超高效液相-串联质谱法同时检测血液和尿液中15种降血糖药

Simultaneously Determining by UPLC-MS into 15 Hypoglycemic Drugs in Blood and Urine

目的建立超高效液相色谱-串联质谱法同时测定血液和尿液中15种降糖药物(二甲双胍、维达列汀、苯乙双胍、罗格列酮、西他列汀、吡格列酮、甲苯磺丁脲、妥拉磺脲、格列吡嗪、格列齐特、格列本脲、格列美脲、那格列奈、格列喹酮、瑞格列奈)的分析方法。方法分别用乙腈对血液和尿液样品沉淀蛋白过膜后,采用超高效液相色谱-三重四级杆质谱联用仪测定。色谱柱为Agilent Eclipse Plus C18色谱柱(2.1 mm×100 mm×3.5μm),有机相为0.1%甲酸甲醇溶液,水相为5 mmol/L乙酸铵和甲酸水溶液,进行梯度洗脱;流速为0.3 mL/min,采用电喷雾正离子模式,选择反应监测方式(SRM)检测。结果15种药物在血液和尿液中在低浓度(0.5~10 ng/mL)至高浓度(200~1000 ng/mL)范围内线性关系良好,R^(2)均大于0.998。二甲双胍提取回收率为28.1%~42.9%,其余14种降糖药物提取回收率在60.8%~119.0%之间;二甲双胍在尿液中检出限为5 ng/mL,其余14种药物在血液和尿液中的检出限均低于2 ng/mL。15种药物在血液和尿液三种添加浓度中日内和日间精密度相对标准偏差RSD均不高于8.9%(n=6)。结论该方法检测灵敏度高、速度快、专属性高、重复性好,可用于生物样品血液和尿液中15种降血糖药物的同时检测。

Objective To establish an ultra-performance liquid chromatography-tandem mass spectrometry(UPLCMS)method for determination of 15 hypoglycemic drugs(metformin,vildagliptin,phenformin,rosiglitazone,sitagliptin,pioglitazone,tolbutamide,tolazamide,glipizide,gliclazide,glyburide,glimepiride,nateglinide,gliquidone,repaglinide)in blood and/or urine.Methods The blank control samples of both blood and urine were spiked with the selected drugs,then purifi ed with acetonitrile to precipitate the proteins,successively having subjected to the analysis of UPLC-triple quadrupole mass spectrometry(UPLC-MS/MS)under the conditions:LC operation was performed through an Agilent Eclipse Plus C18 column(2.1mm×100mm×3.5μm)that was eluted with the mobile phases consisting of water containing 0.1%formic acid plus 5mmol/L ammonium acetate(A)and methanol containing 0.1%formic acid(B)at the fl ow rate of 0.3mL/min;MS operation was run at the selected reaction monitoring(SRM)in electrospray positive ion mode.Results An adequate linearity was afforded at concentrations from the lower limit of quantifi cation(0.5-10ng/mL)to the upper limit of quantifi cation(200-1000 ng/mL),with the coeffi cients of determination(R2)being higher than 0.998.The extraction recoveries of 14 hypoglycemic drugs ranged from 60.8%to 119.0%,with exception of the metformin’s showing from 28.1%to 42.9%.The detection limits of 14 drugs were lower than 2ng/mL in blood and urine,leaving the metformin’s being 5ng/mL in urine.The 15 drugs unveiled their relative standard deviations(RSD)relating to intra-/inter-day precisions were less than 8.9%with the addition of three concentrations into the blood and urine(n=6).Conclusions The method is of high sensitivity,high speed,high specifi city and good reproducibility,capable of detecting the selected 15 hypoglycemic drugs in biological samples of blood and urine.