人血浆中安罗替尼质量浓度测定方法的建立及其应用

Establishment and Application of Method for Concentration Determination of Anlotinib in Human Plasma

目的:建立测定人血浆中安罗替尼质量浓度的方法并应用于临床。方法:以醋酸铵为盐析助剂,以乙腈为溶剂,采用盐析辅助液液萃取对血浆样品进行前处理;以伏立康唑为内标,采用液相色谱-串联质谱法(LC-MS/MS)测定。色谱柱为Waters X Bridge C_(18),流动相为0.2%甲酸水溶液-乙腈(梯度洗脱),流速为1 mL/min,柱温为40℃,进样量为10μL,分流比为3∶7;离子源为电喷雾离子源,以多反应监测模式进行正离子扫描,用于定量分析的离子对分别为m/z 408.3→339.3(安罗替尼)、m/z 350.2→281.3(内标)。结果:安罗替尼检测质量浓度的线性范围为0.2~200 ng/mL(R^(2)=0.996 7);定量下限为0.2 ng/mL;日内、日间精密度试验的RSD均小于12%(n=6或n=3);准确度为90.92%~108.00%(n=6或n=3);平均提取回收率为87.51%~100.00%(RSD<8%,n=6);平均基质效应为96.66%~99.93%(RSD<5%,n=6)。3例使用安罗替尼治疗的非小细胞肺癌患者体内的血药浓度为8.74~65.60 ng/mL。结论:该方法操作简单、准确度高、专属性强,可用于非小细胞癌患者体内安罗替尼的血药浓度监测。

OBJECTIVE:To establish a method for concentration determination of anlotinib in human plasma and apply it in the clinic.METHODS:The plasma samples were pretreated by salting-out assisted with liquid-liquid extraction with ammonium acetate as salting out assistant and acetonitrile as solvent.Using voriconazole as internal standard,LC-MS/MS method was adopted.The separation was performed on Waters X Bridge C_(18 )column with mobile phase consisting of 0.2%formic acid solutionacetonitrile(gradient elution)at the flow rate of 1 mL/min.The column temperature was set at 40℃,and sample size was 10μL.The split ratio was 3∶7.The electrospray ion source and multiple reaction monitoring mode were used for the analysis.The ion pair of anlotinib and internal standard under positive ion mode were m/z 408.3→339.3 and m/z 350.2→281.3,respectively.RESULTS:Anlotinib showed a good linear relationship in the concentration range of 0.2-200 ng/mL(R^(2)>0.9967).The lowest limit of quantitation was 0.2 ng/mL.Intra-day and inter-day RSDs were no more than 12%(n=6 or n=3).Accuracies were 90.92%-108.00%(n=6 or n=3).The average extraction recoveries were 87.51%-100.00%(RSD<8%,n=6).The average matrix effects were 96.66%-99.93%(RSD<5%,n=6).The plasma concentration of 3 patients with NSCLC treated with anlotinib was 8.74-65.60 ng/mL.CONCLUSIONS:The method is simple,accurate and specific,and is suitable for the plasma concentration monitoring of anlotinib in NSCLC patients.