Free Access
Reprod. Nutr. Dev.
Volume 42, Number 3, May-June 2002
Page(s) 217 - 226

Reprod. Nutr. Dev. 42 (2002) 217-226
DOI: 10.1051/rnd:2002020

Comparison between glycerol and ethylene glycol for the cryopreservation of equine spermatozoa: semen quality assessment with standard analyses and with the hypoosmotic swelling test

Roberto Mantovania, Ada Rotab, Maria Elena Falomob, Lucia Bailonia and Leila Vincentic

a  Department of Animal Science, University of Padua, Agripolis, 35020 Legnaro (PD), Italy
b  Department of Veterinary Clinical Sciences, University of Padua, Italy
c  Department of Animal Pathology, Faculty of Veterinary Medicine, University of Turin, Italy

(Received 4 February 2002; accepted 21 May 2002)

The aims of this study were to compare glycerol (G) at customary concentrations and ethylene glycol (EG) as cryoprotectants for stallion semen in a skimmed milk (SM) extender, to test different EG concentrations and to compare the results of manual and computerized analysis with the hypoosmotic swelling (HOS) test. Ejaculates from two stallions were collected over 3 weeks (6 ejaculates per stallion), diluted in a SM based extender, divided into 4 fractions, centrifuged and diluted again to a concentration of 100 $\times$ 10 6 mL -1 progressive motile spermatozoa (PMS) in addition with the cryoprotectant (3% G, 3% EG, 6% EG, 9% EG). Sperm motility was assessed both by microscopy (in raw and frozen-thawed semen immediately after thawing) and with an HTM-IVOS analyzer (Hamilton-Thorne Research, MA, USA), at 0, 1, 4, 6, and 12 h after thawing and storage at 21 °C. Raw and frozen-thawed (0 h) semen samples for G and EG at 3% were also submitted to the HOS test with a 100 mOsm sucrose solution and were evaluated to detect the presence of swollen tails. The higher EG concentrations (i.e. 6 significantly reduced the percentage of motile and PMS, immediately after thawing. At the same concentration, i.e. 3%, G resulted in a higher percentage of PMS than EG (36.2 vs. 30%, P < 0.05), but at 12 h after thawing and storage at 21 °C, no significant differences were detected between G and EG at 3%. The correlations between progressive motility (assessed by direct microscope observation or measured through the HTM analyzer) and the HOS test results for 3%G and EG were r = 0.61 and r = 0.35, respectively. The HOS test confirmed its suitability as a complementary method of analysis for stallion semen. We conclude that with the SM extender used, EG could substitute G as the cryoprotectant for stallion semen if used at the same or lower concentration.

Key words: stallion / semen / cryoprotectant / glycerol / ethylene glycol

Correspondence and reprints: Roberto Mantovani

© INRA, EDP Sciences 2002