Introduction: Pre-clinical evidence suggests that nicotinamide adenine dinucleotide (NAD+) precursors can improve disease-associated mitochondrial dysfunction, however, clinical studies using the precursor nicotinamide riboside have so far failed to recapitulate these mitochondrial-enhancing benefits. Nicotinic acid (NA) is an alternative NAD+ precursor, which enters the NAD+ pool via a different metabolic pathway to nicotinamide riboside, and has proven effective at enhancing mitochondrial function in diabetics. However, the efficacy of NA on skeletal muscle mitochondrial function in ageing muscle remains to be determined. Aim: Using a battery of mitochondrial-focused measurements, we investigated the efficacy of NA supplementation on skeletal muscle and mitochondrial function in healthy, physically inactive, older (65-75 years) males. Methods: In a double-blind, randomised, placebo-controlled fashion, eighteen older males were randomly assigned to 2-weeks of NA analogue supplementation (250 mg acipimox thrice daily, n=8) or placebo supplementation (n=10). Pre, during (week 1) and post (week 2) supplementation, (an)aerobic muscle function was measured via moderate incremental and ramp incremental cycling, 10-second Wingate cycling and the Short Physical Performance Battery Test (SPBBT). Muscle biopsies obtained from the m. vastus lateralis were used to quantify mitochondrial function via high resolution respirometry, mitochondrial content via citrate synthase, electron transport chain content via immunoblotting and mitochondrial and myofibrillar muscle protein synthesis via gas-chomotography-pyrolysis-isotope ratio mass spectrometry. Volunteers provided written informed consent prior to participation and all study procedures were approved by the South West Frenchay Research Ethics Committee (16/SW/0099). Results: NA supplementation reduced respiratory exchange ratio during moderate and ramp exercise (p0.05). Anaerobic minimum power, determined by Wingate, increased from week 1 to week 2 in NA supplemented volunteers (p0.05). NA supplementation tended to increase mitochondrial respiration (p=0.10), with coupled respiration through complex I increased at week 2 and maximal uncoupled respiration increased at week 1 and 2, compared to baseline. NA supplementation also tended to increase citrate synthase (p=0.10) but had no significant impact on electron transport chain content (p>0.05), mitochondrial or myofibrillar fractional synthesis rate (p>0.05). Conclusion: NA supplementation alters substrate utilisation towards fat oxidation and may improve mitochondrial function/content in older healthy adults, however due to the limited n, larger clinical trials are needed to substantiate these findings.