反相高效液相色谱法测定硫唑嘌呤代谢产物6-巯基嘌呤血浆药物浓度

摘要

Objective To establish a reversed phase high -performance liquid chromatography(RP -HPLC)method to determine 6-mercaptopurine (6-MP)in human plasma,thereby providing the foundation for clinical blood concentration monitoring.Methods U-sing Hypersil ODS2 column (250 nm ×4.6 nm,5 μm),the mobile phase was acetonitrile -0.3% glacial acetic acid (3∶97),the flow rate was 0.8 mL · min -1 ,the detection wavelength was set at 323 nm,and plasma samples were extracted by 20 μL 15% perchloric acid.Results A linearity was observed within the rangefrom 0.02 mg ·L -1 to 2.00 mg ·L -1 ,the lowest detection limit was 0.02 mg ·L -1 ,the average method recovery rate of four concentrations were 85.71%,92.93%,103.53% and 96.02%,respectively,and the intra -day and inter -day precision are <15% and in line with the Chinese Pharmacopoeia biological sample analysis regulations.Con-clusions The experiment successfully established a simple,fast,and low cost 6-MP plasma concentration measuring method with high sensitivity and good stability.Particularly,the method needs less blood.It is also in line with the relevant provisions of the AZA blood drug concentration monitoring and pharmacokinetics study.%目的：建立反相高效液相色谱法（RP －HPLC）检测人血浆中硫唑嘌呤（AZA）代谢产物6-巯基嘌呤（6-MP）的浓度，为临床进行血药浓度监测提供基础。方法采用 Hypersil ODS2色谱柱（250 nm ×4．6 nm，5μm），流动相为乙腈-0．3％冰乙酸化（3∶97），流速为0．8 mL·min －1，检测波长323 nm，血浆样品经15％高氯酸去蛋白后加氢氧钠（NaOH）调节 pH 后20μL 进样分析。结果6-MP 血浆样品在0．02～2．00 mg·L －1范围内线性关系良好，最低检测下限（LLOQ）为0．02 mg·L －1；LLOQ、高浓度（1．5 mg·L －1）、中浓度（1 mg·L －1）和低浓度（0．05 mg·L －1）提取回收率分别为85．71％、92．93％、103．53％和96．02％，质控四个浓度的日内与日间精密度均＜15．0％，符合中国药典生物样本分析相关规定。结论该实验成功建立一种简单、快速、高灵敏度的6-MP 血浆浓度测定方法，符合 AZA 血药浓度监测和药代动力学研究的相关规定。