HomePhilippine Journal of Veterinary Medicinevol. 40 no. 1 (2003)

Ethylene Glycol as a Cryoprotectant for the Freezing of Buffalo Spermatozoa: I. Effects of Concentration and Equilibration Time on Post-Thaw Survival

Conrado A. Valdez | Jose Arceo N. Bautista | Alan S. Del Rio | Jesus Augustus Cesar G. Flor | Eduardo B. Torres

Discipline: Veterinary Medicine



This study examined the post-thaw survival of frozen buffalo spermatozoa using two different concentrations of ethylene glycol and three different equilibration time. Semen was collected from the three Murrah buffalo bulls using an artificial vagina. The ejaculate was divided into three, which were then diluted with Tris egg-yolk extender with 7% glycerol and 7% or 10% ethylene glycol. Semen extended with 7% and 10% ethylene glycol were equilibrated for 15-20 minutes, 2 hours or 4 hours. Semen extended with 7% glycerol was equilibrated for 4 hours. These were packaged in 0.5 ml French straws during their equilibration period, after which they were suspended in liquid nitrogen vapor for 7 minutes. The straws were submerged in liquid nitrogen and were evaluated for percent motility after a week of storage in liquid nitrogen tank. Post-thaw survival results show that semen frozen with 7% ethylene glycol was better (p< 0.005) than those frozen with 10% ethylene glycol. However, post-thaw survival produced by 7% or 10% ethylene glycol were significantly lower (p<0.005) than using 7% glycerol. Results further show that a short equilibration time seems to have no detrimental effect on post-thaw survival with the use of ethylene glycol as cryoprotectant regardless of concentration. By shortening the equilibration time, the use of ethylene glycol may save time in processing bubaline semen, which in routine procedure took less than 6 hours to complete. It was also observed, that there was a significant variation in survival of spermatozoa among individual bulls. This study demonstrates that ethylene glycol can be used in freezing bubaline spermatozoa. However, there is still a need to study other factors that affect the survival of spermatozoa frozen with the use of ethylene glycol as cryoprotectant.