Ketogenesis in isolated rat liver mitochondria I. Relationships with the citric acid cycle and with the mitochondrial energy state

1. A method is described to calculate the distribution of acetyl-CoA over the citric acid cycle and ketogenesis during the oxidation of fatty acids in the presence of added malate. 2. Increasing concentrations of added Krebs cycle intermediates lower the rate of ketogenesis both in the low-energy state (State 3; in the presence of glucose and hexokinase (EC 2.7.1.1)) and in the high-energy state (State 4). 3. In State 3 acetyl-CoA is initially used almost exclusively for the synthesis of citrate. Citrate accumulates in the medium, the concentration of malate decreases and a parallel increase in the rate of ketogenesis is observed. 4. The rapid accumulation of citrate in State 3 is also found during oxidation of pyruvate plus malate. 5. Uncouplers have no effect on the distribution of acetyl-CoA and on the accumulation of citrate in State 3. 6. Transition from State 3 to State 4 is accompanied by an inhibition of the Krebs cycle and an increased ketogenesis. 7. Under all conditions tested the relative rate of ketogenesis and the 3-hydroxy-butyrate/acetoacetate ratio were positively correlated. 8. Addition of ATP and oligomycin to uncoupled mitochondria did not affect ketogenesis. 9. Our results indicate that the concentration of added Krebs cycle intermediates and the NADH/NAD+ ratio are the only factors controlling the entry of acetyl groups into the cycle