CHAPTER V.

CREATINE AND CREATININE.

History.

THE juice of flesh contains a crystallizable substance, which was in 1835 discovered by Chevreul1 in beef-tea of the Dutch Company, and by him termed creatine. Creatine and creatinine (in the form of chloride of zinc salt) were subsequently, in 1814, found in the urine contemporaneously by Heintz2 and Pettenkofer, but their identity with the crystallizable substance of the juice of flesh was not then recognised. In 1847 creatine and creatinine were demonstrated by Liebig to be constant ingredients of the juices of the flesh of almost all the classes of vertebrate animals and of the urine of man. Verdeil and Marcet' found creatine in the blood of the ox.

100.00

Atomic weight of dry creatine 131

Crystallized creatine, therefore, corresponds to the formula CH9N3O+2Aq. 1 atom of dry creatine 131

2 atoms of water.

87.92

18

12.08

Creatinine is present in the flesh of man and the mammalia, of birds, amphibia, and fishes. The flesh of fowl yields the

Journal de Pharm.,' t. xxi, p. 236.

(Heintz) Poggendorf's Annal.,' lxii, p. 602; 1xx, p. 460; lxxiii, p. 696; lxxiv, p. 125.

3 Annal. d. Chem. und Pharm.,' Bd. lii, S. 97.

Chemische Untersuchung über das Fleisch und seine Zubereitung zum Nahrungsmittel,' Heidelberg, 1847, p. 47.

5 Journ. de Pharm.,' (3) xx, p. 89. Liebig and Kopp's 'Jahresbericht,' 1851, p. 586.

largest proportion (3.2 per mille); next the ox's heart (1.3 per mille); and the flesh of the cod-fish (0.9 to 17 per mille). The quantity of creatine obtained from the flesh of man is 0.67 per mille, about the same proportion as from beef. Creatine is present in the blood and urine of man and all animals hitherto examined.

Liebig's method of obtaining Creatine.

In order to obtain creatine pure, a quantity, say ten pounds, of flesh of a recently killed animal is freed of fat and finely chopped; five pounds are then mixed with an equal amount of cold distilled water, carefully kneaded through by means of the hands, and then pressed carefully in a bag of coarse linen. The residue is now again carefully mixed with another five pounds of water, and again subjected to pressure. The fluid from the first pressing is put aside for further treatment to be described; the fluid from the second pressing serves towards the extraction of the second portion of the flesh. The first portion of flesh is a third time treated in a similar way with five pounds of water, and the fluid obtained by pressing is used for the second extraction of the second portion of flesh; the latter is treated a third time with pure water, and pressed.

The united fluids are filtered through a clean cloth, and filled into a large balloon of glass; the latter is placed in a kettle with water, which is gradually heated to the boiling point, and kept at that temperature until the extract has lost its colour, and albumen and colouring matter have separated in the form of a coagulum. If a portion of the fluid in a test-tube heated to boiling remains clear, this operation is completed.

The fluid is now separated from the coagula by filtration through a cloth, and subsequent pressing. The united fluids are then filtered through paper.

The colour of the fluid so obtained is reddish, if extracted from the flesh of the ox, doe, hare, or fox; but the fluid from the flesh of the calf, fowl, or fish, is scarcely coloured. The extracts from the flesh of all animals are acid, from the presence of free acid, which must be removed before evaporating the fluid, as it would cause a dark brown colour of the concentrated fluid, and not permit the crystallization of creatine. In order to remove this acid, a concentrated solution of caustic baryta is added to the extract as long as a white precipitate is thereby produced. Neutrality or alkalinity should not deter the operator from adding the baryta solution. as long as it produces any turbidity.

After separation from the precipitate, which contains all the phosphoric acid of the juice of flesh in the form of phosphate of baryta and magnesia, the fluid is evaporated in shallow china dishes on the water- or sand-bath, taking care never to allow it to be heated to ebullition; the upper part of the dish must never get more hot than the fluid, as a ring of dry substance is formed thereby, which afterwards, on the addition of new portions of fluid, dissolves, and on further concentration imparts a brown colour to the fluid. The extracts from the flesh of fowl or fish remain colourless and clear to the last. If an excess of baryta have been added, a pellicle of carbonate of baryta is formed on the surface. The extract from the flesh of the ox, calf, or horse, at certain stages of its concentration, forms pellicles of organic matter on its surface, which must be removed as often as they are formed.

