With the increased cases of multidrug- or rifampicin-resistant tuberculosis and co-infection with HIV globally, it is difficult to achieve ideal clinical responses because of poor drug absorption and drug-drug interactions. Herein, a bioanalytical UPLC-MS/MS method was developed and validated to quantify five anti-TB agents in human plasma samples for detecting blood drug concentrations to improve therapeutic effects. To overcome the matrix effects, stable isotope labeled analogue of each analyte was used for internal standardization. A simple single-step protein precipitation by acetonitrile was employed for the sample preparation, then the analytes including rifampicin, rifabutin, pyrazinamid, ethambutol, isoniazid and their isotope labeled internal standards (ILISs) were implemented on an HILIC silica column with a gradient mode. The linear range for each analyte was covering the peak drug concentration (Cmax) in the 20 times diluted plasma samples. The coefficient of variation of intra- and inter-day precision was less than 17.0 %, and the accuracy ranged between 91.5 and 110.0 %. The extraction recoveries of all agents were ≥90.2 %, and the matrix effects with internal standard-normalization for all agents were 97.1-110.0 %. The optimal blood sampling time was designed basing on the results of stability validation. This UPLC-MS/MS method with a run time of 3.5 min was successfully applied to routine therapeutic monitoring of the five anti-TB agents in patient plasma.

中文翻译：

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通过UPLC-MS / MS对人血浆中的抗结核药异烟肼，乙胺丁醇，吡嗪酰胺，利福平和利福布丁进行快速，高度灵敏的定量。

随着耐多药或耐利福平的结核病病例增加以及全球范围内HIV合并感染，由于不良的药物吸收和药物与药物的相互作用，难以实现理想的临床反应。本文中，开发了一种生物分析UPLC-MS / MS方法，并经过验证可对人血浆样品中的五种抗结核药进行定量，以检测血液中的药物浓度以改善治疗效果。为了克服基质效应，将每种分析物的稳定同位素标记的类似物用于内部标准化。采用简单的一步法通过乙腈沉淀蛋白质进行样品制备，然后在HILIC硅胶色谱柱上以梯度模式对包括利福平，利福布汀，吡嗪酰胺，乙胺丁醇，异烟肼及其同位素标记内标（ILIS）的分析物进行分析。每个分析物的线性范围覆盖了20倍稀释血浆样品中的峰值药物浓度（Cmax）。日内和日间精度的变异系数小于17.0％，且精度在91.5至110.0％之间。所有试剂的提取回收率均≥90.2％，所有试剂经内标标准化的基质效应为97.1-110.0％。根据稳定性验证的结果设计了最佳采血时间。这种运行时间为3.5分钟的UPLC-MS / MS方法已成功应用于患者血浆中五种抗结核药物的常规治疗监测。准确度在91.5和110.0％之间。所有试剂的提取回收率均≥90.2％，所有试剂经内标标准化的基质效应为97.1-110.0％。根据稳定性验证的结果设计了最佳采血时间。这种运行时间为3.5分钟的UPLC-MS / MS方法已成功应用于患者血浆中五种抗结核药物的常规治疗监测。准确度在91.5和110.0％之间。所有试剂的提取回收率均≥90.2％，所有试剂经内标标准化的基质效应为97.1-110.0％。根据稳定性验证的结果设计了最佳采血时间。这种运行时间为3.5分钟的UPLC-MS / MS方法已成功应用于患者血浆中五种抗结核药物的常规治疗监测。