Impact of replacing egg yolk with lecithin on quality of pre-freeze and post-thaw buffalo spermatozoa

Abstract

Objective: To estimate the result of egg yolk replacement with alternative cryopreservatives such as plant-derived lecithin from soybean on sperm quality parameters pre and post freezing in buffalo bulls. Methods: The control cryopreservation extender was tris-citric acid-fructose-egg yolk-glycerol (TCFYG) diluent. Semen samples were extended gradually 1:10 with TCFYG control extender and tris-citric acid-fructose-glycerol (TCFG) extender plus variable concentrations of soybean lecithin (0.5%, 1.0%, 1.5%, 2.0%, 2.5% and 3.0%) to ensure 60 million active spermatozoa/mL of the extended semen. The diluted semen samples were refrigerated slowly (roughly for 2 h) up to 5 °C and equilibrated for 2 h. Semen was filled into 0.25 mL polyvinyl French straws (IMV, France). After equilibration period, the straws were placed horizontally on a rack and frozen in a vapor 4 cm above liquid nitrogen for 10 min and were then dipped stored in liquid nitrogen at -196 °C. Results: The respective overall percentages of forward motile spermatozoa, live spermatozoa, morphologically normal spermatozoa, acrosome integrity and hypo-osmotic swelling reactivity observed primarily in fresh semen, after equilibration (pre-freeze stage) and post freezing (post-thaw stage) in TCFYG (control) extended semen declined progressively and statically (P<0.01) during these periods of study. Pre-freezing stage: replacement of egg yolk into TCFG with soybean lecithin at concentrations of 1.0% and 1.5% significantly (P<0.01) ameliorated the maintenance of (motility, viability, acrosome and membrane integrity %), meanwhile it had significantly (P<0.01) reduced the abnormality % of spermatozoa to the lowest value compared to control TCFYG and to some other concentrations in use. Post-thaw stage: the replacement of egg yolk with 1.0% soybean lecithin (SL) showed significantly (P<0.01) higher percentage of sperm progressive motility compared to 1.5% SL and TCFYG control. These values were significantly (P<0.01) higher than 0.5%, 2.0%, 2.5% and 3.0% SL. The post thawing live sperm percentage mean values were significantly (P<0.01) higher in 1.0% SL and 1.5% SL compared to control. These values were significantly (P<0.01) higher than in 0.5%, 2.0%, 2.5% and 3.0% SL. The mean values of post-thaw morphological normal sperm percentage did not differ between 1.0% SL and control groups but significantly (P<0.01) higher than 0.5%, 1.5%, 2.0%, 2.5% and 3.0% SL. The respective percentage mean values of post-thaw sperm with head, mid-piece and tail abnormalities were significantly (P<0.01) lower in 1.0% SL than all other SL concentrations. Concerning the post-thaw percentages of acrosome and sperm membrane integrity, the respective mean values were significantly (P<0.01) higher in 1.0% SL and 1.5% SL as compared to control. Mean values of both parameters in the 0.5% SL were intermediate between 1.0% and 1.5% SL versus control groups. The previously mentioned mean values in acrosome/membrane integrity were significantly (P<0.01) higher than 2.0% SL, 2.5% SL and 3.0% SL. Conclusions: Lecithin-based diluent can be a potent proper alternative extender for preservation of spermatozoa during pre- and post-freezing process. SL 1.5% extenders have supplied an optimal environment and condition for ameliorating the quality of pre-freezing and post-thaw buffalo spermatozoa by means of improved motility, viability, functional acrosome, sperm membrane integrity and morphologically normal spermatozoa.

Keywords

• Buffalo
• Semen
• Preservation
• Soybean lecithin