280 G. ASHWELL, Z. DISCHE VOL. 4 (1950) 



metabolism. In a second series the influence of various cations and anions on those 

 reactions was investigated. 



1. The aerobic metabolism in the hemolysate 



a) O2 uptake, lactic acid formation in the hemolysate in absence of glucose. The hemo- 

 lysate to which 0.2 ml of liquid per ml was added shows a marked respiration which 

 varied in our experiments between 19 and 92 cmm per i ml and 4 hours. The respiration 

 is in general much higher during the first hour and drops afterwards to a lower but 

 constant level. The R.Q. varies considerably between 0.82 and i (Table I). The erythro- 

 cytes contain very little hexoses soluble in trichloracetic acid. Less than i y/ml of 

 hexose (calculated as glucose) was found in the hemolysate. This amount does not 

 change during the 4 hours of the experiment. On the other hand there is a considerable 

 decrease in the amount of adenosine-5-phosphate. In experiment VI (Table I) 84 y/ml 

 of this compound, corresponding to 35 y/ml pentose, disappeared in 4 hours. If all 

 of this pentose had been oxidized to CO2 half of the total O2 uptake in this experiment 

 would be accounted for. The breakdown of adenosine-5-phosphate can be explained 

 by the fact that it is formed in the hemolysate by the ATPase and dephosphorylated 

 to adenosine which, as was shown for human erythrocytes, can be split, with phosphory- 

 lation, to form triosephosphate and hexosediphosphate. One part of the respiration of 

 the hemolysate in absence of glucose must be due to the oxidation of either fat or pro- 

 tein'. The hemolysate contains from the beginning very small amounts of lactic acid 

 (about 5 y/ml). In some cases small amounts of this acid are formed during incubation, 

 but not more than about 5 y/ml. 



b) The tricarboxylic acid cycle in the hemolysate. The presence of this enzyme system 

 in the hemolysate can be demonstrated after addition of citrate or one of the dicarboxylic 

 acids metabolised by the system. When M/1200 of succinic, fumaric, malic, oxaloacetic, 

 citric and a-ketoglutaric acid is added the O2 uptake increases considerably (Table I). 

 In presence of ketoglutaric and citric acid much more than in that of other acids this 

 additional Og uptake increases with the concentration of the acid. It is about twice 

 as great in presence of M/600 succinate than of M/i 200. At the same time lactic acid 

 is formed in significant amounts. This increases with the concentration of succinate 

 or malate. The amount of lactic acid varies with the nature of the acid in the following 

 sense : malate, fumarate > succinate > a-ketoglutarate > citrate. This can be explained 

 by the assumption that oxaloacetic is formed from malic acid, with reduction of co- 

 enzyme I to dihydrocoenzyme I. One part of the oxaloacetic acid is decarboxylated to 

 pyruvate and COg. As the cytochrome system is not able to oxidize dihydrocozymase 

 rapidly enough, one part of it reduces pyruvate to lactate. The same sequence of reactions 

 was observed by E. A. Evans^* in liver extracts. As the increase in succinate increases 

 the O2 uptake as well as lactic acid formation the cytochrome system apparently com- 

 petes with the pyruvate for dihydrocozymase. Thus the fact that lactic is formed from 

 citrate indicates that the whole series of reactions from citrate to oxaloacetates goes 

 on in the hemolysate. Pyruvic acid also increases the respiration and lactic acid for- 

 mation, though less than any one of the polycarboxylic acids, and the increase is ob- 

 served only during the last 3 hours of the 4 hour period. 



2. Aerobic metabolism in presence of glucose 



When 50 mg % of glucose is added to the hemolysate it is broken down at a rate 

 References p. 292. 



