SUBSTRATES OTHER THAN SUCCINATE 111 



to disprove the theory that glutarate is active because it undergoes /?-oxi- 

 dative decarboxylation to malonate. Little has been done with isolated 

 preparations, but in three cases accumulation of a-ketogkitarate has been 

 observed in the presence of malonate: in locust sarcosomes from malate 

 (Rees, 1954), in suspensions of ascites cells from fumarate (Kvamme, 

 1958 c), and in ascites cells and rat heart mitochondria from glutamate and 

 malate (Borst, 1962), in all instances the malonate concentration being 

 rather high (20-50 mM). 



The mechanism of such accumulation is poorly understood. We have seen 

 that in some tissues the a-ketoglutarate oxidase can be inhibited by malo- 

 nate, especially at concentrations above 10 mM, so that the results can be 

 partially explained in this way (at least for the ascites cells and locust 

 particulate preparations). The moderate increases in a-ketoglutarate excre- 

 tion brought about by the cycle intermediates and substrates might well be 

 due to a greater rate of formation with unchanged utilization rate. However, 

 the possibility of a formation of c-ketoglutarate from glutamate cannot be 

 ignored. It will be recalled that in potato slices the succinate formed in the 

 presence of malonate comes partly from such a source (Romberger and 

 Norton, 1961). Permeability effects causing a leakage of a-ketoglutarate 

 and other anions from the tissues must also be considered. El Hawary (1955) 

 found that several inhibitors (arsenite, maleate, iodoacetate, alloxan, and 

 fluoride) increase the serum a-ketoglutarate levels in rats, and simulta- 

 neously raise pyruvate levels. It may well be that any severe metabolic 

 disturbance causes a release of cycle substrates from the tissues and an 

 increased excretion, as well as secondary changes such as hyperglycemia. 



Effects on the Levels of Other Cycle Substrates 



The tissue concentrations of all the cycle substrates are probably altered 

 by malonate and it is sufficient here to mention the results with potato tuber 

 slices (Table 1-19) and tobacco leaves (Table 1-20). The reduction in the 

 incorporation of C^** from glucose into malate in the former and the marked 

 falls in malate level in the latter would be anticipated from a block of suc- 

 cinate oxidation. Fumarate and oxalacetate levels probably are changed 

 similarly. In this connection, one wishes that more information were avail- 

 able on the factors that control the tissue pools of cycle intermediates, since 

 it is evident that these substances do not occur only in the mitochondria 

 in kinetic equilibria depending on the relative rates of the cycle reactions, 

 but must also be present in cellular compartments. The transfer of the 

 substances between these compartments and the active cycle regions must 

 depend on processes that could be affected by inhibitors. Such compart- 

 ments are well known in plant cells but it is probable that similar situations 

 are applicable to animal cells. 



