Introduction: It is widely but erroneously believed that drugs get into cells by passing through the phospholipid bilayer portion of the plasma and other membranes. Much evidence shows, however, that this is not the case, and that drugs cross biomembranes by hitchhiking on transporters for other natural molecules to which these drugs are structurally similar. Untargeted metabolomics can provide a method for determining the differential uptake of such metabolites. Objectives: Blood serum contains many thousands of molecules and provides a convenient source of biologically relevant metabolites. Our objective was to measure them. Methods: We develop an untargeted LC-MS/MS method to detect a broad range of compounds present in human serum. We apply this to the analysis of the time course of the uptake and secretion of metabolites in serum by several human cell lines, by analysing changes in the serum that represents the extracellular phase (the exometabolome or metabolic footprint). Results: Our method measures some 4,000-5,000 metabolic features in both ES+ and ES- modes. We show that the metabolic footprints of different cell lines differ greatly from each other. Conclusion: Our new, 15-minute untargeted metabolome method allows for the robust and convenient measurement of differences in the uptake of serum compounds by cell lines following incubation in serum, and its relation to differences in transporter expression. Key words: human serum, untargeted metabolomics, transporters, LC-MS/MS, Orbitrap, cell culture

中文翻译：

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一种非靶向代谢组学策略，用于测量哺乳动物细胞系代谢物吸收和排泄的差异

简介：人们普遍但错误地认为药物是通过血浆和其他膜的磷脂双层部分进入细胞的。但是，许多证据表明并非如此，药物通过搭便转运蛋白上与这些药物结构相似的其他天然分子的转运蛋白而跨过生物膜。非靶向代谢组学可以提供一种确定此类代谢物差异吸收的方法。目标：血清包含成千上万个分子，并提供了生物相关代谢产物的便捷来源。我们的目标是测量它们。方法：我们开发了一种无目标的LC-MS / MS方法，可检测人血清中存在的多种化合物。我们通过分析代表细胞外相的血清变化（exometabolome或代谢足迹），将其应用于几种人类细胞系对血清代谢物摄取和分泌的时间过程的分析。结果：我们的方法在ES +和ES-模式下测量了大约4,000-5,000个代谢特征。我们表明，不同细胞系的代谢足迹彼此差异很大。结论：我们的新型15分钟无靶代谢组学方法能够可靠，方便地测量在血清中孵育后细胞系对血清化合物摄取的差异及其与转运蛋白表达差异的关系。关键词：人血清，非靶向代谢组学，转运蛋白，LC-MS / MS，Orbitrap，细胞培养 通过分析代表细胞外相（exometabolome或代谢足迹）的血清变化。结果：我们的方法在ES +和ES-模式下测量了大约4,000-5,000个代谢特征。我们表明，不同细胞系的代谢足迹彼此差异很大。结论：我们的新型15分钟无靶代谢组学方法能够可靠，方便地测量在血清中孵育后细胞系对血清化合物摄取的差异及其与转运蛋白表达差异的关系。关键词：人血清，非靶向代谢组学，转运蛋白，LC-MS / MS，Orbitrap，细胞培养 通过分析代表细胞外相（exometabolome或代谢足迹）的血清变化。结果：我们的方法在ES +和ES-模式下测量了大约4,000-5,000个代谢特征。我们表明，不同细胞系的代谢足迹彼此差异很大。结论：我们的新型15分钟无靶代谢组学方法能够可靠，方便地测量在血清中孵育后细胞系对血清化合物摄取的差异及其与转运蛋白表达差异的关系。关键词：人血清，非靶向代谢组学，转运蛋白，LC-MS / MS，Orbitrap，细胞培养 我们表明，不同细胞系的代谢足迹彼此差异很大。结论：我们的新型15分钟无靶代谢组学方法能够可靠，方便地测量在血清中孵育后细胞系对血清化合物摄取的差异及其与转运蛋白表达差异的关系。关键词：人血清，非靶向代谢组学，转运蛋白，LC-MS / MS，Orbitrap，细胞培养 我们表明，不同细胞系的代谢足迹彼此差异很大。结论：我们的新型15分钟无靶代谢组学方法能够可靠，方便地测量在血清中孵育后细胞系对血清化合物摄取的差异及其与转运蛋白表达差异的关系。关键词：人血清，非靶向代谢组学，转运蛋白，LC-MS / MS，Orbitrap，细胞培养