HSCT患儿血浆中白消安的浓度测定及暴露量研究

Study on the Concentration Determination and Exposure of Busulfan in Children with Hematopoietic Stem Cell Transplantation (HSCT)

目的:建立测定造血干细胞移植(HSCT)患儿体内白消安(BU)血药浓度的液相色谱-串联质谱(LC-MS/MS)法,并进一步估算非清髓预处理(RIC)方案下患儿体内BU暴露情况。方法:以白消安D8(BU-D8)为内标,乙腈蛋白沉淀,色谱柱为Waters ACQUITY UPLC BEHC18(50 mm×2.1 mm,1.7μm);柱温:40℃;流动相以含0.1%甲酸的乙腈为有机相,以含10 mmol·L^(-1)甲酸铵和0.1%甲酸的水溶液为水相,梯度洗脱;流速:0.4 mL·min^(-1);进样量:2μL。采用电喷雾离子源(ESI)和多反应监测(MRM)模式检测,BU和BU-D8离子通道分别为m/z 264.10→150.90和m/z272.10→159.10。使用NextDose贝叶斯回归模型分别计算患儿体内BU暴露量,以药-时曲线下面积(AUC)表示。结果:BU在0.1~10.0μg·mL^(-1)范围内具有良好的线性关系,定量下限为0.1μg·mL^(-1)。定量下限以及低、中、高浓度质控品的批内和批间精密度分别为4.3%~5.1%和3.2%~6.9%,平均提取回收率为(111.2±11.8)%,经内标归一化的基质效应均<15%。21例患儿经模型估算的BU AUC中位值为1191.23(775~2511.57)μmol·min·L^(-1)),个体间差异较大。结论:该方法灵敏度、准确度高,操作简便快速,适用于临床开展BU血药浓度监测。RIC方案下患儿体内BU暴露存在个体差异性,有待进一步开展相关研究。

Objective:To develop a liquid chromatography-tandem mass spectrometry(LC-MS/MS)method for the determination of plasma concentration of busulfan(BU),and to further estimate the exposure of BU in children with reduced-intensity conditioning regimen(RIC)during hematopoietic stem cell transplantation(HSCT).Methods:The plasma samples were precipitated by acetonitrile with busulfan-D8(BU-D8)as internal standard.HPLC separation was performed on a Waters ACQUITY UPLC BEH C_(18) column(50 mm×2.1 mm,1.7μm)at 40℃.The mobile phase consisting water(10 mmol·L^(-1)ammonium formate-0.1%formic acid)and acetonitrile(0.1%formic acid)was applied for gradient elution at 0.4 mL·min^(-1).The injection volume was 2μL.The electro spray ionization mode was performed and multiple reaction monitoring was conducted at a transition of 264.1→150.90(m/z)for BU and 272.1→159.1(m/z)for BU-D8.The NextDose Bayesian regression model was used to calculate the BU exposure as Area Under Curve(AUC).Results:Linearity was good ranged from 0.1μg·mL^(-1)to 10.0μg·mL^(-1).Limit of quantification was 0.1μg·mL^(-1).The intra-and inter-day RSD were 4.3%-5.1%and 3.2%-6.9%,respectively.The average recovery rate of extraction was(111.2±11.8)%.The matrix effects normalized by internal standard were all less than 15%.The median value of BU AUC estimated by the model in 21 children was 1191.23(775-2511.57)μmol·min·L^(-1),which varied greatly among individuals.Conclusion:The method is sensitive,accurate,simple and rapid,and suitable for clinical monitoring of BU plasma concentrations.There are individual differences in BU exposure in children under RIC regimen,and further studies need to be conducted.