104 



thus obtained was filtered through musHn.^° These clear solu- 

 tions were made up to 50, 100 or 200 cubic-centimeters, depending 

 upon the amount of material used in the preparation of the 

 extracts. 



The tests were carried out in the following manner: 5 c.c. 

 of the plant extract were placed in each of a series of test-tubes 

 and to each such portion of extract ten drops of reagent were 

 added from a dropping bottle. This was a test for the oxygen- 

 ases (direct oxidases) and was repeated in every detail, except 

 for the addition of five drops of i per cent, pure hydrogen peroxide 

 solution, ^"^ when testing for peroxidase. The latter treatment 

 caused an increase of coloration, when compared with the corre- 

 sponding oxygenase effects, if peroxidase were present. Boiled 

 portions of the enzyme solutions were tested in precisely the 

 same manner for control purposes. Portions of the extracts 

 were tested again after standing one hour, and once more after 

 the lapse of twenty-four hours, to reveal any subsequent change 

 in the action of the oxidases. The presence of catalase was 

 shown by the evolution of gas when five drops of i per cent, 

 hydrogen peroxide solution were added. Any change of color 

 indicating chromogens or any peculiar appearance of the plant 

 juices were noted. 



It became evident very early in our work that failure to obtain 

 a positive test for oxidases usually indicated the presence of 

 acids; so we determined the acidity of many of the extracts by 

 titrating ten cubic-centimeter portions with Njio potassium hy- 

 droxide solution, using phenolphthalein as the indicator. To 

 serve as a further check on our results, all of these tests were 

 made on another day with another sample of the material to 

 obviate the effects of any psychological differences on the ob- 

 server's part, or individual variations in the plants examined. 



s" This muslin had previously been treated with boiling dilute hydrochloric acid 

 solution. It was then washed with water, treated with boiling dilute ammonium 

 hydroxid solution, washed with distilled water until neutral, and finally dried in a 

 dust-free place. 



61 The best hydrogen peroxide is the "Perhydrol" of Merck, containing 30 per 

 cent, of H2O2. It was diluted with twenty-nine volumes of water. This product 

 is practically neutral and contains no preservative. 



