INDOL, PHENOL, ETC. 63 
for five minutes (70° to 80° C.). The browning of media due to excess of sodium 
nitrite must not be mistaken for this pink or red reaction. Uninoculated tubes 
should be included in the test, which may be made on the second and tenth day. 
For methods of determining phenol see Lewandowski in Deutsche Med. 
Wochenschrift, 1890, p. 1186, and Chester’s Manual, p. 33. Schmidt (Bd. I, 
p- 1008) gives the following as a qualitative reaction for tyrosin: Dissolve by 
boiling in water and add a solution of mercuric nitrate. The red reaction is 
sharper if a little fuming nitric acid diluted in water is added. ‘Try also the violet 
reaction with neutral iron chloride.* Leucin crystallizes in white soft scales. 
REDUCTION OF NITRATES (AND MORE COMPLEX NITROGEN COMPOUNDS) TO 
NITRITES, TO AMMONIA, AND TO FREE NITROGEN. 
For the pathologist the iodine-starch reaction is the most satisfactory test for 
nitrites, because it is not superlatively sensitive and consequently does not indicate 
traces of nitrite absorbed from the air. It is made as 
—_ —_— follows: ‘Twenty-five cubic centimeters of distilled 
water are added to one-half gram (more or less) of 
pure potato starch and the fluid boiled. One cubic 
centimeter or more of this starch-water and 1 cc. of 
Jreshly prepared potassium-iodide water (1 : 250) are 
now put into the culture fluid, to which is then added 
a few drops of strong sulphuric-acid water (2:1). If 
any appreciable quantity of nitrite is present the culture 
immediately becomes blue-black from the liberation of 
free iodine, which acts upon the starch. Old potassium 
iodide water should never be used without first testing 
carefully, as it usually contains some free zodine. It 
is always best to first make a trial test without the 
bacteria. Commercial starch frequently contains prod- 
ucts of bacterial decomposition and starch prepared 
aseptically should be substituted. 
At least one-third of the organisms which have 
fallen under the writer’s observation in recent years 
give the nitrite reaction when grown in peptonized 
beef-bouillon containing potassium nitrate. 
a) 
Fig. 56.+ 
*Mann (p. 323) gives the following as a specific tyrosin reaction: Denigés has recommended 
the well-known phenol aldehyde reaction for the detection of tyrosin. Nasse, in repeating Denigés’ 
observations, has found the following to be a very delicate test for tyrosin, as neither proteids 
nor peptones give the color-reaction. Proceed thus: Add a few drops of formol solution to con- 
centrated sulphuric acid, when, on warming with tyrosin, a brown-red color is obtained, which, on 
addition of acetic acid, becomes green, 
+Fic. 56.—Bacterium syringae (van Hall). Nitrate bouillon cultures 5 days old, to each of 
which has been added boiled starch water, potassium iodide water, and sulphuric acid. In tube a 
the potassium nitrate was reduced to the nitrite, and on addition of the reagents free iodine was 
liberated, and the starch blued. In the other no nitrite had formed, no iodine was liberated, and the 
starch remained colorless. For discrepancy see text. 
