BACKGROUND: Quercetin is a flavonoid with high reactive oxygen species (ROS) scavenging and ion chelating activity. It also enhances the activity of antioxidant enzymes and reduces enzymatic activity such as NADPH oxidase and NADH-dependent oxido-reductase. Tempol, as a superoxide dismutase mimetic agent, converts superoxide to less toxic hydrogen peroxide (H2O2), but cannot reduce highly toxic hydroxyl radicals in Fenton or Haber-Weiss reactions mediated with free iron or cupper. OBJECTIVE The study was to compare the effect of Quercetin and Tempol in an optimized commercial cryo-protective media on ROS induced cryoinjury for the first time. METHODS Following administration of these compounds during freezing process, sperm motility, viability, ROS production and DNA integrity were assessed before and after freezing/thawing process. RESULTS Data showed that 10 µM Quercetin and 5 µM Tempol significantly improved sperm motility and viability, but they together had no additive effect. Supplementation with Quercetin alone or combination of Quercetin with Tempol decrease the ROS concentration, but the reduction was not significant for Tempol alone compared to control group. Quercetin and Tempol significantly decrease DNA fragmentation. CONCLUSIONS The supplementation of Quercetin or Tempol, but not their combination improves the quality of cryopreserved human semen.