Comparative evaluation between the extenders TES-TRIS and ACP-112® and the association of Sálva Marajó oil (Lippia origanoides) in the quality of cryopreserved buffalo sperm

Abstract

For artificial insemination, it is essential to use frozen semen, however the freezing process causes deleterious changes to the structure and integrity of sperm membranes that compromise the function of sperm. To avoid this cellular damage, extenders and suitable substrates must be used to recover the highest possible number of viable cells post-thaw. To this end, in the first experiment, we evaluated three different extenders: TES-TRIS, which is widely used for buffaloes; and an extender composed of powdered coconut water-based (ACP-112®) with or without milk (ACP-112®-milk) for buffalo semen freezing. In the second experiment, we evaluated the effect of Lippia origanoides oil extract on protecting buffalo sperm against cryoinjury arising from freezing semen. Semen was collected from ten buffalo bulls (10 ejaculates/bull) and diluted in TES-TRIS (control), ACP-112® or ACP-112®-Milk in the first experiment. In the second experiment, the samples were diluted in the diluent with the best results for sperm quality obtained in experiment I, and 2.5 ?g mL-1, 5 ?g mL-1 or 10 ?g mL-1 of the plant extract was added to treatments; and a control group containing only the diluent was also included. The fresh semen was analyzed for conventional features such as motility, concentration, morphology and viability. After thawing, the samples were evaluated again for motility, vigor and supra-vital staining, and then, were performed the of thermal-resistance test, hypoosmotic test and evaluated sperm membrane integrity with the fluorescent probes PI, FITC-PSA and JC-1 using flow cytometry. The data were submitted to ANOVA, and the results were compared by Tukey’s test at a significance of 5%. In the first experiment, the extender TES-TRIS showed better results for the various characteristics evaluated compared to ACP-112® and ACP-112®-Milk (P < 0.05), demonstrating greater protection of the buffalo sperm structures during cryopreservation. In the second experiment, the treatments with different concentrations of Lippia origanoides essential oil extract showed no differences among the assessed variables regarding the protection of sperm structures during cryopreservation (P > 0.05). Based on these data, we demonstrated the beneficial effects of TES-TRIS for post-thaw buffalo sperm quality; however, no protective effect was observed for buffalo sperm cryopreserved with the different tested concentrations of Lippia origanoides extract oil.