158 METHODS OF ANALYSIS [Chap. 



to separate the layers. Designate the residual aqueous layer as A. Combine the 

 amyl alcohol extracts and wash with 4-5 portions of N/4 hydrochloric acid or until 

 this solvent extracts very little color. These washings will contain any Indigo 

 Carmine, Amaranth and Tartrazine present, the Indigo Carmine being removed in 

 somewhat larger proportion in the first washings than the other 2. With ordinary 

 concentration very little Ponceau will be removed. Designate these combined 

 washings as B. 



6 ORANGE I AND ERYTHROSINE. 



Measure, if necessary, the amyl alcohol extract, under 5, then (1) Dilute with an 

 equal volume of petroleum ether or low boiling gasoline, and again wash several 

 times with N/4 hydrochloric acid to extract Ponceau 3R and Naphthol Yellow S; 

 or, (2) Without dilution with gasoline, wash with 5% salt solution until these 2 dyes 

 are extracted. Designate these washings as C. The Ponceau and Yellow having 

 been removed, the amyl alcohol, containing an equal volume of gasoline, is washed 

 a few times with water which will extract Orange I. This dye having been removed, 

 shake the solution, although the latter may appear almost colorless, with very dilute 

 sodium hydroxid solution to remove Erythrosine. If considerable Orange I is present, 

 some of it may contaminate the washings containing the Ponceau 3R and Naphthol 

 Yellow S, especially when these have been separated by means of N/4 hydrochloric 

 acid after the addition of gasoline. 



7 INDIGO CARMINE, AMARANTH AND TARTRAZINE. 



The presence of 2 or all 3 of these dyes is usually indicated by the appearance 

 of the N/4 hydrochloric acid washings, B, under 5. Evaporate the combined N/4 

 hydrochloric acid washings to dryness to remove the excess of hydrochloric acid and 

 dissolved amyl alcohol. Dissolve the residue in water, divide the solution and iden- 

 tify the constituent colors in the portions. To a portion of the slightly acidified 

 solution add a few decigrams of urea, warm and add 1 or 2 drops of sodium nitrite 

 solution. Indigo Carmine is converted into the pale yellow isatin sulphonate while 

 the other dyes are but little affected. The isatin compound is not ordinarily present 

 in sufficient concentration to tint the solution but it differs from Tartrazine also in 

 being much less readily extracted by amyl alcohol from strong acid solutions (less 

 than one half from 4N acid). The solution now contains the Amaranth or Tar- 

 trazine, or both, practically unaffected. Amaranth is much more quickly attacked 

 by most reducing agents than Tartrazine. Treat the solution, which should be neu- 

 tral or faintly acid (in the presence of sodium carbonate the reduction of the tar- 

 trazine takes place still more slowly), at room temperature with a dilute solution of 

 sodium hyposulphite (Na2S204), adding the latter very carefully, drop by drop, and 

 allowing sufficient time after the addition of each drop for the reduction to take 

 place. When the color shows that the Amaranth has been destroyed completely, 

 shake the mixture at once with air to oxidize the slight excess of hyposulphite before 

 it can react further on the Tartrazine. 



To separate the Indigo Carmine heat to boiling another portion of the solution, 

 which should be neutral or faintly acid, and add dilute sodium hyposulphite solution, 

 drop by drop, until all the dyes are reduced. On shaking with air the Indigo Carmine 

 is quickly re-formed. 



8 NAPHTHOL YELLOW S AND PONCEAU 3R. 



Treat the N/4 acid solution or the salt solution, C, under 6, as the case may be, 

 containing the Ponceau and Naphthol Yellow S, with enough hydrochloric acid to 

 make it about 2N and shake out a few times with washed ethyl acetate*. Remove 



