Effect of compatible solutes and diluent composition on the post-thaw motility of ram sperm

The effect of the compatible solutes proline, glycine betaine and trehalose in Tris-based diluents at varying pH, concentrations of egg yolk or glycerol on the post-thaw motility characteristics and fertility of ram sperm was examined. In addition, the amino acid glycine was compared with proline, glycine betaine and a standard Tris-based diluent. Post-thaw motility was assessed using a Hamilton–Thorn motility analyser. In the presence of glycerol and egg yolk, proline and glycine betaine improved the post-thaw motility characteristics of ram sperm. Regardless of the pH of the diluent at which semen was frozen, the percentage of motile sperm was higher when frozen in the presence of proline or glycine betaine than in their absence, whereas proline and glycine betaine only improved the progressive and rapid percentages of sperm for semen frozen in diluents at pH lower than 7.0. When semen was frozen in the absence of egg yolk or glycerol all the motility characteristics were reduced. Increasing the concentration of egg yolk in the diluent from 5% to 10, 15 or 20% had no effect on the post-thaw motility of sperm. The addition of 27 mM of proline or glycine betaine to the diluent also improved post-thaw motility. However, at a concentration of 81 mM, proline and glycine betaine had a detrimental effect on the percentage of motile sperm. Trehalose had no effect on the motility of sperm frozen in glycerol-containing diluents, but motility was lower after cryopreservation in glycine than in Tris-, proline- or glycine betaine-based diluents. There were no differences in the fertility of sperm frozen in Tris-, proline or glycine betaine diluents after cervical or laparo-scopic insemination of ewes.