Energy expenditure (EE) and substrate oxidation were measured in six male and four female adult humans during day (10.00-23.00) and night (23.00-07.00). All subjects received diets of the same composition, but four subjects (Group I) had a higher intake of metabolizable energy (ME) than FE, while ME was near EE in three subjects (Group II) and below EE in three subjects (Group III). The EE varied from 388 kJ/h in Group I to 540 kJ/h in Group III during day and from 248 to 351 kJ/h during night. However, the reduction from day to night was identical for all subjects with a night value of 64% of the day value. Oxidation of carbohydrate (OXCHO) was fairly constant with 223 kJ/h during day and being reduced to 100 kJ/h during night in all subjects. Oxidation of fat (OXF) varied from 121 kJ/h in Group I to 223 kJ/h in Group III during day and from 98 to 190 kJ/h by night. The decreased EE from day to night was of the same magnitude independent of the ME intake, while the contribution from OXCHO and OXF to EE depended of ME intake. When ME intake was above EE (Group I), relatively more carbohydrate (56%) than fat (31%) was oxidized during day, leaving a surplus of dietary fat for fat retention. Nevertheless, during night the contributions from OXCHO and OXF were similar (40%). When energy supply was below EE (Group III), the contributions from OXCHO and OXF to EE were similar (45%) during day. However, during night the contribution from OXCHO was reduced (29%) while OXF increased (54%). This investigation indicates that during day the major oxidative fuel is carbohydrate, while fat oxidation reflects the difference between OXCHO and EE; however, during night changes in the energy status are accommodated by increasing OXF.