Screening of mammalian target of rapamycin inhibitors in natural product extracts by capillary electrophoresis in combination with high performance liquid chromatography-tandem mass spectrometry

Journal of Chromatography. a
Yanmei ZhangJingwu Kang

Abstract

In this study, capillary electrophoresis (CE) combined with HPLC-MS/MS were used as a powerful platform for screening of inhibitors of mammalian target of rapamycin (mTOR) in natural product extracts. The screening system has been established by using 5-carboxyfluorescein labeled substrate peptide F-4EBP1, a known mTOR inhibitor AZD8055, and a small chemical library consisted of 18 natural product extracts. Biochemical screening of natural product extracts was performed by CE with laser induced fluorescence detection. The CE separation allowed a quantitative measurement of the phosphorylated product, hence the quantitation of enzymatic inhibition as well as inhibition kinetics. The hits are readily identified as long as the peak area of the phosphorylated product is reduced in comparison with the negative control. Subsequent assay-guided isolation of the active natural product extract was performed with HPLC-MS/MS to track the particular active components. The structures of the identified active components were elucidated by the molecular ions and fragmentation information provided by MS/MS analysis. The CE-based assay method only requires minute pure compounds, which can be readily purified by HPLC. Therefore, the combination ...Continue Reading

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Citations

Jan 8, 2016·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Hanqing PangJin-ao Duan
Dec 8, 2015·Analytical Chemistry·Rachel K HarstadMichael T Bowser
Jun 1, 2017·The Analyst·Nicolas GuichardSandrine Fleury-Souverain
May 22, 2019·Electrophoresis·Wei-Feng Wang, Jun-Li Yang

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