Flying insects achieve the highest known mass-specific rates of O(2) consumption in the animal kingdom. Because the flight muscles account for >90% of the organismal O(2) uptake, accurate estimates of metabolic flux rates (J) in the muscles can be made. In steady state, these are equal to the net forward flux rates (v) at individual steps and can be compared with flux capacities (V(max)) measured in vitro. In flying honeybees, hexokinase and phosphofructokinase, both nonequilibrium reactions in glycolysis, operate at large fractions of their maximum capacities (i.e., they operate at high v/V(max)). Phosphoglucoisomerase is a reversible reaction that operates near equilibrium. Despite V(max) values more than 20-fold greater than the net forward flux rates during flight, a close match is found between the V(max) required in vivo (estimated using the Haldane relationship) to maintain near equilibrium and this net forward flux rate and the V(max) measured in vitro under simulated physiological conditions. Rates of organismal O(2) consumption and difference spectroscopy were used to estimate electron transfer rates per molecule of respiratory chain enzyme during flight. These are much higher than those estimated in mammalian muscles. Current evidence indicates that metabolic enzymes in honeybees do not display higher catalytic efficiencies than the homologous enzymes in mammals, and the high electron transfer rates do not appear to be the result of higher enzyme densities per unit cristae surface area. A number of possible mechanistic explanations for the higher rates of electron transfer are proposed.