Orlandi Carla, Jacques Carine, Duplan Hélène, Debrauwer Laurent, Jamin Emilien L
Toxalim (Research Centre in Food Toxicology), INRAE UMR 1331, Paul Sabatier University (UPS), ENVT, INP-Purpan, Toulouse 31062, France.
MetaboHUB-Metatoul, National Infrastructure of Metabolomics and Fluxomics, Metatoul-AXIOM, Toulouse 31077, France.
Anal Chem. 2023 Feb 7;95(5):2822-2831. doi: 10.1021/acs.analchem.2c04196. Epub 2023 Jan 30.
Liquid chromatography-mass spectrometry (LC-MS)-based metabolomics usually combines hydrophilic interaction liquid chromatography (HILIC) and reversed-phase (RP) chromatography to cover a wide range of metabolomes, requiring both significant sample consumption and analysis time for separate workflows. We developed an integrated workflow enabling the coverage of both polar and nonpolar metabolites with only one injection of the sample for each ionization mode using heart-cutting trapping to combine HILIC and RP separations. This approach enables the trapping of some compounds eluted from the first chromatographic dimension for separation later in the second dimension. In our case, we applied heart-cutting to non-retained metabolites in the first dimension. For that purpose, two independent miniaturized one-dimensional HILIC and RP methods were developed by optimizing the chromatographic and ionization conditions using columns with an inner diameter of 1 mm. They were then merged into one two-dimensional micro LC-MS method by optimization of the trapping conditions. Equilibration of the HILIC column during elution on the RP column and vice versa reduced the overall analysis time, and the multidimensionality allows us to avoid signal measurements during the solvent front. To demonstrate the benefits of this approach to metabolomics, it was applied to the analysis of the human plasma standard reference material SRM 1950, enabling the detection of hundreds of metabolites without the significant loss of some of them while requiring an injection volume of only 0.5 μL.
基于液相色谱-质谱联用（LC-MS）的代谢组学通常结合亲水作用液相色谱（HILIC）和反相（RP）色谱来覆盖广泛的代谢组，但这需要大量的样品消耗以及针对不同工作流程的分析时间。我们开发了一种集成工作流程，通过使用中心切割捕集技术将HILIC和RP分离相结合，在每种电离模式下仅进样一次样品就能覆盖极性和非极性代谢物。这种方法能够捕集从第一维色谱洗脱出来的一些化合物，以便在第二维中进行后续分离。在我们的案例中，我们将中心切割应用于第一维中未保留的代谢物。为此，通过使用内径为1 mm的色谱柱优化色谱和电离条件，开发了两种独立的小型化一维HILIC和RP方法。然后通过优化捕集条件将它们合并为一种二维微LC-MS方法。在RP柱上洗脱时对HILIC柱进行平衡，反之亦然，这减少了整体分析时间，并且多维性使我们能够避免在溶剂前沿期间进行信号测量。为了证明这种方法对代谢组学的益处，将其应用于人类血浆标准参考物质SRM 1950的分析，仅需0.5 μL进样体积就能检测到数百种代谢物，同时不会导致其中一些代谢物的显著损失。
