State Key Laboratory of Quality Research in Chinese Medicine, Macau Institute for Applied Research in Medicine and Health , Macau University of Science and Technology , Macao , China.
Guangdong Key Laboratory for Genome Stability & Disease Prevention, Department of Pharmacology and Carson International Cancer Research Centre , Shenzhen University School of Medicine , Shenzhen 518060 , China.
Anal Chem. 2018 Oct 2;90(19):11210-11215. doi: 10.1021/acs.analchem.8b01873. Epub 2018 Sep 20.
Carboxyl-containing metabolites (CCMs) widely exist in living systems and are the essential components for life. Global characteristics of CCMs in biological samples are critical for the understanding of physiological processes and the discovery for the onset of relevant diseases. However, their determination represents a challenge due to enormous polarity differences, structural diversity, high structural similarity, and poor ionization efficiency in mass spectrometry. Herein, 5-(diisopropylamino)amylamine (DIAAA) derivatization coupled with liquid chromatography-mass spectrometry (LC-MS) was developed for mapping the CCMs. With this methodology, the sensitivity was significantly enhanced. More importantly, the hydrophobicity of polar CCMs, amino acids, TCA cycle intermediates, and short-chain fatty acids and the hydrophilicity of low-polar CCMs, long-chain fatty acids, and bile acids were significantly increased, resulting in a remarkable separation efficiency for which 68 CCMs can be simultaneously determined. Furthermore, the polarity-tuning effect was confirmed to be induced by the different impacts of aliphatic chains and nitrogen atom in DIAAA, the latter existing as a cation in the acidic mobile phase, using different derivatization reagents. Finally, this derivatization method was utilized to hunt for the potential biomarkers in colorectal cancer (CRC) patients and 52 CCMs, related with several key metabolic pathways, including amino acids metabolism, TCA cycle, fatty acid metabolism, pyruvate metabolism, and gut flora metabolism were identified. This innovative polarity-tuning derivatization-LC-MS approach was proved to be a valuable tool for probing global metabolome with high separation efficiency and sensitivity in various biological samples.
含羧基代谢物（CCMs）广泛存在于生命体系中，是生命的基本组成部分。了解生物样本中 CCMs 的全局特征对于理解生理过程和发现相关疾病的发病机制至关重要。然而，由于其在质谱中存在巨大的极性差异、结构多样性、高度结构相似性和较差的电离效率，其测定具有挑战性。在此，开发了 5-（二异丙基氨基）戊胺（DIAAA）衍生化结合液相色谱-质谱（LC-MS）方法来绘制 CCMs。采用该方法，灵敏度显著提高。更重要的是，极性 CCMs、氨基酸、三羧酸（TCA）循环中间产物和短链脂肪酸的疏水性以及低极性 CCMs、长链脂肪酸和胆汁酸的亲水性显著增加，从而实现了显著的分离效率，可以同时测定 68 种 CCMs。此外，通过使用不同的衍生化试剂，证实了 DIAAA 中脂肪链和氮原子的不同影响会产生极性调谐效应，后者在酸性流动相中以阳离子形式存在。最后，该衍生化方法用于寻找结直肠癌（CRC）患者的潜在生物标志物，鉴定出与氨基酸代谢、TCA 循环、脂肪酸代谢、丙酮酸代谢和肠道菌群代谢等几个关键代谢途径相关的 52 种 CCMs。这种创新的极性调谐衍生化-LC-MS 方法被证明是一种在各种生物样本中具有高分离效率和灵敏度的探索全局代谢组学的有价值工具。
