Motility and fertility of bull sperm in whole milk extender containing antioxidants

Bull sperm are exposed to aerobic conditions during processing before freezing, and they have little endogenous antioxidant to protect them against reactive oxygen species that may be present. Seventeen laboratory studies and two field trials were conducted with 174 semen collections from bulls in an artificial breeding cooperative. More than 250 combinations and concentrations of reduced glutathione (GSH), superoxide dismutase (SOD), ascorbic acid, hypotaurine (HPT), 2,2,6,6-tetramethylpeperidine-1-oxyl (Tempo) and 4-hydroxy-2, 2, 6, 6-tetramethylpeperidine (Tempol) were tested by adding these compounds to fresh semen, and to a whole milk (WM) glycerol extender. Semen packaged in straws in the WM extender was frozen with liquid nitrogen. The motility of frozen-thawed sperm during storage at 25 or 5 °C after freezing was compared with semen stored without freezing. Antioxidants generally were not beneficial, except the percentage of motile sperm was improved by 6–11% units (P<0.05) when sperm were stored unfrozen or after freezing when 0.5 mM of GSH with or without SOD was added. In two field trials, non-return rates were 71.9, 69.5 and 70.9% (P>0.05) with WM containing 0.0, 0.5 and 1.0 mM of GSH, respectively, and 74.0 and 73.9% with WM and WM plus 0.5 mM of GSH and 100 U/ml of SOD (P>0.05). WM contains an abundant supply of casein which is an antioxidant, and additional antioxidants were ineffective in improving motility of sperm immediately after freezing and thawing or in affecting fertility. However, sperm responses were different in egg yolk-Tris extender. Sperm in this egg yolk extender tolerated substantial concentrations of Tempo and Tempol compared with toxic effects in WM (P<0.05). Therefore, optimal combinations of antioxidants tested here may have more useful applications in organizations using an egg yolk-based semen extender.