Reduction of dissolved oxygen in semen extender with nitrogen gassing reduces oxidative stress and improves post-thaw semen quality of bulls

The objective of the present study was to investigate the effects of two different concentrations of dissolved oxygen (DO, 4 and 8) ppm in the extender on oxidative stress affecting plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), and deoxyribonucleic acid (DNA) damage of bull spermatozoa following cryopreservation. For the experiment, nitrogen (N2) gassing of the extender for varied time intervals yielded extender with DO concentration of 4 ppm and 8 ppm (Groups II and III, respectively). For the Control (Group I) without N2 gassing, a DO concentration of 11.7 ppm was recorded. Following sample selection, ejaculates were divided into three aliquots and were extended to have 80 × 106 spermatozoa/mL of extender in the three groups. Semen samples were evaluated for reactive oxygen species (ROS), lipid peroxidation (LPO), total antioxidant capacity (TAC) and superoxide dismutase (SOD) at the fresh, pre-freeze, and post-thaw stages. Evaluation of PMI, MMP, and DNA damage were conducted on frozen-thawed samples. There were greater (P < 0.05) increase in ROS and LPO and decrease in TAC concentrations in Group I than Groups II and III. Mean values of SOD at the post-thaw stage was greater (P < 0.05) in Group II than Group I. There was a similar trend in the PMI in Groups II and III; MMP and DNA integrity in Group II was greater compared with Group I. In conclusion, results indicate there was a beneficial effect of maintaining DO concentrations at 4 rather than of 8 or 11.7 ppm in extender for sustaining post-thaw semen quality.