Antioxidant status of chromatographic fractions of boar seminal plasma (SP) and mitochondrial functions of SP-treated spermatozoa following cryopreservation

In this study we investigated the antioxidant status of fractionated seminal plasma (SP1, >40 kDa; SP2, <40 kDa), obtained by gel filtration chromatography (Wasilewska-Sakowska et al. 2019a), and mitochondrial functions in SP-treated spermatozoa during pre-freeze equilibration and post-thaw storage periods. The Total Antioxidant Status (TAS), represented by enzymatic and non-enzymatic antioxidants was markedly affected by the treatments, being significantly higher in either SP1 or SP2 than in the whole seminal plasma (wSP). It was observed that ATP content was markedly higher in SP1- or SP2-treated samples following pre-freezing equilibration. Although there were no marked differences in the ATP content of frozen-thawed (FT) spermatozoa among treatments, regardless of the cryopreservation stage, mitochondrial membrane potential (MMP) of FT spermatozoa differed significantly among treatments. Furthermore, significant differences in the sperm MMP at post-thaw were not reflected in the ATP levels, suggesting that variations in the surrounding environments of FT spermatozoa might have varying effects on the sperm mitochondrial functions following cryopreservation. It seems that high TAS levels in the fractionated SP could reaffirm the protective role of antioxidants in sperm cryo-damage.