Date of Award:
Master of Science (MS)
Animal, Dairy, and Veterinary Sciences
Thomas D. Bunch
Free radicals are short-lived molecules that can cause decreased embryonic development in vitro. Antioxidants are molecules that block free radical formation or guard against their harmful effects. Many studies have linked exposure of media to light and culturing of embryos in high (20%) oxygen concentrations to free radical production. Some of the antioxidants used in culture media are superoxide dismutase (SOD), catalase, zinc (II), ethylenedinitrilo tetraacetic acid (EDTA), mannitol, vitamin E, dimethyl sulfide, and taurine. Most research involving antioxidants and embryonic development has been conducted on non-farm animals, particularly mouse and rabbit. Studies have shown that antioxidants in vitro culture improved embryo development to the blastocyst stage.
In this study, we evaluated the effects of SOD and catalase on bovine embryo development. Four concentrations of SOD (0, 1500, 3000, 6000 IU/ml) and catalase (0, 75, 100, 125 μg/ml) and combinations of the two antioxidants were evaluated through maturation, fertilization, and culture. SOD and catalase were first reconstituted in water and then diluted to their final concentrations. Oocytes were matured in M-199 plus 0.5 μg/ml LH, 5 μg/ml FSH, and 10% FBS at 39°C in 5% CO2 for 24 hours. They were then placed in fertilization-TALP with heparin and 1 x 106/ml sperm. Embryos were cultured in CR2 medium supplemented with alanine, glycine, and 3 mg/ml of fatty-acid free bovine serum albumin in modular incubators with 5% CO2, 5% O2, and 90% N2. Embryo development was evaluated on day 8. Three replicates with approximately 50 embryos per treatment were used to evaluate the effects of SOD and catalase. The control had better embryo development than all treatments. The treatment that was most similar to the control was treatment 2, which consisted of no SOD and 75 μg/ml catalase.
Based on these observations, levels of both SOD and catalase were lowered to 0, 100, 250, and 500 IU/ml and 0, 10, 25, and 50 μg/ml, respectively. Although these levels appeared to improve embryo development, there were no statistical differences. Based on the culture system and media currently used along with the precautions against light and oxygen concentration, we did not find any beneficial effects of supplementing medium with SOD or catalase.
Anderson, Bret L., "Effects of Antioxidants on Development of In Vitro Fertilized Bovine Embryos" (1995). All Graduate Theses and Dissertations. 3920.