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Objective: The aim of this study was to examine the effect of rectal ozone on the histopathological healing of the colonic mucosa, tissue oxidative stress, and bacterial translocation in the experimental colitis model. Materials and Methods: Three groups of rats were randomly formed [Group 1: Sham group, group 2: Control group, group 3: Ozone treatment group]. Microscopic and macroscopic scoring were done histopathologically in all groups. Glutathione-s-transferase (GST), superoxide dismutase (SOD), glutathione (GSH), and malondialdehyde (MDA) levels were measured in the colon and liver tissue. Blood cultures were taken for the detection of bacterial translocation. Results: Microscopic damage scores were found as 0.0 (0.0-2.0) in the sham group, 3.0 (3.0-3.0) in the control group, 2.0 (0.0-2.0) in the ozone treatment group (p=0.001). Macroscopic damage scores were found as 0.0 (0.0-1.0) in the sham group, 3.0 (0.0-4.0) in the control group, 0.0 (0.0-1.0) in the ozone treatment group; the scores of ozone treatment and sham groups were found to be statistically different (p=0.004). Compared to the control group, the bacterial translocation in the liver, mesenteric lymph node, portal vein, spleen, and systemic blood was fewer in the ozone treatment group. Statistically significant differences were also observed between the groups’ SOD and GST levels in colon tissue and MDA, SOD, and GST levels in liver tissue. Regarding MDA values in the liver tissue of the groups, it was 1.95±0.43 in Group I, 3.63±0.81 in Group II, and 1.19±0.72 U/mg in Group III (p=0.017). When the liver tissue SOD levels of the groups were examined, it was 8.21±0.76 U/mg in Group I, 4.57±0.58 U/mg in Group II, and 8.62±1.13 U/mg in Group III (p=0.029). When GST values in liver tissue belonging to the groups were examined, 0.35±0.16 in Group 1, 0.23±0.03 in Group II, 0.49±0.13 U/mg in Group III (p=0.036). Conclusion: Rectal ozone application plays a role in increasing the organism’s antioxidant activity and has an effective role in increasing the enzyme activities of antioxidant defenses. In addition, rectal ozone application has a positive effect on wound healing at a histopathological level and reduces bacterial translocation in various tissues.