Effect of Some Alternative Components of Egg Yolk in Tris-Extender on Sperm Characteristics of Ram Semen Frozen with Two Methods of Packaging Semen

Abstract

This study aimed to evaluate the effect of alternative components of egg yolk in Tris extender and semen packaging methods during cryopreservation on sperm characteristics and antioxidant system in seminal plasma of frozen-thawed semen of Finnish Landrace rams. Semen was collected from 5 sexually matured Finnish rams (50-70 kg LBW and 2-4 years old) by artificial vagina once weekly for 7 weeks. Only semen with mass motility of ≥70% was pooled and diluted with Tris-citric extender containing 15% egg yolk (Tris-EY) or 1% soybean lecithin (Tris-SBL) or 2 mM butylated hydroxytoluene (Tris-BHT).Semen was extended at a rate of 1:5 (semen/extender) with three extender types. After semen extension, semen was placed for cooling in the refrigerator (5 o C) for 4 hours as equilibration period and packaged in 0.25 ml French straws or 0.25 ml pellets in liquid nitrogen. Semen was thawed at 37°C for 30 s. Semen was evaluated after dilution, equilibration and thawing, for progressive motility, livability and abnormality of spermatozoa and curled tail spermatozoa responded to a solution of osmolarity of 75 mOsm for 30 min. The concentration of total antioxidants (TAC), malondialdehyde (MDA) and lactic dehydrogenase (LDH) activity in post-thawed seminal plasma were determined. Results showed that sperm characteristics, including percentages of progressive motility, livability, abnormality and curled tail in post-diluted, post-equilibrated or post-thawed semen were not affected significantly by the type of extender. Progressive motility and curled tail percentages in post-thawed semen were higher (P<0.001; P<0.05) in straws than in pellets. Livability and abnormality percentages were insignificantly better at straws than in pellets. The recovery rate of motility and livability was higher at straws than in pellets. All sperm characteristics indicated insignificant effect of interaction between type of extender and semen packaging method. TAC was higher (P<0.05), while MDA concentration was lower (P<0.05) in Tris-SBL and Tris-BHT than in Tris-EY. The activity of LDH was insignificantly the highest in Tris-BHT than in other extenders. The TAC, MDA concentration and LDH activity in post-thawed semen were not affected significantly by semen packaging method. Effects of interaction between type of extender and semen packaging method on each of TAC, MDA concentration and LDH activity were not significant. The current study concluded the successful usage of Tris-SBL or Tris-BHT in comparing with the possible disadvantages of using egg yolk in Tris-based extender of ram semen.