16 
could no longer be extracted, heated with a 1 per cent solution of 
sodium carbonate for one hour at 40° C. The mixture was passed 
through flannel and the filtrate moderately acidulated with acetic acid 
to precipitate nucleo-proteids. The clear filtrate now obtained had a 
considerable power of catalyzing hydrogen peroxid, although it could 
not contain a-catalase any longer. 
In a second trial the exhausted tobacco was heated for two hours at 
from 45° to 50° C., with a solution of 0.2 per cent sodium carbonate, 
the filtrate acidulated with acetic acid, again filtered, and the filtrate 
now saturated with ammonium sulphate at the ordinary temperature. 
By this treatment the active principlé was salted out, as is the case with 
f-catalase, the precipitate showing powerful action while the filtrate 
showed only slight traces of activity. It may also be stated that the 
salted-out product was easily soluble again in water and can not there- 
fore have consisted of nucleo-proteids. 
Since there is a zymogen for every well-known enzym, the supposi- 
tion seems justifiable that there is also a zymogen of catalase. Various 
observers have shown that animal or vegetable cells, before they form 
the enzyms, are rich in small granules, which disappear in a measure as 
the enzyms are formed. The substance first secreted by the nucleus, 
forming the granules in the cytoplasm, is distinguished by Macallum 
as pro-zymogen. Vines, in his investigation on the presence of diastase 
in leaves, observed that the turbid unfiltered extracts saccharified 
more starch than the clear filtered ones. Brown and Morris found, 
furthermore, that powdered leaves are more powerful in diastasic 
action than an aqueous extract prepared from the same amount of leaf. 
Whether the action here is caused by a zymogen which is gradually 
transformed into diastase, or whether the diastase itself is retained to 
a certain extent by the solid compounds of the leaf, has not been 
decided. It has been observed that highly dilute acids bring on the 
transformation of zymogen intoenzym. J. Reynolds Green has found 
that certain rays of light—located chiefly in the red, orange, and blue 
regions of the spectrum—and a prolonged exposure to a temperature 
of 38° C. can also cause this change." 
The existence of a zymogen of catalase in tobacco leaves appears 
probable from an observation of the writer that in the process of 
sweating tobacco in bulk at a temperature of from 40° to 50° C. the 
power of catalyzing hydrogen peroxid is increased, not only for the 
soluble but also for the insoluble catalase. Digestion with 0.1 per 
cent oxalic acid at from 25° to 36° C. proved a failure in this direction, 
since the catalytic power was decreased thereby. Acids therefore 
seem unsuited to transform zymogen into enzym in the case of catalase. 
1Philosoph. Transactions, vol. 188 (1897). On the other hand, diastase itself is 
gradually destroyed by white light, as the same author has observed. 
a = 
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