TESTS FOR INDOLE 31 



peptone must, therefore, be used. As the dilute solution 

 of sodium nitrite is unstable, a stock 5 per cent, solution 

 may be kept ; 2 c.c. of this solution are diluted to 100 c.c. 

 with distilled water at the time of making the test, and 

 1 c.c. of this dilution is added to every 10 c.c. of the 

 culture. The addition of the acid liberates free nitrous 

 acid, which reacts with any indole present and yields 

 a pink colour. Sometimes when the reaction is appa- 

 rently absent or feeble, it may be obtained or intensified 

 by placing the tube in the blood-heat incubator for half 

 an hour. The sulphuric acid should be pure and free 

 from oxides of nitrogen, hence hydrochloric acid is often 

 preferable. 



A more delicate method of testing is to run a little 

 hydrochloric acid down the side of the tube, so that a 

 layer forms at the bottom, the nitrite having been pre- 

 viously added to the culture if required. A pink ring 

 at the juncture of the hydrochloric acid and culture 

 indicates the presence of indole. The pink pigment, the 

 product of the reaction, may be extracted by shaking 

 with a little amylic alcohol. 



Another good method is to add 1 c.c. of ether to the 

 culture, shake, pipette off a little of the ether, and mix 

 with a few drops of water in a small tube and then add 

 the test reagents. 



Other delicate and more certain reagents for the detection of 

 indole are paradimethylamidobenzaldehyde (4 grin., dissolved in 

 absolute alcohol 380 c.c., hydrochloric acid 8 c.c.). To about 

 10 c.c. of culture 5 c.c. of this solution are added, and then 5 c.c. 

 of a saturated aqueous solution of potassium persulphate ; 

 indole gives a pink or red colour. Another test is /3-naphthaquin- 

 one - sodium - mono - sulphonate (2 per cent, aqueous solution), 

 which gives, when the mixture is rendered alkaline with caustic 

 potash, a blue or blue-green colour or precipitate. The coloured 

 compound may be extracted with chloroform, in which it yields 

 a red solution. 



