Impact of quercetin, carnosine, and ozone in the cryopreservation on Nellore (Bos indicus) semen

Abstract

The objective of this study was to reduce the effects of cryoinjury caused in bovine semen by cryopreservation. Ejaculates were collected from Nellore bulls and subjected to freezing in C (control), ozone (15, 30, and 60 µg mL^-1 of ozone), quercetin (25, 50, and 100 µg mL^-1 of quercetin), and carnosine groups (100, 200, and 300 ng mL^-1 of carnosine). Samples were evaluated post-thaw (M0) and post-rapid thermoresistance test (M30) for sperm kinetics (total motility, progressive motility, curvilinear speed, linearity and amplitude of lateral head displacement) and cell structure viability (plasma membrane integrity, acrosomal integrity, mitochondrial potential, membrane fluidity, and lipid peroxidation). There were no differences (P > 0.05) between the control, quercetin, and carnosine-treated groups for the parameters evaluated at M0 and M30. In turn, supplementation with ozone resulted in lower values for sperm kinetics (P < 0.05) and lower mitochondrial potential at M30 (P < 0.05). Quercetin and carnosine at the concentrations used did not promote significant gains in frozen semen, nor did they demonstrate cytotoxicity. We expected to obtain positive results with the use of ozone. Nonetheless, the addition was harmful to the parameters of sperm kinetics, and its effect was not observed as a possible pro-antioxidant. We believe that the fact that the gas did not harm the sperm structure opens avenues for tests with lower dosages, since, by reducing its concentration, we could minimize the damage to sperm kinetics.

Keywords: lipid peroxidation; reactive oxygen species; spermatozoa.

Figure 1. Flow cytometry results (values expressed in percentage) at post-thaw (M0) and post-RTT (M30) moments. PIPNA: intact plasma membranes with intact acrosome; PIMST: intact plasma membrane and high mitochondrial potential; YOMERO: intact and non-fluid plasma membrane; PERO: percentage of peroxidized cells; CONTROL: without the addition of ozone, quercetin or carnosine; CAR100: 100 ng mL-1 of carnosine; CAR200: 200 ng mL-1 of carnosine; CAR300: 300 ng mL-1 of carnosine; O15: 15 µg mL-1 of ozone; O30: 30 µg mL-1 of ozone; O60: 60 µg mL-1 of ozone; Q25: 25 µg mL-1 of quercetin; Q50: 50 µg mL-1 of quercetin; Q100: 100 µg mL-1 of quercetin.