An ultra-performance liquid-chromatography mass-spectrometry (UPLC-MS/MS) method for simultaneous quantitation of metronidazole and 2-hydroxymetronidazole in human plasma was developed and validated. Metronidazole and 2-hydroxymetronidazole were extracted from a small volume of human plasma (10 μL) by hydrophilic lipophilic balanced solid phase extraction on 96-well μ-elution plates. Chromatographic separation of analytes was achieved on an Acquity UPLC BEH C18 column (1.7 μm, 2.1 × 100 mm) using gradient elution with a blend of 0.1% formic acid in water and 0.1% formic acid in methanol at a flow rate of 0.25 mL/min. Mass spectrometric detection was achieved using multiple reaction monitoring (MRM) in positive-ion electrospray-ionization (ESI) mode. Ion transitions were optimized at m/z 171.85->127.9 for metronidazole and m/z 187.9->125.9 for 2-hydroxymetronidazole. The assay was linear for both analytes over the concentration range of 0.1–300 μM; intra- and inter-assay precisions and accuracies were <13%. Recoveries for metronidazole and 2-hydroxymetronidazole ranged from 88 to 99% and 78 to 86%, respectively. Matrix effects for metronidazole and 2-hydroxymetronidazole in plasma ranged from 102 to 105% and 99 to 106%, respectively. The method was successfully applied to determine metronidazole and 2-hydroxymetronidazole plasma concentrations in a pharmacokinetic study conducted in adults administered an oral dose of 500 mg metronidazole. Pharmacokinetic parameters were comparable to previously reported values. By design, this method is amenable to high sample throughput and has the potential to be automated.

建立并验证了同时定量人体血浆中甲硝唑和2-羟基甲硝唑的超高效液相色谱质谱法（UPLC-MS / MS）。通过在96孔μ-洗脱板上进行亲水亲脂平衡固相萃取，从少量人体血浆（10μL）中提取甲硝唑和2-羟基甲硝唑。在Acquity UPLC BEH C18色谱柱（1.7μm，2.1×100 mm）上进行色谱分离，使用0.1％甲酸水溶液和0.1％甲酸甲醇溶液的混合液以0.25 mL / mL的流速进行梯度洗脱分钟 使用正离子电喷雾电离（ESI）模式下的多反应监测（MRM）实现了质谱检测。离子跃迁以m / z优化甲硝唑和m / z为171.85-> 127.92-羟基甲硝唑为187.9-> 125.9。在0.1–300μM的浓度范围内，两种分析物的测定均为线性；批内和批间精密度和准确性均低于13％。甲硝唑和2-羟基甲硝唑的回收率分别为88％至99％和78％至86％。血浆中甲硝唑和2-羟基甲硝唑的基质效应分别为102％至105％和99％至106％。在成人口服500毫克甲硝唑的药代动力学研究中，该方法已成功应用于测定甲硝唑和2-羟基甲硝唑的血浆浓度。药代动力学参数与先前报道的值相当。通过设计，该方法适合高样品通量，并且具有自动化的潜力。