This study aimsto evaluate the efficacy of fresh pineapple juice and bromelain consumption on acetic acid-induced ulcerative colitis in rats. Furthermore the study aims to determine the amount of bromelain in fresh pineapple juice by HPLC as well as some of the bioactive constituents of fresh pineapple juice. Sixty male albino rats were divided into six groups; each group consisted of 10 rats. Group (1) served as normal control (Healthy) and group (4) served as positive control (Colitis). Healthy rats of group (2) and colitis rats of group (5) treated with 0.5ml fresh pineapple juice/100g body weight by stomach tube as well as healthy rats of group (3) and colitis rats of group (6) treated with 0.5ml bromelain/100g body weight daily using stomach tube. The results of chemical analysis of pineapple revealed that each 100ml of the fresh prepared pineapple juice contains 425±3.00mg as gallic acid equivalent (GAE) for total phenols, 584±4.00mg as catechin equivalent (CE) for total flavonoids, 25±1.00 mg for ascorbic acid and 57±1.00% as ascorbic acid equivalent (AAE) for total antioxidant capacity. Induction of ulcerative colitis by acetic acid (0.25ml of 16% acetic acid/ 100g body weight) markedly increased the oxidative stress which caused significant reduction in nonenzymatic antioxidant such as blood reduced glutathione (GSH), enzymatic antioxidants as serum catalase (CAT) and colon superoxide dismutase (SOD) activities, also resulted in increased lipid peroxidation (LPO) of membranes as colon malondialdehyde (MDA) was significantly increased. Furthermore, serum inflammatory marker levels as tumor necrosis factor-α (TNF-α), myeloperoxidase (MPO) activity and C-reactive protein (CRP) were significantly increased in colitis rats (p>0.05). While treatment of colitis rats by fresh pineapple juice or bromelain ameliorated the oxidative stress status and decreased inflammatory markers which alleviated the colitis. The microscopic examination of colon sections illustrated the modulating effect of both fresh pineapple juice and bromelain on colon cells. In conclusion, fresh pineapple juice had a better effect than bromelain as its high content of antioxidants as well as bromelain.