On Digestive Proteolysis / Being the Cartwright Lectures for 1894

by R. H. Chittenden

COMPOSITION OF ANTIPEPTONES.

	From blood-fibrin.[172]	From blood-fibrin.[173]	From antialbumose.[174]	From casein.[175]	From myosin.[176]
C	47.30	49.59	48.94	49.94	49.26
H	6.73	6.92	6.65	6.51	6.87
N	16.83	15.79	15.89	16.30	16.62
S	0.73	—	—	0.68	1.16
O	28.41	—	—	26.57	26.09

From these data it is evident that, while each individual peptone may have a composition peculiar to itself, they are all alike in possessing a relatively low content of carbon. The antialbumid, however, split off in these hydrolytic changes, like the antialbumid formed by the action of dilute acids at 100° C., is characterized by a correspond­ingly high content of carbon and a low content of nitrogen. As an illustration, may be mentioned the myosin-antialbumid formed in the digestion of myosin from muscle-tissue by an alkaline trypsin-solution. This body contains 57.48 per cent. of carbon, 7.67 per cent. of hydrogen, 13.94 per cent. of nitrogen, 1.32 per cent. of sulphur, and 19.59 per cent. of oxygen.[177] It is only necessary to compare these figures with those expressive of the composition of myosin-antipeptone, to appreciate how wide a gap there is between these two products of trypsin-proteolysis, and both members of the anti-group. Antialbumid, however, is a peculiar product, one which is liable to crop out somewhat unexpectedly, and with varying shades of resistance toward the proteolytic ferments. As formed in pepsin-proteolysis, it is more or less readily soluble in sodium carbonate, and in part readily convertible into antipeptone by trypsin. Still, the same substance, or at least a closely related body, makes its appearance in the form of an insoluble residue whenever a native proteid is digested by trypsin. At times, the amount of this insoluble product may be quite large, even reaching to one-fourth of the total proteid matter;[178] but when so formed in the intestine it must entail a heavy loss of nutriment, for whenever the anti-group is split off after this fashion it becomes very resistant to the further action of the ferment. Separating in this manner from an artificial digestive mixture, it may be dissolved in dilute caustic alkali, reprecipitated by neutralization, and then once again brought into solution with dilute sodium carbonate. In this form, it will yield some antipeptone by the further action of trypsin, although even then a large amount of the antialbumid is prone to separate out as a gelatinous coagulum, more or less resistant to the further action of the ferment.

The peculiar action of trypsin, however, as a proteolytic enzyme is shown in the production of a row of crystalline nitrogenous bodies of simple constitution whenever the ferment is allowed to continue its action for any length of time, either on native proteids or on proteolytic products containing the hemi-group. This, to be sure, is a fact long known, but it gains added significance as year by year new bodies are discovered as products of trypsin-proteolysis with various forms of proteid matter. The very character of the bodies originating in this manner gives evidence of the far-reaching decompositions involved; decompositions which are perhaps attributable as much to the innate tendencies of the proteid material as to the specific action of the ferment. As represen­ta­tives of this peculiar line of cleavage, we have first the well-known bodies, leucin and tyrosin; leucin, a body belonging to the fatty acid series, long known as amido-caproic acid, but now generally considered as amido-isobutylacetic acid, (CH₃)₂ CH CH₂ CH (NH₂) COOH; and tyrosin, a body belonging to the aromatic group, having the formula

C6H4		OH
		CH2 CH (NH2) COOH

These two bodies are therefore representatives of two distinct groups or radicals present in the hemi-portion of the proteid molecule; the first belonging to the fatty acid series, the second to the aromatic group from which come such well-known bodies as indol, skatol, benzoic acid, and other substances prominent in proteid metabolism. Moreover, these two hydrolytic products of trypsin-proteolysis are formed in considerable quantity, at least in an artificial digestion. Thus, Kühne has reported the finding of 9.1 per cent. of leucin and 3.8 per cent. of tyrosin as the result of a typical digestion, and I have tried many similar experiments with like results. Further, we know from observations made by different investigators that both leucin and tyrosin may be formed in considerable quantities in trypsin-proteolysis as it occurs in the living intestine. But to this point we shall return later on.

