75 
that tyrosinase is easily destined or rendered inactive by alcohol, 
and hence if the ferment is prepared by this method it is necessary 
to filter off the alcohol as soon as practicable. 
In order to prepare tyrosinase from the ink sac of the cuttle fish, 
Gessard ( 184 ) macerated the ink sac with chloroform water and filtered 
through a Chamberland filter. A clear solution was thus obtained 
which gives with tyrosin the same color changes as those shown with 
an aqueous extract of Russula delica. In the preparation of the animal 
tyrosinase employed in their latest investigations Yon Furth and 
Jerusalem ( 178 ) employed the hemolymph of the pupae of DeicipJiila 
euphorbiae. This was half-saturated with ammonium sulfate, and 
the washed and pressed precipitate thus obtained dissolved in 0.04 
per cent soda solution. This solution exhibited strong tyrosinase 
reactions. 
In order to obtain the ferment from wheat bran, Bertrand and 
Mutermilch ( 65 ) recommend the following method: One part of wheat 
bran is mixed with four parts of water and the mixture allowed to 
stand for several hours. The mixture is then centrifugalized, and the 
solution thus obtained is mixed with three volumes of 95 per cent 
alcohol and again centrifugalized. The precipitate is then washed 
with 80 per cent alcohol, mixed with distilled water, and again cen- 
trifugalized. The solution thus obtained is then mixed with three to 
four times its volume of alcohol, and the precipitate thus formed is 
collected, washed with strong alcohol, and dried in vacuo over sul- 
furic acid. The substance thus obtained, the yield of which is about 
0.8 per cent, contains no laccase. On the other hand, when dissolved 
in water and filtered through a Chamberland filter, a clear solution is 
obtained which of itself undergoes no alteration on exposure to the 
air. On the addition of small amounts of tyrosin, however, it passes 
rapidly through a succession of colors — rose, cherry red, and finally, 
brownish black. On the other hand, if all gaseous and dissolved 
oxygen be removed, or if the filtered solution be heated to 100° C. 
for five minutes, no coloration with tyrosin is observed. 
In the preparation of vegetable tyrosinase from Agaricus melleus, 
Von Furth and Jerusalem ( 178 ) rubbed up two kilograms of the fungus 
with sand, and extracted the mass with two liters of chloroform water. 
After two or three hours the supernatant liquid was poured off and 
mixed with twice its volume of 96 per cent alcohol. The precipitate 
was filtered off on raw silk. 
All observers seem to be agreed that tyrosinase is a true enzyme. 
Thus it appears to conform to Portier’s ( 33 °) definition of an oxidizing 
ferment, according to which, first, gaseous or dissolved oxygen is 
necessary for its action; second, its action is accompanied by a 
notable absorption of oxygen; third, it is destroyed by heat; and, 
fourth, it is nondialyzable. 
