Sixty-day oral antioxidant supplementation enhances sperm morphology and seminal plasma antioxidant capacity in stallions

While dietary supplementation with antioxidants has been identified as an effective strategy for mitigating oxidative stress (OS) damage in stallion sperm, specific protocols and the duration of its efficacy remain poorly defined. This study aimed to evaluate the effects of a 60-d dietary supplementation with a commercial antioxidant combination on semen quality and OS biomarkers in stallions. Stallions were divided into two groups: the control group (n = 5; CTRL) re- ceived 30 g/500 kg of placebo while the treated group (n = 5; OXY) received 30 g/500 kg of Oxyliver daily for 60 d. Semen was evaluated for volume, concentration, viability, morphology, membrane integrity, motility, and kinetic parameters every 15 d for the 60-d supplementation period and for an additional 60 d after treatment cessation (D0, D15, D30, D45, D60, DP15, DP30, DP45, DP60). Total antioxidant capacity (TAC), ferric reducing/antioxidant power (FRAP), and malon- dialdehyde (MDA) were measured at 30-d intervals. The change in sperm parameters over time differed significantly be- tween groups, specifically for sperm membrane integrity (P = 0.004), total motility (P = 0.003), morphological abnormali- ties (P = 0.028), and OS parameters (TAC P = 0.031, FRAP P = 0.044, MDA P = 0.048). Compared to CTRL, the OXY group showed greater sperm membrane integrity (P = 0.004) and total motility (P = 0.039) at D60. Conversely, total morpholog- ical abnormalities were lower in OXY at D60 (P = 0.003), DP15 (P = 0.009), and DP30 (P = 0.018), mainly due to a reduction in tail defects at the same time points (P=0.008, P=0.014, P=0.020, respectively). Moreover, TAC and FRAP were greater in the OXY group at D30 (P = 0.004 and P = 0.012, respectively) and D60 (P = 0.001 and P = 0.009, respectively), while MDA concentrations were lower in the OXY group at D60 (P < 0.001), DP30 (P = 0.033) and DP60 (P = 0.003). In con- clusion, despite the small sample size, this longitudinal study provides preliminary evidence that a 60-d supplementa- tion with this combined formula enhances seminal plasma antioxidant capacity during supplementation and reduces lipid peroxidation even after the end of supplementation. It also induces a transient improvement in sperm membrane functionality and total motility at specific time points, whereas sperm morphology showed a more robust improvement beyond the supplementation period.