1912.] Studies on the Reductase of Liver and Kidney. 491 



tion of intermediate compounds ; and, indeed, it would appear that the first 

 stage of the reduction is more readily effected by the reductase of the press 

 juice than the second stage. We intend to investigate the chemical nature 

 of such intermediate compounds. 



A number of experiments was carried out with a view to determining the 

 relation of the reducing activity of the press juice to temperature. In these 

 experiments, the results of which are given below, 3 c.c. of a 24-hour-old 

 juice were mixed with 10 c.c. of 0'05-per-cent. solution of soluble Prussian 

 blue. Both the soluble Prussian blue solution and the press juice were brought 

 to the temperature of the experiment before mixing. 



I 



Temperature. 



Time required for the mixture to become 

 light green-grey. 



°C. 



mins. 







40 



14 



10 



25 



6 



40 



2 -5 to 3 



55 



1 -25 



67 



less than 1 



72 



2 



Press juice and a solution of soluble Prussian blue that had been cooled to 



— 2° C. were mixed and placed in a freezing mixture at —14° C. It was 

 found at the end of two hours, although the mixture quickly became solid 

 after immersion in the freezing mixture, that the colour had turned from blue 

 to green. That reductase is not permanently inhibited by a temperature of 



— 14° C. is shown by the fact that when this mixture was melted and 

 warmed, the green colour faded rapidly to light green-grey. It is most 

 probable that, at —14° C, the extremely weak action of the reductase is due 

 not so much to inhibition of the enzyme as to the formation of the solid 

 phase. 



Experiments were also carried out at 100° C. The press juice and the 

 solution of soluble Prussian blue were separately brought to this temperature 

 and then mixed. It was invariably observed that the blue colour of the mixture 

 faded to the light green-grey in about one minute. The blue colour could be 

 re-established, however, by shaking up the cooled colourless mixture with a 

 few cubic centimetres of dilute hydrochloric acid. Since it has been shown 

 that the strong activity of a fresh juice is completely destroyed by heating to 

 100° C. for two or three minutes, it is evident that at this temperature the 

 decolorisation of soluble Prussian blue cannot be due to the action of 

 reductase, but that it must be ascribed to some other cause. In a recent 



