The oxygen flux into the mitochondria of skeletal muscle increases with exercise. However, the extent of oxidative damage to mitochondrial proteins of skeletal muscle has only been estimated. We studied the alteration of reactive carbonyl derivatives (RCD) in mitochondrial and cytosolic fractions of skeletal muscle following 9 weeks of swimming training in rats. The RCD content of mitochondria was significantly elevated compared with the cytosolic fraction of both control and exercised animals. Accumulation of RCD in muscle mitochondria of the exercised group was also significantly elevated (P < 0.05). On the other hand, alteration of the accumulation of RCD was not apparent in the cytosolic fraction of skeletal muscle. The activity of proteasome complex, however, was increased in the cytosolic fraction of exercised muscle (P < 0.05). The data suggest that mitochondria of skeletal muscle accumulate significantly larger amounts of RCD than the cytosolic fraction and the tendency of the accumulation varies in cell fractions. Exercise training increases the accumulation of protein damage in mitochondria of skeletal muscle but cytosolic proteins are protected by increased activity of proteasome complex and possibly by other antioxidant enzymes.