The aim of this study was to evaluate the effect of date seed oil (DSO) on epididymal sperm characteristics and testicular antioxidant enzymes in male mice. DSO was diluted into isotonic saline solution (0.9%) and different doses (5, 10, 15 and 20%) were prepared. Fifty male mice were divided into five groups; in four groups DSO was given by intraperitoneal injection of oil solution for 28 days. The control group was injected by isotonic saline solution without DSO. Body and reproductive organ weights, sperm characteristics (count, motility, viability and morphology) were assessed. In addition, levels of malondialdehyde (MDA), activities of superoxide dismutase (SOD) and catalase (CAT) were investigated in testes. A significant increase in sperm count, motility and viability of all treated animal groups was observed when compared with the control group (P < 0.05). Unlike, the percentage of abnormal sperm was significantly lower in all treated groups than in the control group (P < 0.05). A significant decrease in MDA levels and marked increase in SOD and CAT activities in mice treated with high doses of DSO (15 and 20%) were also noted. We suggest that DSO can improve the epididymal sperm quality and could ameliorate the testicular strategy defences.