I20 



BRIXTON CHANCE 



succinate would deprive DPNH of the oxidizing capacity of the chain and 

 lead to a greater degree of pyridine-nucleotide reduction. However, many 

 features of the reaction called our attention to a need for closer study. For 

 example, the rate at which succinate produced increased reduction of 

 pyridine nucleotide appeared to be slow compared with that at which 



K 50 sec 



-M 



Spectrophotometric troce 



"^ 340-374m//, 



log Iq/I = 010 



7mM succinate 

 Aerobic — >• . — "^ 

 mitochondria ' 



330/xM 



0-5fxW\ / 

 PNH/sec 



Platinum microelectrode trace 



Pyridine nucleotide red 



uction I 



Fig. 2. Illustrating increased reduction of pyridine nucleotide in a suspension 

 of rat-liver mitochondria caused by addition of 7 mM succinate. Absorbancy changes 

 measured spectrophotometrically by double-beam spectrophotometer and respira- 

 tion by vibrating platinum microelectrode. Downward deflection upon addition of 

 reagent indicates increased light absorption at 340 m/x relative to 374 m/x. Diagram 

 indicates final concentrations of reagents added, respiratory rate in /umoles/l./sec. 

 and increment of oxygen taken up during phosphorylation of 330 [jlm ADP. 

 Rates of pyridine-nucleotide reduction also given in /xmoles/l./sec. The metabolic 

 states of mitochondria are indicated by numerals 1-3-4. Rat-liver mitochondria 

 diluted in isotonic salt meciium to concentration of approximately 2 mg. protein/ 

 ml. at pH 7-4, temperature 25' (Expt. 332-2). 



succinate could intercept oxidizing equivalents from the respiratory chain 

 and was no more rapid than the rate at which pyridine nucleotide could be 

 reduced by DPNH-linked substrates. Another puzzling feature of the re- 

 action was that it w^as slowed by addition of very low concentrations of 

 uncoupling agents and completely inhibited by larger concentrations [3]. 

 This too seemed inconsistent with a simple competitive reaction which 

 should also occur at higher respiratory rates caused by addition of the un- 

 coupling agents. Further doubts as to a simple explanation of the reaction 



