Effects of k-carrageenan supplementation or in combination with cholesterol-loaded cyclodextrin following freezing-thawing process of rooster spermatozoa

Abstract This experimental research purposely seeks to explore the effect of supplementing k-carrageenan (k-CRG) or CLC (cholesterol-loaded cyclodextrins) or the combined effect of k-CRG and CLC as supplements of antioxidants to an extender for rooster semen freezing. A total of 75 neat pooled ejaculates were collected twice a week from twenty-five (25) arbor acres broiler roosters (30 wks) during the experimental period. In each replicate, semen samples (n= 15, three ejaculates for each rooster) were pooled and divided into nine equal aliquots, and each aliquot was diluted with one of the following extender supplemented with k-CRG, CLC, and k-CRG + CLC after which it was subjected to cryopreservation process using the “pellet” method. In study I, the supplementation of extenders with k-CRG was in five equal aliquots as follows; (0.2, 0.4, 0.6, 0.8) mg/mL and control group (k-CRG 0) mg/mL while in Study II, there was a combination of both k-CRG + CLC (0.4 mg/mL + 1.5 mg/mL, respectively), 0.4 mg/mL k-CRG, 1.5 mg/mL CLC and control group. Sperm quality parameters, endogenous antioxidant enzymes, lipid peroxidation (MDA) and ROS were all assessed after the freeze-thaw process. Our findings in study I indicated that at post-thaw, an optimum 0.4 mg/mL k-CRG supplementation in the extender improved semen quality parameters, endogenous enzymes, MDA and ROS in comparison to the control group. Interestingly prior to the freeze-thaw process, it was depicted in study II that combined k-CRG + CLC (0.4 mg/mL+1.5 mg/mL) inclusion in the extender provided maximum protection to sperm quality parameters, endogenous enzymes, MDA and ROS in comparison to 1.5 mg/mL CLC and control group at post-thaw. Besides, there was also a significant difference observed in the extenders supplemented with combined k-CRG + CLC (0.4 mg/mL +1.5 mg/mL) when compared to 0.4 mg/mL k-CRG for semen quality parameters and endogenous antioxidant enzymes (SOD, CAT, and GPx) but no significant difference was observed for MDA and ROS. Also, there was a significant difference observed in the extender supplemented with 1.5 mg/mL CLC when compared to the control group for semen quality parameters, SOD, CAT, and MDA but no significant difference for GPx and ROS at post-thaw. In conclusion, k-CRG at an optimal dosage of 0.4 mg/mL proved effective for improving post-thaw sperm quality but its combined addition k-CRG + CLC at an optimal concentration of (0.4 + 1.5) mg/mL in the extender provided greater protection to the rooster spermatozoa at post-thaw.