Effect of monosaccharides and disaccharides in Tris-based diluents on motility, acrosome integrity and fertility of pellet frozen ram spermatozoa

Abstract

In semen freezing diluents, sugars may be incorporated as metabolites, as cryoprotectants or to maintain diluent osmolality. This study examined the cryoprotective effects of mono- and disaccharides in Tris-glucose-egg yolk-glycerol based diluents on the post-thawing motility, acrosome integrity and fertility of pellet frozen ram spermatozoa. In Experiment 1, five concentrations of Tris (300, 250, 200, 150, 100 mM) and five concentrations of glucose (30, 75, 120, 165, 210 mM) were incorporated into freezing diluents. There was an interaction between Tris and glucose concentration on post-thawing motility of spermatozoa ( P < 0.001), and the best Tris:glucose combinations (300:30, 250:75, 200:120, 150:165 and 100:210 mM) were isotonic (330 mosmol). These optimal combinations were used in Experiment 2 with either fructose, glucose, lactose, sucrose or trehalose as the sugar component of the diluent. There was no effect of Tris:sugar combination or sugar type on post-thawing motility of spermatozoa. Glucose, sucrose and trehalose were further examined in combinations with Tris of 300:30, 200:120, 100:210 (Tris:sugar) and glycerol (0, 2.5, 5.0% v/v) in the diluent in Experiment 3. Recovery of spermatozoa was better in diluents containing glycerol than without ( P < 0.001). In diluents without glycerol, motility was higher in the presence of sucrose or trehalose than glucose ( P < 0.01). In Experiment 4, the five optimal Tris:sugar combinations were examined using glucose alone in diluents containing 5.0% v/v glycerol, but there was no effect of Tris:glucose concentration on post-thawing motility, acrosome integrity or fertility of spermatozoa following intrauterine and cervical insemination. We conclude that a range of sugar concentrations (30–210 mM) may be successfully incorporated into ram semen freezing diluents.