Chemical labeling (CL) in combination with liquid chromatography–mass spectrometry (LC-MS) analysis has been demonstrated to be a promising technology in metabolomic analysis. However, identification of chemically labeled metabolites remains to be challenging. Retention time (RT) is one of the most important parameters for the identification of metabolites, but it could vary greatly in LC-MS analysis. In this work, we developed a chemical labeling-based HPLC retention index (CL-HPLC RI) strategy to facilitate the identification of metabolites. In this CL-HPLC RI strategy, a series of 2-dimethylaminoethylamine (DMED)-labeled fatty acids were used as calibrants to establish RIs for DMED-labeled carboxylated compounds and a series of 4-(N,N-dimethylamino)phenyl isothiocyanate (DMAP)-labeled fatty amines were used as calibrants for DMAP-labeled amine compunds. To calculate the RIs, the whole LC chromatogram was divided into 24 time intervals by 23 DMED-labeled fatty acid standards or 15 time intervals by 14 DMAP-labeled fatty amine standards. Then, we established the RIs of 854 detected DMED-labeled carboxylated metabolites and 1057 DMAP-labeled amine metabolites in fecal samples and demonstrated that RIs were highly reproducible under different elution gradients, columns, and instrument systems. Finally, we applied this strategy to the identification of metabolites in human serum. Using RIs, 267 DMED-labeled carboxylated metabolites and 273 DMAP-labeled amine metabolites in human serum matched well with the fecal metabolome database. Taken together, the developed CL-HPLC RI strategy was demonstrated to be a promising method to facilitate the identification of metabolites in metabolomic analysis.

中文翻译：

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基于化学标记的代谢组学分析液相色谱保留指数的建立

化学标记（CL）与液相色谱-质谱（LC-MS）分析相结合已被证明是代谢组学分析中的一项有前途的技术。然而，鉴定化学标记的代谢物仍然具有挑战性。保留时间（RT）是鉴定代谢物的最重要参数之一，但在LC-MS分析中可能会有很大差异。在这项工作中，我们开发了一种基于化学标记的HPLC保留指数（CL-HPLC RI）策略，以促进代谢物的鉴定。在此CL-HPLC RI策略中，使用一系列2-二甲基氨基乙胺（DMED）标记的脂肪酸作为校正剂，以建立DMED标记的羧化化合物的RI和一系列4-（N，N-二甲基氨基）苯基异硫氰酸酯（DMAP）标记的脂肪胺被用作DMAP标记的胺化合物的校准物。为了计算RI，将整个LC色谱图按23个DMED标记的脂肪酸标准液分为24个时间间隔，或按14个DMAP标记的脂肪胺标准液分为15个时间间隔。然后，我们在粪便样品中建立了854个检测到的DMED标记的羧化代谢产物和1057 DMAP标记的胺代谢产物的RI，并证明在不同的洗脱梯度，色谱柱和仪器系统下，该RI具有高度的重现性。最后，我们将此策略应用于人血清中代谢物的鉴定。使用RIs，人血清中的267种DMED标记的羧化代谢物和273种DMAP标记的胺代谢物与粪便代谢组数据库很好地匹配。在一起