The effects of acute endurance running on the metabolic profiles of rat skeletal muscle were studied. Male Wistar strain rats were continuously run on a treadmill for 1 h (speed, 35 m/min; grade, 0 degrees). Soleus (SOL) and extensor digitorum longus (EDL) were removed after 30-min running, and a 0, 1, 6, 24, 48, and 72 h post-exercise, and enzymes activity (CK, LDH, PFK, PK, SDH, and MDH) and substrates contents (glycogen and pyruvate) were measured biochemically. The time course of the enzyme activities showed two distinct patterns: CK, LDH, SDH, and MDH showed two peaks, at 0 and 24 h post-exercise, while PFK and PK showed one peak at 0 h post-exercise. The activities of glycolytic enzymes and CK in EDL and oxidative enzymes in SOL showed marked changes after exercise. The glycogen level was lowest at 0 h postexercise in both muscles and recovered to resting level by 24 h postexercise. Pyruvate increased with running and showed the highest value at 1 h post-exercise. Increased oxidative capacity of skeletal muscle in response to the acute endurance exercise dropped gradually to the resting level by 48 h post-exercise. An endurance exercise may induce a flexible adaptation on the oxidative capacity within skeletal muscle. We conclude that the respective time course of the enzyme activities must be considered when discussing metabolic changes that occur with acute endurance exercise.