Myocardial ischemia/reperfusion (I/R) injury is the major problem that aggravates cardiac damage. Several established animal models fail to explain the similarity in disease mechanism and progression as seen in humans; whereas guinea pig shows high similarity in cardiovascular parameters. Hence, current study is aimed to develop an animal model using guinea pigs that may best correlate with disease mechanism of human myocardial I/R injury. Male guinea pigs were randomized into three groups: normal diet (ND), high fat diet (HFD) and sham; fed with respective diets for 90 days. Myocardial infarction (MI) was induced by ligating left anterior descending artery (LAD) for 30 min followed by 24 h and 7 days of reperfusion in ND and HFD groups. Electrocardiogram (ECG) showed the alterations in electrical conduction during myocardial I/R injury. Elevated levels of lactate dehydrogenase (LDH) and creatine kinase-MB ((CK-MB)) were higher in HFD compared to ND. Inflammatory markers such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were up-regulated in I/R injury animals compared to sham. Fold change of these protein expression levels were higher in HFD compared to ND. Elevated lipid profile and increased aortic wall thickness in HFD animals depicts the risk of developing cardiovascular complications. ECG analysis strongly confirmed MI through changes in sinus rhythm that are reflected in infarcted tissue as verified through TTC staining. Thus the combination of HFD followed by I/R injury proved to be an efficient model to study pathophysiology of myocardial I/R injury with minimal tissue damage and surgical mortality.