AbstractThe fatty acid composition and metabolic activity of cholesterol esters in milk and mammary tissue (cow, sow and goat) were investigated. Cholesterol esters of freshly secreted milks incubated at 40 C for 2 hr showed no change in fatty acid composition and no incorporation of 1‐14C‐palmitate. The fatty acid composition of cholesterol esters in the milk of all three species exhibited elevated levels of a unique group of fatty acids when compared to other milk ester lipid classes. This group was comprised of monounsaturated acids and acids with odd numbers of carbons. Tissue cholesterol esters contained lower levels of these acids. Evidence from experiments in which an odd carbon acid (C15) was infused into the lactating mammary gland indicated that the group of unique acids is preferentially retained in the cholesterol ester fraction which is secreted with milk. These infusion experiments also provided evidence that cholesterol esters accumulate in developing milk fat globules in a manner paralleling triglyceride accumulation, and that acyl moieties of cholesterol esters may be desaturated in the form of the intact ester. Our results are compatible with the hypothesis that acyl moieties of cholesterol esters in lactating mammary tissue are turning over rapidly.