66 E. C. Slater and W. C. Hulsmann 



with the phosphorylation coupled with the cytochrome 

 oxidase reaction has been supported by experiments which 

 have shown that this phosphorylative step is more sensitive to 

 low concentrations of PNP (Table II). 



Table II 



Uncoupling of oxidative phosphorylation by p-nitrophenol 



Rat heart sarcosomes ; pH 7. It should be noted that the sarcosomes were not 

 given a prehminary hypotonic pretreatment in the experiments with ascorbate 

 and cytochrome c (see Slater, 1955). 



Expt, Substrate 



1 Glutamate 

 Glutamate 

 Succinate 

 Succinate 



Ascorbate + cytochrome c 

 Ascorbate + cytochrome c 



2 Glutamate 

 Glutamate 

 Succinate 

 Succinate 



Ascorbate + cytochrome c 

 Ascorbate + cytochrome c 



Ageing of mitochondria 



The oxidative phosphorylation system in mitochondria 

 becomes "uncoupled" if the mitochondria are allowed to 

 stand for a few hours at room temperature (Kielley and 

 Kielley, 1951). The "uncoupling", as revealed by the ATPase 

 activity, reaches a maximum if the aged mitochondrial 

 suspension is then subjected to repeated freezing and thaw- 

 ing (Myers and Slater, 1957a). Fragmented mitochondrial 

 preparations, such as the Keilin and Hartree heart-muscle 

 preparation (Myers and Slater, 1957a) or the digitonin 

 particle prepared from liver mitochondria by Cooper and 

 Lehninger (1956), have similar properties (Purvis, 1959). 

 Unlike intact mitochondria] preparations, these preparations 

 require the addition of Mg2+ for optimal ATPase activity. 

 DNP has no effect. 



Since these preparations oxidize substrates in the absence 



