HPLC-MS/MS法测定血液病患者血浆中伊布替尼血药浓度的方法学研究及临床应用

Methodological Study and Clinical Application on the Determination of the Plasma Concentration of Ibrutinib in Plasma of Patients with Hematological Diseases by HPLC-MS/MS

目的:建立测定血液病患者血浆中伊布替尼血药浓度的HPLC-MS/MS法,并将其应用于临床治疗药物监测。方法:以伊布替尼-d 5为内标,血浆样本经含内标的甲醇沉淀蛋白处理。HPLC色谱柱为Ultimate XB-C18(50 mm×4.6 mm,5μm),柱温60℃,流动相为含0.1%甲酸+2 mmol/L乙酸铵的水溶液与含0.1%甲酸的甲醇溶液,流速0.8 ml/min,梯度洗脱。质谱检测方式为电喷雾正离子(ESI+)模式,多离子反应监测(MRM)扫描,同时监测伊布替尼m/z 441.3~138.1(定量离子),伊布替尼m/z 441.2~304.1(定性离子);伊布替尼-d 5 m/z 446.2~138.1。以伊布替尼和伊布替尼-d 5峰面积比为定量依据,计算血浆中伊布替尼浓度,并对其性能进行考察。结果:伊布替尼在10~2000 ng/ml范围内线性较好(r^(2)=0.9997),最低检测限为1.65 ng/ml;批间、批内RSD均小于10%;回收率为101.31%~102.54%。5例临床血液病患者伊布替尼血药浓度检测结果范围为3.07~254 ng/ml,在1.65~2000 ng/ml之间。结论:本方法可简便、准确、快速地检测伊布替尼浓度,适用于伊布替尼血药浓度的日常监测。

Objective:A HPLC-MS/MS method for the determination of the plasma concentration of ibrutinib in plasma of patients with hematological diseases was established and applied to clinical therapeutic drug monitoring.Methods:With ibutinib-d 5 as internal standard,plasma samples were treated with methanol precipitated protein containing internal standard.The HPLC column was Uitimate XB-C18(50 mm×4.6 mm,5μm)and the column temperature was 60℃.The mobile phase was aqueous solution containing 0.1%formic acid+2 mmol/L ammonium acetate and methanol solution containing 0.1%formic acid.The flow rate was 0.8 ml/min and gradient elution was adopted.The mass spectrometric detection method was electrospray positive ion(ESI+)mode,multiion reaction monitoring(MRM)scanning,and simultaneous monitoring of ibrutinib m/z 441.3~138.1(quantitative ions),ibrutinib m/z 441.2~304.1(qualitative ions),ibrutinib-d 5 m/z 446.2~138.1.The peak area ratio of ibrutinib to ibrutinib-d 5 was used as the quantitative basis.The concentration of ibrutinib in plasma was calculated and its properties were investigated.Results:The linear of ibrutinib was good in the range of 10~2000 ng/ml(r^(2)=0.9997)and the limit of detection was 1.65 ng/ml.The RSD between batches and within batches were both less than 10%.The recovery of ibrutinib was 101.31%~102.54%.The plasma concentration of ibrutinib in 5 clinical patients with hematologic diseases ranged from 3.07 ng/ml to 254 ng/ml,all of which ranged from 1.65 ng/ml to 2000 ng/ml.Conclusion:The HPLC-MS/MS method is simple,accurate and rapid to detect the concentration of ibrutinib,and is suitable for daily monitoring of the plasma concentration of ibrutinib.