Objective — to assess the changes in biochemical ejaculatory sperm membrane specific to apoptotic cells after exposure to different concentrations of H2O2 oxidative stress inducer for various periods of incubation, as well as to evaluate the effect of antioxidants in vitro both separately and in combination to protect male gametes from H2O2-induced damage. Material and methods. Oxidative stress induced by incubating the sperm of fertile men (n=18) to 25 and 200 μM H2O2. As the antioxidant defense cells pre-incubated with ascorbic acid and/or α-tocopherol. Externalization of phosphatidylserine (PS) were detected by staining sperm Annexin-V (AnV+) and propidium iodide (PI+). Results. Only incubation with 200 μM H2O2 for 2 hours leads first to a transition of viable cells into cells that bind only AnV, indicating the breaking of the symmetry of the cell membrane and externalize PS and 4 hours there was a transition from (AnV+) — cells to cells, stained with PI, indicating membrane damage and loss of its integrity. Prior adding ascorbic acid or α-tocopherol alone to sperm resulted in complete protection of spermatozoa from H2O2-induced PS externalization, but the addition of a mixture of antioxidants has the opposite effect. Conclusion. Oxidative stress induced by H2O2 irreversible biochemical changes in the cell membrane, thereby beginning PS expressed on its surface, which leads to death ejaculatory sperm. The direction of the effect of the impact depends on the concentration of H2O2 and the incubation time. Adding to the sperm fertile men ascorbic acid or α-tocopherol does not have any influence on the change in the distribution of PS, but protects the cells against H2O2-stimulated PS externalization.