Besides leucin and tyrosin, aspartic acid and glutamic acid have long been known as decomposition-products of the vegetable proteids. Thus, both acids were discovered by Ritthausen and Kreusler[179] in the cleavage of such proteids by boiling dilute acid. Hlasiwetz and Habermann[180] likewise obtained aspartic acid in large quantity by the breaking down of animal proteids under the influence of bromine. Further, Siegfried[181] has recently obtained glutamic acid as a product of the decomposition of the phosphorus-containing proteid, reticulin, from adenoid tissue. As products of trypsin-proteolysis, Salkowski and Radziejewski[182] found aspartic acid in the digestion of blood-fibrin; and v. Knieriem[183] likewise obtained it in the digestion of gluten from wheat. Both of these acids belong to the fatty acid series, the aspartic acid being a dibasic acid, COOH. CH₂CH (NH₂). COOH, or amido-succinic acid, while glutamic acid, COOH. C₃H₅ (NH₂). COOH, is likewise a dibasic acid, known as amido-pyrotartaric acid.

Of more interest physiologically, are the recently discovered nitrogenous bases lysin and lysatinin, or lysatin. These two bodies were first identified by Drechsel[184] and his co-workers as products of the decomposition of various proteids, when the latter are boiled with hydrochloric acid and stannous chloride. They were first obtained by Drechsel as cleavage products of casein.[185] Later, Ernst Fischer,[186] working under Drechsel’s direction, separated them as decomposition-products of gelatin; while Siegfried[187] obtained them as products of the cleavage of conglutin, gluten-fibrin, hemiprotein, and egg-albumin, by boiling with hydrochloric acid and stannous chloride. In all of these cases it is obvious, from the method of treatment pursued, that the two bodies result from a simple hydrolytic cleavage of the proteid molecule. Hence, it might be assumed that these two bases would likewise be formed in trypsin-proteolysis. This assumption, Hedin,[188] working in Drechsel’s laboratory, has proved to be correct, and furthermore he has shown that the amount of these bases formed in pancreatic digestion is not inconsiderable. Thus, as products of the digestion of three kilos. of moist blood-fibrin with an alkaline solution of trypsin, 28 grammes of pure platino-chloride of lysin were obtained, and sufficient lysatinin to establish its identity.

Lysin has the composition of C₆H₁₄N₂O₂, being a diamido-caproic acid, a homologue of diamido-valerianic acid. Hence, this body, like leucin or amido-caproic acid, is a representative of the fatty acid group, the chemical relationship between the two bodies being plainly apparent from their constitution. The constitution of lysatinin is less definitely settled, but apparently it has the composition of a creatin, its formula being C₆H₁₃N₃O₂, in which case it might be more appropriately termed lysatin. The special point of interest, however, connected with this latter body as a product of trypsin-proteolysis is the fact that by simple hydrolytic decomposition, all chance of oxidation being excluded, it can break down into urea.[189] For years, chemists have been seeking to trace out the line of cleavage or decomposition by which urea results in proteid metabolism. In the nutritional changes of the body, nearly all the nitrogen of the ingested proteid food is excreted in the form of urea, but chemists working with dead food-albumin have been heretofore unable to break down proteid matter directly into urea. This, however, Drechsel has now succeeded in doing, and it is to be especially noted that the line of decomposition or cleavage is simply one of hydration, in which the proteid molecule, either through the action of boiling dilute acids, or through the more subtle influence of the hydrolytic enzyme, trypsin, is gradually broken down into cleavage products, from one or more of which comes lysatin. The very resemblance of this body to creatin suggested that, since the latter breaks down into urea and sarcosin when boiled with baryta water, lysatin might possibly behave in a similar manner. This, as has been previously stated, was found to be the case, and Drechsel obtained from ten grammes of a double salt of lysatin and silver one gramme of urea nitrate, by simple boiling with baryta water.