When the extract has been evaporated to about th part of its volume, and has assumed a syrupy consistence, it is put into a moderately warm place, and evaporation slowly allowed to go on; very soon there appear on its surface small, short, colourless needles, which increase in numbers by standing and cooling of the fluid, so that the walls of the vessel gradually become covered by them. These crystals are creatine. They are freed from the mother-liquor by filtration, washed with water, lastly with alcohol, and dissolved in boiling water. Should this solution be coloured, it is boiled with a little animal charcoal, and, after filtration, will be as clear as water. On cooling, it deposits creatine in perfectly pure crystals.

According to Gregory, creatine is obtained cheaply from cod, which, when chopped, well mixed with little more than its own weight of water, and pressed out, yields a fluid which, when neutralized (after the coagulation of albumen) by baryta, filtered to separate the phosphate of baryta, and gently evaporated till, on cooling, it forms a thin jelly, deposits, on standing, creatine in large crystals, nearly pure.

The heart of the ox is another convenient material, rich in creatine. The cheapest material from which to obtain creatine is urine. The method of obtaining creatine from urine I shall describe under creatinine.

Physical properties.

Creatine crystallizes in the clinorhombic system: all axes

1 Quarterly Journ. of the Chem. Soc.,' i, p. 25. Chemistry,' 3rd edit., 1852, p. 444.

Handbook of Organ.

unequal, two at right angles to each other in a horizontal plane, the third axis inclined in a plane cutting either of the other two axes. The clinodiagonal inclined to the principal axis in an angle of 70° 20′; inclination of the planes ∞ P: P in the orthodiagonal principal section = about 133° 2'. Specific gravity of the crystals 135 to 1:34. The crystals are colourless, perfectly transparent, and lustrous. They are connected with each other in tufts and groups, and then resemble acetate of lead (vide plate ii, fig. 5).

Chemical properties.

Creatine is easily soluble in boiling water; a solution saturated at that temperature becomes on cooling a mass of fine lustrous needles. From a dilute solution, however, creatine crystallizes very slowly in rather large crystals, which may attain a length of from one fourth to three eighths of an inch, and a thickness of one eighth of an inch, and will further increase in size if left in the mother-liquor for some time.

1000 parts of water at 64°4 F (18°C.) dissolve 13:44 creatine, or one part of creatine dissolves in 74-4 water.

In cold alcohol creatine is almost insoluble, one part requiring 9410 parts of alcohol for solution. It is more soluble in spirits of wine containing some water.

The watery cold solution of creatine, which contains a very small amount of that substance, possesses for that reason a weak, bitterish taste, and causes a sensation of irritation in the pharynx. If the solution contains a trace of a foreign organic substance, it changes very easily, mouldy vegetations form in it, and it assumes a disgusting odour.

Creatine even in the largest quantity does not neutralize the acid reaction of the weakest acid; it does not possess a basic character; it is soluble in baryta water at a higher temperature, but crystallizes out of the cooling solution without having undergone any change. The crystals so obtained contain no baryta, and from the solution the whole of the baryta may be precipitated by carbonic acid.

Decompositions.-On being boiled with baryta water, creatine is decomposed, ammonia being evolved on the one hand, and carbonate of baryta in crystalline granules formed on the other. This decomposition will take place even though the air be entirely excluded from influencing the substance.

Heintz, Poggend. Ann.,' lxxiii, p. 595.

In solution there remains an organic base of the composition C&H-NO1, sarcosine. This formula, when deducted from the elements of creatine, leaves a formula which exactly corresponds to the composition of urea.

It is, therefore, evident that ammonia and carbonic acid are products of a secondary decomposition and derived from the urea.

A solution of permanganate of potash in which creatine is dissolved loses its red colour only after standing some length of time in a higher temperature. No gas is evolved during this process. The fluid after decoloration does not any longer contain creatine, but yields white crystals on evaporation, and the potash is in part combined with carbonic acid.

The influence of strong mineral acids is very remarkable. A solution of creatine, to which at the ordinary temperature hydrochloric acid has been added, gives on spontaneous evaporation crystals consisting of unchanged creatine. If, however, the solution is heated with strong hydrochloric acid, creatine cannot be obtained any longer from the solution. The same effect is obtained by either sulphuric, phosphoric, or nitric acid. If creatine is dissolved in one of these acids, and the solution is evaporated at a gentle heat, crystals are obtained which are easily soluble in alcohol, a property which does not belong to creatine. These crystals contain a portion of the acid in chemical combination; they are, in fact, the salts of the acid with an organic base, creatinine. This transformation of creatine into creatinine under the influence of mineral acids consists essentially in the elimination from the former of four equivalents of water.

Creatine being present in the muscles, striated and organic,1

Gorup-Besanez, loc. cit., p. 121.