Animal Reproduction (AR)
Animal Reproduction (AR)
Original Articles

Cat epididymal semen cryopreserved with and without vitamin E: effect on sperm parameters and lipid peroxidation

Beatrice Ingrid Macente, Raquel Ribeiro Gutierrez, Maricy Apparício, Cristiano de Carvalho Balieiro, Cleber Fernando Menegasso Mansano, Marcelo Maia Pereira, Juliana Corrêa Borges-Silva, Eliandra Antonia Pires-Buttler, André Luis Batista Galvão, Gilson Hélio Toniollo, Gaia Cecília Luvoni, Maria Giorgia Morselli, Wilter Ricardo Russiano Vicente

Downloads: 3
Views: 966


The aims of this study were to investigate: 1) if the addition of α-tocopherol (vitamin E) in three concentrations (0.3, 0.6 and 0.9 mM) is able to preserve spermatozoa integrity after thawing and 2) the effect of a-tocopherol supplementation on lipid peroxidation. Fifty four domestic cats were used in this study constituting 18 pools (3 cats per pool). Each pool was submitted at four experimental groups: group 0 (control) – epididymal sperm were frozen with a commercial Botucrio® extender; group 0.3, group 0.6 and group 0.9 – the extender was supplemented with 0.3, 0.6 and 0.9 mM of a-tocopherol, respectively. Each semen sample was evaluated for motility, progressive forward motility, morphology, sperm viability (plasma membrane integrity-PMI), hypo-osmotic swelling test (HOST), before and after thawing. The evaluation of lipid peroxidation reaction by Thiobarbituric Acid Reactive Substances (TBARS) test was performed on thawed semen only. Results demonstrated that there was no significant difference between control and the three a-tocopherol groups with regards to motility and progressive motility after thawing (P > 0.05). As expected, in fresh samples viability was significantly higher than in all the cryopreserved groups in which there was no positive influence of any of the a-tocopherol concentration used. Lipid peroxidation was higher in the supplemented groups 0.6 and 0.9 mM of a-tocopherol than in control and in 0.3 mM group. In conclusion, the addition of a-tocopherol to the commercial extender had no positive influence on reduction of lipid peroxidation. This topic deserves further investigations to better understand the effect of cryopreservation procedures on epididymal spermatozoa and to establish adequate strategies to counteract sperm cryodamages.


antioxidant, feline, oxidative stress, sperm, a-tocopherol.


Axnér E, Hermansson U, Linde-Forsberg C. 2004. The effect of Equex STM paste and sperm morphology on post-thaw survival of cat epididymal spermatozoa. Anim Reprod Sci, 84:179-191.

Baumber J, Ball BA, Linfor JJ, Meyers SA. 2002. Reactive oxygen species and cryopreservation promote deoxyribonucleic acid (DNA) damage in equine sperm. Theriogenology, 58:301-302.

Belala R, Fatmi S, Kaidi R, Iguer-Ouada M. 2016. Benefits of cholesterol and α-tocopherol loaded cyclodextrins in dog semen cryopreservation. Revue Méd Vét, 167:22-27.

Breininger E, Beorlegui NB, O’Flaherty CM, Beconi MT. 2005. Alpha-tocopherol improves biochemical and dynamic parameters in cryopreserved boar semen. Theriogenology, 63:2126-2135.

Buege JA, Aust SD. 1978. Microsomal Lipid peroxidation. Methods Enzymol, 52:302-310.

Buranaamnuay K. 2017: Protocols for sperm cryopreservation in the domestic cat: A review. Anim Reprod Sci, 183:56-65.

Castellini C, Lattaioli P, Dal Bosco A, Minelli A, Mugnai C. 2003. Oxidative status and semen characteristics of rabbit buck as affected by dietary vitamin E, C and n-3 fatty acids. Reprod Nutr Dev, 43:91-103

Cocchia N, Ciani F, El-Rass R, Russo M, Borzacchiello G. 2009. Cryopreservation of feline epididymal spermatozoa from dead and alive animals and its use in assisted reproduction. Zygote, 18:1-8.

Comercio EA, Monachesi NE, Loza ME, Gambarotta M, Wanke MM. 2013. Hypo-osmotic test in cat spermatozoa. Int J Androl, 45:310-314.

