LC-MS/MS法测定人血浆中沙丙蝶呤的浓度  

Determination of Sapropterin in Human Plasma by LC-MS/MS

作　　者：王月玥 林明弘 WANG Yueyue;LIN Minghong(Xuzhou Medical University,Xuzhou 221000,China;PPC Group,Xuzhou 221009,China)

机构地区：[1]徐州医科大学,徐州221000 [2]徐州立兴佳正医药科技有限公司,徐州221009

出　　处：《药学与临床研究》2021年第3期169-174,共6页Pharmaceutical and Clinical Research

摘　　要：目的:建立LC-MS/MS法测定人血浆中沙丙蝶呤的浓度。方法:经抗氧化剂(抗坏血酸,VC)预处理过的血浆样品,通过甲醇蛋白沉淀等一系列操作后进行LC-MS/MS分析。以沙丙蝶呤-^(13)C ^(15)N3为内标,使用的色谱柱为Kinetex■EVO C_(18)100A(4.6 mm×100 mm,5μm),流动相为含0.1%1 mol·L_(-1)乙酸铵的水溶液-乙腈(93∶7,v/v),等度洗脱分离,柱温为30℃,流速为0.6 mL·min_(-1),进样量为15μL。质谱检测采用电喷雾离子源(ESI),正离子多反应监测(MRM)模式扫描分析。沙丙蝶呤和沙丙蝶呤-^(13)C^(15)N_(3)的目标离子对分别为m/z 242.3→166.1和m/z 246.1→170.2。结果:沙丙蝶呤血药浓度在1~300 ng·mL^(-1)范围内线性关系良好;质控样本批内精密度(RSD)在1.2%~4.7%,批间精密度(RSD)在1.5%~4.3%;沙丙蝶呤的平均提取回收率为98.0%~101.2%;无明显空白血浆基质效应,不影响待测物的定量分析;血浆样本稳定性符合接受标准;制备后样本稳定性符合接受标准;方法重现性良好;浓度超出定量上限的血浆样本经空白血浆稀释5倍后可以准确测定。结论:本方法能够快速、简便、准确地测定人血浆中沙丙蝶呤的浓度。该方法经验证后,成功用于沙丙蝶呤临床样本的血药浓度的检测。Objective:To establish an efficient and sensitive LC-MS/MS method for the determination of sapropterin in human plasma.Methods:Sapropterin and internal standard(Sapropterin-13C15N3)were extracted from human plasma,which had been pretreated with antioxidant(ascorbic acid,VC),using methanol as the protein precipitating agent.Chromatographic separation was performed using a Kinetex■EVO C_(18)100A column(4.6 mm×100 mm,5μm)at 30℃.The mobile phase was consisted of 0.1%1 mol·L^(-1) ammonium acetate solution-acetonitrile(93∶7,v/v)at a flow rate of 0.6 mL·min^(-1).Electrospray ionization(ESI)sources were applied and operated in the positive ion mode.A multiple reaction monitoring(MRM)mode with the transition of m/z 242.3→166.1(Sapropterin)and m/z 246.1→170.2(Sapropterin-_(13)C_(15)N_(3))was used for quantification.Results:Determination of sapropterin showed good linearity between 1-300 ng·mL^(-1);The intra-and inter-day precisions(RSDs)were within 4.7%;The average extraction recovery of sapropterin were 98.0%-101.2%;There was no obvious blank plasma matrix effect of sapropterin;Sapropterin was stable during the study;The stability of prepared samples conformed to the accepted standard;The method had good reproducibility;Plasma samples with concentrations exceeding the upper limit of quantitation can be accurately determined after being diluted 5 times by blank plasma.The validated method was successfully applied to the detection of sapropterin concentrations in clinical samples.Conclusion:The method established in this study can quickly,simply and accurately determine the concentrations of sapropterin in human plasma.

关 键 词：沙丙蝶呤 LC-MS/MS 抗坏血酸 甲醇蛋白沉淀剂 人血浆 

分 类 号：R969.1[医药卫生—药理学]