Abstract: In order to study the effect of punicalagin on the sheep semen quality, semen samples of Duolang sheep were stored at room temperature after being diluted in Tris-glucose-fructose-based extender with different concentrations（0, 10.0, 12.5, 15.0, 17.5, 20.0 μmol/L） of punicalagin, and used for evaluating the sperm viability, motility, acrosome integrity, plasma membrane integrity, and antioxidant superoxide dismutase（SOD） activity and catalase（CAT） content in the semen after 24, 48 and 72 h, respectively. The results showed that at room temperature, The sperm motility（55.95%）, motility（42.82%）, acrosome integrity（48.61%）, plasma membrane integrity（44.40%）, CAT activity（4.61 U/mL） and SOD activity（62.70 U/mg） were the highest in the semen samples diluted with extender containing 15.0 μmol/L punicalagin after 72 hours, which were significantly higher than those in the control group（P＜0.05）. In conclusion, the quality of Duolang sheep semen stored at room temperature could be improved when diluted with Tris-glucose-fructose-based extender containing 15.0 μmol/L punicalagin.