Pomegranate seed oil shows positive effects by limiting neutrophil activation and lipid peroxidation through its antioxidant and anti-inflammatory activities. This study evaluated the possible ameliorative effects of pomegranate seed oil, its actions on proinflammatory cytokines, and its antioxidant activity using an acute acetic acid-induced colitis model in rats. 32 male Sprague-Dawley rats were divided into 4 groups: control, colitis, 0.4 ml/kg, and 0.8 ml/kg pomegranate seed oil treatment after colitis. At the end of the experiment, histopathological and biochemical analyses of intestinal tissues and blood were performed. The study revealed that administering different doses of pomegranate seed oil dramatically reduced total oxidant levels, nuclear factor kappa B, proinflammatory cytokines, and myeloperoxidase activity and appreciably reduced colitis injury. These findings suggest that pomegranate seed oil may alleviate colitis symptoms effectively and exert protective effects through antioxidant, anti-inflammatory mechanisms.