CHAPTER III.THE CHEMICAL UNITY OF LIVING BEINGS.
The varieties and essential unity of the protoplasm—Its affinity for oxygen—The chemical composition of protoplasm—Its characteristic substances.—§ 1. The different categories of albuminoid substances—Nucleo-proteids—Albumins and histones—Nucleins.—§ 2. Constitution of nucleins.—§ 3. Constitution of histones and albumins—Schultzenberger’s analysis of albumin—Kossol’s analysis—The hexonic nucleus.
The chemical unity of living beings corresponds to their morphological unity.
The Varieties and Essential Unity of the Protoplasm.—One essential feature of the living being is that it is composed of matter peculiar to it, which is calledliving matter, orprotoplasm. But this is a somewhat incorrect way of expressing the facts. There is no unique living matter, no single protoplasm; their number is infinite, there are as many as there are distinct individuals. However like one man may be to another, we are compelled to admit that they differ according to the substance of which they are constituted. That of the first offers a certain characteristic personal to the first, and found in all his anatomical elements; similarly for the second. With Le Dantec we shall say that the chemical substance of Primus is not only of the substance of man, but in all parts of his body and in all his constituent cells it is the exclusive substance of Primus; and, in the same way, the living matter of another individual Secundus will carry everywhere his personal impress, which differs from that of Primus.
But it is none the less true that this absolute specificity is based with certainty only on differences which from the chemical point of view are exceedingly slight. All these protoplasms have a very analogous composition. And, if we regard as negligible the smallest individual, specific, generic, or ordinal variations we may then speak in a general manner ofprotoplasmorliving matter.
Experiment shows us, in fact, that the real living substance—apart from the products it manufactures and can retain or reject—is in every cell tolerably similar to itself. The fundamental chemical resemblance of all protoplasms is certain, and thus we may speak of their typical composition. We may sum up the work of physiological chemistry for the last three quarters of a century by affirming that it has established the chemical unity of all living beings—that is to say, a very notable analogy in the composition of their protoplasm.
This living matter is essentially a mixture of the proteid or albuminoid substances, to which may be added other categories of immediate principles, such as carbohydrates and fatty matters. But the latter are of secondary importance. The essential element is the proteid substance. The most skilful chemists have tried for more than half a century to discover its composition. Only during the last few years—thanks to the researches of Kossel, the German chemist, following on those of Schultzenberger and Miescher—we are beginning to know the outer wallsor the framework of the albuminoid molecule; in other words, its chemical nucleus.
Physical Characters of Protoplasm.—About 1860 Ch. Robin thought that he had defined living matter sufficiently—or, at least, as perfectly as could be expected at that time—by attributing to it three physical characteristics. They were:—Absence of homogeneity, molecular symmetry, and the association of three orders of immediate principles—albuminoids, carbohydrates and fats. These characteristics assist, but do not suffice, to define the organization.
No doubt the characteristics must be completed by the addition of a certain number of more subtle physical features.
One of them refers to the structure of protoplasm as revealed by the microscope. Throughout the whole of the living kingdom, from the bacteria studied by Kunstler and Busquet to the most complicated protozoa, protoplasmic matter presents the same constitution, and in consequence, this structure of the protoplasm must be considered as one of its distinctive characters. It is not homogeneous; it is not the last term of the visible organization: it is itself organized. Experiment shows that it does not resist breaking up or crushing. Mutilations cause it to lose its properties. As for the kind of structure that it presents, it may be expressed by saying that it is that of a foamy emulsion.
We saw above that our knowledge as to the physical condition of protoplasm has been completed by the theories of Bütschli’s micellæ or Pfeffer’s tagmata.
