Aflatoxin is a potent carcinogen often found in animal feedstuffs. Although it reportedly impairs development of the preimplantation pig embryo, it is not known whether it adversely affects development of the preimplantation bovine embryo. We conducted 3 experiments to investigate this possibility and determine whether deleterious effects of aflatoxin were caused by increased production of reactive oxygen species (ROS). Experiments were conducted with embryos produced in vitro and cultured after fertilization with various concentrations of aflatoxin. For experiment 1, embryos were treated with 0 (control), 40, 400, or 4,000 µg/L of aflatoxin B (AFB). Treatment at all concentrations of AFB tended to reduce cleavage rate, with the 2 highest concentrations having significant effects. As compared with the control, 40 µg/L AFB reduced the percentage of oocytes becoming blastocysts and the percentage of cleaved embryos becoming blastocysts (19.7 vs. 8.1% and 30.3 vs. 14.3%, respectively). Complete inhibition of blastocyst formation occurred at concentrations of 400 and 4,000 µg/L of AFB. Experiments 2 and 3 involved a 2 × 2 factorial design with effects of AFB (0 and 40 µg/L), the antioxidant Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid, a water-soluble analog of vitamin E; 0 and 5 µM), and their interaction on production of ROS in putative zygotes (experiment 2) and development to the blastocyst stage (experiment 3). Production of ROS was increased by AFB, and this effect was reversed by Trolox. However, Trolox did not prevent the reduction in development to the blastocyst stage caused by AFB. Thus, the anti-developmental effects of AFB are not caused solely by increased ROS production. Rather, other underlying mechanisms exist for the adverse effects of aflatoxin on embryonic development. Overall, results indicate the potential for feeding aflatoxin-contaminated feed to cause embryonic loss in cattle.