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Liquid or frozen storage of poultry semen negatively affects the sperm motility and has crucial role in reducing fertility. Phospholipids in chicken sperm membrane are composed of high proportions of polyunsaturated fatty acids which are susceptible to lipid peroxidation. One of possible ways to improve semen quality is supplementing the ration with antioxidant compounds such as melatonin. In this study, seventy-two roosters were randomly divided into three equal groups. The first group (control; Group C) was exposed to 14 h light and 10 h darkness. The second group (Group M) was exposed to the same lighting period, but in combination with melatonion supplementation (3 mg/kg body weight daily). The third group (Group L) was exposed to 24 h constant light. Semen was collected using abdominal massage and stored for 6 and 12 h at 5°C. Motility, viability, malondialdehyde (MDA) concentration, fertility, and hatchability were evaluated before and after storage. The results showed that the percentage of viable and motile sperms was significantly (P < 0.05) decreased in the control samples after 6 h storage but in Group M these parameters were significantly (P < 0.05) decreased after 12 h. The MDA concentration was significantly (P < 0.05) decreased in Group M compared with Group C after 6 and 12 h storage. The sperm membrane lipid analysis showed that the percentage of unsaturated fatty acids in Group M was significantly (P < 0.05) lower than in Group C. Fertility and hatchability did not change significantly (P < 0.05) both in M and L Groups compared with C Group. Melatonin administration improved semen quality and decreased lipid peroxidation during liquid storage. It also reduced the percentage of polyunsaturated fatty acids in the sperm membrane lipid composition but it did not affect fertility and hatchability.