The inhibitors usually associated with the activity of the cytochrome oxidase system—cyanide and carbon monoxide—are also effective in reducing the oxidation of H2 by intact cells of Azotobacter vinelandii. The hydrogenase system is more sensitive to CO than is the respiratory system.

Oxidation of a carbon source and of hydrogen by Azotobacter cells is inhibited in a quantitatively different manner by the following compounds: sodium azide, hydroxylamine, sodium iodoacetate, and sodium fluoride. In every case, a concentration range which is definitely inhibitory for respiration has little or no effect on the hydrogenase activity.

The differential inhibition by hydroxylamine explains certain observations in the literature which have been erroneously interpreted as demonstrating a specific inhibition by NH2OH of biological nitrogen fixation. This supposed demonstration has been offered as support for the hypothesis that NH2OH is an intermediate in the fixation reaction.

The differential inhibitors can be used for detection of hydrogenase in cultures possessing a high endogenous respiration. The method is illustrated by an experiment with root nodule bacteria from pea and cowpea nodules. No hydrogenase was found in either.

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