240 E. Margoliash and A. Schejter 



Replacing K^ in equation (12) by its value shown in equation (11) gave: 

 1 i + K' 7 + K' 



Combining the values of « given in equation (1) and (10) one obtains the 

 following relation: 



^- = ^ "^) 



Introducing this value for Rj^ into equation (13) gave: 



X k,K'i k.K'i k^s ^ ^ 



It should be noted that Rj^, as used in this derivation, is not a constant, since 

 like n, in the system considered, it will vary with the ratio of a and s and the 

 rate of reaction of the donor with Cat. H2O2 1. However, as stated above for 

 n, Rj^ will remain constant for any particular set of experimental conditions. 

 Equation (13) also shows that a plot of 1/A as a function of « or 5 will be a 

 straight hne. 



EXPERIMENTAL 



Rat blood was obtained by heart puncture under ether anaesthesia and 

 defibrinated with glass beads. The erythrocytes were packed by centrifuga- 

 tion and washed three times with 0-9 % NaCl in the centrifuge. The packed 

 cells were lysed by adding three volumes of glass-distilled water. After com- 

 plete haemolysis the red cell ghosts and debris were centrifuged off. The 

 final concentration of all haemolysates was calculated in terms of the original 

 volume of packed cells. 



Catalase activity was estimated by the method of Feinstein (1949) with 

 sodium perborate as substrate at 37°C. Ascorbate was used as a source of 

 hydrogen peroxide in the presence of air (Margohash and Novogrodsky, 

 1958). N-ethylmaleimide was a commercial preparation and the 3-amino- 

 1 :2:4-triazole used was a highly purified sample kindly given by Prof. D. 

 Appleman. 



RESULTS 



Effect of Dilution of Haemolysates on the Rate of Inhibition of Catalase by 

 3-amino- 1:2: 4-triazole 



The rate of the irreversible inhibition of catalase by AT in rat blood 

 haemolysates was found to increase with the dilution of the haemolysate 

 (Table 1). 



It should be noted that these results were obtained with 2 x 10~^ M 

 ascorbate as hydrogen peroxide source. When the concentration of ascorbate 

 was increased to 2 x 10~^ m 98 % inhibition of catalatic activity was obtained 



