There is increasing evidence that individual isomers of conjugated linoleic acid (CLA) may have unique biological or biochemical effects. A primary objective of this study was to determine whether there might be differences in the anticancer activity of 9,11-CLA and 10,12-CLA. This was achieved by evaluating the reduction in premalignant lesions and carcinomas in the mammary gland of rats that had been treated with a single dose of methylnitrosourea and given 0.5% of either highly purified CLA isomer in the diet. Our results showed that the anticancer efficacies of the two isomers were very similar. At 6 wk after carcinogen administration, the total number of premalignant lesions was reduced by 33-36%. At 24 wk, the total number of mammary carcinomas was reduced by 35-40%. The concentration of each CLA isomer and its respective metabolites was analyzed in the mammary fat pad. Tissue level of 10,12-CLA was much lower than that of 9,11-CLA. The pool of metabolites from each isomer was very similar between the two groups and represented only a small fraction of total conjugated diene fatty acids. Feeding of 9,11-CLA resulted in minimal changes in other unsaturated fatty acids. In contrast, feeding of 10,12-CLA produced a wider spectrum of perturbations. Small but significant increases in 16:1 and 16:2 were detected; these were accompanied by decreases in 20:2, 20:3, 20:4, 22:4, and 22:6. The above observation suggests that 10,12-CLA might be more potent than 9,11-CLA in interfering with elongation and desaturation of linoleic and linolenic acids. In summary, our study showed that, at the 0.5% dose level, the anticancer activity of 9,11-CLA and 10,12-CLA was very similar, even though accumulation of 10,12-CLA in the mammary tissue was considerably less than that of 9,11-CLA. These confounding changes of the other unsaturated fatty acids in contributing to the effect of 10,12-CLA need to be clarified.