66 



F. EDMUND HUNTER, JR. 



of the mitochondria disappear. The differential centrifugation yields 

 pellets which demonstrate the disintegration of most of the mitochondria 

 into smaller particles and soluble protein. Essentially all of the pyridine 

 nucleotide is released into the soluble fraction during ascorbate lysis. 



Protein Distribution after Mitochondrial Swelling 



Mitochondria diluted i :25 for treatment with 0-5 mM ascorbate, or 5 mM 

 PO4, or 3 mM succinate, centrifuged 8 000 x g for 10 min., 20 000 x g for 20 min., 

 100 000 X g for 60 min. 



Fig. 19. Protein distribution after mitochondrial swelling. 



Figure 20 outlines some possible interrelationships between swelling 

 inducing agents, inhibitors, and the electron transport chain. While the 

 evidence for active electron transfer or a closely associated high energy 

 intermediate conditioning the membrane response is strong in the case of 

 a great many swelling-inducing substances, this question cannot be 

 answered completely jusi now for ascorbate-lysis. We must determine 

 whether inhibitors like cyanide, antimycin A, and SN 5949 act by preven- 

 ting electron transport or by chelation of metal ions. There is, of course, 

 the possibility that ascorbate-lysis is dependent on two conditions : (a) some 

 electron transfer, and (b) an action of an ascorbate-metal complex. 



The failure of dehydroascorbate (DHA) to act more like ascorbate 

 suggests that ascorbate is not acting as a simple oxidation-reduction 

 couple with DHA. Conceivably a half-oxidized radical [11] (rather than 

 DHA) might be involved. Another possibility which must be given 

 serious consideration is that ascorbate catalyzed formation of lipid per- 

 oxides [12, 13] may be responsible for disintegration of the mitochondrial 

 membranes. Additional possibilities include alterations in lipoproteins, 

 activation of some lytic enzyme like lecithinase, release of lysolecithin, 

 lysoplasmalogen, or a fatty acid. 



