Ultrafast Quantitative 2D NMR: An Efficient Tool for the Measurement of Specific Isotopic Enrichments in Complex Biological Mixtures

Two-dimensional nuclear magnetic resonance (2D NMR) is a promising tool for studying metabolic fluxes by measuring C-13-enrichments in complex mixtures of C-13-labeled metabolites. However, the methods reported so far are hampered by very long acquisition durations limiting the use of 2D NMR as a quantitative tool for fluxomics. In this paper, we propose a new approach for measuring specific C-13-enrichments in a very fast way, by using new experiments based on ultrafast 2D NMR. Two homonudear 2D experiments (ultrafast COSY and zTOCSY) are proposed to measure C-13-enrichments in a single scan. Their advantages and limitations are discussed, and their high analytical potentialities are highlighted. Both methods are characterized by an accuracy of 1-2%, an average precision of 3%, and an excellent linearity. The analytical performance is equivalent or better than any of the conventional methods previously reported. The two ultrafast experiments are applied to the measurement of C-13-enrichments on a biomass hydrolyzate, showing the first known application of ultrafast 2D NMR to a real biological extract. The experiment duration is divided by 200 compared to the conventional methods, while preserving 80% of the quantitative information. This new approach opens new perspectives of application for fluxomics and metabonomics.