18 
process of curing; in other words, whether the abnormal softening of 
the leaves was accomplished without the action of bacteria. Parts 
of cured tobacco leaves and of fresh leaves, the latter previously 
dried at 45° C., were therefore moistened with water and left for twelve 
days at from 45° to 50° C.in test tubes with as little 20 per cent alcohol 
as sufficed to cover them. Bacterial development was thus excluded, 
but not the action of enzyms. Vapor of chloroform in closed flasks was 
also used in place of the 20 per cent alcohol. The result was that the 
green leaves (healthy as well as mosaic diseased leaves) did not show 
any decrease of tenacity, but the cured leaves did, although only to a 
slight degree. Krom sweated leaves a concentrated extract was now 
made at the ordinary temperature, a second portion of leaves being 
extracted with the first extract obtained, and so on until the fourth 
portion was extracted with the same water. The concentrated extract 
thus obtained was saturated with ammonium sulphate, whereby a 
copious precipitate of proteins of a very dark brown color was formed. 
‘This precipitate was freed of most of the adhering salt solution by 
pressure between filter paper, dissolved in a little water, and in pres- 
ence of chloroform permitted to act for five days on a small piece of 
fermented leaf at 50° C. Although bacterial growth was here suc- 
cessfully prevented, the leaf was very much softened and lost most 
of its former tenacity. Nevertheless, the writer is not yet fully pre- 
pared to give a decisive answer as to the nature of the cellulose injur- 
ing agency in the sweating leaf.' 
TEST FOR OXIDASE AND PEROXIDASE IN THE FRESH TOBACCO LEAF, 
Although in report No 59 (p. 28) the way of testing for the oxidiz- 
ing enzyms of tobacco was mentioned, some further remarks on this 
subject will not be superfluous for those who intend to make further 
observations on these enzyms. ‘The principal reagent, as has been 
mentioned, is guaiac resin in absolute alcoholic solution, which gives a 
blue coloration when oxidase is present and, in connection with hydro- 
gen peroxide, a blue coloration when peroxidasé is present. 
While the test for oxidase and peroxidase in the stalk and ribs of 
leaves can be directly applied to the objects themselves, there is some 
difficulty encountered in the lamina, since the green color of the chloro- 
phyll interferes with the recognition of the pure blue reaction.” Further- 
more, when the juice is prepared by rubbing the leaf to a pulp and 
pressing this through flannel, it will contain so many chlorophyll gran- 
ules in suspension that the juice has to be filtered through filter paper. 
'Bumke and Wolffenstein have recently found that cellulose is comparatively 
easily changed, under certain conditions, hydralcellulose being formed. Perhaps 
a Similar action takes place in the sweating leaf when too much moisture is present 
and the oxidizing enzyms act upon the cellulose walls. 
*The blue reaction may be distinguished to some extent in the leaf by applying 
the reagent to the spongy tissue after the removal of the lower epidermis. 
