Bacterial Substrate Transformation Tracked by Stable-Isotope-Guided NMR Metabolomics: Application in a Natural Aquatic Microbial Community

The transformation of organic substrates by heterotrophic bacteria in aquatic environments constitutes one of the key processes in global material cycles. The development of procedures that would enable us to track the wide range of organic compounds transformed by aquatic bacteria would greatly improve our understanding of material cycles. In this study, we examined the applicability of nuclear magnetic resonance spectroscopy coupled with stable-isotope labeling to the investigation of metabolite transformation in a natural aquatic bacterial community. The addition of a model substrate ((13)C(6)–glucose) to a coastal seawater sample and subsequent incubation resulted in the detection of >200 peaks and the assignment of 22 metabolites from various chemical classes, including amino acids, dipeptides, organic acids, nucleosides, nucleobases, and amino alcohols, which had been identified as transformed from the (13)C(6)–glucose. Additional experiments revealed large variability in metabolite transformation and the key compounds, showing the bacterial accumulation of glutamate over the incubation period, and that of 3-hydroxybutyrate with increasing concentrations of (13)C(6)–glucose added. These results suggest the potential ability of our approach to track substrate transformation in aquatic bacterial communities. Further applications of this procedure may provide substantial insights into the metabolite dynamics in aquatic environments.