The purpose of this study was to examine the effect of different rutin concentrations on rams epididymal sperm. A local slaughterhouse provided 50 pair of testes from 25 rams. The testes were sent to the lab at room temperature. Spermatozoa were extracted by suspending portions of cauda epididymis in tris solution. Ram sperm was cryopreserved (in liquid nitrogen) in a tris extender containing rutin at 0, 0.5, 0.75, 1, and 1.25 mM. Rutin showed superior sperm total and progressive motility, beat cross frequency, straight line velocity, velocity average pathway and membrane integrity values at 0.75 and 1 mM. The morphology of the sperm and the superoxide dismutase levels did not significantly change with different treatments. Moreover, rutin at 0.75 and 1 mM was also shown to have the highest level of mitochondrial activity. The results showed ATP, total antioxidant capacity, and glutathione peroxidase levels were significantly greater in the rutin 0.75 and 1 mM groups (P< 0.05). Rutin at 0.75 and 1 mM levels had the lowest reactive oxygen species concentrations. Rutin at 0.75 and 1 mM substantially increased the proportion of viable sperm (P< 0.05). The lowest amount of apoptosis was observed in 0.75 and 1 mM rutin. Rutin at 0.75 and 1 mM yielded the least significant percentage of dead sperm. It may be inferred that adding 0.75 and 1 mM to the sperm extender can enhance the quality of the epididymal sperm in rams after the cryopreservation process.