The aim of this study was to develop and validate a UHPLC–MS/MS assay to quantify cyclosporin (CYC), tacrolimus (TAC), sirolimus (SIR) and everolimus (EVE) in human whole blood for therapeutic drug monitoring. Analytes were extracted from 50 μL human whole blood by protein precipitation. The separation of the drugs was performed on an Acquity UPLC BEH C18 column. Analytes were eluted with a mobile phase consisting of 2 mM ammonium acetate with 0.1% formic acid (v/v) in deionised water and 2 mM ammonium acetate with 0.1% formic acid (v/v) in methanol at a flow rate of 300 μL/min in gradient elution. The method performance was evaluated by analysing patient blood samples and/or external quality control samples [proficiency testing (PT) scheme]. The method was linear from 23.75 to 1094.0, 1.3 to 42.4, 1.3 to 47.0 and 1.2–41.6 μg/mL for CYC, TAC, SIR and EVE, respectively. The within‐ and between‐assay reproducibility results were ˂ 11%. Results from PT and patient sample quantification were comparable to those obtained previously by an in‐house validated method using protein precipitation and liquid–liquid extraction. This method showed good analytical performance for quantifying CYC, TAC, SIR and EVE in whole blood over their respective calibration ranges.

中文翻译：

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通过UHPLC-MS / MS同时定量全血中环孢菌素，他克莫司，西罗莫司和依维莫司，用于治疗药物监测

这项研究的目的是开发和验证一种UHPLC-MS / MS测定方法，以定量监测全血中环孢菌素（CYC），他克莫司（TAC），西罗莫司（SIR）和依维莫司（EVE）来监测治疗药物。通过蛋白质沉淀从50μL人全血中提取分析物。在Acquity UPLC BEH C18色谱柱上进行药物分离。用流动相洗脱分析物，流动相由去离子水中的2 mM乙酸铵和0.1％甲酸（v / v）和2 mM乙酸铵和0.1％甲酸（v / v）的甲醇溶液组成，流速为300μL / min梯度洗脱。通过分析患者的血液样本和/或外部质量控制样本[能力验证（PT）方案]，评估了方法的性能。对于CYC，TAC，该方法的线性范围为23.75至1094.0、1.3至42.4、1.3至47.0和1.2–41.6μg/ mL。SIR和EVE分别。测定内和测定间重现性结果为˂11％。PT和患者样品定量的结果与以前使用蛋白质沉淀和液-液萃取通过内部验证方法获得的结果相当。该方法显示出良好的分析性能，可在各自的校准范围内定量全血中的CYC，TAC，SIR和EVE。