Objective To investigate the effect of pancreatic kininogenase on myocardial injury, oxidative stress, and fibrosis in rats with myocardial ischemia-reperfusion injury. Methods SD rats were divided into sham, model and drug- treated groups. The heart rate (HR), left ventricular systolic pressure (LVSP), and short-axis contraction rate (FS) of rats were measured in each group. HE staining was used to detect myocardial pathological lesions. ELISA were used to detect the contents of creatine kinase isoenzyme (CK-MB), myoglobin (Mb) and cardiac troponin I (cTnI). Assay kits were used to detect the contents of superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione (GSH). Western blot was used to assay the expression levels of mitochondrial pathway apoptosis-related and fibrosis marker proteins. Masson staining was used to observe myocardial tissue fibrosis. RT-PCR was used to assay the expression levels of α-smooth muscle actin (α- SMA), transforming growth factor-β1 (TGF-β1) and fibronectin (FN). Results Compared with the model group, HR, LVSP and FS in 3 and 6 U/ kg PKase groups were significantly increased (P<0.05), pathological damage of myocardial tissue was significantly improved, the contents of CK-MB, Mb, MDA and cTnI were significantly reduced (P<0.05), SOD and GSH contents were increased (P<0.05), the degree of myocardial fibrosis was significantly improved, Bax / Bcl-2, cas-9, cas-3, α-SMA, TGF-β1 and FN levels were reduced significantly (P< 0.05). Conclusions Pancreatic kininogenase relieves myocardial injury, oxidative stress and fibrosis in rats with myocardial ischemia-reperfusion injury.