In a previous study, we showed that lean subjects are capable of rapidly adjusting fat oxidation to fat intake on a high-fat (HF) diet when glycogen stores are lowered by exhaustive exercise. However, it has been proposed that obese subjects have impaired fat oxidation. We therefore studied the effect of low glycogen stores on fat oxidation after a switch from a reduced-fat (RF) diet to an HF diet in obese subjects. Ten healthy, obese male and female subjects (26 +/- 2 yr, body mass index 31.8 +/- 1.4, maximal power output 228 +/- 14 W) consumed an RF diet (30, 55, and 15% of energy from fat, carbohydrate, and protein, respectively) at home for 3 days on four occasions (days 1-3). On two occasions, subjects came to the laboratory on day 3 at 1500 to perform an exhaustive glycogen-lowering exercise test (Ex), after which they went into a respiration chamber for a 36-h stay. On the other two occasions, subjects directly entered the respiration chamber at 1800 for a 36-h stay. In the respiration chamber, they were fed, in energy balance, either an HF diet (60, 25, and 15% of energy from fat, carbohydrate, and protein, respectively) or an RF diet. All diets were consumed as breakfast, lunch, dinner, and two or more snacks per day. Twenty-four-hour respiratory quotient was 0.91 +/- 0.01, 0.89 +/- 0.01, 0.84 +/- 0.01, and 0.81 +/- 0.01 with RF diet, RF + Ex, HF, and HF + Ex treatments, respectively. With the HF treatment, fat oxidation was below fat intake, indicating the slow change of oxidation to intake on an HF diet. After the HF + Ex treatment, however, fat oxidation matched fat intake. In conclusion, obese subjects are capable of rapidly adjusting fat oxidation to fat intake when glycogen stores are lowered by exhaustive exercise.