UPLC-MS/MS测定人痰液中异烟肼、吡嗪酰胺和左氧氟沙星的浓度

目的建立超高效液相色谱串联质谱法(UPLC-MS/MS)同时检测痰液中3种药物异烟肼(isoniazid,INH)、左氧氟沙星(levofloxacin,LFX)和吡嗪酰胺(pyrazinamide,PZA)浓度的方法,冒在为实现临床个体化用药提供帮助。方法痰液样品离心后经甲醇/乙腈蛋白沉淀法预处理后进样液质联用仪进行分析。实验采用的流动相A为含0.1%甲酸的水溶液,流动相B为含0.1%甲酸的乙腈;流速0.35mL·min^-1;采用ACQUITY UPLAC HSS T31.8μm column(2.1mm×100mm,Waters公司)分离;采用电喷零电离源,采用多反应监测(multiple reaction monitoring,MRM)模式对待测物进行正离子扫描检测。此外,本试验盲态收集5例患者的痰液,用该法进行定量分析。结果测得痰液中INH、PZA和LFX分别在48~6000 ng·mL^-1(r=0.9988)、480~60000 ng·mL^-1(r=0.9993)、120~15000 ng·mL^-1(r=0.9995)内表现出良好的线性关系。INH、PZA和LFX 3种药物的日内和日间精密度均低于15%,且3种药物的提取回收率均处于97.21%~107.80%之间。7例受试者体内均检测到药物INH,质量浓度分别为44.74、120.1、301.5、481、595.5、1220及1570 ng·mL^-1;PZA的质量浓度分别为104.2、6273.34和3185 ng·mL^-1;1例受试者检出LFX,质量浓度为199.86 ng·mL^-1。结论本试验建立了痰液中INH、PZA及LFX的检测方法,具有灵敏度高、测定结果准确、快速等诸多优点,可以应用于临床治疗药物的监测。

Simultaneously Quantifying Isoniazid、Pyrazinamide and Levofloxacin in Human Sputum by HPLC-MS/MS

OBJECTIVE To develop an UPLC-MS/MS method for simultaneous determination of 3 anti-tuberculosis drugsisonia-zid(INH)、pyrazinamide(PZA)and levofloxacin(LFX)]in sputum by HPLC-MS/MS.METHODS The drugs(INH,PZA、and LFX)were extracted by methanol/acetonitrile(V-V=1:1)、All of them separated on ACQUITY UPLC HSS T31.8μm column(2.1 mm×100 mm,Waters Corporation)with the mobile phase of aqueous solution(containg O.1%formic acid)-acetonitrile(containing 0.1%formic acid)at a flow rate of 0.35 mL·min^-1.Multiple reaction monitoring(MRM)mode was performed combined with electrospray ionization source operating in the positive ionization mode.Furthermore,it was blindly collected 7 sputum samples,and analyzed the drug concentration by the built method in this work.RESULTS The liner calibration curves of INH、PZA、LFX were obtained in the concentration range of 48-6000 ng·mL^-1(r=0.9988)、480-60000 ng·mL^-1(r=0.9993)^120-15000 ng·mL^-1(r=0.9995),respectively.The CY of intra-and inter-day precision was less than 15%and the absolute recovery was 97.21%-107.80%.In the 7 clinical samples,the concentration of INH is 44.74,120.1,301.5,481,595.5,1220 and 1570 ng·mL^-1,104.2,6273.34 and 3185 ng·mL^-1 for PZA,and 199.86 for LFX,respectively.CONCLUSION In this work,it is built a LC-MS method to simultaneously quantify INH,PZA and LFX,which is simple,sensitive and accurate.This method is successfully applied to analyze the clinical samples.