Potential of Brown Macroalgae Extract (Sargassum ilicifolium (Turner) C. Agardh) on the Quality and Protamine-1 (PRM1) Protein Levels of Frozen-thawed Semen in Simmental Bulls

Semen cryopreservation provides numerous advantages, but it also results in reduced semen quality following thawing. In order to mitigate these negative consequences, antioxidants are often added to semen extenders. Brown macroalgae has recently been shown to be a natural source of antioxidants. The purpose of this study was to investigate whether supplementation of extender with four concentrations of macroalgae extract (Sargassum ilicifolium (Turner) C. Agardh) would improve the quality of sperm post thawing. A total of 30 semen samples collected twice weekly over a period of four weeks from four Simmental bulls were used in this study. These samples were divided into five parts, diluted using extender supplemented with S. ilicifolium extract at 0, 250, 500, 1000 and 2000ppm, and cryopreserved in liquid nitrogen. Post-thaw sperm motility, viability, membrane integrity, DNA fragmentation, reactive oxygen species (ROS), malondialdehyde (MDA) levels, and protamin-1 (PRM-1) protein concentration were analyzed. Results demonstrated that semen samples frozen in extender supplemented with Sargassum ilicifolium extract exhibited reduced levels of spermatozoa damage, as indicated by significantly better sperm progressive motility, viability, membrane integrity, and reduced DNA fragmentation due to lower ROS production compared to the control group (P<0.05). The best results in terms of sperm motility, viability, membrane integrity, and lower DNA fragmentation were observed in the 1000ppm treatment group. Reduced ROS levels in the treatment groups were associated with lower MDA and increased PRM1 levels. Moreover, DNA fragmentation was inversely related to PRM-1 levels. Overall, the best results across multiple parameters were observed in the 500– 1000ppm treatment range. Further studies are needed to fully explore the potential of S. ilicifolium extract in maintaining semen quality during the freeze–thaw process. © 2025, University of Agriculture. All rights reserved.