Expansion of the fat stores has been proposed as a prerequisite for increasing fat oxidation in response to a high-fat diet in individuals with a predisposition to obesity. In a cross-sectional design we measured 24-h substrate oxidations on a standardized diet in 38 overweight or obese and 35 nonobese women. Fat oxidation (g/d) was mainly a function of total energy requirements (r = 0.71, P < 0.0001). To account for this we used for further analysis oxidative fat energy (%), a counterpart to dietary fat energy (%). After differences in fat energy of consumed food (%), age, and 24-h energy balance were adjusted for, obese women had higher oxidative fat energy than did nonobese women [40.2% (37.8-42.6) vs 36.0% (33.6-38.5), P < 0.02]. Adjusted oxidative fat energy (%) increased with increasing size of fat mass (r = 0.31, P < 0.01). This relation suggests that a 10-kg change in fat mass may be caused by a change in dietary fat energy of > 1.6% (0.4-2.7%). The study supports the concept that in susceptible individuals the expansion of fat stores is a prerequisite to increase the oxidative fat energy to an amount commensurate with a high percentage of dietary fat energy.