Effect of idebenone supplementation in a semen extender on boar spermatozoa quality during liquid storage

In swine production, over 99% of artificial insemination (AI) procedures utilize boar semen preserved in liquid form at 17°C for up to 5 days. However, spermatozoa are highly susceptible to oxidative stress during storage, which impairs motility, membrane integrity, and overall fertility. Reactive oxygen species-induced lipid peroxidation (LPO) compromises sperm structure and function. Although antioxidants are used to mitigate oxidative damage, idebenone (IDB) - a short-chain benzoquinone with potent mitochondrial antioxidant properties - has not been studied in boar semen preservation. This study aimed to investigate the effects of IDB supplementation in a semen extender on boar spermatozoa quality during 120 h of liquid storage at 17°C.

Ejaculates from 25 Duroc boars were diluted with Beltsville Thawing Solution to a final concentration of 30 × 10⁶ sperm/mL. In Experiment 1, semen samples (n = 13) were treated with 0, 78, 156, 312, 625, or 1250 nM of IDB and assessed at 24-h intervals for motility (computer-assisted sperm analysis), viability (eosin-nigrosin), and membrane integrity (hypo-osmotic swelling test). In Experiment 2, 78 nM IDB was selected for its optimal effects and further tested (n = 12) for acrosome integrity (fluorescein isothiocyanate-conjugated peanut agglutinin), capacitation status (chlortetracycline staining), LPO (thiobarbituric acid reactive substances), and kinematic parameters.

After 120 h, the 78 nM IDB group showed significantly improved total motility (81.4%) and membrane integrity (60.3%) compared to control (72.9% and 46.7%, respectively; p < 0.05). Viability was higher in IDB-treated groups, and high concentrations (625-1250 nM) reduced motility. In Experiment 2, 78 nM IDB improved total and progressive motility (84.0% and 46.8%), preserved acrosome integrity (64.06% vs. 58.75%), reduced capacitated sperm (64.72% vs. 59.11%), and decreased malondialdehyde levels (51.84 nmol vs. 59.61 nmol) relative to controls (p < 0.05).

IDB at 78 nM effectively preserves boar semen quality during 5-day liquid storage by enhancing motility, viability, membrane and acrosome integrity, and reducing oxidative stress. These findings highlight its potential as a novel antioxidant additive in AI protocols.