Department of Laboratory Medicine, Seoul National University Bundang Hospital, Gyeonggi-do, South Korea.
J Antimicrob Chemother. 2013 Sep;68(9):2066-73. doi: 10.1093/jac/dkt154. Epub 2013 May 8.
Therapeutic drug monitoring (TDM) of anti-tuberculosis (TB) drugs is beneficial for patients responding slowly to treatment and those with multidrug-resistant TB. We used ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to develop a rapid method for simultaneously measuring the blood concentrations of nine second-line anti-TB drugs: streptomycin, kanamycin, clarithromycin, cycloserine, moxifloxacin, levofloxacin, para-aminosalicylic acid, prothionamide and linezolid.
Serum samples were extracted with acidified methanol and neutralized with NaOH. A Waters Acquity HSS T3 column and gradients of ammonium formate and acetonitrile in 0.1% formic acid were used for UPLC separation. Drug concentrations were determined by multiple reaction monitoring in positive ion mode, and assay performance was evaluated. We applied this method to TDM, analysing random serum samples from 85 patients treated with second-line drugs.
Sample preparation using acidified methanol extraction followed by neutralization yielded good recovery and ionization efficiency, with chromatographic separation achieved within 3 min per sample. Within-run and between-run precisions were 1.7%-7.5% and 1.7%-12.4%, respectively, at concentrations representing low and high levels for the nine drugs. Lower limits of detection and quantification were 0.025-0.5 and 0.25-5.0 μg/mL, respectively. Linearity was acceptable at five concentrations for each drug. No ion suppression was observed at the retention time for most compounds, except for streptomycin, kanamycin and cycloserine, which were eluted close to the void volume of the column. In a limited pilot study, all quantifiable human samples had values within the validated assay ranges.
The performance of our MS/MS detection technique was generally acceptable. The method provided rapid, sensitive and reproducible quantification of nine second-line anti-TB drugs and should facilitate drug monitoring during treatment.
抗结核（TB）药物的治疗药物监测（TDM）有益于治疗反应缓慢的患者和耐多药结核患者。我们使用超高效液相色谱-串联质谱（UPLC-MS/MS）建立了一种快速测定方案，可同时测量 9 种二线抗 TB 药物的血药浓度：链霉素、卡那霉素、克拉霉素、环丝氨酸、莫西沙星、左氧氟沙星、对氨基水杨酸、丙硫异烟胺和利奈唑胺。
血清样品用酸化甲醇提取，并用 NaOH 中和。采用 Waters Acquity HSS T3 柱和 0.1%甲酸中的甲酸铵和乙腈梯度洗脱进行 UPLC 分离。采用正离子模式下的多重反应监测法测定药物浓度，并评价分析性能。我们将该方法应用于 TDM，分析了 85 例二线药物治疗患者的随机血清样本。
使用酸化甲醇提取和中和的样品制备方法可获得良好的回收率和离子化效率，每个样品的色谱分离时间在 3 分钟内。9 种药物的低、高浓度时，批内和批间精密度分别为 1.7%-7.5%和 1.7%-12.4%。检测限和定量限分别为 0.025-0.5 和 0.25-5.0 μg/mL。每种药物的 5 个浓度点均具有可接受的线性。除链霉素、卡那霉素和环丝氨酸外，大多数化合物在保留时间处未观察到离子抑制，而这些化合物的洗脱时间接近色谱柱的空隙体积。在一项有限的初步研究中，所有可定量的人样本值均在验证后的测定范围内。
我们的 MS/MS 检测技术的性能总体上是可接受的。该方法可快速、灵敏和重现地定量 9 种二线抗 TB 药物，有助于治疗期间的药物监测。
