The present study was designed to determine the long-term effects of orotic acid (OA), a multi-organ tumor promoter, in rats not exposed to any carcinogen. Male Fischer 344 rats (130-150 g) were divided into two groups and given either a semisynthetic basal diet (BD) or the same diet containing 1% OA. Animals from both groups were killed after 1 or 2 years of treatment. Foci of placental glutathione-S-transferase (GST 7-7) positive hepatocytes were observed in the livers of both BD and OA fed rats killed after 1 year. However, they were more in number in animals receiving OA (156 +/- 21 versus 51 +/- 11/cm3). After 2 years, hepatic nodules were seen in almost all the animals given OA and in approximately 30% of the rats given BD. The nodules were of two main types: (i) a reddish-brown type, present in 85% of rats exposed to OA and in 27% of rats given BD, and (ii) a greyish-white type, found in 50% of animals fed OA and in none of the animals fed BD. These two types of lesions were also histologically different. Reddish-brown nodules were composed of slightly enlarged hepatocytes resembling normal surrounding tissue, while greyish-white nodules were similar in structure and are indistinguishable from hepatic nodules induced by genotoxic chemical carcinogens. The results are interpreted to suggest that the foci/nodules seen in OA-fed rats are due to a promoting effect of OA on spontaneously arising and/or diet-induced altered cells.