Effects of Curcumin Oil Nano-Emulsion on Buffalo Sperm Cryopreservation: Insights on Sperm Quality, Kinematic Parameters, Sperm Ultrastructure, Oxidative Stress, Apoptosis, Microbiota, and Molecular Docking Analysis and Fertilising Capacity.

This study investigated the potential of incorporating curcumin oil nano-emulsion (CONE) into a Tris-based freezing extender to relieve cryodamage effects and enhance sperm cryo-resistance in buffalo bulls. Pooled semen from five fertile buffalo bulls (n = 40 ejaculates) was divided and supplemented with 0.5% CONE (CONE0.5), 1% CONE (CONE1), or without CONE (CONE0, control), then extended and packaged. Post-thaw assessments included sperm kinematic parameters, oxidative stress markers, apoptosis levels, microbial load, ultrastructural integrity, molecular docking interactions, and conception rates. The CONE0.5 group exhibited significantly higher sperm viability, progressive motility, and membrane integrity, as well as significantly lower nitric oxide and MDA levels, compared to the CONE0 and CONE1 groups (p < 0.05), indicating reduced oxidative stress. Compared to the CONE0 group, the CONE0.5 group exhibited a significantly higher percentage of viable and early apoptotic sperm, and a significantly lower percentage of late apoptotic sperm (p < 0.05). Both CONE-treated groups demonstrated significantly lower post-thaw total bacterial, spore-forming bacterial, and coliform bacterial counts (p < 0.05). Conception rates were higher in the CONE0.5 group (80.0%) compared to the CONE0 group (68.0%) (p > 0.05). Molecular docking analysis revealed strong binding affinities between curcumin and caspase-3 (-7.87 kcal/mol), HSP70 (-8.52 kcal/mol), and PRDX-1 (-8.90 kcal/mol), suggesting potential mechanisms of action. In summary, the inclusion of 0.5% CONE as a cryoprotective additive in buffalo semen freezing media demonstrates a potential for improving post-thaw semen quality parameters, suggesting a possible enhancement of reproductive efficiency.