Supplementation with Different Concentrations of Dithiothreitol and Glutathione During Cryopreservation of Semen from Large White Boars

Abstract

The study aims to compare different concentrations (0, 2.5, 5, 7.5 and 10 mM) of antioxidants, Dithiothreitol (DTT) and Glutathione (GSH), on sperm parameters following the freeze-thawing of semen from large white boars. Three large white boars were selected for semen collection and the collected semen was sent to the lab for analysis. Following dilution with Beltsville thawing solution and 120 min of equilibration at 17°C, the semen was centrifuged at 800 g for 10 min at 15°C. Following supplementation of semen with DTT or GSH at different doses (0, 2.5, 5, 7.5 and 10 mM), the semen was transferred into 0.25 mL freezing straws. After 20 min in Liquid Nitrogen (LN₂) vapor, the straws were submerged in the LN₂ tank, which was at a temperature of -196°C. Following thawing at 37°C, the semen was assessed for several sperm characteristics. Data were analyzed using the GenStat^® statistical program with the generalized linear model procedure. Treatment means were separated using Fisher’s protected t-test least significant difference at a 0.05 level of significance (Mean ± SD). When compared to all treatments, the proportion of sperm with total motility and progressive motility was higher in control (51.1±12.7; 27.1±10.8), 5 mM GSH (45.2±16.1; 24.7±11.3) and 5 mM DTT (43.8±10.3; 23.9±10.7) (p<0.05). When compared to all treatments, sperm rapid motility was highest in the control (21.3%) and 5 mM DTT (21.5%) treatments (p<0.05). The control (46.8±6.5), 5 mM GSH (44.8±8.2) and 5 mM DTT (40.3±2.20) had a high percentage of normal live sperm following thawing as compared to all the treatments (p<0.05). In the hypo-osmotic swelling test, the percentage of the swollen tail was greater in the control treatment (38.8±6.1) than in the treatments with 5 mM DTT (33.6±4.9) and 5 mM GSH (32.8±4.5) (p<0.05). However, the differences were not statistically significant. In conclusion, the quantities of 5 mM GSH and 5 mM DTT added to freezing extenders were appropriate for cryopreserving large white boar semen.