| Popis: |
Covering a wide spectrum of molecules is essential for global metabolome assessment. While metabolomics assays are most frequently carried out in microbore LC-MS analysis, reducing the size of the analytical platform has proven its ability to boost sensitivity for specific -omics applications. In this study, we elaborate the impact of LC miniaturization on exploratory small-molecule LC-MS analysis, focusing on chromatographic properties with critical impact on peak picking and statistical analysis. We have assessed a panel of small molecules comprising endogenous metabolites and environmental contaminants covering three flow regimes—analytical, micro-, and nano-flow. Miniaturization to the micro-flow regime yields moderately increased sensitivity as compared to the nano setup, where median sensitivity gains around 80-fold are observed in protein-precipitated blood plasma extract. This gain resulting in higher coverage at low µg/L concentrations is compound dependent. At the same time, the nano-LC-high-resolution mass spectrometry (HRMS) approach reduces the investigated chemical space as a consequence of the trap-and-elute nano-LC platform. Finally, while all three setups show excellent retention time stabilities, rapid gradients jeopardize the peak area repeatability of the nano-LC setup. Micro-LC offers the best compromise between improving signal intensity and metabolome coverage, despite the fact that only incremental gains can be achieved. Hence, we recommend using micro-LC for wide-target small-molecule trace bioanalysis and global metabolomics of abundant samples. |
