This study compared measured serum [Na⁺] (S_[Na⁺]) with that predicted by the Nguyen-Kurtz equation after manipulating ingested [Na⁺] and changes in body mass (∆BM) during prolonged running in the heat. Athletes (4 men, 4 women; 22-36 yr) ran for 2 h, followed by a run to exhaustion, and 1 h recovery. During exercise and recovery, subjects drank a 6% carbohydrate solution without Na⁺ (Na⁺0), 6% carbohydrate solution with 18 mmol/L Na⁺ (Na⁺18), or 6% carbohydrate solution with 30 mmol/L Na⁺ (Na⁺30) to maintain BM (0% BM), increase BM by 2%, or decrease BM by 2% or 4% in 12 separate trials. Net fluid, Na⁺, and K⁺ balance were measured to calculate the Nguyen-Kurtz predicted S_[Na⁺] for each trial. For all beverages, predicted and measured S_[Na⁺] were not significantly different during the 0%, -2%, and -4% BM trials (-0.2±0.2 mmol/L), but were significantly different during the +2% BM trials (-2.6±0.5 mmol/L). Overall, Na⁺ consumption attenuated the decline in S_[Na⁺] (-2.0±0.5, -0.9±0.5, -0.5±0.5 mmol/L from pre- to post-experiment of the 0% BM trials for Na⁺30, Na⁺18, and Na⁺]0, respectively) but the differences among beverages were not statistically significant. Beverage [Na⁺] did not affect performance; however, time to exhaustion was significantly shorter during the -4% (8±3 min) and -2% (14±3 min) vs. 0% (22±5 min) and +2% (26±6 min) BM trials. In conclusion, when athletes maintain or lose BM, changes in S_[Na⁺] can be accurately predicted by changes in the mass balance of fluid, Na⁺, and K⁺ during prolonged running in the heat.