The importance of either carbohydrate or energy restriction in initiating the metabolic response to fasting was studied in five normal volunteers. The subjects participated in two study protocols in a randomized crossover fashion. In one study the subjects fasted for 84 h (control study), and in the other a lipid emulsion was infused daily to meet resting energy requirements during the 84-h oral fast (lipid study). Glycerol and palmitic acid rates of appearance in plasma were determined by infusing [2H5]glycerol and [1-13C]palmitic acid, respectively, after 12 and 84 h of oral fasting. Changes in plasma glucose, free fatty acids, ketone bodies, insulin, and epinephrine concentrations during fasting were the same in both the control and lipid studies. Glycerol and palmitic acid rates of appearance increased by 1.63 +/- 0.42 and 1.41 +/- 0.46 mumol.kg-1.min-1, respectively, during fasting in the control study and by 1.35 +/- 0.41 and 1.43 +/- 0.44 mumol.kg-1.min-1, respectively, in the lipid study. These results demonstrate that restriction of dietary carbohydrate, not the general absence of energy intake itself, is responsible for initiating the metabolic response to short-term fasting.