•PM suffered from higher levels of ROS earlier than LT.•The mitochondria swelled and ruptured preferentially in PM compared to LT.•Faster metabolism of ATP-related compounds appeared in PM compared to LT.•The caspase-9 activation was earlier than caspase-3 activation in both muscle types.•MFI may be related to energy metabolism and caspases activities involved in mitochondria. The aim of this study was to investigate the differences in the effects of mitochondria-involved energy metabolism and caspases activation on postmortem tenderness in different muscle fiber types. Beef Longissimus thoracis (LT) and Psoas major (PM) muscles showed significant difference in mitochondrial function. Our data revealed that PM suffered from higher levels of reactive oxygen species (ROS) earlier than LT, causing faster mitochondrial swelling and rupture. Additionally, faster metabolism of ATP-related compounds and activation of caspase-9 appeared in PM, but the activity of caspase-3 in PM was lower than that in LT. Differences in myofibril fragmentation index (MFI) of LT and PM at different aging stages suggested that energy metabolism and caspases activities may play a role in tenderness at different aging stages. These results indicated that oxidative stress-mitochondria-mediated tenderization process could be muscle-specific.