Properties of the Protoplasm. Its Affinity for Oxygen.—From the chemical point of view, living matter presents a very remarkable property—namely,a great affinity for oxygen. It absorbs it so greedily that the gas cannot remain in a free state in its neighbourhood. Living protoplasm, therefore, exercises a reducing power. But it does not absorb oxygen in this way for its own advantage; oxygen is not absorbed, as was supposed thirty years ago, to supply fuel wherewith to burn the protoplasm. The products are not those of its oxidation, of its own disintegration. They are the products of combustion of the reserve-stuff which is incorporated in it. These substances have been supplied to it from without, like the oxygen itself, with the blood. This was proved by G. Pflüger in 1872 to 1876. The protoplasm is only the focus, the scene, or the factor of combustion. It is not its victim, it does not itself furnish the fuel. It works like the chemist, who obtains a reaction with the substances that are given to him.
As for the reducing power of protoplasm, A. Gautier in 1881 and Ehrlich in 1890 have given fresh proofs. A. Gautier in particular has insisted that the phenomena of combustion take place, so to speak, outside the cell, and at the expense of the products which surround it; while on the contrary the really active and living parts of the nucleus and of the cellular body, work protected by the oxygen, as in the case of anaerobic microbes.
This result is of great importance. Burdon Sanderson, the late learned professor of physiology at the University of Oxford, has not hesitated to compare it to the discovery of respiratory combustion by Lavoisier. There is no doubt some exaggeration in the comparison; but there is, on the other hand, no less exaggeration in supposing that it is not of greatimportance. We may no longer in these days speak without reservation of the vital vortex of Cuvier, and of the incessant twofold movement of assimilation and dissimilation which is ever destroying living matter and building it up again. In reality, the living protoplasm varies very little; it only undergoes oscillations of very slight extent; it is the materials, the reserve stuff on which it operates, which are subject to continual transformations.
Chemical Composition of Protoplasm.—One of the the three characters attributed by Ch. Robin to living matter was its chemical composition, of which little was known in his time. He insisted on the constant presence in the living elements of three orders of immediate principles—proteid substances, carbohydrates, and fatty bodies. In reality the proteid substances, or albuminoids, alone are characteristic. The two other groups, carbohydrates and fatty bodies, are rather the signs and the products of the vital activity, than constituents of the matter on which it is exercised.
It is therefore on the knowledge of the proteid substances that all the sagacity of biological chemists has been exercised. Their efforts for thirty years, and particularly in the last few years, have not been barren; they enable us to give a first rough sketch of the constitution of these substances.
The Different Categories of Albuminoid Substances.—Albuminoid or proteid substances are extremely complex compounds, much more so than any of thosewhich are being constantly studied by the chemist. They also are to be found in great variety. It has been difficult to separate them one from the other, to characterize them rigorously, or, in other words, to classify them. However, it has been done now, and we distinguish three classes which are differentiated at once from the physiological and from the chemical points of view. The first comprises the complete or typical albuminoids. They are theproteidsornucleo-albuminoids. They are to be found in the most active and most living parts of the protoplasm, and therefore in the spongioplasm of the cell and around the nucleus. The second group is formed ofalbuminsandglobulins, compounds already simpler, fragments derived from the destruction of the preceding, into which they enter as constituent elements. In the isolated state they do not belong to the really living protoplasm; they exist in the cellular juice, in the interstitial and circulating liquids in the blood and in the lymph. The third category comprises real but incomplete albuminoids. They are to be found in the portions of the economy which have a specialized or attenuated life, and are destined to serve as a support to the more active elements—i.e., they contribute to the building up of the bony, cartilaginous, conjunctive, elastic tissues. They are calledalbumoids. It is naturally the first group, that of the proteids—i.e., of the complete and characteristic compounds of the living substance—upon which the attention of the physiologists must be fixed. It is only quite recently that the clear definition of these substances has been given, and proteid compounds detected in the confused mass.
