After the medicinal or surgical use of carbolic acid the amount of the phenolsulphate in the urine is increased; two substances are also formed by the breaking up of carbolic acid, called pyrocatechin and hydroquinon. These become in alkaline urine dark brown on exposure to the atmospheric oxygen, and it is this that produces the well-known colour of the urine in so-called carboluria.'

Cresol sulphate of potassium.-This is not so abundant as the phenol-salt, but may be obtained in the following way from the urine of an herbivorous animal like the horse: 3 or 4 litres are evaporated to a syrup; this is extracted with absolute alcohol, filtered, and the filtrate precipitated with an alcoholic solution of oxalic acid; the precipitate is removed by filtration, and the filtrate made slightly alkaline with caustic potash. This produces a precipitate, which is filtered off; the filtrate is evaporated to a thin syrup, which is then kept at a temperature of 0° C. Leafy crystals of the potassium cresol sulphate separate out, and may be purified by recrystallisation out of absolute alcohol (Krukenberg'). The formula for cresol is C,H,(HO); that for the potassium sulphate of cresol is C,H,.SO,K. Cresol, like phenol, gives a red colour with Millon's reagent, but no violet colour with ferric chloride.

Of the three isomerides called cresol, that known as paracresol is the most abundant; ortho- and metacresol also combined as sulphates are present in mere traces. (The meaning of the prefixes ortho-, meta-, and para- in connection with aromatic substances is explained on p. 76.)

Some of the paracresol formed in the intestine is further changed into phenol and excreted as the phenol-sulphate of potassium (Baumann).

3

Catechol-sulphate of potassium.-Catechol or pyrocatechin, CHO, has two isomerides, named hydroquinon and resorcin. These were first found as ethereal sulphates in the urine of dogs after they had been fed on dihydroxyl combinations of benzene.? This salt of pyrocatechin has since been found normally in traces in human urine, especially in children, but more abundantly in that of the horse. It becomes darker when the urine putrefies. This, however, is only apparent when catechol appears in abnormally large quantities (see Carboluria, Alkaptonuria). Protocatechuic acid both free and as an ethereal sulphate is also present (Baumann,' Preusse3).

Indoxyl-sulphate of potassium.-The parent of this substance, named indole (C,H,N), is formed in the intestine; indoxyl, a radicle derived from this, has the formula C,H,NO; this united with SOK forms the indoxyl-sulphate which is found in the urine (C,H,NO.SO,K). This substance forms white glancing tablets and plates, easily soluble in water, less so in alcohol. By oxidation, indigo-blue is formed from it (2C,H,KNSO1+0,=2C ̧H,NO+2HKSO1).

[indox. sulph, of potassium]

[indigo-blue] [pot, hyd. sulphate]

The indoxyl-sulphate of potassium has received the unfortunate name of indican, under the mistaken idea that it is identical with plant indican. This latter substance is a glucoside, and only resembles the indican of urine, in that one of its decomposition products is 1 Grundriss der med. chem. Anat. p. 87.

2 Baumann and Herter, Zeit. physiol. Chem. i. 244; ii. 335.

5 Ebstein and Müller, Arch. path. Anat. lxii. 554; Furbringer Berlin, klin. Woch. 1875, Nos. 24 and 28; Fleischer, Ibid. Nos. 39 and 40.

4 Pflüger's Archiv, xii. 63; xiii. 16.

5 Zeit. physiol. Chem. ii. 329.

indigo blue (see pp. 78, 79). The following methods have been devised for obtaining indigo from urine :

(1) Jaffe's method':-Equal parts of urine and hydrochloric acid are mixed; to this mixture a few drops of a saturated solution of 'bleaching powder' are added cautiously till the maximum of blue colour appears. The mixture is then agitated with chloroform, which takes up the blue pigment; on evaporating off the chloroform the indigo is left. If this is weighed an approximate quantitative estimation of the amount of indigo in the volume of urine originally taken can be made. Albumin, if present, must be separated before performing this test, as it develops a blue colour with hydrochloric acid.

(2) MacMunn's method":—Equal parts of urine and hydrochloric acid with a few drops of nitric acid are boiled together, cooled, and agitated with chloroform. The chloroform is generally violet, and shows an absorption band before D, due to indigo blue, and another after D, due to indigo red (see fig. 97, spectrumı 4, p. 748). This method is preferable to Jaffe's, as 'bleaching powder' destroys small quantities of indigo.

