Effect of silymarin antioxidant on the membrane integrity, viability and motility of ram frozen sperm

The need for artificial insemination in the ewe is to access fresh and frozen sperm. Cryopreservation is a suitable method for long-term storage of semen and sperm, but may cause a minor irreversible damage to the sperm cell, especially its membrane region (Khorramabadi et al., 2017). The oxidation of fatty acids leads to the production of free oxygen radicals (ROS). These radicals are necessary in normal conditions for certain physiological activities and sperm processes, but excessive production of ROS in the sperm can reduce membrane fluidity, DNA fracture, damage to proteins, and ultimately reduced sperm motility and fertility. (Bakhshayesh et al., 2017). Silymarin, the scientific name of Silbum marianum, is the English name Milk Thistle and is a potent inhibitor of oxidative stress as a potent antioxidant. Due to the nature of its friend fat, silymarin is firmly attached to the plasmid membrane compounds, thus preventing fracture and its collapse by increasing membrane strength. Increasing the amount of active oxygen species and lipid peroxidation disrupts mitochondrial membrane, decreases ATP and damage to sperm acesomes, which ultimately reduces the progression of sperm motility. (Kvasnikova et al., 2003). The mechanism of action of salimarin is through stimulation of the ribosomal RNA and protects the membrane from oxidative damage. They also stated that silymarin stimulates the activity of the antioxidant enzymes of superoxide dismutase (SOD) and glutathione peroxidase (Wellington and Jarvis, 2001) . Silymarin improves the sperm motility and survival, sperm abnormalities, and preservation of the sperm membrane after freezing-thawing. Improvement of semen quality in both freezing and cooling is due to the strong silicomain antioxidant capacity (Fakorzai et al., 2008; Longpyram et al., 2013). .

Regarding the anti-oxidant effects of silymarin, this study aimed to investigate the effect of silymarin on storage of ram sperm.

In this research, four rams of pure ghee were used 2-3 years old and each ram was 6 times sperm. Sperm was performed twice a week by artificial vagina. After the sample was transferred to the laboratory, a series of preliminary evaluations including sample size, sample color, wave motion, total mobility, in-situ motion, progressive motion and live weight, and abnormal sperm and density were performed. If standards were met, Necessary (scaling over 2.5 billion sperm and progressive movement above 70%) dilution was performed with treatments. The diluents were prepared prior to sampling and placed in a bin Marie temperature of 37 ° C. Tris-based diluent was used to contain 2.73 g of tries, 1.4 g of fructose, 1 g of citric acid and 100 mg streptomycin in 100 ml sterile distilled water. To prepare the diluent, 73% of the prepared solution will be mixed with 20% egg yolk and 7% glycerol. In order to dilute the sprays from trace based diluent, egg yolk, semen collected with diluent without antioxidants (control) and silymarin 5 and 10 μg / ml mix After cooling for 90 minutes in the refrigerator and reaching 5 ° C, it was placed in 4 cm above the liquid nitrogen for 8-10 minutes and then immersed in liquid nitrogen. The post-mortem examinations of sperm quality properties included liveliness, total motion, progressive motion, integrity of the plasma membrane (tests) on days 0, 15, 30 of the experiment.

According to the results of the experiment, the diluents containing the levels of silymarin significantly increased the total motion, progressive, live survival of ram sperm after the freeze-thawing process compared to the control group Were opened. Addition of different concentrations of silymarin was 5 and 10 μg with mean of 74.45% and 67.33%, respectively, compared to the control group, significantly increased the total sperm motility with mean of 68.07 and 60.95%, significantly Progressive motility of frozen ram sperm were frozen (p <0.01). According to the results, the in situ movement in the control group (5.92 ± 0.92) was lower than that of the 5 and 10 μg silymarin (6.38 ± 0.92) and (8.9 ± 0.92) respectively In the least amount.

Application of silymarin as an antioxidant in the ram sperm diluent system reduces oxidative damage and improves sperm quality properties such as mobility, viability and the health of acrosome and plasma membrane during freezing process. Was opened. Adding 5 μg of silymarin to diluted ram minus 10 μg and control group reduced fat peroxidation and also improved the structure and performance of sperm during storage.