We studied the effects of preexercise meal composition on metabolic and performance-related variables during endurance exercise. Eight well-trained cyclists (maximal oxygen uptake 65.0 to 83.5 ml . kg-1 . min-1) were studied on three occasions after an overnight fast. They were given isoenergetic meals containing carbohydrate (CHO), protein (P), and fat (F) in the following amounts (g/70 kg body wt): high-carbohydrate meal, 215 CHO, 26 P, 3 F; high-fat meal, 50 CHO, 14 P, 80 F. On the third occasion subjects were studied after an overnight fast. Four hours after consumption of the meal, subjects started exercise for 90 min at 70% of their maximal oxygen uptake, followed by a 10-km time trial. The high-carbohydrate meal compared with the high-fat meal resulted in significant decreases (P < 0.05) in blood glucose, plasma nonesterified fatty acids, plasma glycerol, plasma chylomicron-triacylglycerol, and plasma 3-hydroxybutyrate concentrations during exercise. This was accompanied by an increase in plasma insulin (P < 0.01 vs. no meal), plasma epinephrine, and plasma growth hormone concentrations (each P < 0.05 vs. either of the other conditions) during exercise. Despite these large differences in substrate and hormone concentrations in plasma, substrate oxidation during the 90-min exercise period was similar in the three trials, and there were no differences in performance on the time trial. These results suggest that, although the availability of fatty acids and other substrates in plasma can be markedly altered by dietary means, the pattern of substrate oxidation during endurance exercise is remarkably resistant to alteration.