UPLC-MS/MS法同时检测大鼠体内7-乙基-14-氨基喜树碱及其代谢产物
Simultaneous Determination of 7-ethyl-14-Aminocamptothecin and Metabolite in Rat by UPLC-MS/MS Method
作者：王万(四川大学 化学工程学院, 四川 成都 610065;四川赛诺唯新生物技术有限公司, 四川 成都 610023)；廖立东(四川赛诺唯新生物技术有限公司, 四川 成都 610023)；干玉娟(四川赛诺唯新生物技术有限公司, 四川 成都 610023)；李晖(四川大学 化学工程学院, 四川 成都 610065)
Author：WANG Wan(College of Chemical Eng.,Sichuan Univ.,Chengdu 610065,China;Sichuan Sino-innovation Bio-technology Co.,Ltd,Chengdu 610023,China)；LIAO Lidong(Sichuan Sino-innovation Bio-technology Co.,Ltd,Chengdu 610023,China)；GAN Yujuan(Sichuan Sino-innovation Bio-technology Co.,Ltd,Chengdu 610023,China)；LI Hui(College of Chemical Eng.,Sichuan Univ.,Chengdu 610065,China)
收稿日期：2017-05-15 年卷（期）页码：2018,50(2):226-230
期刊名称：工程科学与技术
Journal Name：Advanced Engineering Sciences
关键字：7-乙基-14-氨基喜树碱;内酰胺代谢产物;大鼠血浆;超高效液相色谱-串联质谱;同时定量检测
Key words：7–ethyl–14–Aminocamptothecin;lactam metabolite;rat plasma;UPLC-MS/MS;simultaneous quantitative analysis
基金项目：省科技厅应用基础研究；2014JY0042；仿制药QbD与制药过程分析技术的整合
中文摘要
7-乙基-14-氨基喜树碱（EAC）是具有良好抗肿瘤活性的新型喜树碱衍生物，为测定静脉注射EAC后原形化合物及其酰胺型代谢产物（EAC-M）在体内的浓度，揭示体内动力学性质，现研究建立了同时定量测定SD大鼠血浆中EAC和EAC-M的UPLC-MS/MS方法。以7-乙基-14-硝基喜树碱（ENC）、吉咪替康（CMT）为内标，血浆样品加入内标溶液和乙腈沉淀蛋白后，以Phenyl-Hexyl柱（2.1 mm×50 mm，1.8 μm）为分离柱，5 mmol/L醋酸铵水溶液（含0.1%甲酸）-乙腈为流动相梯度洗脱，流速0.5 mL/min。洗脱物经电喷雾离子源（ESI）正离子化，在三重四级杆质谱仪上以多反应监测模式（MRM）测定EAC（m/z 392→291）和EAC-M（392→290）。方法学研究结果显示，内源性物质不干扰待测物和内标的测定。在0.500～250 ng/mL范围内，EAC和EAC-M的浓度与响应值的线性关系良好，定量下限均为0.500 ng/mL。方法的日内、日间精密度（RSD）≤7.6%，准确度（RE）为-4.2%～5.9%。提取回收率为84.4%～98.7%，基质效应为94.8%～104%。高于定量上限浓度的样品，用空白血浆稀释6倍至线性范围，测得的精密度（RSD）为1.8%，准确度（RE）为-10%～-3.0%。样品经设定的稳定性考察条件后，测得的准确度（RE）为-8.6%～11%，RSD
英文摘要
7-ethyl-14-Aminocamptothecin (EAC) is a novel derivate of camptothecin which exhibits favorable antitumor activity.To determine the concentrations of the parent drug and lactam form metabolite (EAC-M) in vivo after intravenous administration of EAC,an UPLC-MS/MS method was established to simultaneously quantify EAC and EAC-M in SD rat plasma.7-ethyl-14-Nitrocamptothecin (ENC) and Chimmitecan (CMT) were used as internal standards.After protein precipitation by adding solution of internal standards and acetonitrile,plasma samples were separated on a Phenyl-Hexyl colum (2.1mm×50 mm,1.8 μm) with the mobile phase consisting of 0.5 mmoL/L ammonium acetate solution (containing 0.1% formic acid)-acetonitrile.Gradient elution was employed at a flow rate of 0.5 mL/min.Eluted compounds were ionized by an electrospray ionization source (ESI) in positive mode.The parent to production transitions for both the parent drug (m/z392→291) and the metabolite (m/z392→290) were monitored on a triple quadrupole mass spectrometer,using the multiple reaction monitoring (MRM).Based on the results from method validation,no interference was observed in blank plasma samples.The linear ranges for EAC and EAC-M were 0.500~250 ng/mL,the limits of quantitation were 0.500 ng/mL.The inter- and intra-day precisions (RSD) were not more than 7.6%,the accuracies (RE) were -4.2%~5.9%.The extraction recovery values were 84.4%~98.7%,and the matrix effects were 94.8%~104%.Samples beyond the upper limits of quantification were analyzed after a 6-fold-dilution with blank plasma,the precisions (RSD) were 1.8%,and the accuracies (RE) were -10%~-3.0%.In stability tests,the accuracies (RE) were -8.6%~11% and the precisions (RSD) were blow 7.2%.Data from pharmacokinetic tests indicated that EAC is moderately cleaned and is widely distributed in tissues.EAC-M was rapidly detected in plasma after intravenous administration which showed a 5% exposure of EAC.No accumulation of EAC or EAC-M was observed after an administration for 5 consecutive days.In summary,the provided method is convenient,sensitive,selective,and suitable for investigation of EAC and EAC-M in pharmacokinetic research.
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