The average degree of reduction of mitochondrial NAD has been measured in the intact toad sartorius by a fluorometric technique. It has been shown that cytoplasmic NADH does not interfere materially with these measurements. The percentage reduction of this respiratory coenzyme has been determined in a number of physiological steady states which are well correlated with fluorometrically determined levels of NADH in suspensions of mitochondria from the hind leg musculature of the toad. In addition, these findings are closely comparable to similar, spectrophotometric measurements on mitochondria from other sources. In the presence of an adequate O2 level a single twitch produces a decrease in fluorescence from the resting steady state which is followed by a slow return to the base line condition. This cycle indicates the intensity and the time course of the oxidative recovery metabolism. The area under this curve is directly related to the number of twitches up to three or four. Greater activity produces a curtailment of oxidative recovery due to glycolysis. In the presence of iodoacetate the linear relation holds for five to seven twitches. At still higher levels of activity a curtailment of the change in NAD level sets in, probably due to the removal of AMP by catabolic reactions.

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