Influence of tannin-rich extract from commercial Acacia mearnsii and gallocatechin on ovine cryopreserved semen viability.

The objective of this study was to evaluate the effect of a tannin-rich extract from commercial Acacia mearnsii (MTE_0), and gallocatechin, a flavonoid compound derived from Acacia mearnsii, on the long-term viability and motility of cryopreserved ovine semen. Six fresh ejaculates obtained from six adult merino rams twice per week for three weeks were allocated to five aliquots (0, 12.5, 25, 50, and 100 µM gallocatechin added into the Optidyl™ extender) before cooling and cryopreservation. Effects of MTE_0 and gallocatechin on post-thawed motility characteristics were analyzed using computer-assisted semen analysis (CASA), and viability (LIVE/DEAD^® kit, Molecular Invitrogen, Waltham, MA), oxidative stress (2,7-dichlorodihydrofluorescein diacetate (H2DCFDA, Thermo Fisher^®, Waltham, MA)) for reactive oxygen species (ROS), mitochondrial membrane potential (JC-1 MitoTracker, Molecular Invitrogen, Waltham, MA), acrosomal integrity (lectin PNA), and capacitation using merocyanine 540 (M540) and YO-PRO-1 dyes in flow cytometry. Data were analyzed using one-way ANOVA (IBM SPSS 21.0 for Windows, Armonk, NY). Gallocatechin at 25 µM positively affected (p ≤ .001) kinematic parameters including average path velocity (VAP), progressive velocity (VSL), and beat cross frequency (BCF) of cryopreserved semen. Similarly, gallocatechin at 25 µM^- improved sperm motility (live 21.99 ± 2.06%), reduced ROS levels (26.45 ± 1.10%), and mitigated premature capacitation (viable and stable 20.08 ± 1.48%) compared to other treatments. Gallocatechin addition to semen resulted in a significant (p ≤ .001) positive effect compared with the MTE_0 extract. It is concluded that gallocatechin inclusion at 25 µM significantly reduces semen deterioration following cryopreservation. This study is the first to introduce gallocatechin as an efficient antioxidant additive to ovine semen to improve its quality during storage. Our findings will help improve post-thaw ovine semen quality and longevity. Future studies to elucidate the mechanism of anti-oxidative stress action of gallocatechin and its derivatives on semen motility and longevity are recommended.