The Nucleo-proteids.—This progress in the characterization and specification of the proteids required in the first place a knowledge of two particular compounds, thenucleinsand thehistones. This did not become possible until after the researches of Miescher and Kossel on the nucleins, which went on from 1874 to 1892, and those of Lilienfeld and d’Yvor Bang on the histones, from 1893 to 1899. The complete albuminoids are constituted by the combination of two kinds of substances—albumins or histones on the one hand, and nucleins on the other. By combining solutions of albumins or histones with solutions of nuclein, the synthesis of the proteid is effected. The study of the properties and characteristics of these nucleo-albumins and nucleo-histones is going on at the present moment. It is being carried out with much method and with wonderful patience by the German school.
All the proteids contain phosphorus in addition to the five chemical elements, carbon, oxygen, hydrogen, nitrogen, and sulphur, which are common to the other albuminoids. Another interesting feature in their history is that the action of the gastric juice divides them into their two constituents:—the nuclein, which is deposited and resists the destructive action of the digestive liquid, and the albumin or histone, which on the contrary experiences this action with the usual consequences. Thus the gastric juice furnishes a process which is very simple and very convenient in the analysis of the proteids.
Localization of the Nucleo-Proteids.—What we said before as to the important physiological rôle of the cellular nucleus may arouse the expectation that in it will be found the living matter which is chemically the most differentiated, the albuminoids of highestrank—i.e., the nucleo-proteids and their constituents. Not that they would not be found in the protoplasm of the rest of the cell, but there is certainly a risk that they would be less concentrated there and more blended with accessory products; they are there connected with much more secondary vital functions. This conclusion inspired the early researches of Professor Miescher, of Basle, in 1874, and, twenty years later, those of Professor Kossel, one of the most eminent physiological chemists in Germany.
In fact, these compounds have been found in all tissues which are rich in cellular elements with well-developed nuclei. The white globules of the blood furnished to Lilienfeld the first nucleo-histone ever isolated. The red globules themselves, when they possess a nucleus, which is the case in birds and reptiles as well as in the embryo of mammals, contain a nucleo-proteid which was easily isolated by Plosz and Kossel. Hammarsten, the Swedish chemist, who has acquired a great reputation from his researches in other domains of biological chemistry, prepared the nucleo-proteids of the pancreas in 1893. They have been obtained from the liver, from the thyroid gland (Ostwald), from brewers’ yeast (Kossel), from mushrooms, and from barley (Petit). They have been detected in starchy bodies and in bacteria (Galeotti).
Constitution of Nucleins.—Our path is already marked out if we wish to penetrate farther into the constitution of these proteids, which are the immediate principles highest in complexity among those which form the living protoplasm. We must analyze the two components, the albumins and the histones on the one hand, and the nucleins on the other. As for the nucleins, this has already been done, or very nearly so.
Kossel, in fact, decomposed the nuclein by a series of very carefully arranged operations, and has reduced it step by step to its crystallizable organic radicals. At each stage that we descend in the scale of simplification a body appears which is more acid and more rich in phosphorus. At the third stage we come to phosphoric acid itself. The first operation divides the nuclein into two substances: the new albumin and nucleinic acid. After separating these elements they can be reunited: a solution of albumin with a solution of nucleinic acid reconstitutes the nuclein. A second operation separates the nucleinic acid in its turn into three parts. One is a body of the nature of the sugars—i.e., a carbohydrate. The appearance of a sugar in this portion of the molecule of nucleinic acid is an interesting fact and fertile in results. The second part is constituted by a mixture of nitrogenous bodies, well known in organic chemistry under the name ofxanthic bases(xanthin, hypoxanthin, guanin, and adenin). The third part is a very acid body and full of phosphorus—thymic acid. If in a third and last operation the thymic acid is analyzed, it is finally separated into phosphoric acid and into thymene, a crystallizable base, and thus we are brought back to the physical world, for all these bodies incontestably belong to it.
