( 518 ) 
the fermentation liquid and in the liquid prepared with enzyme and 
indican solution, the indigo is formed from indoxyl. This is con- 
firmed by me, being successful in obtaining from those liquids three 
typical reaction-products of indoxyl, namely the indigonides of isatin, 
benzaldehyde and pyro-uvie acid. 
I applied these reactions in the following manner: To 25 c.c. of 
indican solution was added 1 gram of the leaf powder (This is pre- 
ferable to using the solution, as during the action of the enzyme 
red resinous products are formed which are not retained by paper 
filters. These products, however, adhere to the leaves, otherwise 
they might cause wrong conclusions to be drawn from the reactions 
particularly as we will see later on that indoxyl under some circum- 
stances does not yield indirubin). After sufficient action 20 c.c. of 
or saturated solution of isatin in 50 pCt. alcohol and 50c.c. of a satu- 
rated solution of sodium carbonate were added, or a little benzal- 
dehyde or pyro-uvie acid and then for every 100 parts of liquid 40 
parts of hydrochloric acid of 1.13 sp. gr. 
The substances which were formed all had the qualitative pro- 
perties which are ascribed in the literature to the three indigonides. 
As one of the components is known and the other exhibits all 
the properties of indoxyl and no other ones, whilst the formed pro- 
ducts agree, qualitatively, completely with the substances which the 
said reagents form from indoxyl, it is proved that indoxyl is the 
other component. 
I have not made any ultimate analyses of the substances as it 
was extremely difficult to purify them. Moreover, I think that 
their identity has been sufficiently established. 
The fact that the fermentation product is indoxyl, explains the 
formation of large quantities of indigo-red in some processes of 
indigo manufacture. 
I may call attention to the fact that during the so-called fermen- 
tation no indican ever passes into the surrounding liquid. This was, 
of course, easily proved by rendering the liquid so strongly alkaline 
that no enzyme-action could take place, then passing a current of 
air to oxidize my indoxyl and testing the filtrate after neutralisation 
with enzyme powder or solution of the same. 
I noticed some phenomena which must not be lost sight of when 
testing for indoxyl. 
If to a solution of indican is added a little phenol-phtalein, then 
so much lime-water that the solution has a decided alkaline reaction 
and then some enzyme powder, the red colour gradually vanishes. 
This may be caused by the presence of some acid contained in the 
