Excessive production of reactive oxygen species (ROS) during semen cryopreservation can induce structural and functional changes in spermatozoa. It is well known that antioxidants can mitigate the effect of ROS. Moreover, the application of antioxidants in freezing media is an appropriate strategy for protecting spermatozoa against deleterious effects of ROS during the cryopreservation process. As an example, oregano is a medicinal plant with important activities, with antiseptic, antibacterial, antithrombotic, and antioxidant properties. This study aimed at evaluating the antioxidant effects of oregano extract on cryopreserved human spermatozoa. In the first phase, 13 semen samples with different concentrations of oregano extract (0.0, 50, 100, 150, 300, and 500 μg/mL) were cryopreserved to achieve an optimal dose of oregano extract. Then, motility, viability, and plasma membrane integrity were evaluated. In the second phase, 20 samples were cryopreserved in freezing media supplemented with or without the optimal concentration of oregano (100 μg/mL). After thawing, motility, the levels of ROS, lipid peroxidation, and translocation of phosphatidylserine (PS) were evaluated. The results showed that 100 μg/mL oregano extract significantly increased the total motility in frozen-thawed spermatozoa in comparison with the control group (28.2 ± 4.3 vs. 42.4 ± 1.6, p < 0.05). This concentration significantly decreased the percentage of 2',7'-dichlorofluorescein-positive cells (25.53 ± 1.2 vs. 21.48 ± 1.2) and the malondialdehyde level (4.25 ± 0.7 vs. 0.82 ± 0.4 μM) (p < 0.05). In the oregano group, the percentage of vital spermatozoa without PS externalization was significantly higher than that in the control group (25.88 ± 1.6 vs. 16.8 ± 1.9, p < 0.001), while the percentage of dead spermatozoa with PS externalization spermatozoa was significantly lower than that in the control group (51.65 ± 1.4 vs. 60.36 ± 1.9, p < 0.05). In general, the addition of oregano extract to sperm freezing extender has protective effects against oxidative stress and apoptosis.
Keywords: Origanum vulgare; antioxidant; cryopreservation; reactive oxygen species; spermatozoa.