Nanopartículas de lipoproteína de baixa densidade na criopreservação do sêmen canino

Abstract

Structures on the manometric scale have unique functional properties, not found on the macro scale, mainly due to the high surface area of contact, which results in an intense interaction with the matrix in which the nanoparticles are inserted. In seminal cryopreservation diluents based on egg yolk, the main cryoprotective component is low density lipoprotein (LDL). However, egg yolk has other constituents that are harmful to gametes, in this context, the extraction of the micelles of LDL for seminal freezing, through the plasma fraction of the egg yolk (EYP) has been showing better results. Given the above, this study aimed to evaluate different LDL nanoparticles production processes, on the quality of cryopreserved canine semen. EYP was submitted to three systems: ultrasound bath (LDL-B) - 30min at 40A; ultrasound probe (LDL-P) - 50% of amplitude, 30min; high pressure homogenization (LDL-P) - 10,000PSI for 6 cycles. Diameter properties, polydispersity index (PDI) and zeta potential of the diluents were evaluated. Sperm quality after thawing was measured using computer-assisted sperm analysis (CASA) and flow cytometry. The nano techniques applied in EYP modified properties of diameter, PDI and zeta potential of LDL particles. LDL-P was characterized as a monodisperse system (PDI = 0.37) and stable, while in the other experimental groups polydisperse solutions were obtained. The processing of EYP formed freezing diluents that were more effective in controlling the production of reactive intracellular oxygen species (P < 0.05). The ultrasound systems (LDL-B and LDL-P) produced micelles capable of maintaining the highest total sperm motility, after thawing (P < 0.05). Compared to the EYP control, LDL-P was more efficient in maintaining progressive sperm motility, integrity and membrane fluidity (P < 0.05). The LDL-P nanoparticles obtained an average size of approximately 250nm, with 0.3 of PDI and -1.15mV of zeta potential. Therefore, ultrasound using an EYP probe has proven to be an efficient method for the production of LDL nanoparticles, increasing cryoprotection during the freezing of canine semen.