Metabolic Pathways of Bacterial Nitrification 397 



Reaction ( 1 ) can take place using methlene blue as the hydrogen 

 acceptor with the presumption that the hypothetical nitroxyl 

 (NOH) is the unstable and unidentified intermediate at the 

 oxidation state of plus 1 for tlie nitrogen atom. Reaction (2) re- 

 quired O2 and is completely inhibited by cyanide whereas re- 

 action (3) is ascribed to the unstabihty of (NOH). With the 

 exception of hydroxylamine, the intermediates during the enzyma- 

 tic conversion of ammonia to nitrite by Nitrosomonas are not 

 known. 



Although in vivo ammonia oxidation is highly sensitive to 

 various metal binding agents, there is no information regarding 

 the participation of any metal in the ammonium or hydroxylamine 

 metabolism of these bacteria. Similarly there is virtually no in- 

 formation about the electron transport system and the energy 

 coupling mechanisms (other than the recent preliminary reports 

 of Nicholas and Jones (34), and Delwiche (15) cited above) 

 as well as assimilatory processes in these micro-organisms, 



NITRITE OXIDATION BY NITROBACTER 



Nitrobacter sp. as strict chemoautotrophs derive their entire 

 energy for growth and metabolism from nitrite oxidation (with 

 an energy efficiency of 5 to 8%) according to the equation (10) : 



NO2- + /2 O2 -^ NO,- + 17.5 K.Cal. 



Optimal nitrite oxidation by intact cells occurs at pH 8.0 and 

 at a substrate concentration of 3 to 5 X IQ-^ M at 28-30° C (2). 

 The optimal nitrite concentration for the Nitrobacter strain used 

 by Lees and Simpson (28) was somewhat lower, namely, 4 X 

 10^^ M. The bacterium requires 5 ppm of each magnesium and 

 phosphorus, and 7 /'g of iron per liter for the optimal growth 

 as studied in a modified growth medium devoid of calcium car- 

 bonate particles (2). Although a slight stimulation in growth 

 was observed with traces of molybdenum and zinc (1 /'g/L), 

 the results were variable. Neither vitamins, amino acids nor any 

 other organic conf actors are required for its normal growth (2). 

 Zavarzin (39) also reported that although nitrite oxidation was 

 stimulated by molybdenum or tungstate in the presence of iron, 

 it was somewhat depressed when both molybdenum and tungstate 



