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The diverse characteristics and large number of entities make metabolite separation challenging in metabolomics. To date, there is not a singular instrument capable of analyzing all types of metabolites. In order to achieve a better separation for higher peak capacity and accurate metabolite identification and quantification, we integrated GC × GC-MS and parallel 2DLC-MS for analysis of polar metabolites. To test the performance of the developed system, 13 rats were fed different diets to form two animal groups. Polar metabolites extracted from rat livers were analyzed by GC × GC-MS, parallel 2DLC-MS (-) and parallel 2DLC-MS (+), respectively. By integrating all data together, 58 metabolites were detected with significant change in their abundance levels between groups (p ≤ 0.05). Of the 58 metabolites, three metabolites were detected in two platforms and two in all three platforms. Manual examination showed that discrepancy of metabolite regulation measured by different platforms was mainly caused by the poor shape of chromatographic peaks resulting from low instrument response. Pathway analysis demonstrated that integrating the results from multiple platforms increased the confidence of metabolic pathway assignment.

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Integrating comprehensive two-dimensional gas chromatography mass spectrometry and parallel two-dimensional liquid chromatography mass spectrometry for untargeted metabolomics