FERRICYANIDE 677 



when the ferricyanide is washed out. Ferricyanide slowly injected intraven- 

 ously into mice (1.5 g/kg of sodium salt) produces no disturbance of tumor 

 metabolism, but when the various tissues are treated with re++"'", only the 

 tumor turns blue. The anaerobic glycolysis of no other tissue is depressed 

 by ferricyanide, which in this respect differs from other glycolytic inhibitors 

 (Mendel and Strelitz, 1937). Renal medulla was studied particularly because 

 it has a significant rate of aerobic glycolysis and ferricyanide was found to 

 have no effect (and in some cases even stimulated somewhat). This action 

 was not investigated further until Birkenhager (1959, 1960) attempted to 

 locate the site of inhibition. He confirmed that 10 mM ferricyanide does 

 indeed inhibit both aerobic and anaerobic glycolysis in Walker and Crocker 

 tumors, but not in Ehrlich ascites cells, and further showed that it inhibits 

 the extra glycolysis brought about by dinitrophenol. The respiration in the 

 presence of glucose rises 30-60% in the presence of ferricyanide in the two 

 former tumors, but not in the ascites cells. The problem of what happens 

 to the glucose taken up, since this is not depressed as much as lactate for- 

 mation, remains unsolved. A small accumulation of pyruvate was found 

 under aerobic conditions but not anaerobically, and no other glycolytic in- 

 termediates could be detected. Use of glucose- 1-C^* and glucose-6-C^* point- 

 ed to the conclusion that ferricyanide either directly or indirectly inhibits 

 glycolysis at the level of phosphohexose isomerase or phosphohexokinase; 

 this would make more hexose phosphate available for the pentose phosphate 

 shunt. However, aldolase was found to be very sensitive to ferricyanide 

 (88% inhibition at 0.5 mM) and addition of aldolase to a tumor extract in 

 which glycolysis has been abolished by ferricyanide leads to recovery. Phos- 

 phoglyceraldehyde dehydrogenase is not sensitive to ferricyanide nor does 

 its addition reverse the glycolytic inhibition. Birkenhager ascribed the dif- 

 ference between cells and extracts in susceptibility to ferricyanide as due 

 to permeability factors. Certainly one might expect an ion such as ferri- 

 cyanide to enter cells with difficulty. However, the initial observation of 

 Mendel that tumor tissue is specifically sensitive to ferricyanide remains to 

 be explained. If such a difference exists, it must be due to ferricyanide 

 penetrating into tumor cells more readily, since none of the enzymes con- 

 sidered to be the point of attack differs markedly in tumor cells compared 

 with normal tissues. 



Inasmuch as ferrocyanide is presumably formed in tissues during the 

 reduction of ferricyanide, the effects of ferrocyanide on the tricarboxylate 

 cycle may play a role in any over-all action. Martin (1955) noted that 

 growth of Aspergillus niger is inhibited by ferrocyanide at concentrations 

 below 0.002 mM. However, acid production may not be simultaneously in- 

 hibited, and is depressed 50% only at 0.4 mM. An accumulation of citrate 

 is actually observed at 1 mM ferrocyanide and, at this concentration, iso- 

 citrate dehydrogenase is inhibited 100% (Ramakrishnan et al., 1955). In 



