Use of lipid peroxidation in the evaluation of antioxidant action in cryopreserved semen

Abstract

The cryopreservation process causes oxidative stress to the sperm cell and the addition of antioxidants to the semen refrigeration and freezing media can help to protect the sperm against the damage induced by the reactive oxygen species (ROS), including irreversible loss of motility, lipid peroxidation and DNA fragmentation, interfering in the sperm fertilization capacity. The objective of this work was to evaluate the effect of the addition of the antioxidants Hydroxy butyltoluene (BHT) and ethanolic extract of the stem shell of the Cumbretum leprosum to the Botucrio® diluent for cryopreservation of equine semen. Four ejaculates of four Quarter Horse stallions were collected through artificial vagina. After thawing the samples in a water bath at 37ºC, the lipid peroxidation rate of spermatozoa was estimated by measuring the level of malonaldehyde. The experimental design was a randomized block design. The variables of seminal volume, sperm motility, sperm vigor and HOST were submitted to analysis of variance (ANOVA) using the procedure general linear models (Proc GLM) and for Tukey test, in the probability of 5%. The malonaldehyde quantification was submitted to Analysis of Variance (ANOVA), followed by Tukey as post hoc test, in the probability of 5%. There was no difference, since the increase of BHT in concentrations 5, 1.0 and 2.0 mM of BHT and of Cumbretum leprosum extract in doses of 30, 60 and 120 mg to Botucrio diluent did not improve in sperm quality. In this way the antioxidants were not determined in promoting the decrease free radical formation and, consequently, did not improve the quality of the semen after the cryoinjury causing damage to the spermatozoa membrane. Supplementation at 0.5, 1.0 and 2.0 mM doses of BHT and Cumbretum leprosum extract at doses of 30, 60 and 120 mg to the botucrio diluent did not influence the seminal quality after cryopreservation.