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Objective: To evaluate the effects of reduced glutathione on the quality of cryopreserved Boer buck spermatozoa. Methods: The current study was carried out on five Boer bucks from which semen samples were collected by artificial vagina. After microscopical evaluation at 37 ℃, semen samples that fulfill the ideal requirements for extension were diluted in a tris–based extender including different concentrations of reduced glutathione (2, 5, 7 and 10 mM) and those without glutathione served as a control. Sperm motility, viability, acrosome integrity, DNA integrity, total antioxidant capacity and lipid peroxidation were assessed post-thawing. Results: The current results revealed that post-thawing motility, viability and acrosomal integrity were significantly improved [(66.67±5.50)%, 168.30±18.59 and (12.75±2.45)%, respectively] when 5 mM glutathione was added to semen extender; especially as compared with the control [(40.00 ±2.88)%, 95.00±8.90 and (25.75±3.46)%, respectively]. Similarly, at this concentration (5 mM) sperm DNA damage, tail length and tail moment of cryopreserved semen were significantly (P<0.05) reduced [(2.32±0.27)%, (1.64±0.49) μm and 3.55±0.63, respectively] compared with the control extender [(6.66±0.84)%, (4.09±0.47) μm and 26.47±0.51, respectively]. Moreover, addition of 5 mM glutathione to buck semen extender significantly (P<0.05) increased total antioxidant capacity [0.51±0.07) mμ/mL] and decreased lipid peroxidation of cryopreserved spermatozoa [(8.68±2.72) nmol/mL] compared with the control [(0.18±0.02) mμ/mL and (24.92±5.80) nmol/mL, respectively]. Conclusions: The addition of 5 mM glutathione to semen diluent improve freezability of Boer buck spermatozoa through DNA protection from deterioration and oxidative stress reduction. Moreover, 10 mM of glutathione exerts cytotoxic effects on Boer buck semen.