390 VIII. HEMATIN ENZYMES, I. CYTOCHROME SYSTEM 



and the reaction between hemoglobin and nitrite in the absence of oxygen: 

 NOT + 2 Hb + H+ -^ HiOH + HbNO 



6.3.3. Oxidation of Groups in Globin. The oxidant may, in 

 some instances, not only oxidize the ferrous iron of hemoglobin to 

 ferric iron, but may also oxidize groups in the globin. While Conant 

 and co-workers {^79), in their investigations of the reaction between 

 hemoglobin and ferricyanide, did not find evidence for any other 

 reaction but the oxidation of the hemoglobin iron by one equivalent 

 of ferricyanide (at pH 7 or below) (cf. also 19 J/.!), Schiiler (2471) 

 found that in the reaction between guinea pig carbon monoxide 

 hemoglobin and ferricyanide at pH 9.5 more ferricyanide is reduced 

 than corresponds to the oxidation of four atoms of iron per mole — 

 64,000 — of hemoglobin; two additional moles of ferricyanide are 

 used for the oxidation of the sulfhj'dryl groups of the globin {2^71). 

 Mirsky and Anson (1963) found that at a pH above 7 maximally two 

 active sulfhydryl groups are present in hemoglobin, which disappear 

 by oxidation with ferricyanide. At a pH below 7, however, the 

 sulfhydryl groups are nonreactive and ferricyanide oxidizes only the 

 iron. According to this two different hemoglobins exist, one of which, 

 produced by ferricyanide at pH 6-8, contains unaltered globin, while 

 the other, produced by ferricyanide in more alkaline solution, con- 

 tains a dehydroglobin. It will be shown below that these observations 

 are of particular interest for the theory of autoxidation of hemoglobin 

 to hemoglobin. 



On the basis of differences in the catalytic effect on glucose oxidation in 

 the erythrocyte, Warburg and his school {2941<2943,294i) claimed that 

 different hemzglol)ins existed. The hem/globin formed in the erythrocyte 

 by amyl nitrite caused only an induced oxidation of glucose, the hemoglobin 

 being then converted by oxygen to oxyhemoglobin which did not react 

 further. The hem/globin produced by phenylhydroxylamine, however, was 

 found to cause a true catalysis, many moles of glucose being oxidized per 

 mole of phenylhydroxylamine. Warburg assumed that in the latter case an 

 abnormal hem/globin was reduced by glucose to an abnormal hemoglobin 

 which with oxygen, instead of yielding oxyhemoglobin, was reconverted to 

 the hem/globin. The investigations of Jung (144^) and of Keilin and Hartree 

 {IJ^OG) have shown, however, that the catalytic effect is one of phenylhydroxy- 

 lamine rather than of hem/globin. Phenylhydroxylamine in reacting with 

 oxyhemoglobin is itself oxidized to nitrosobenZene in a coupled oxidation {cf. 

 Chapter X). Nitrosobenzene can unite with hemoglobin to nitrosobenzene 

 hemoglobin, which Keilin and Hartree believed to be autoxidizable. The 

 catalytic effect is better explained by the observation of Jung that nitroso- 



