色谱

色谱

• 研究论文 • 上一篇    下一篇

超高效液相色谱-串联质谱法测定大鼠血浆中的磷酸西他列汀

汤瑶1, 李响2, 闻镍1, 孙旭1, 朱凌1, 于敏1, 李佐刚1*, 李波1   

  1. 1. 中国食品药品检定研究院国家药物安全评价监测中心, 北京 100176; 2. 吉林大学生命科学学院, 吉林 长春 130012
  • 收稿日期:2011-01-25 修回日期:2011-03-10 出版日期:2011-06-28 发布日期:2011-07-25
  • 通讯作者: 李佐刚,研究员,主要从事临床前药代动力学研究和GLP（良好实验室规范）管理. Tel: (010)67872233-8312
  • 基金资助:

    科技部“863计划”项目(No. 2006BAI14B06)

Determination of sitagliptin phosphate in rat plasma by ultra high performance liquid chromatography-tandem mass spectrometry

TANG Yao1, LI Xiang2, WEN Nie1, SUN Xu1, ZHU Ling1, YU Min1, LI Zuogang1*, LI Bo1   

  1. 1. National Center for Safety Evaluation of Drugs, National Institutes for Food and Drug Control, Beijing 100176, China; 2. College of Life Sciences, Jilin University, Changchun 130012, China
  • Received:2011-01-25 Revised:2011-03-10 Online:2011-06-28 Published:2011-07-25

PDF

744

被引次数

26

摘要： 建立了大鼠血浆中磷酸西他列汀含量检测的超高效液相色谱-串联质谱(UPLC-MS/MS)分析方法。以大鼠空白血浆为基质,通过添加标准品的方法配制含磷酸西他列汀和内标物氟西汀的样品,选用甲醇为沉淀剂,经离心除去血浆中的蛋白质,上清液用于目标物的检测。采用Thermo Hypersil Gold C18柱(50 mm×2.1 mm, 1.9 μm)为分析柱,Phenomenex Security Guard C18(4 mm×3.0 mm)为预柱,以乙腈和0.05%(v/v)甲酸水溶液为流动相进行梯度洗脱,流速为200 μL/min, 5 min内实现了快速分离。采用电喷雾正离子(ESI+)模式电离,选择反应监测(SRM)模式检测,确定了磷酸西他列汀和氟西汀的监测离子对分别为m/z 408.0→235.0和m/z 310.0→148.0,用基质匹配标准溶液法进行定量。结果表明: 大鼠血浆中磷酸西他列汀的质量浓度在1～1000 μg/L范围内时线性关系良好(r=0.9991),检出限(信噪比为3)为0.2 μg/L;其平均回收率为85%～115%;日内及日间的相对标准偏差(RSDs)均小于15%,满足生物样品检测的要求。将该方法初步用于大鼠静脉注射后的血浆样品中磷酸西他列汀的检测。该方法快速、灵敏度高、操作简便、重现性好,能够用于磷酸西他列汀药代动力学等方面的研究。

关键词: 超高效液相色谱-串联质谱法, 大鼠, 电喷雾离子化, 磷酸西他列汀, 血浆

Abstract: an ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the determination of sitagliptin phosphate in rat plasma was established. The blank rat plasma sample added with sitagliptin phosphate and the internal standard (fluoxetine) standard solution were prepared. Methanol was added in the sample for the deproteinization. Then the sample was vortex-mixed and centrifuged. The clear supernatant was used for the analysis of UPLC-MS/MS. A Thermo Hypersil Gold C18 column (50 mm×2.1 mm, 1.9 μm) was employed with a guard column of Phenomenex Security Guard C18 column (4 mm×3.0 mm), and the column temperature was set at 35 ℃. The gradient elution of acetonitrile and water (containing 0.05% (v/v) formic acid) as mobile phases was performed at a flow rate of 200 μL/min, and a rapid separation was completed in 5 min. The electrospray was operated in the positive ionization mode and the sitagliptin phosphate and fluoxetine were identified by selected reaction monitoring (SRM) mode, and the monitoring ions of them were m/z 408.0→235.0 and m/z 310.0→148.0, respectively, which were used to qualify and quantity the targets by the method of matrix-matched standard solution. The calibration curve showed good linearity within the concentrations of 1 to 1000 μg/L (r=0.9991); the limit of detection was 0.2 μg/L. The mean recoveries were from 85% to 115% at the spiked levels of 5, 50 and 500 μg/L; the relative standard deviations (RSDs) of intra- and inter-day of variation were both less than 15%, which can meet the determination requirements of biological samples. Then the method was initially used for the determination of sitagliptin phosphate in SD rat plasma after the administration of a single intravenous injection dose of sitagliptin phosphate. The method is rapid, sensitive, convenient and reproducible in the determination of sitagliptin phosphate, and can be used for the pharmacokinetics research of sitagliptin phosphate in plasma.

Key words: electrospray ionization (ESI), plasma, rat, sitagliptin phosphate, ultra high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS)