INTEGRATED OXIDATIONS IN ISOLATED MITOCHONDRIA 79 



in the vicinity of the succinate dehydrogenase and that intact mitochondria 

 are relatively impermeable to oxaloacetate. 



With particulate preparations derived from liver mitochondria by 

 lysis with phosphate and washing with KCl [13] and from kidney by the 

 method of Slater [14], concentrations of oxaloacetate of the order of 10 juM 

 had a profound inhibitory action on succinate oxidation. With these 

 preparations permeability effects would be expected to be far less pro- 

 nounced. However the order of addition of substrate and inhibitor had a 



Kp Kp OAA 

 100 n-7 -t :. 



^P i^P Malonate 



T T 



12 3 4 5 

 Time (mm) 



Fig. 9. Effect of oxaloacetate (12-5 /^m) and malonate (125 /<m) on the oxida- 

 tion of succinate (10 niM) by a Slater kidney preparation. The medium contained 

 20 mM tris, pH 7 45. 



marked effect on the inhibition of oxygen uptake. This is illustrated for a 

 kidney preparation in Fig. 9. If 10 to 50 /xM oxaloacetate were added 

 before the succinate, no oxygen uptake occurred for approximately i min., 

 after which the steady-state of inhibited respiration was observed. On the 

 other hand, when the oxaloacetate was added after the succinate several 

 minutes elapsed before the steady-state was established. In the latter case, 

 the lower the oxaloacetate concentration the longer was the time taken 

 before the final rate was observed. In contrast the order of addition of sub- 

 strate and inhibitor was unimportant when malonate and pyrophosphate 

 were used. 



