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工程科学与技术:2018,50(2):226-230
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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
(1.四川大学 化学工程学院, 四川 成都 610065;2.四川赛诺唯新生物技术有限公司, 四川 成都 610023)
Simultaneous Determination of 7-ethyl-14-Aminocamptothecin and Metabolite in Rat by UPLC-MS/MS Method
(1.College of Chemical Eng.,Sichuan Univ.,Chengdu 610065,China;2.Sichuan Sino-innovation Bio-technology Co.,Ltd,Chengdu 610023,China)
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投稿时间:2017-05-15    修订日期:2017-12-01
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<7.2%。药代动力学结果显示,静脉注射后EAC在大鼠体内的清除速率中等,组织分布高。给药后EAC-M在血浆中迅速检出,暴露量约相当于EAC的5%。连续给药5 d,EAC和EAC-M在大鼠体内无明显蓄积。所建立的方法简便、灵敏、选择性强,适用于SD大鼠血浆中EAC及代谢产物EAC-M的测定和药代动力学研究。
Abstract: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/z 392→291) and the metabolite (m/z 392→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.
文章编号:201700661     中图分类号:R917    文献标志码:
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
作者简介:王万(1980-),男,博士生.研究方向:生物化工.E-mail:myrid@foxmail.com
引用文本:
王万,廖立东,干玉娟,李晖.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[J].工程科学与技术,2018,50(2):226-230.
WANG Wan,LIAO Lidong,GAN Yujuan,LI Hui.Simultaneous Determination of 7-ethyl-14-Aminocamptothecin and Metabolite in Rat by UPLC-MS/MS Method[J].Advanced Engineering Sciences,2018,50(2):226-230.