Metabolic analysis.

Abstract

Analysis of the metabolome with coverage of all of the possibly detectable components in the sample, rather than analysis of each individual metabolite at a given time, can be accomplished by metabolic analysis. Targeted and/or nontargeted approaches are applied as needed for particular experiments. Monitoring hundreds or more metabolites at a given time requires high-throughput and high-end techniques that enable screening for relative changes in, rather than absolute concentrations of, compounds within a wide dynamic range. Most of the analytical techniques useful for these purposes use GC or HPLC/UPLC separation modules coupled to a fast and accurate mass spectrometer. GC separations require chemical modification (derivatization) before analysis, and work efficiently for the small molecules. HPLC separations are better suited for the analysis of labile and nonvolatile polar and nonpolar compounds in their native form. Direct infusion and NMR-based techniques are mostly used for fingerprinting and snap phenotyping, where applicable. Discovery and validation of metabolic biomarkers are exciting and promising opportunities offered by metabolic analysis applied to biological and biomedical experiments. We have demonstrated that GC-TOF-MS, HPLC/UPLC-RP-MS and HILIC-LC-MS techniques used for metabolic analysis offer sufficient metabolome mapping providing researchers with confident data for subsequent multivariate analysis and data mining.