LC-MS/MS法同时测定Beagle犬血浆中阿托伐他汀钙和盐酸吡格列酮的浓度

目的:建立LC-MS/MS方法测定Beagle犬血浆中阿托伐他汀钙和盐酸吡格列酮的浓度,并验证方法。方法: 采用蛋白沉淀法提取血浆中药物,以地西泮为内标;使用Shim-pack XR-ODS(2.0 mm×100 mm,2.2 μm)色谱柱,Shim-pack GVP-ODS(2.0 mm×5 mm,2.2 μm)保护柱,柱温40℃;流动相A为5 mmol·L^-1醋酸铵(pH 5.55),B为乙腈,采用梯度洗脱(0~ 0.6 min,10%B;0.6~1.2 min,10%→90%B;1.2~3.5 min,90%B;3.5~4.1 min,90%→10%B,5.5 min结束);流速为0.3 mL·min^-1;进样体积20 μL。质谱均采用电喷雾离子源(ESI),以多反应 离子监测(MRM)模式进行定量分析。结果: 阿托伐他汀钙、盐酸吡格列酮分别在0.05~50、0.5~1000 ng·mL^-1范围内有良好的线性关系,最低检测限分别为0.05、0.5 ng·mL^-1,日内、日间精密度RSD均<15%(n=5)。含药血浆室温放置24 h、处理后4℃放置24 h、反复冻融3次及-70℃下放置2个月稳定。结论: 本方法简便、准确,能用于Beagle犬血浆中阿托伐他汀钙、盐酸吡格列酮的药物浓度测定和药动学研究。

Objective: To establish an LC-MS/MS method to determine the concentrations of atorvastatin calcium and pioglitazone hydrochloride in Beagle dog plasma,and to verify the method. Methods: The drugs were extracted by precipitating protein,with diazepam as an internal standard.The Shim-pack XR-ODS(2.0 mm×100 mm,2.2 μm)column at 40℃ and the Shim-pack GVP-ODS(2.0 mm×5 mm,2.2 μm)pre-column were used.The mobile phase consisted of phase A(5 mmol·L^-1 ammonium acetate,pH 5.55)and phase B(acetonitrile),the phase B accounted for 10% of the total mobile phase ratein the first 0.6 min,then changed to 90% from 0.6 min to 1.2 min,and kept at 90% until 3.5 min,then returned to 10% from 3.5 min to 4.1 min.The analysis was stopped until 5.5 min,the total mobile phase rate was 0.3 mL·min^-1,injected volume was 20 μL.Electrospray ion source was applied in mass spectrometry,and quantitative analysis was undertaken in multiple reaction monitoring. Results: Atorvastatin calcium and pioglitazone hydrochloride had a good linear relation between 0.05-50 ng·mL^-1,0.5-1000 ng·mL^-1,respectively.The intra-and inter-day precision RSD were both less than 15%(n=5).Drugs in dog plasma were stable at room temperature for 24 h,and at 4℃ for 24 h after being extracted,even after freezing and thawing three times and stored at -70℃ for two months. Conclusion: The developed method is simple and accurate,which can be used to determine the concentration and the pharmacokinetics of atorvastatin calcium and pioglitazone hydrochloride in Beagle dogs.