NAD+ and NADH act as coenzymes in metabolic reactions. The reduction of NAD+ to NADH is linked with generation of ATP through glycolysis and oxidative phosphorylation. The consumption of NAD+ by various signaling proteins regulates protein modification, cell fate and survival. Therefore, NAD+ and NADH measurements have the potential to inform about tissue energetics and health. Recently, some investigators have suggested that NADH and NAD+ may be detected in human muscle using 31-phosphorus MRS. However, the utility and reliability of this measure is not clear. The goals of this project were to 1) determine whether the NADH+NAD peak can be resolved in human skeletal muscle at 3T, 2) compare peak resolution with and without a decoupling technique, and 3) evaluate the reliability of this measure. Interpretation of these data and their potential for studying alterations in NAD+ and NADH homeostasis in human muscle remain to be determined.