Two-dimensional electrophoresis was used to generate the aqueous protein expression patterns of rat extensor digitorum longus muscle (EDL, fast twitch muscle) and solues muscle (SOL, slow twitch muscle) of different ages. Two specific protein spots S1 and S3, were identified from EDL muscles at the ages of 12 and 18 months onward respectively. In the EDL muscles of aged rat (24 months) after intensive exercise training, S3 was still detected while S1 disappeared. In addition, diaphragm muscle (DIA, fast twitch muscle), which retains physically active throughout the life span, was used as nondisuse control. The results showed that the expressions of S1 and S3 in 24-month DIA muscle were identical with the trained aged EDL muscle. It is suggested that exercise might delay the onset of S1 expression. However, the expression of S3 over age seemed to be progressive and exercise independent. Another protein spot, S2 was identified to express only in young EDL and SOL muscles, but its expression decreased over age. Furthermore, exercise has no effect on S2 expression since S2 could not be detected in aged DIA as well as trained aged EDL and SOL muscles. These results indicated that aqueous protein expression patterns of skeletal muscle undergo changes during aging. Some of these changes such as S2 and S3 appear progressively, and some such as S1 could be delayed by exercise. S3 was identified as ubiquitin, which might play an important role in protein degradation during skeletal muscle aging process.