OXIDATIVE MECHANISMS IN ANIMAL TISSUES 19 



blue is autoxidizable, it was possible in some cases to carry out the 

 air oxidation of a substrate by the addition of its specific dehydro- 

 genase and methylene blue. Such an oxidation was not, however, 

 affected by cyanide or carbon monoxide and in this respect did not 

 resemble the oxidation of the substrate by the living cell. As knowl- 

 edge concerning the cytochrome system increased, it was soon 

 realized that these iron porphyrin compounds played the role of 

 methylene blue within the cells. Thus it was generally agreed about 

 ten years ago that activation of the substrate was brought about by 

 a specific dehydrogenase and that then the substrate reacted with 

 oxygen through the cytochrome chain. 



This, then, was the state of affairs in 1930 when Professor War- 

 burg came to this country to deliver lectures on his work on what 

 we now call cytochrome oxidase. Barron and Harrop (9) had shortly 

 before published experiments showing that the addition of methyl- 

 ene blue to non-nucleated red blood cells brought about an oxygen 

 consumption if glucose was present as a substrate. While Professor 

 Warburg was at Johns Hopkins, Dr. Barron obligingly repeated his 

 experiments at the request of his distinguished visitor, who watched 

 the proceedings carefully. Upon his return to Germany, Warburg 

 himself repeated the experiments and with his collaborators began 

 the isolation of the red blood cell constituents responsible for this 

 effect. Thus was begun a series of studies which brought forth some 

 of the most noteworthy advances ever made in this field. As you 

 know, these experiments led to the discovery of the vitamin- 

 containing coenzymes essential to the functioning of most dehydro- 

 genase systems. They showed that the activated hydrogen of the 

 substrate did not react directly with the cytochrome system, but 

 that at least two reversible oxidation-reduction systems were inter- 

 posed. 



So we have today the following general picture of the pathway 

 of oxidations from the substrate side. In the presence of a specific 

 protein and of a particular organic compound of low molecular 

 weight, often called a coenzyme, the substrate loses two electrons 

 and two hydrogen ions. In the majority of cases so far studied this 

 coenzyme is one of the pyridine nucleotides. In these cases two elec- 

 trons and one hydrogen ion are accepted by the pyridine nucleotide, 

 the other hydrogen ion being released to the environment. We do 

 not know whether both the pyridine nucleotide and the protein are 

 concerned in the activation of the substrate molecule. If the protein 

 alone is responsible for this activation, the pyridine nucleotide may 



