THE AROMATIC CONSTITUENTS OF THE URINE. 253 



The isolation of indican from urine as such is a rather complicated 

 process, and need not be described at this place. 



Quantitative Estimation. WANG'S METHOD. This method is 

 based upon the decomposition of the indican by strong hydrochloric 

 acid, and the oxidation of the resulting indoxyl to indigo-blue. 

 This is then transformed into indigo-sulphuric acid, and estimated 

 as such by titration with a solution of potassium 'permanganate of 

 known strength. To this end, a stock solution of the salt is kept on 

 hand, which contains about 3 grammes to the liter. Five c.c. of 

 this solution are diluted with 195 c.c. of water, when the titre is 

 ascertained before each titration by comparing it with a dilute solu- 

 tion of oxalic acid. The amount of indigo-blue which each cubic 

 centimeter represents is ascertained by multiplying the correspond- 

 ing amount of oxalic acid by 1.04. 



The amount of urine which is necessary varies with the amount 

 of indican present. If a preliminary test gives an intense reaction, 

 from 25 to 50 c.c. are sufficient ; otherwise it is better to use larger 

 amounts, as from 200 to 250 c.c. The urine is then precipitated 

 with a 20 per cent, solution of basic lead acetate, care being taken 

 to avoid an excess. A large portion of the filtrate, representing 

 a known amount of urine, is then treated with an equal volume 

 of Obermayer's reagent, and extracted with chloroform by shaking. 

 This is continued with portions of 30 c.c. until the chloroform takes 

 up no more coloring-matter. The combined extracts are freed from 

 chloroform by distillation. The residue is dried for a few minutes 

 on a water-bath, and is then washed with a mixture of equal parts 

 of water, ether, and alcohol (96 per cent.), to remove the reddish- 

 brown pigment which is present together with the indigo-blue. 

 The solution is passed through a small filter, so as to collect any 

 particles of the blue pigment which may be present in suspension. 

 The filter is dried, extracted with boiling chloroform, and the 

 resulting solution filtered into the flask containing the residual 

 indigo-blue. The chloroform is distilled off, and the residue treated 

 with 3 or 4 c.c. of concentrated sulphuric acid, while still warm. 

 This solution is allowed to stand for twenty-four hours. It is then 

 poured into 100 c.c. of water, the bottle is washed out with a little 

 more water, when the solution and washings are filtered and titrated 

 with the permanganate solution. The color at first changes to 

 green, and finally becomes yellowish or colorless. The calculation 

 is conducted as outlined above. 



Skatoxyl Sulphate. Skatoxyl sulphate, like indoxyl sulphate, 

 occurs in the urine in combination with potassium and sodium. Its 

 amount, however, is normally small, and it may at times be absent 

 altogether. Larger quantities are found under pathological condi- 

 tions associated with an increased degree of intestinal putrefac- 

 tion, and it may then happen that more skatoxyl sulphate is found 

 than indican. This, however, is uncommon, and in disease also 

 more indican is usually present. Like indican, it is decomposed 



