokinetic parameters of the analytes were evaluated using WinNonlin 8.1 software.Results The method showed the good linearity within the calibration range of 1.11-534.00μg·L^(-1)for vildagliptin in human plasma.The relative standard deviation(RSD)of intra-day/inter-day precision was within ±15%,and the relative error(RE)of accuracy was within 15% at each quality control level.The RSDs of internal standard normalized matrix factor values were below 5% for normal,hemolyzed and hyperlipidemic plasma.In addition,other validation results(selectivity,extraction recovery,carry-over,dilution integrity and stability)also met the corresponding criterion.Conclusion The established method is successfully applied in pharmacokinetic studies of vildagliptin.