The effects of dietary fat on antioxidant mechanisms in colon mucosa were examined using male Sprague-Dawley rats. Animals were fed one of four AIN-76A-based test diets differing in amount and type of fat. The basal diet (BD) contained 5% corn oil; the menhaden oil diet (MO) contained 1% corn oil and 19% menhaden oil; the corn oil diet (CO) contained 20% corn oil; and the beef tallow diet (BT) contained 1% corn oil and 19% beef tallow; all were adjusted to provide equal amounts of other nutrients per kilojoule. Homogenates of colon mucosa were assayed after 2 wk and 1, 3, 6 and 9 mo for activities of catalase (CAT), glutathione peroxidase (GSHPx), superoxide dismutase (SOD), glutathione-S-transferase, glutathione reductase (GSSGRx), and total glutathione (GSH) content. Activity of GSHPx was depressed in rats fed high fat diets compared with rats fed the low fat BD. Animals fed the BD and MO diets had higher levels of CAT and GSSGRx as compared with rats fed the CO and BT diets. Activities of SOD, GSSGRx and GSHPx, as well as GSH concentration, decreased over time. Ornithine decarboxylase activity was not induced by the high fat diets. Thiobarbituric acid-reacting substances were virtually undetectable. Thus, both type and amount of fat affected antioxidant mechanisms in colon mucosa without producing overt oxidative damage in the tissue.