396 ENZYMES 



4. Peroxidases set free oxygen from hydrogen peroxide 

 and other peroxides. 



For comparative experiments with peroxidase, Gruess uses 

 I gram of pulverized powder, prepared as above, which is 

 mixed and ground with sand with 2 5 c.c. of water. For the 

 test, 5 c.c. of the nitrate is mixed with -5 c.c. of guaiaconic 

 acid dissolved in alcohol, and 'I c.c. of a -5 per cent solution 

 of hydrogen peroxide. 



Appleman, for comparative tests, allows a definite quantity 

 of the extract (see above) to act on a definite volume of 

 guaiaconic acid solution in the presence of hydrogen per- 

 oxide, the test tube being kept at a constant temperature 

 whilst the experiment is going on. For the comparison, a 

 standard blue aqueous solution of indigo carmine is made ; 

 the time required for the blue of the guaiacum mixture to 

 match the colour of the standard blue is taken as the index of 

 the peroxidasic activity. 



It should be remarked that, according to Aso, the presence 

 of certain substances, e.g., tannin or sodium fluoride, interferes 

 with the colour reactions normally given by oxidases. 



Estimation. 



Willstatter and Stoll have elaborated a method, depending 

 on the production of purpurogallin from pyrogallol and hy- 

 drogen peroxide in the presence of the peroxidase, for esti- 

 mating peroxidase. It is termed the " purpurogallin number," 

 and represents the number of milligrams of purpurogallin 

 which would be produced by I mgm. of the vacuum-dried 

 preparation. This number is about 0*25 for well-pounded 

 horse-radish, 360 for the crude enzyme, before purification by 

 means of mefturic chloride, and about 670 for the purest 

 sample of the enzyme so far obtained. 



GENERAL CONSIDERATIONS. 



Up to comparatively recent times an oxidase was con- 

 sidered to be a single enzyme, but according to Bach and 

 Chodat,* what used to be termed oxidase is really a mixture 



* Bach and Chodat : " Biochem. Centrbl.," 1903, I, 416 ; Bach : " Ber. deut. 

 chem. Gesells.," 1906, 39, 2126; 1907, 40, 230; 1908, 41, 216. 



