98 1. MALONATE 



Succinate Accumulation in the Whole Animal 



Some of the most interesting and suggestive experiments on succinate 

 accumulation resulting from malonate inhibition have been performed with 

 whole animals and, although such work is often difficult to interpret in a 

 quantitative fashion, the results have demonstrated that malonate can 

 partially block the succinate oxidase in various tissues of the living animal. 

 Such inhibition has obvious implications for developments in the study of 

 drug actions and chemotherapy. The first work of this type was done by 

 Krebs et al. (1938), who determined the urinary excretion of succinate, cit- 

 rate, and a-ketoglutarate following injections of cycle intermediates and mal- 

 onate into rats and rabbits. Some of their results are shown in Table 1-17. 

 The effects on citrate and a-ketoglutarate will be discussed later (see page 

 104). Although malonate increases the succinate excretion some 5-fold, 

 only 2.9% of the injected fumarate is recovered as succinate compared to 

 0.6% in the controls. The injection of 10 millimoles of a substance into a 

 rabbit will lead to a maximal extracellular concentration of approximately 

 30 TdM, so that reasonably high concentrations of malonate were probably 

 achieved. The effect of the malonate had mainly disappeared after 24 hr 

 due to the excretion and destruction of the malonate. An almost 10-fold 

 increase in the urinary succinate was seen in the more recent experiments 

 of Thomas and Stalder (1958), in which 3.7 millimoles/kg of sodium mal- 

 onate were fed to rats, the succinate over a 40-hr period rising from a 

 control value of 2.35 mg to 28.0 mg. 



The blood concentration of succinate is increased in rabbits following 

 the injection of malonate (Forssman, 1941). Intravenous injection of 2.8 

 millimoles/kg of malonate leads to a slow rise in the blood succinate to 

 around 0.20 mM at 3 hr, while injections of 3.5-5.1 millimoles/kg give 

 levels as high as 0.77 mM. A lethal dose of 8.25 millimoles/kg produces 

 death in 35 min and at the time of death the succinate concentration is 

 1.1 mM. The lower doses produce no obvious effects on the animals. The 

 normal values for blood succinate are about 0.025 milf. 



The succinate found in the urine and blood in these studies originated 

 mainly in the tissues of the animals. Are malonate inhibition and succinate 

 accumulation especially related to a particular tissue, or do all the tissues 

 contribute to the metabolic disturbance? Can differences in the metabolic 

 patterns of the various tissues be demonstrated by their responses to the 

 administration of malonate? How do tumors compare with normal tissues in 

 their susceptibility to malonate? It was to answer such questions as these 

 that Busch and Potter (1952 a, b) at the McArdle Memorial Laboratory 

 at Wisconsin undertook their excellent series of studies on the accumulation 

 of succinate in various tissues of rats following injections of malonate. 

 Analyses for malonate and succinate were made by anion exchange chrom- 

 atography (Busch et al., 1952) at various times after the subcutaneous 



