Effect of butylated hydroxytoluene concentration on Saanen buck sperm freezability.
Project no. 23299 · Status: Kapanmış (Yayın bekleniyor) · 2021
Summary
<p>The main objective of this study was to define the effect of cryopreservation with different concentrations of butyl hydroxytoluene (BHT) at 0.6 mM, 2.0 mM, and 5.0 mM in Tris-citric acid-glucose egg yolk glycerol (TCGYG) based extender on semen quality of Saanen buck. Also, the effect of washed and unwashed sperm by centrifugation before cryopreservation was examined. Therefore, an optimal concentration of BHT could be found to develop a successful cryopreservation of buck sperm. The study was carried out at Ege University, Faculty of Agriculture, Department of Animal Science, Small Animals Unit. A total number of 4 Saanen bucks with 1-2 years old and 50-60 kg body weight were used in this study. Semen was collected twice a week during the mating season (November-January) using an artificial vagina in the presence of a female goat. The collected semen was mixed in test tubes and the mixture was divided into two equal main groups. One portion was diluted in tris-citric acid-glucose (TCG) solution (1:5) and washed twice by centrifugation at 600 x g for 10 minutes, while the other portion was kept unwashed.The supernatant was carefully removed after washing and the residue was divided into four equal parts, while the unwashed semen was divided into two equal parts. Afterwards, washed and unwashed semen were re-suspended in one of the different TCGbased media with 5% glycerol and 15% powdered egg yolk supplemented or not with different concentrations of BHT according to the treatments: 1) washed semen; 2) washed semen + 0.6 mM BHT; 3) washed semen + 2.0 mM BHT; 4) washed semen + 5.0 mM BHT; 5) unwashed semen and 6) unwashed semen + 5.0 mM BHT. Following supplementation or not with BHT, semen treatments were packed into 6 xii different 0.25 ml straws then were left to equilibrate at +4°C for 2 hours. After equilibration, the straws were kept in liquid nitrogen vapor at 4 cm above the liquid nitrogen level for 15 minutes and frozen by storing them in liquid nitrogen. The total number of the freezing process was repeated 20 times. The amount of mixed ejaculate (ml) and spermatozoa concentration (x109 /ml) obtained by mixing the collected semen were recorded, while the evaluation of total, progressive, local sperm motility (%) and dead spermatozoa (%) was determined by Computer Assisted Semen Analysis (CASA).The obtained results showed that the maximum amount of semen was 2.5 ml; the minimum amount was 1.0 ml, and the average was 1.75 ml. The average of sperm concentration was 2,320x109 /ml. In terms of total and progressive sperm motility in TCGYG extender after thawing, BHT groups were significantly (P<0.05) different than the control group in both washed and unwashed semen treatments. The highest percentages of total and progressive sperm motility were recorded with 2.0 mM of BHT with an average of 71.658and 42.44 %, respectively in washed semen, while the lowest percentages were recorded in the control group. Moreover, the difference between the washed and unwashed semen was significant (P<0.05). The washed semen was significantly (P<0.05) higher in the percentages of total and progressive sperm motility compared to unwashed semen.In the term of local sperm motility in TCGYG extender, there was no significant (P>0.05) difference between the control group and BHT groups in both washed and unwashed semen. The difference between the washed and unwashed semen was also insignificant (P>0.05). On the other hand, the percentage of dead spermatozoa in TCGYG extender, was significantly (P<0.05) different in BHT groups than the control group in both washed and unwashed semen. The lowest percentage of dead spermatozoa was recorded with 2.0 mM of BHT with an average of 28.34 % in washed semen, while the highest percentage was recorded in the control group. Moreover, the difference between the washed and unwashed semen was significant (P<0.05). The washed semen was significantly (P<0.05) lower in the percentage of dead spermatozoa compared to unwashed semen.In conclusion, BHT increased significantly (P<0.05) both sperm motility and viability in Saanen bucks. This study showed that BHT concentration ranging from 0.6 to 2.0 mM/mL could be the optimum concentration for freezing semen of Saanen buck. The highest percentages of sperm motility and viability were recorded after thawing of washed semen in TCGYG extender containing 2 mM of BHT. Consequently, the addition of BHT with a concentration of 2.0 mM could be recommended as an optimum concentration to improve many parameters of washed and unwashed sperm cryopreserved in an egg yolk-based medium. Also, the highest sperm motility and viability were recorded in washed semen compared to unwashed semen in this study. Therefore, it could be recommended to make sperm washing before cryopreservation by initial centrifugation of the semen to improve sperm cryosurvival.</p>