同时测定小鼠血浆中氯吡格雷及其代谢产物浓度的LC-MS/MS法及其药代动力学研究  

Determination of clopidogrel and its metabolites in plasma by LC-MS/MS and its application in the pharmacokinetics study in mice

作　　者：张梦然 邰婷[1] 米琼宇[1] 吉金子 黄贝贝 贾雨萌 谢红光[1] 

机构地区：[1]南京医科大学附属南京医院中心实验室,江苏南京210006

出　　处：《南京医科大学学报（自然科学版）》2017年第8期955-960,共6页Journal of Nanjing Medical University(Natural Sciences)

基　　金：国家自然科学基金(81473286;81503144)

摘　　要：目的 :建立并验证小鼠血浆中氯吡格雷及其代谢物的LC-MS/MS检测方法 ,研究氯吡格雷在小鼠体内的药代动力学特征。方法:以吡罗昔康为内标,经乙腈沉淀蛋白后,采用电喷雾离子源(ESI)正离子扫描模式多反应监测(multiple reaction monitoring,MRM)方式检测氯吡格雷及其活性代谢物、羧酸代谢物和葡萄糖醛酸代谢物的血药浓度。色谱柱为安捷伦Poroshell120 SB-C18柱(100 mm×2.1 mm,2.7μm);流动相为水(含0.1%甲酸)和乙腈(含0.1%甲酸)并按梯度洗脱。小鼠单次灌胃给予氯吡格雷10 mg/kg,测定氯吡格雷及其3种主要代谢物的血药浓度,并研究其药代动力学特征。结果:氯吡格雷及其代谢产物在一定测定范围内均呈现良好的线性关系(r>0.99),提取回收率均大于80%,未见明显基质效应,其日内、日间精密度均较好(RSD<15%),准确度相对误差(RE)为-2.40%~5.00%,稳定性较好。在小鼠的药代动力学研究中,该方法能达到预期的实验要求。结论:所建立的LC-MS/MS分析方法能快速、准确地测定小鼠血浆中氯吡格雷及其代谢产物浓度,其专属性、回收率、基质效应、稳定性、精密度和准确度等均符合生物样品测定要求。Objective： To develop and validate a sensitive high performance liquid chromatography-tandem mass spectrometry method for the simultaneous determination of clopidogrel and its metabolites and subsequently to use it to investigate pharmacokinetic properties of clopidogrel in mice. Methods： Clopidogrel and its metabolites were determined at an electron spray ionization（ESI）source using multiple reaction monitoring（MRM） in positive ion mode after protein precipitation by acetonitrile,in which piroxicam was added as an internal standard. Chromatographic separation was achieved on a Poroshell 120 SB-C18 column（100×2.1 mm, 2.7μm, Agilent Technologies） using a mobile phase that consisted of both solvent A（0.1% formic acid in water） and solvent B（0.1%formic acid in acetonitrile） in a gradient elution manner. The pharmacokinetics of clopidogrel was investigated in mice receiving a single dose of clopidogrel（10 mg/kg） by gavage administration. Results： The simultaneous assay of clopidogrel and its metabolites achieved good linearity（r 0.99） within the range of the spiked plasma concentrations. The recoveries were greater than 80%, and no significant matrix effect was found. The both intra-and inter-day RSDs were less than 15%, and the accuracy ranged from-2.40% to5.00%. The stability of the method was good. In addition, a series of pharmacokinetics parameters were obtained in mice after receiving a single oral dose of clopidogrel through this analytical method. Conclusion： The modified methodology was highly sensitive, accurate, and reproducible for the simultaneous determination of clopidogrel and its three major metabolites in mouse plasma and therefore, it was successfully applied in pharmacokinetic studies of clopidogrel in mice.

关 键 词：氯吡格雷 LC-MS/MS 药代动力学 药物代谢 

分 类 号：R969.1[医药卫生—药理学]