Melatonin and canthaxanthin enhances sperm viability and protect ram spermatozoa from oxidative stress during liquid storage at 4°C.

Antioxidants are used to minimize oxidative stress during liquid semen storage. The main aim of current study was to elucidate effect of supplementing melatonin and canthaxanthin in Tris-based extender could enhance seminal quality of ram at 4°C up to 72 h. A total of 48 ejaculates were collected from breeding Magra rams (n = 8) and were preliminarily subjected for various macroscopic and microscopic semen evaluation tests. These ejaculates were pooled and divided into three equal aliquots. Two aliquots were diluted (1:10) using extender encompassing final concentration of 1mM melatonin and 25 µM canthaxanthin and stored at 4°C. Third aliquot with extender only was kept as control. Structural and functional seminal changes were observed at different time points of preservation. Results revealed that mean values for progressive sperm motility, viability and total antioxidant capacity were significantly higher (p < 0.05) in melatonin group while hypo-osmotic swelling test was significantly (p < 0.05) higher in canthaxanthin group. Total sperm abnormalities and malondialdehyde levels were significantly (p < 0.05) lower in both treatment groups indicating their antioxidant efficacy in protection of spermatozoa from oxidative stress. Results of study indicated that supplementation of these antioxidants to ram semen could be used to enhance storage life of liquid semen at 4°C up to 72 h.

Cholesterol Loaded Cyclodextrin Supplementation Enhances the Cholesterol-to-Phospholipid Ratio and Diminishes Oxidative Stress in Jack Spermatozoa During Cryopreservation.

The present study was conducted with the hypothesis that addition of cholesterol to the extender would stabilize the sperm membranes by increasing the cholesterol-to-phospholipid (C:P) ratio and would result in an improved post-thaw semen quality and reduce oxidative stress in the jack (Martina franca) semen. Forty-eight ejaculates from six jacks were collected and analyzed for the present study. The freshly collected semen sample of each jack stallion was divided into five equal fractions after addition of the primary extender without cholesterol-loaded cyclodextrin (CLC) (C) and with 1, 1.5, 2, and 3 mg/mL CLC to obtain 120 × 106 sperm/mL spermatozoa concentration. The semen was cryopreserved using customized freezing protocols. Evaluation of seminal parameters, the C:P ratio, and the oxidative status of jack spermatozoa was analyzed at all stages of cryopreservation. The oxidative status in the jack semen was evaluated by measuring malondialdehyde, glutathione and total antioxidant capacity levels. The results indicated that the mean percent values for various seminal quality parameters and the oxidative parameters were found to be significantly higher (P < .05) in CLC-treated groups with the highest values for 2 mg of CLC/120 × 106 spermatozoa. In conclusion, the present study revealed that the supplementation of CLC before cryopreservation has significantly reduced the oxidative stress and also increased the C:P ratio during semen cryopreservation process. Furthermore, a reduction in lipid peroxidation levels, reduced damage to the sperm plasma and acrosome membranes and improvement in the post-thaw sperm integrity as well as stability were recorded.

Ameliorative Effect of Ascorbic Acid and Glutathione in Combating the Cryoinjuries During Cryopreservation of Exotic Jack Semen.

The present study was designed to study the adverse effects of cryopreservation and evaluation of the cryoprotective effect of reduced glutathione (GSH) and ascorbic acid (AA) supplementation on exotic jack semen in combination or alone. For this, 24 semen samples from four adult and fertile jacks were collected via artificial vagina using an estrus jenny as dummy. After semen collection, the semen was evaluated for various qualitative and quantitative parameters in fresh, cooled, and frozen-thawed semen. The semen pellet was extended with the freezing extender containing either AA (0.9 g/L), GSH (2.5 mM), or combination of both (AA 0.9 g/L + GSH 2.5 mM), and another aliquot was kept as control without adding the antioxidants. The jack semen underwent cryodamage, which was evident by the observation of significant (P < .05) decline in the seminal quantitative parameters at various stages of cryopreservation process. Prefreeze and postthaw semen evaluation revealed that the values of plasma membrane, acrosome integrity, and chromatin integrity were found to be significantly higher (P < .01) in the group of samples supplemented with the combination (0.9 g/L AA +2.5 mM GSH) than AA- and GSH-alone or control groups. Supplementation of antioxidants to the freezing extender improved jack prefreeze and postthaw semen quality with the superiority of GSH over AA alone. From the present study, it was inferred that, exotic jack spermatozoa are susceptible to injuries because of cryopreservation, but these cryo-induced damage can be ameliorated significantly (P < .05) with the use of antioxidants and contribute to the improvement of semen cryopreservation procedures.