112 1. MALONATE 



Sequential Inhibition with Malonate and Fluoroacetate on Citrate Levels 



Before leaving the subject of accumulation of cycle intermediates, a 

 few words must be said on the effects of malonate on the increases in citrate 

 brought about by fluoroacetate. This was discussed in Volume I (page 502) 

 and the results obtained by Potter (1951) presented (Fig. 1-10-5). The prin- 

 ciple of the experiments is simply that fluoroacetate blocks the utilization 

 of citrate so that the tissue citrate levels rise at different rates and to dif- 

 ferent degrees. If malonate is administered to the animals prior to the 

 fluoroacetate, the accumulation of citrate may be modified. These studies 

 thus provide information on the effects of malonate on the rates of forma- 

 tion of citrate and supplement the results discussed above. 



The tissues differ greatly in their response to malonate in the presence of 

 fluoroacetate. In thymus, for example, malonate blocks the formation of 

 citrate completely and no citrate accumulation at all occurs. The kidney 

 behaves similarly but some citrate begins to accumulate an hour after 

 the fluoroacetate is injected. In spleen and brain the inhibition of citrate 

 formation is around 50%. Heart responds quite differently from the other 

 tissues. Here malonate actually increases the citrate formation somewhat. 

 Potter believed these results to indicate that in heart there are pathways 

 other than the cycle for the synthesis of oxalacetate, and suggested the con- 

 version of ketone bodies (malonate induces ketonemia) to citrate by a 

 pathway involving oxalacetate, the ketone bodies arising from fatty acids. 

 Liver was not studied but if similar reactions occur in this tissue, they 

 could,. at least in part, account for the increases in serum citrate and urinary 

 excretion of citrate. In any event, these experiments illustrate very well 

 the inherent differences between tissues with respect to their metabolic 

 pathways. The stimulation of citrate formation in the heart in the presence 

 of fluoroacetate was confirmed by Fawaz and Fawaz (1954), whereas in the 

 kidney only a depression was observed. The use of fluoroacetate to block 

 the utilization of citrate is a useful technique by which to study the effects 

 of malonate uncomplicated by possible effects on the rate of disappearance 

 of citrate. 



ANTAGONISM OF MALONATE INHIBITION 

 WITH FUMARATE 



The overcoming of an inhibition by the addition of an intermediate nor- 

 mally arising distal to the site of the block is often excellent evidence 

 for the locus of action of the inhibitor and for the specificity of the inhi- 

 bition. For this reason, fumarate has frequently been used in malonate- 

 inhibited preparations and a reversal of the inhibition taken as proof for 

 the specific action of malonate on succinate oxidase. The first acceptable 



