Derivatization of the tricarboxylic acid cycle intermediates and analysis by online solid-phase extraction-liquid chromatography-mass spectrometry with positive-ion electrospray ionization
Derivatization of the tricarboxylic acid cycle intermediates and analysis by online solid-phase extraction-liquid chromatography-mass spectrometry with positive-ion electrospray ionization
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DOI：
10.1016/j.chroma.2011.07.095
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发表时间：
2012-04-06
影响因子：
4.1
通讯作者：
Giera, M.
中科院分区：
文献类型：
--
作者：
Kloos, D.;Derks, R. J. E.;Giera, M.
细胞代谢过程的分析是基本的生物学兴趣。细胞代谢物，如三羧酸（TCA）循环的中间产物，提供了有关细胞代谢状态的重要信息。TCA循环不仅是有氧细胞能量调节的关键因素，它还可能在细胞信号传导中发挥作用。本文描述了一种新的衍生化策略，使用经验选择的N-甲基-2-苯基乙胺作为衍生化试剂，碳二亚胺作为共试剂，用于羧酸的选择性衍生化，例如TCA循环的二羧酸和三羧酸。描述了衍生化方案的优化。这一程序使得使用在线固相萃取和反相液相色谱结合灵敏的正离子电喷雾电离质谱的衍生物的分析。完整的程序，包括使用核壳硅胶柱材料，允许快速分析样品基质中的TCA循环中间体，这里显示的是猪心组织提取物，在3-4个数量级上具有良好的线性。检测限范围从12到1000 nM，取决于分析物。(C)2011爱思唯尔有限公司版权所有。
The analysis of cellular metabolic processes is of fundamental biological interest. Cellular metabolites, such as the intermediates of the tricarboxylic acid (TCA) cycle, provide essential information about the metabolic state of the cell. Not only is the TCA cycle a key factor in the energy regulation within aerobic cells, it possibly also plays a role in cell signaling. This paper describes a novel derivatization strategy, using the empirically selected N-methyl-2-phenylethanamine as derivatization reagent with a carbodiimide as co-reagent, for the selective derivatization of carboxylic acids, such as the di- and tri-carboxylic acids of the TCA cycle. Optimization of the derivatization protocol is described. This procedure enables analysis of the derivatives using on-line solid-phase extraction and reversed-phase liquid chromatography in combination with sensitive positive-ion electrospray ionization mass spectrometry. The complete procedure, involving the use of core-shell silica column material, allows the rapid analysis of TCA cycle intermediates in sample matrices, here shown for pig heart tissue extracts, with a good linearity over 3-4 orders of magnitude. Detection limits range from 12 to 1000 nM, depending on the analyte. (C) 2011 Elsevier B.V. All rights reserved.