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,旨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0.35 mL·min-1;采用ACQUITY UPLC HSS T3 1.8 μm column (2.1 mm×100 mm,Waters公司)分离;采用电喷雾电离源,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48~6 000 ng·mL-1(r=0.998 8)、480~60 000 ng·mL-1(r=0.999 3)、120~15 000 ng·mL-1(r=0.999 5)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、1 220及1 570 ng·mL-1;PZA的质量浓度分别为104.2、6 273.34和3 185 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。
Abstract
OBJECTIVE To develop an UPLC-MS/MS method for simultaneous determination of 3 anti-tuberculosis drugs [isoniazid (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 T3 1.8 μm column (2.1 mm×100 mm,Waters Corporation) with the mobile phase of aqueous solution (containing 0.1% 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 48-6 000 ng·mL-1(r=0.998 8)、480-60 000 ng·mL-1(r=0.999 3)、120-15 000 ng·mL-1(r=0.999 5), respectively. The CV 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, 1 220 and 1 570 ng·mL-1, 104.2, 6 273.34 and 3 185 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.
关键词
异烟肼 /
吡嗪酰胺 /
左氧氟沙星 /
液质联用 /
痰液
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Key words
isoniazid /
levofloxacin /
aziazinone /
LC-MS/MS /
sputum
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中图分类号：
R917
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参考文献
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