L. G. M. BAAS BECKING AND H. C. HAMPTON 
263 
precaution, we distrust the value of the numbers we obtained with the 
permanganate method. 
3. We come now to the manometrical methods. These lack the advan- 
tage of the titration method in working under normal pressure. Further- 
more, the fluid may reach the unfavorable condition of over-saturation 
with gas (P. Waentig and A. Steche, I.e.). Of the authors whose work has 
been done by this method we name: C. H. Appleman,^^ W. W. Bonns,^^ 
H. H. Bunzel (I.e.), W. E. Burge,!^ c. Foa,^' W. B. Magath,!^ W. Zaleski 
and Anna Rosenberg. The chief objections to this method are: (i) The 
pressure becomes higher during the reaction. We feel justified in disre- 
garding the effect of over-pressure as our check experiments have shown 
this to be negligible. (2) The solution contains a great part of the oxygen. 
This is true only for narrow vessels, in which the surface is small in propor- 
tion to the volume of air. Even in the apparatus of H. H. Bunzel (I.e.) 
we feel that there is danger of the fluid becoming oversaturated. Bunzel 
tried to avoid this danger by shaking. But R. O. Hertzog (I.e.) cites a list 
of cases in which enzyms are destroyed by shaking. For instance, P. Waentig 
and A. Steche (I.e.) proved the destructive action of shaking on catalase. 
It seemed clear to us, therefore, that if we chose the lesser of two evils, 
namely the manometrical method, we should take a container with a very 
broad bottom and a shallow layer of fluid. Experiments have shown us 
that the effect of shaking on the exchange of gas in such a column is minimal. 
In the short time of the catalase reaction the enzym is not injured by shaking. 
In more prolonged experiments (with oxidases for instance) it may be. 
There is another advantage in experimenting with a shallow layer of fluid 
since R. O. Hertzog (I.e.) proved that the catalase reaction is subject to 
the laws of diffusion, which is the most complete in thin layers. 
II. 
One more important criticism of nearly all methods of enzym deter- 
mination is possible. To detect the fault we must start at the very be- 
ginning, at the definition of the word enzym. An enzym is a substance 
that ehanges the veloeity of a reaction. Peroxide of hydrogen will decompose 
spontaneously but slowly. It will oxidize a certain amount in one month. 
Catalase changes the reaction time from one month to one minute. The 
only method theoretically justified would therefore be to determine the 
the time in which a reaction is completed under the influence of an enzym. 
That time is the measure of the enzym action. If the reaction is monomo- 
12 Bot. Gaz. 50. 1910. 
13 Ann. Mo. Bot. Garden 5. 1918. " , 
1^ Amer. Journ. Physiol. 44. 1917. 
1^ Biochem. Zeitschr. II. 1908. 
1^ Journ. Biol. Chem. 24. 1918. 
17 Biochem. Zeitschr. 33. 191 1. 
1^ On the assumption that the reaction time without enzym is very much greater than 
that time with enzym. 