The quantity of indigo in the urine of starving dogs was found by Salkowski to be 4 to 5 milligr. in three days. After abundant meat meals it rose to 16 to 17 milligr. per diem. In 1500 c.c. of normal human urine, Jaffe found from 4 to 19 milligr. of indigo. Horse's urine contains twenty-three times as much. The greater abundance of indigo, like that of other aromatic substances in the urine of herbivora, depends on the diet. It is absent in the urine of new-born children.+

When indol is injected under the skin, or given by the stomach, it appears in the urine as indoxyl sulphate of potassium (Jaffe, Nencki and Masson," Christiani)

This salt is also increased in the urine in intestinal obstruction, peritonitis, typhus, cholera, cancer of the liver, long-standing suppuration, and Addison's disease. No doubt in many of these cases this is due to increased absorption of putrefactive products. Many aromatic drugs, like turpentine, oil of bitter almonds, and creosote, also increase its amount in the urine.

Sometimes urine shows when decomposing a bluish red pellicle of microscopic crystals of indigo blue and red, owing to the decomposition of the indoxylsulphate (Hill-Hassal, Stirling). A calculus composed of such crystals has been cnce described (Ord).

Skatoxyl sulphate of potassium. -Skatole is methyl-indole CH(CH3)N. Like indole, it is formed by the putrefaction of proteids in the intestine; some of it is absorbed, and passes into the urine as the skatoxyl-sulphate of potassium (C,H,NO.SO,K). It is rather more abundant in human urine than the indoxyl-salt.8

1 Pflüger's Archiv, iii. 448.

5 Ber. d. deutsch, chem. Ges. ix. 138.

5 Centr. med. Wiss. 1872, No. 1.

7 Zeit. physiol. Chem. ii. 273.

2 Clinical Chem. of Urine, p. 97.

4 Senator, Zeit. physiol. Chem. iv. 1. 6 Maly's Jahresb. 1874, p. 221.

8 G. Hoppe-Seyler, Zeit. physiol. Chem. xii. 1. F. Hoppe-Seyler, however, states that he has found great variations in the relative and actual amounts of both salts without assignable cause (Physiol. Chem. p. 846).

Otto obtained a red pigment which he considered to be formed from the skatoxyl-sulphate as indigo is from the indoxyl-sulphate. Mester, however, who fed a dog on skatole, found only traces of the skatoxyl salt in the urine, but abundance of the skatoxyl-pigment. He gives certain reactions of this pigment, and considers it identical with those previously described under the names of urorubin, urorosein, uroerythrin, &c. In the urine it exists as a chromogen of unknown nature, perhaps a compound of skatoxyl with glycuronic acid, analogous to indoxylglycuronic acid. On adding mineral acids to the urine containing it, it became red or reddish violet, especially on warming. The pigment is probably an oxidation product of the chromogen.

The following table (adapted from Hoppe-Seyler) will be found convenient for the separation of some of these substances :

Urine is evaporated to one-third of its volume, and shaken with ether; this takes up pyrocatechin and hydroquinon

The following table gives in a concise way some important reactions of these substances:

5 Using G. Hoppe-Seyler's method of isolating these substances (Zeit. physiol. Chem.

vii. 423).

CHAPTER XLI

THE PIGMENTS OF THE URINE

THE pigments of the urine have been described under different names by different observers, and in the following account of them I shall follow MacMunn1 very closely. It will also be convenient here to describe, not only the normal pigments, but also those occurring in disease.

NORMAL UROBILIN

This is the principal colouring matter of normal urine. It may be obtained from the urine by adding neutral and then basic lead acetate until there is no further precipitate. The precipitate consists of the chloride, sulphate, and urate of lead, and it carries down with it most of the pigment; it is filtered off; the filtrate is clear and almost colourless. The pigment is extracted from the precipitate by alcohol acidulated with sulphuric acid; the extract is filtered off from the remainder of the precipitate which is insoluble in this reagent. The extract has a deep yellowish colour; it is agitated with chloroform. This reagent dissolves out the pigment, which is obtained in an approximately pure condition by evaporating the chloroform from the chloro

formic extract.

Normal urobilin thus obtained is amorphous, yellowish brown in colour, freely soluble in alcohol, chloroform, acids, acidulated water, and partly soluble in ether and benzene. An acid solution of it shows spectroscopically one absorption-band close to and enclosing the F line. If the solution be made neutral by alkalis, the band disappears. If the absorption spectrum of normal urobilin (fig. 97, spectrum 1, p. 748) be compared with that of choletelin (fig. 88, spectrum 2, p. 685), it will be found that the two are practically identical. It would, however, be premature to say that the two substances are identical, until spectroscopic analysis is supported by other analytical methods. When normal urobilin is separated out and dissolved in alcohol, and treated with zinc chloride and ammonia, the solution shows a green fluorescence, which, however, is not nearly so well marked as that obtained

1 Clinical Chemistry of Urine, pp. 104-112.