Sustained military operations (SUSOPs) are associated with performance decrements and cognitive dysfunction. β‐Alanine (BA) supplementation may have a role in increasing soldier resiliency by enhancing muscle‐buffering capacity and reducing oxidative stress. The purpose of this study was to examine the effects of BA on physical performance, cognition, endocrine function, and inflammation during a 24 h simulated SUSOP. Nineteen males were randomized into one of two groups: BA (n = 10) or placebo (n = 9; PLA) (12 g/day) for 14 days preceding the 24 h SUSOP. Assessments were performed at 0 h (0H), 12 h (12H), and 24 h (24H) during the SUSOP. No changes in visual tracking ability, jump power, or upper‐body muscular endurance were observed between groups or time points (P's > 0.05). Increases in subjective feelings of soreness and fatigue were noted at 12H compared to 0H (P < 0.05) in PLA, but not in BA. Visual reaction time for PLA was slower at 24H compared to 0H (P = 0.035), and PLA made more errors on reaction time testing at 12H compared to BA (P = 0.048), but motor reaction time was faster (P = 0.016) for PLA. Simulated litter carry and 1 km run completion times increased at 24H compared to 0H in both groups (P < 0.05), however, PLA had a longer 1 km time compared to BA at 24H (P = 0.050). Increases in inflammatory and endocrine markers were observed over the SUSOP, with no differences between groups. BA supplementation appears to maintain some aspects of cognition and physical performance during a 24 h SUSOP, with no effects on endocrine function or inflammation.