Constitution of Histones.—But we are only half-way through our task. We are acquainted in its origin with one of the genealogical branches of the proteid, the nucleinic branch. We must also learn something of the other branch, the albumin or histone branch. But on this side the problem assumes a character of difficulty and complexity which is admirably adapted to discourage the most untiring patience.
The analysis of albumin for a long time baulked the chemist “Here,” said Danilewsky, “we come to a closed door which resists all our efforts.” We know how vastly interesting what is taking place on the other side must be, but we cannot get there. We get a mere glimpse through the cracks or chinks which we have been able to make.
This analysis of albuminous matter at first requires great precautions. The chemist finds himself in the presence of architecture of a very subtle kind. The molecule of albumin is a complex edifice which has used up several thousand atoms. To perceive the plan and structure, it must be dismantled and separated into parts which are neither too large nor too small. Such careful demolition is difficult. Processes too rough or too violent will reduce the whole to the tiniest of fragments. It is a statue which may be reduced to dust, instead of being separated into recognizable fragments, easily fitted in place along their fractured faces.
Analysis of Albumin by Schützenberger.—Schützenberger, a chemist of great merit, attempted (about1875) this thankless task. Others before him had experimented in various ways. Two Austrian scientists, Hlasitwetz and Habermann, in 1873, and a little later Drechsel in 1892, had used concentrated hydrochloric acid to break down albumin. They also employed bromine for the same purpose. More recently Fuerth had used nitric acid with a similar object. Schützenberger tried another way. The battering ram which he used against the edifice of albumin was a concentrated alkali, baryta. He warmed the white of an egg with barium hydrate in a closed vessel at a temperature of 200°. The albumin of egg then divides into a certain number of simpler groups. The difficulty is to isolate and to recognize each part in this mass of the materials of demolition. That can be done by the aid of the processes of direct analysis. By mentally combining these different fragments, the original building is reconstructed. This method of demolition is certainly too rough and violent. Schützenberger’s operation gives us very fine fragments—small molecules of free hydrogen, of ammonia, of carbonic, acetic, and oxalic, acids which reveal extreme pulverization. These products represent about a quarter of the total mass. The other three-quarters are formed of larger fragments, the examination of which is most instructive. They belong to four groups. The first comprises five or six bodies, amido-acids orleucins. It proves the existence in the molecule of albumin of compounds of the series of fats—i.e., arranged in an open chain. The second group is formed by tyrosin and kindred products—i.e., by the bodies of the aromatic series, which force us to acknowledge the presence in the molecule of albumin of a benzene nucleus. The thirdgroup forms around the nucleus known to chemists under the name of pyrrol. The fourth comprises bodies such as the glucoproteins, connected with the sugars, or carbohydrates.
Does the fact that the molecule of albumin is destroyed in producing these compounds raise the question as to whether it implies the idea that in reality they pre-exist in it? Chemists are rather inclined to admit this. However, the conclusion does not appear to be permissible. Duclaux considers it doubtful. It is not certain that all these fragmentary bodies pre-exist in reality, and it is no more certain that a simple bringing of them together represents the primitive edifice. Materials of demolition from a house that has been pulled down give no idea of its natural architectural character. There is only one way of justifying the hypothesis, and that is to reconstitute the original molecule of albumin by bringing the fragments together. We have not got to that stage yet. The era of syntheses of such complexity is more or less near, but it has certainly not yet begun.
Moreover, it is not correct to say that the simple juxtaposition of the surfaces of fracture will reproduce the initial body. The fragments, so far as analysis has obtained them, are not absolutely what they might have been in the original structure. There they adhered the one to the other, not only by the mere contact of their surfaces of fracture, as is supposed, but in a slightly more complex manner. The fragments of the molecule are joined by bonds. We can picture them to ourselves by supposing these bonds to be like hooks. The hooks, which could only be broken by violence, are called by the chemistssatisfied atomicities. These atomicities, set free by the breaking up, cannot remain in this condition; they must be satisfied anew. The hook tries to attach itself. In Schützenberger’s experiment the addition of water provides for this necessity. A molecule of water (H2O) splits into two, the hydrogen (H) on the one side and the hydroxyl (OH) on the other. These two elements cling to the liberated bonds of the fragments of the molecule of albumin, and thus the bodies were found complete. Schützenberger’s experiment was too violent, too radical, and it gave too large a number of fragments, with their free hooks and atomicities unsatisfied, for rather a large proportion of the water added disappeared during the experiment. In one case this quantity was as much as 17 grammes per 100 grammes of albumin. The molecules of this water were employed in the reparation of the incomplete fragmentary molecules of the albumin.
