Investigation of the effect of Nepeta italica Subsp. cadmea extract on frozen-thawed buck spermatozoa

Sperm cryopreservation adversely affects semen quality and subsequent fertilization ability due to the overproduction of reactive oxygen species (ROS). The addition of cryoprotectants to the extender can protect sperm against the harmful effects of oxidative stress (OS). Studies are carried out to use natural antioxidants such as plant extracts rich in flavonoids and phenolic compounds as cryoprotectants. In this study, the effects of three different doses of antioxidant and flavonoids rich Nepeta italica subsp. cadmea oil (0.75 µl/10 ml, 1.5 µl/10 ml and 6 µl/10 ml) on motility, acrosomal integrity and DNA fragmentation on frozen-thawed Ankara buck spermatozoa were investigated. The plants were harvested during the flowering stage on Honaz Mountain (Denizli, Turkey) and air dried in the shade at room temperature. Essential oil of the powdered plant was extracted by hydrodistillation using a Clevenger apparatus system, then stored at 4⁰C in an amber colored glass bottle. The ejaculates taken from the Ankara bucks during the breeding season with an electroejaculator were combined and diluted with tris diluent and three different doses of plant oil, and then frozen. It was determined that the plant oil at the dose of 0.75 µl/10 ml was improved the post-thawed sperm motility compared to control group and other groups (p<0.05). Similar results were obtained from the analysis of acrosomal integrity and DNA fragmentation. In the acrosomal membrane integrity after freeze–thawing, 0.75 µl/10 ml dose of essential oil was found to be higher than the doses of control and 1.5 µl/10 ml, 6 µl/10 ml (p<0.05). In DNA fragmentation analysis, 0.75 µl/10 ml dose of essential oil was found to be lower than the doses of control and 1.5 µl/10 ml, 6 µl/10 ml (p<0.05). As a result, the cryoprotective effect of 0.75 µl/10 ml dose of essential oil on motility, acrosomal integrity and DNA fragmentation parameters in Ankara bucks sperma after freeze-thawing was determined. It has been shown that the most appropriate dose of Nepeta italica subsp. cadmea essential oil having antioxidant effects and containing flavonoids can be used as a cryoprotectant in sperma cryopreservation. However, there is a need to reveal the possible cryoprotective effects of determined doses of plant essential oil in sperm cryopreservation by studying with different parameters.