Primary cultures of rat hepatocytes were used to determine unscheduled DNA synthesis (UDS) and cytotoxicity of purified 4-deoxynivalenol (vomitoxin), a trichothecene mycotoxin produced on cereal grains by fungi of the genus Fusarium. Nontoxic and toxic doses of deoxynivalenol, 0.1 to 1000 micrograms/ml, did not significantly increase UDS as measured by net grains per nucleus, net grains per nuclear area or percentage of cells incorporating greater than or equal to 5, 6, 10 or 20 grains per nucleus. Evidence of cytotoxicity, manifested as a reduction in cell number in autoradiographs, pyknotic nuclei or vacuolated cytoplasm, was observed in hepatocytes treated with deoxynivalenol concentrations of 5 micrograms/ml and above. These findings suggest that the cellular toxicity of deoxynivalenol may not be mediated by a DNA-damaging event in cultured hepatocytes. An increased percentage of large-sized nuclei was also found to be associated with toxic doses of deoxynivalenol as well as 2-acetylaminofluorene used as the positive control.