Background and Aim: Freezing and thawing processes increase the susceptibility of spermatozoa to oxidative stress, which negatively impacts sperm cells functions and reduces its potential fertilizing capacity. The current study aimed to explore the ameliorative effects of various proportions of GTE on post-thawing buffalo bull spermiograms, DNA integrity, and antioxidant activities. Methods: Phytochemical analysis of GTE was done by high-performance liquid chromatography (HPLC). Seventy-two ejaculates were collected from fertile buffalo bulls weekly (n = 12) for 6 weeks, diluted in a tris-based extender supplemented with GTE (0%, 0.1%, 0.5%, 0.75%, 1.0%, and 2.0%) and assessed for sperm motilitiy characterestics, acrosome, plasma membrane and DNA integrities, as well as for the activities of antioxidant enzymes (superoxide dismutase, glutathione reduced, catalase, and total antioxidant activity) and lipid peroxidation (MDA). Results and conclusion: Phytochemical analysis of GTE by high-performance liquid chromatography (HPLC) indicated the presence of five high polyphenol content with high accounting for epigallocatechin-3 gallate and epicatechin. Computer-assisted sperm analysis (CASA) showed the highest total and progressive motility values, and wobble coefficient (WOB) was obtained with 0.75% GTE. At the same time, the distance of average path (DAP) was enhanced with 2.0% GTE compared to the control. Incorporation of 0.75% GTE significantly (P< 0.001) increased  acrosome and plasma membrane integrities, and reduced the percentage of DNA-damage, tail length, and tail moment of frozen-thawed buffalo bull spermatozoa. Activities of antioxidant enzymes (superoxide dismutase, glutathione reduced, catalase, and total antioxidant activity) improved with 0.75% GTE, whereas lipid peroxidation (MDA) was reduced significantly (P˂ 0.001) with 0.5%, 0.75%, and 1.0% GTE compared to control. In conclusion, the addition of 0.75% GTE in buffalo bull semen diluted with a tris-based extender ameliorated the cryodamage influence on sperm parameters as well as reduced DNA damage with optimum antioxidant capacity.