Differential Attenuation of NMR Signals by Complementary Ion-Exchange Resin Beads for De Novo Analysis of Complex Metabolomics Mixtures.

A primary goal of metabolomics is the characterization of a potentially very large number of metabolites that are part of complex mixtures. Application to biofluids and tissue samples offers insights into biochemical metabolic pathways and their role in health and disease. 1D1 H and 2D13 C-1 H HSQC NMR spectra are most commonly used for this purpose. They yield quantitative information about each proton of the mixture, but do not tell which protons belong to the same molecule. Interpretation requires the use of NMR spectral databases, which naturally limits these investigations to known metabolites. Here, a new method is presented that uses complementary ion exchange resin beads to differentially attenuate 2D NMR cross-peaks that belong to different metabolites. Based on their characteristic attenuation patterns, cross-peaks could be clustered and assigned to individual molecules, including unknown metabolites with multiple spin systems, as demonstrated for a metabolite model mixture and E. coli cell lysate.