超高效液相色谱串联质谱法同时测定人血清中利奈唑胺及其代谢产物PNU-142300的浓度

Simultaneous determination of linezolid and its metabolite PNU-142300 in human serum by ultra-performance liquid chromatography-tandem mass spectrometry

目的建立超高效液相色谱串联质谱法(UPLC-MS/MS)同时测定血清中利奈唑胺及羟乙基甘氨酸代谢产物(PNU-142300)的浓度,并用于重症肺炎患者血药浓度监测。方法以d3-利奈唑胺为内标,血清样品采用乙腈沉淀蛋白,色谱柱Acquity UPLC BEH C18 column色谱柱(50.0 mm×2.1 mm,1.7μm),流动相:0.1%甲酸水溶液和0.1%甲酸乙腈溶液,梯度洗脱,流速:0.4 mL·min^(-1),柱温:40℃;进样量:5μL。采用电喷雾电离源,正离子多反应方式监测。考察该方法的专属性、标准曲线、精密度、准确度、提取回收率、基质效应和稳定性。将该分析方法应用于不同肾功能状态的重症肺炎患者利奈唑胺及PNU-142300含量测定。结果利奈唑胺及PNU-142300均在0.1~40.0μg·mL^(-1)的浓度范围内线性关系良好,准确度相对误差(RE)范围为-8.05%~8.33%,日内、日间精密度相对标准差(RSD)分别≤4.33%和≤7.73%。利奈唑胺、PNU-142300的提取回收率在94.37%~102.92%范围,不受基质效应的影响,稳定性良好。肾功能不全组利奈唑胺及PNU-142300的中位谷浓度分布是肾功能正常组的2.15和3.89倍。结论本方法快速、简单、准确,适用于不同肾功能状态患者血清中利奈唑胺及其代谢产物PNU-142300浓度的测定。

Objective To establish a validated ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) assay for the simultaneous determination of linezolid and its metabolite PNU-142300 in human plasma,which may be further applied for the therapeutic drug monitoring in patients with severe pneumonia.Methods After protein precipitation with acetonitrile containing an internal standard(d3-linezolid),chromatographic separation of the analytes was accomplished using a Acquity UPLC BEH C18 column under gradient elution,where the mobile phase consisted of water with 0.1% formic acid and acetonitrile with 0.1% formic acid at a flow rate of 0.4 mL·min^(-1),a temperature of 40 ℃,and a 5 μL sample injection.The mass spectrometer was equipped with an electrospray ion source using positive multiple reaction monitoring(MRM) mode.The specificity,standard curve,precision,accuracy,extraction recovery,matrix effect and stability were investigated.The assay was also employed to determine the concentrations of linezolid and PNU-142300 in patients with severe pneumonia and different categories of renal function.Results Standard curves were linear at 0.1 to 40.0 μg · mL^(-1) for linezolid and PNU-142300.The accuracy ranged from-8.05% to 8.33% and the intra-day and inter-day precisions were below than 4.33% and7.73%,respectively.Extraction recoveries of the analytes were within the range of 94.37%-102.92% and no matrix effect was observed.The analytes during the assay and storage in serum samples were observed to be stable.Compared with patients with normal renal function,median trough concentrations of linezolid and PNU-142300 in patients with renal impairment were 2.15-fold and 3.89-fold higher,respectively.Conclusion This assay is rapid,accurate,simple,and it is suitable for the determination of linezolid,PNU-142300 in serum samples obtained from patients with different kidney function.