Objective: We previously showed that brief intermittent ischemia applied during the onset of reperfusion (i.e., postconditioning) is cardioprotective in a canine model of ischemia–reperfusion. This study tested the hypothesis that the early minutes of reperfusion (R) during which postconditioning (Post-con) is applied are critical to its cardioprotection. Methods: In anesthetized open-chest rats, the left coronary artery (LCA) was occluded for 30 min and reperfused for 3 h. All rats were randomly divided into six groups: Control ( n =8): no intervention at R; Ischemic preconditioning (IPC) ( n =8): the LCA was occluded for 5 min followed by 10 min of R before the index occlusion; Post-con 1 ( n =8): after LCA occlusion, three cycles of 10 s R followed by 10 s LCA re-occlusion were applied during the first minute of R; Post-con 2 ( n =8): Six cycles of 10 s R and 10 s re-occlusion were applied during the first 2 min of R; Delayed Post-con ( n =8): the ligature was loosened for full reflow for the first minute of R, after which the three-cycle Post-con algorithm was applied; Sham ( n =6): the surgical procedure was identical to other groups, but the LCA ligature was not ligated. Results: Infarct size (TTC staining) was 23% smaller in Post-con 1 (40±2%*) than in Control (52±3%), confirmed by plasma creatine kinase activity (18±2* vs. 46±6 IU/g protein). There was no further reduction in infarct size with 6 cycles of Post-con (40±2.9%, p >0.05 vs. Post-con 1). Meanwhile, infarct size reduction was significantly greater in the IPC group (17±3%) than in Post-con1 ( p <0.01). The plasma lipid peroxidation product malondialdehyde (MDA, μM/ml) was less after R in IPC and Post-con 1 (0.8±0.07* and 0.8±0.06*) vs. Control (1.21±0.08), consistent with a visual decrease in superoxide anion generation (dihydroethidium staining) in the AAR myocardium after 3 h of reperfusion. Neutrophil accumulation (myeloperoxidase activity, MPO, U/100 g tissue) in the AAR was less in IPC (1.4±0.3*) and Post-con 1 (2.5±0.3*) vs. Control (5.5±0.6). The reductions in infarct size, creatine kinase, MDA and DHE staining were lost with delayed Post-con, while MPO activity remained lower than in Control (3.2±0.4*). Conclusions: (1) Post-con at onset of R reduces myocardial injury; (2) cardioprotection may be mediated, in part, by inhibiting oxidant generation and oxidant mediated injury; (3) the first minute of R in the rat model is critical to cardioprotection by Post-con; and (4) cardioprotection by Post-con may be independent of neutrophil accumulation in AAR. * p <0.05 Post-con vs. Control.