It follows that Schützenberger’s experiment gave too large a number of very small pieces corresponding to far too great a pulverization. The very small fragments are the molecules of acids such as acetic acid, oxalic acid, carbonic acid, molecules of ammonia, and even of hydrogen, which we know we are setting free.
But, apart from these products which represent a quarter of the molecule of albumin submitted to analysis, the other three quarters represent larger fragments which may be considered as the real constituents of the building. Thus we find four kinds of groups which may be accepted as natural. The first of these groups is that of the leucins or amido-acids. It proves the existence in the moleculeof albumin of compounds of the fatty series. There is also an aromatic group—a pyridine group—and a group belonging to the category of sugars. Imagine a certain grouping of these four series. This would be the nucleus of the molecule of albumin. If we graft on to this nucleus, on to this framework as it were, so many annexes, or lateral chains, the building will be loaded with embellishments; it will have been made unstable andipso factoappropriate for the part that it plays in the incessant transformations of the organism.
Kossel’s Analysis. Hexonic Nucleus.—Kossel has approached the problem in another fashion. He did not attempt to attack the albumin of the egg. This body is, in fact, a heterogeneous mixture as complex as the needs of the embryo of which it forms the food. Kossel tried a physiologically simpler albuminoid. He got it from an anatomical element having no nutritive rôle, of a very elementary organization and physiological functional activity, and yet one of energetic vitality—the male generating cell. Instead of the hen’s egg he therefore analyzed the milt of fish, and, in the first place, of salmon. As was to be expected from what has been said of the proteids, this living matter gives a combination of the nuclein, already known, with an albumin. The latter is abundant, forming a quarter of the total mass. Its reaction is strongly alkaline, which is the general characteristic of the variety of albumin known by the name of histones. Miescher, the learned chemist of Basle, who had noticed this basic albumin when working on the Rhine salmon, gave it the name of protamin. This is the substance submitted by Kossel to analysis in preference to the albumin ofegg, so dear to the chemists who had preceded him. The disintegration of this molecule, instead of giving the series of bodies obtained by Schützenberger, gave but one, a real chemical base,arginin. At the first trial the albumin examined was reduced to a simple crystallizable element. The conclusion was obvious. The protamin of salmon is the simplest of albumins. To form this elementary proteid substance a hexonic base with water is all that is required.
Continuing on these lines other male generating cells were examined and a series of protamines constructed on the same type was found, and these albuminous bodies proved to be formed of a base or mixture of analogous hexonic bases: arginin, histidin, and lysin—all bodies closely akin in their properties and entirely belonging to the physical world.
Once aware of the existence of this fundamental nucleus, chemists found it in the more complex albumins where it had been missed. It was found in the albumin of egg hidden under the mass of other groups. It was recognized in all animal or vegetable albumins. The nuclei of Schützenberger may be missing. Hexonic bases are the constant and universal element of all varieties of albumins. They prevail in the chemical nucleus of the albuminous molecule, and perhaps as is suggested by Kossel, they may form it exclusively. All the other elements are superadded and accessory. The essential type of this molecular edifice, sought for so long, is known at last.
Conclusion.—To sum up, the chemical unity of living beings is expressed by saying that living matter, protoplasm, is a mixture or a complex of proteid substances with an hexonic nucleus.