The effect of pH on the Km values for ammonia was studied in its oxidation by Nitrosomonas cells and cell-free extracts. The Km values decreased markedly with increasing pH, suggesting (NH3) rather than (NH,+) as the actual substrate for oxidation. Nitrosomonas europaea and other nitrifying bacteria are normally considered to use ammonium ion (NH,+) as substrate for oxidation to

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Conditions essential for the preparation of active ammonia-oxidizing extracts of Nitrosomonas europaea were studied. The extracts were unstable during storage and required specific assay conditions for ammonia oxidation. Bovine serum albumin, spermine, or MgCl2 was required for ammonia oxidation and the concentration of phosphate determined the relative effectiveness of each activator, i.e., albumin being most effective in 0.1 M phosphate and spermine or MgCl2 at lower phosphate concentrations. Kinetic experiments showed a partial reduction of cytochrome c preceding the initiation of oxygen consumption due to ammonia oxidation.

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