An evaluation of cryoprotective compounds on bovine spermatozoa

The potentially cryoprotective properties of 72 higher-molecular-weight polymeric additives and 69 low-molecular-weight compounds were evaluated. The polymeric compound selection was based on solubility in semen extender, toxicity and finally, on the cryoprotective effect on bull spermatozoa as measured by progressive motility. Five compounds showed cryoprotection to the cell, but with no significant improvement over that of TESNaK yolk, TEST yolk, or TEST yolk glycerol extenders used as controls. Low-molecular-weight compounds were selected on the basis of colligative properties particularly that of freezing-point depression. Elimination was based on precipitation of proteins in the extender, toxicity, and cryoprotection to bovine spermatozoa as measured by progressive motility. Nineteen compounds that yielded protection during cryopreservation of bovine spermatozoa were compared using post-thaw motility and membrane integrity using glutamic-oxaloacetic transaminase enzyme retained in the spermatozoa after freezing as an indicator. Semen was diluted with extender containing selected compounds at 35 or 5 °C to determine the effect of temperature at which the cryoprotective compound was added. Glycerol yielded the highest recovery. Diethylene glycol, dimethylsulfoxide, N-methylacetamide, and triethylene glycol appeared not to be different in freezing bovine spermatozoa. The temperature or method of addition of cryoprotective compound did not reveal a significant difference.