The follicular fluid (FF) is crucial for providing oocytes with an ideal environment that promotes their development and maturation. Not only does this fluid supply nutrients and hormones, but also other components that protect both follicular cells and the oocyte itself from potential harmful factors, such as those inducing oxidative stress (OS). The FF has also been suggested to have beneficial effects on sperm when they reach the oviduct. The objectives of this study were to describe the presence and activity levels of redox biomarkers in the preovulatory follicular fluid (PFF), and to analyze the impact of PFF on the motility, viability and reactive oxygen species (ROS) levels of donkey sperm. For this purpose, ten PFF samples obtained from Catalan jennies and nine ejaculates collected from Catalan donkeys were used. Redox biomarkers -including enzymatic and non-enzymatic antioxidants, and oxidative biomarkers- were analyzed in jennies' PFF. After collection, each semen sample was split into two aliquots of equal volume. The two aliquots were centrifuged to remove the seminal plasma
one pellet was resuspended in Tris Buffered Medium (TBM), and the other was resuspended in TBM supplemented with 20% PFF. Motility parameters (CASA) and other semen quality biomarkers (flow cytometry) were assessed after 0, 60 and 120 min of incubation at 38 °C. Exposure of donkey sperm to PFF reduced intracellular ROS levels and helped maintain sperm motility and viability. These findings suggest that, in the oviduct, the PFF components protect donkey sperm from oxidative stress. Furthermore, knowing better the composition of donkey PFF in terms of antioxidant biomarkers may be used to improve the formulation of media for oocyte maturation and fertilization in this and other species.