PHENOLS 69 



purification by extracting with very dilute sodium hydroxide 

 (1:1000). Remove all traces of alkali by washing with water. 

 Now filter the combined chloroform extracts through a dry filter 

 paper into a dry Erlenmeyer flask. Distill off the chloroform, 

 heat the residue on a boiling water-bath for five minutes in the 

 open flask, and wash the dried residue with hot water. Add 

 10 c.c. of concentrated sulphuric acid to the washed residue, heat 

 on the water-bath for five to ten minutes, dilute with 100 c.c. of 

 water, and titrate the blue solution with a very dilute solution of 

 potassium permanganate. The end-point is indicated by the dissi- 

 pation of all the blue color from the solution and the formation 

 of a pale yellow color. 



Beautiful plates of indigo blue sometimes appear in the chloro- 

 form extract of urines containing abundant indican. In urines 

 preserved by thymol the determination of indican is interfered 

 with unless great care is taken in washing the chloroform extract 

 with dilute alkali. Care should be taken, therefore, to make the 

 indican determination upon fresh urine, before the addition of the 

 preservative. 



Plasencia has suggested a method which is shorter than 

 Ellinger's and according to its sponsor, just as accurate. 



Calculation. One cubic centimeter of the diluted permanganate 

 solution is equivalent to about 0.15 mg. of indigo. Ellinger 

 claims that one-sixth of the amount determined must be added to 

 the value obtained by titration in order to secure accurate data. 

 This correction should always be made. 



PHENOLS 



Colorimetric Method of Folin and Denis x 



Principle. This method is based upon the fact that phenols 

 yield with a solution of phosphotungstic-phosphomolybdic acid 

 and alkali a deep blue color the depth of which is proportional to 

 the amount of such substances present. Traces of protein, which 

 may be present in the urine, and uric acid give a blue color with 

 the reagent and are removed by precipitation with an ammoniacal 

 silver solution and colloidal iron as a preliminary to the determina- 

 tion of the phenols. 



1 Folin and Denis: Jour. Biol. Chem., 1915, 22, 305. 



