Caco-2细胞中^(13)C-D-葡萄糖含量测定方法的建立

Establishment of Assay Method for Determination of ^(13)C-D-Glucose in Caco-2 Cells

【目的】建立检测Caco-2细胞中13C-D-葡萄糖含量的液相色谱—质谱联用(LC-MS)方法。【方法】样品经前处理后,通过Atlantis HILIC色谱柱分离,以乙腈与体积分数0.1%甲酸为流动相,质谱为电喷雾离子源,在负离子模式下以选择反应监测(SRM)方式进行检测。【结果】在选定的色谱和质谱条件下,各指标成分峰分离良好,13C-D-葡萄糖在定量范围内呈现良好的线性关系,R2=0.999 5,高、中、低浓度日内精密度和日间精密度相对标准偏差(sR)<20%,回收率在100.61%～108.34%之间,样品检测的重复性及样品稳定性良好。【结论】本方法适用于Caco-2细胞中13C-D-葡萄糖含量的测定。

Objective To establish a method for the determination of^13C-D-glucose in Caco-2 cells by liquid chromatography/tandem mass spectrometry （LC-MS） . Methods The pretreated samples were analyzed with LC-MS/MS system. LC with Atlantis HILIC column was adopted, and the mobile phase consisted of acetonitrile and 0.1% formic acid at flow rate of 1.0 mL/min. Eleetrospray ionization （ESI） was applied and operated in negative ion mode by means of selected reaction monitoring. Results^13C-D-glucose was well separated from internal standard. ^13C-D-glucose showed a good linear relationship in the fixed quantity, R2=0.999 5. The relative standard deviation （SR） of intra-day and inter-day precision at high, medium and low concentration were all less than 20%, and the recoveries were in the range of 100.61% to 108.34% with good repeatability and stability. Conclusion The developed method in this experiment can be used for the detection of ^13C-D-glucose in Caco-2 cells.