A Triple-Resonance NMR Strategy for the Selective Detection of NAD+ and NADH Derived from a 13C/15N-Nicotinamide Riboside Probe in the Liver Extracts of Mice

Alterations in the ratio of oxidized and reduced nicotinamide adenine dinucleotide (NAD+/NADH) reflect the intracellular redox state and have been implicated in a broad spectrum of pathological conditions, including neurogenetic disorders, heart failure, and liver diseases. In the present study, we demonstrate the selective detection of probe-derived NAD+ and NADH signals in mouse liver extracts by means of a triple-resonance nuclear magnetic resonance (NMR) spectroscopy technique. We prepared 13C/15N-enriched nicotinamide riboside (13C/15N-NR), which undergoes enzymatic conversion to NAD+ and NADH in the liver, and detected these labeled metabolites by 1H–{13C–15N} triple-resonance NMR measurements. This study provides a methodological proof-of-concept for the selective detection of NAD-related signals derived from a stable-isotope labeled probe in mouse liver extracts.