The objective of this study was to elucidate the mechanism of beef tenderization. Purified μ-Calpain, calpastatin, myofibrils and leupeptin were used in six different reaction mixtures. Myofibrillar SDS-PAGE and western blotting analysis were conducted after being incubated for a period of time in mixed salt solution(MSS) formulated on the basis of the post rigor condition. The results showed that desmin and troponinT were degraded by μ-Calpain into smaller fragments which were similar to the polypetides degraded by aged beef, while calcium(100 μM) alone had no effects on myofibrils. It was also found that Calpastatin was not capable of completely inhibiting μ-Calpain activity and no degradation occurred for actin. It is concluded that cytoskeletal proteins were degraded by μ-Calpain during aging process, which might be the major reason for improving beef tenderness. Nevertheless, there was little contribution of Lysosomal cathepsins to beef tenderness at early period of aging.