Endogenous compounds of mammalian (boar) sperm sensitizing the formation of reactive oxygen species

Interest in identifying the type of endogenous compounds capable of initiating the formation of reactive oxygen species (ROS) in boar semen upon photoexcitation is driven by the use of photobiomodulation methods in artificial insemination of pigs and the unclear mechanisms underlying the stimulating effect of light on sperm function. One hypothesis regarding the mechanism of biological activity of light suggests the crucial role of ROS in mediating the effects of photobiomodulation. This study examined endogenous fluorophores in boar semen characterized by absorption in the UV-A and visible spectral regions and capable of generating ROS upon photoexcitation. For the first time, weak fluorescence of endogenous porphyrin sensitizers was detected in freshly collected boar ejaculate (native sperm) alongside intense fluorescence of NADH (NAD(P)H) coenzymes and flavins (flavin mononucleotide, FMN, and flavin adenine dinucleotide, FAD). Using steady-state and kinetic spectrofluorimetry, the types of porphyrins localized in the sperm were identified, and the presence of uroporphyrin III, coproporphyrin III, and their zinc complexes was established. Using a chemiluminescent assay for monitoring ROS levels and specific ROS quenchers (scavengers), it was shown that exposure of boar sperm to radiation of blue spectral region leads to the formation of singlet oxygen and hydrogen peroxide. It was concluded that light-induced generation of ROS (primarily singlet oxygen) by endogenous porphyrins and flavins can alter the redox state of sperm and, at low ROS concentrations, exert a mild regulatory effect on their functional activity.