Effects of moderate physical activity (90 min at 45-50% of maximal O₂ uptake 2 times daily) and 'high, (2.5 g protein · kg^-1 · day^-1, n = 6) or 'normal' protein intake (1.0 g protein · kg^-1 · day^-1, n = 8) on the pattern and rate of 24-h macronutrient utilization in healthy adult men were compared after a diet-exercise-adjustment period of 6 days. Energy turnover (ET) was determined by indirect and direct (suit) calorimetry, and 'protein oxidation' was determined by a 24-h continuous intravenous infusion of [1- ¹³C]leucine. Subjects were in slight positive energy balance during both studies. Protein contributed to a higher (22 vs. 10%) and carbohydrate (CHO) a lower (33 vs. 58%) proportion of total 24-h ET on the high- vs. normal- protein intake. The highest contribution of fat to ET was seen postexercise during fasting (73 and 61% of ET for high and normal, respectively). With the high-protein diet the subjects were in a positive protein (P < 0.001) and CHO balance (P < 0.05) and a negative fat balance (P < 0.05). The increased ET postexercise was not explained by increased rates of urea production and/or protein synthesis.