The modifying effect of dietary tuna (Thunnus thynnus orientalis) orbital oil rich in docosahexaenoic acid (DHA) and vitamin D3 (VD3) on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) was investigated in male F344 rats. Animals were given three weekly subcutaneous injections of AOM (15 mg/kg body weight) to induce ACF. The rats were fed the experimental diet containing 5% tuna orbital oil (low fish oil), 23.5% tuna orbital oil (high fish oil), 5% corn oil (low corn oil) or 23.5% corn oil (high corn oil) for 5 weeks, starting 1 week before the first dose of AOM. Animals were sacrificed 2 weeks after the last AOM injection to count colonic ACF and assay the expression of cyclooxygenase (COX)-1 and -2. High corn oil diet significantly increased the development of ACF, when compared with low corn oil diet (P<0.005). High fish oil diet also increased ACF formation compared with low fish oil diet (P<0.01), but the increase was smaller than high corn oil diet. The frequency of ACF was significantly lower in the rats fed high fish oil diet than high corn oil diet (P<0.02). Moreover, frequency of ACF consisted of 4 or more crypts in rats fed the high fish oil diet was significantly lower than that of rats given high corn oil diet. COX-1 and COX-2 expression did not significantly differ among the groups. These results suggest that fish oil derived from tuna, which contains high amounts of DHA and VD3, suppresses the formation and growth of ACF without affecting COX-1 and COX-2 expression, and may have a preventive effect on colon carcinogenesis.