Donnelly ET, McClure N, Lewis SE. 1999. The effect of ascorbate and α-tocopherol supplementation in vitro on DNA integrity and hydrogen peroxide-induced DNA damage in human spermatozoa. Mutagenesis, 14:505-512.

Hatamoto LK, Sobrinho BCA, Nichi M, Barnabe VH, Barnabe RC. 2006. Effects of dexamethasone treatment (to mimic stress) and Vitamin E oral supplementation on the spermiogram and on seminal plasma spontaneous lipid peroxidation and antioxidant enzyme activities in dogs. Theriogenology, 66:1610-1644.

Jeyedran RS, Van der Ven HH, Perez-Pelaez M. 1984. Development of an assay to assess the functional integrity of human sperm membrane and its relationship to other semen characteristics. J Reprod Fertil, 70:219-228.

Lopes BV, Monteiro GA, Ovidio PP, Jordão Jr AA, Lopes MD. 2010. Avaliação do estresse oxidativo no plasma seminal de cães férteis e subférteis após suplementação oral com vitamina C e E. Vet Zootec, 18:452-461.

Luvoni GC, Kalchschmidt E, Marinoni G. 2003. Conservation of feline semen. Part I and II. J Feline Med Surg, 5:203-208 and 257-263.

Macente BI, Mansano CFM, Pereira MM, Martins MIM, Gioso MM, Savi PAP, Gutierrez RR. 2012. Congelação de espermatozóides epididimário de gatos utilizando o diluidor Botucrio após refrigeração por 24hs em contêiner de transporte de sêmen Botu-tainer. Acta Vet Bra, 6:112-117.

Martins MIM, Padilha LC, Souza FF, Lopes MD. 2009. Fertilizing capacity of frozen epididymal sperm collected from dogs. Reprod Domestic Animals, 44:342-344.

Melo CM, Zahn FZ, Martin I, Orlandi C, Dell´Acqua Jr JA. 2007. Influence of semen storage and cryoprotectant on post-thaw viability and fertility of stallion spermatozoa. J Equine Vet Sci, 27:171-175.

Michael AJ, Alexopoulos C, Pontiki EA, Hadjipavlou-Litina DJ, Saratsis P. 2007. Effect of antioxidant supplementation on semen quality and reactive oxygen species of frozen-thawed canine spermatozoa. Theriogenology, 68:204-212.

Paya M, Halliwel B, Hoult JR. 1992. Interactions of a series of coumarins with reactive oxygen species: scavenging of superoxide, hypochlorous acid and hydroxyl radicals. Biochem Pharmacol, 44:205-214.

Satorre MM, Breininger E, Beconi MT. 2012. Cryopreservation with α-tocopherol and Sephadex filtration improved the quality of boar semen. Theriogenology, 78:1548-1556.

Silva SV, Soares AT, Batista AM, Almeida FC, Nunes JF. 2013. Vitamin E (Trolox) addition to Tris-egg yolk extender preserves ram spermatozoon structure and kinematics after cryopreservation. Anim Reprod Sci, 137:37-44.

Statistical Analyses System SAS Institute (SAS Institute). 2008. SAS/STAT 9.2 User’s guide. SAS Institute Inc, Cary, NC.

Towhidi A, Parks JE. 2012. Effect of n-3 fatty acids and α-tocopherol on post-thaw parameters and fatty acid composition of bovine sperm. J Assist Reprod Genet, 29:1051-1056.

Thuwanut P, Chatdarong K, Johannisson A, Bergqvist A-S, Söderquist L. 2010. Cryopreservation of epididymal cat spermatozoa: effects of in vitro antioxidative enzymes supplementation and lipid peroxidation induction. Theriogenology, 73:1076-1087.

Thuwanut P, Chatdarong K, Techakumphu M, Axnér E. 2008. The effect of antioxidants on motility, viability, acrosome integrity, and DNA integrity of frozen-thawed epididymal cat spermatozoa. Theriogenology, 70:233-240.

Zambelli E, Prati F, Cunto M, Iacono E, Merlo B. 2010. Quality and fertilizing ability of electroejaculated cat spermatozoa frozen with or without Equex STM Paste. Theriogenology, 73:886-892.

5c07d9630e882580680e27c1 animreprod Articles
Links & Downloads

Anim Reprod

Share this page
Page Sections