Effect of Propolis Ethanolic Extract Supplementation to Ram Semen Extenders on Sperm Characteristics, Lipid Peroxidation and some Enzymatic Activities in Seminal Plasma in Chilled Semen

Abstract

This study designed to examine effects of adding different concentrations of propolis ethanolic extract (PEE) as antioxidant to ram semen extenders; tris egg yolk (TEY) or tris-soybean lecithin (TSBL) on sperm characteristics, lipid peroxidation and enzymatic activities in seminal plasma of ram semen preserved at 5°C for 48 hours. Semen was collected from five adult Rahmani rams using an artificial vagina twice/week for 5 weeks. Pooled semen was extended with TEY or TSBL supplemented with different concentrations from PEE (0, 0.1, 0.5 and 1.0 mg/ml). Polyphenols and flavonoid compounds in propolis ethanolic extract were identified. In each treatment, percentage of progressive motility, livability, and abnormality were determined. Also, percentages of sperm membrane integrity or chromatin damage were determined. Concentrations of total antioxidant (TAO) and malondialdehyde (MDA), and activity of acid phosphatase (ACP), lactate dehydrogenase (LDH), and asprtate (AST) and alanine (ALT) transaminase, were determined. Results showed that the most effective polyphenolic compounds in the prolopolis ethanolic extract was the chlorogenic acids, catechein, protocatchuic and pyrogallol (120.623, 53.006, 23.907 and 11.048 mg/100g extract, respectively). Luteo.6-arbinose 8-glucose was the predominant identified flavonoid component in propolis extract (5268.78ppm), followed by acacetin, hisperidin, apig.6-rhamnose8- glucose, apig.6-glucose8-rhamnose, kamp.3-(2-p-comaroyl) glucose and luteo.6-glucose 8-arbinose with concentrations )647.53, 561.09, 366.47, 150.46, 144.51 and 109.42 ppm, respectively. The effect of propolis ethanolic extract supplementation to ram semen extenders was significant (P<0.05) only on the overall percentages of sperm progressive motility and sperm chromatin damage while, sperm livability, abnormality and membrane integrity were not affected significantly (P<0.05). Concentration of TAC was numerically (P= 0.82) higher in TEY extender with different concentrations of PEE than TSBL. However, MDA was lower (P<0.05) in free TEY, followed by TSBL with PEE. Activity of LDH, AST and ALT tended to be lower in seminal plasma of semen extended with TSBL than TEY. These results showed beneficial effects of soybean lecithin and propolis ethanolic extract in tris based extender on progressive motility and chromatin integrity of spermatozoa up to 48 h of preservation at 5°C. Also, lipid peroxidation and enzyme activity were lower in TSBL than TEY. In addition, soybean lecithin can effectively replace egg yolk as a protective additive with propolis ethanolic extract for preservation extender without any detrimental effects on post-chilling semen quality in rams.