Impact of silymarin enriched semen extender on bull sperm preservability

Objective:To explore the effect of silymarin on bull spermatozoa during cooling and cryopreservation. Methods: Pooled bull semen were diluted by Tris-Citrate-Fructose egg yolk diluents, purified silymarin powder (obtained from the milk thistle silybum marianum), purchased from Unipharma, Al Obour city, Egypt, was soaked in Tris-citric acid-fructose diluent for 48 h at 10℃ making a stock solution (70 mg/mL), from this stock solution we obtained concentrations of 0.18 mg/mL, 0.36 mg/mL, 0.54 mg/mL, 0.72 mg/mL, 0.90 mg/mL in addition to the control (0.0 mg/mL) reaching a final volume of 5 mL in each tube. Egg yolk was added to each tube to obtain silymarin enriched semen extender (SEE) with 20% egg yolk, cooled slowly up to 5℃ and equilibrated for 4 h. Semen was packed into 0.25 mL polyvinyl French straws (IMV, France). After equilibration periods, 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 in liquid nitrogen. Extended semen was subjected to evaluation (motility, alive %, abnormality %, intact sperm membrane (HOST)% and conception rate) in both chilled and frozen semen.Results: Table 1 revealed that Sperm motility of the concentrations 2, 3 and 4 after 8 d of chilling were significantly (P<0.02) higher than control. Sperm motility of the concentration 2 (45.00%±2.89%) after 9 d of chilling was higher than control and the other concentrations. Addition of SEE in concentration 1 and 2 gave post thawing sperm motility as high as the control (47.50±2.81 and 45.00±2.58, respectively) while other concentration have lower effects on motility as compared to the control. Addition of silymarin improved post thawing alive% and was significantly higher (P<0.0001) than the control. SEE decreased significantly (P<0.0001) the % of post thawing abnormal sperm in concentration 3 and 4 (11.83±0.65 and 16.00±0.58, respectively). SEE improved significantly (P<0.018) the % of post thawing intact spermatozoa membranes (HOST%) in concentrations 2, 4 and 5 (71.17±0.83, 71.83±0.91 and 75.00±3.42 respectively) (Table 2).Conclusion:It could be concluded that silymarin as a natural additive to semen extenders improved preservability in both chilled and frozen bull semen.

Preservability of bull spermatozoa in Tris-egg yolk extender enriched with different concentrations of butylated hydroxytoluene

Objective:To explore the effect of BHT on cattle spermatozoa during cooling and cryopreservation.Methods: Pooled bull semen were diluted by Tris-Citrate-Fructose egg yolk (TCFY) diluent considered as control (0 BHT) and different concentrations of BHT (1.0, 2.0, 3.0, 4.0, 5.0 and 6.0 mM were prepared in ethanol in prewarmed (37℃) test tubes. The ethanol was allowed to evaporate so that, a thin crystallized layer of BHT was deposited on the inner surface of the tubes. Then extended semen was added into the tubes and incubated at 37℃ for 5 min to allow uptake of BHT by spermatozoa. The tubes were cooled slowly (approximately for 2 h) up to 5℃ and equilibrated for 4 h. After equilibration, semen freezing process was carried out. Extended semen was subjected to evaluation (motility, alive sperm, intact sperm membrane (HOST) % and acrosome integrity) in both cooled and cryopreserved semen. Results:The result revealed that sperm motility of post-cooled spermatozoa improved (P<0.05) by the use of BHT concentrations (1, 2 and 3 mM) in Tris semen extender if compared to the control (85.00±1.09), (83.33± 0.63), (81.67± 0.63) and (78.33± 0.63), respectively. Alive sperm percent was significantly higher in all concentrations of BHT. Sperm abnormalities percent were significantly lower in concentrations of BHT 1 and 2 (11.2±0.2), (11.8±0.2)and (13.4±0.4), respectively. Sperm membrane integrity were significantly higher in BHT concentrations (1, 2, 3, 4 and 5 mM). It is exhibited that improved sperm motility in post-thawed frozen semen in the concentrations of BHT (1, 2, 3 and 4 mM) if compared to the control. The sperm membrane integrity were significantly improved at all concentrations of BHT. Acrosome integrity was significantly higher at BHT concentration 1 mM (81.80±0.57) and (76.00±2.05), respectively.Conclusions: It could be concluded that some concentrations of BHT improved bull semen quality post-cooling and post-freezing.