Friday, October 20, 2006

CHAPTER III.: The Blood and the Third Anatomical Element by Antoine Bechamp


The experiments which demonstrated that the fibrin was of a complete whole, formed of a special albuminoid matter belonging to it, and of microzymas, and not a proximate principle, did not solve the problem raised by Dumas, to ascertain under what condition a substance thus composed existed in the blood; neither did it solve the problem whether such a substance pre-existed in it or if it was the result of a chemical transformation accomplished after the bleeding.
It was not at the first attempt that I solved the problem in the sense of the conclusion arrived at in this chapter, viz., that the blood really contains the fibrin in the state of microzymian molecular granulations where the microzyma and the special albuminoid matter are closely associated in a very special anatomical element. The solution of this problem could only be completely given after the collection of the observations which have been summarized in the first two chapters and of those which I have yet to describe.
Let us recall first what, in 1869, was the state of opinion regarding, first, the pre-existence of the fibrin in the blood; second, its production in the blood after it had issued from the vessels. On the pre-existence of the fibrin there were two opinions: According to one, which was that of Hewson, of Milne-Edwards and of Dumas, fibrin existed in the blood in a condition of extreme division, in fine molecules, which, after the bleeding, became consolidated to constitute the ordinary
fibrin. According to the other, which was also that of Hewson, and, in a certain manner, of Dumas, it existed in it in a state of solution or of quasi-solution. Cl. Bernard1 allied himself thereto by admitting that the blood contained an albumino-fibrinous liquid which could only remain liquid in the economy, taking on the form of fibrin after the bleeding.
1. CL Bernard, "Liquides dc 1'organisme." Vol. I, p. 152 (1859).
These opinions, nevertheless, have had no part in the solution of the problem. They had been so thoroughly lost to sight that Estor and I had tacitly ranged ourselves among those who did not admit the pre-existence of the fibrin. Indeed, after having proved the presence of the microzymas of the blood in the fibrin, by the fact of their vibrionian evolution in the substance itself, we said, in February, 1869:
"That which is called the fibrin of the blood is a false membrane formed by the microzymas of the blood, associ­ated with a substance which they secrete in an acid state from the albuminoid elements of this liquid."1
It was because we had sought for and discovered the molecular granulations in the blood, before proving that there were microzymas in the fibrin, that we compared them to the microzymas of the liver, finding them smaller and more transparent than the latter.2
Nevertheless, these microzymas in the blood had not been isolated by us. The consideration that, in the liver, the microzymas are especially included in the hepatic cellules led us to seek in like manner for the microzymas in the globules of the blood. It was on this occasion that we made an experiment suggested by that observation, which is as follows:
1. C. R.,Vol. LXVIII.p. 408. 2. lbid.. Vol. LXIV. p. 713.
Dr. Clement Combescure, who, alongside of me, had experimented on the blood from the point of view of its supposed coagulation in the vessel by an excessive consumption of alcoholic drinks, had long ago discovered that the blood, received directly from the vessels into alcohol at 40% to 45%, far from coagulating, dissolved in it.1
The experiment being repeated under the same conditions we found that the mixture of blood and alcohol remained liquid, appearing limpid, depositing neither globules, nor fibrin, but that by degrees it made an abundant deposit, which the microscope showed was almost exclusively formed of molecular granulations animated by the brownian movement.2 It was from this result that we finally reached the solution of the problem, but only long after I had resumed the study of the fibrin and of its changes.
As I had done in 1869,1 at first regarded the molecular granulations of the deposit from the alcoholized blood as being the microzymas of the blood or the microzymas of the globules. But when I had isolated the fibrinous microzymas of an extreme minuteness and after the study of the molecular granulations of spontaneously changed fibrin, liquified, without becoming fetid, I doubted. Here are the consequences of this doubt:
1. Dr. C. Combescure, Theses sur les effets therapeutiques des ammoniacaux. p. 82. Theses of the Faculty of Medicine of Montpellier (1861). 2. C.R.. Vol. LXX, p. 265 (1870).
The third anatomical element of the blood and the molecular granulations of the blood globules.
The conditions of the experiment for isolating the third anatomical element of the blood are as follows: Take alcohol, rigorously rectified, free from acid and from alkali, and dilute it with distilled water, to bring it to from 35 to 40 per cent. Into two volumes of alcohol thus diluted, one volume of blood is made to flow directly and without interruption as it comes from the vessels. So much for the blood as a whole.
For examining blood already defibrinated it must be passed through a fine linen cloth, to remove the fibrin which
might be held in suspension, and it also is then poured into twice its volume of like diluted alcohol.
The mixtures, dark red, being left to themselves in a cool place, there is formed by degrees a clear red deposit which takes at least 24 hours to be completed.
The deposit is much more abundant for the entire blood than for the defibrinated.
The deposit is first washed by decantation in alcohol at 35 per cent, then on a filter with alcohol at 30 per cent., until it is perfectly white. Under the microscope the matter resolves itself into an infinite number of very fine molecular granula­tions. These granulations are mixed with remains of cellules, more abundant in a deposit furnished by the defibrinated blood.
I made several determinations of these molecular granulations. The following were made upon sufficiently large volumes of sheep's blood, by bleeding from the jugular vein.
800 c.c. of the whole blood gave 37.4 gr. of humid granulations completely drained, containing 5.76 grammes of matter dried at 120° (= 248° F.); that is, 7.07 grammes of dried granulations per litre of the whole blood.
2675 c.c. of the same blood, first defibrinated, gave 22.1 gr. of humid granulations, drained, and containing 4.87 gr. of matter dried at 120° C.; that is, 1.82 gr. per litre of defibrinated blood.
But these quantities are far from being constant, even for the blood of the same animal. For example, one litre of sheep's blood, in another experiment, gave only 5.70 gr. of granulations dried at 120° C., and another by whipping only, gave 3.15 grammes of fibrin, dried at 120° C., per litre.
However it maybe with regard to the molecular granulations of defibrinated blood let us consider them as representing (as will be hereafter demonstrated) the molecular granulations and the envelopes of the destroyed globules; the difference 7.07 gr.— 1.82 gr. = 5.25 gr. will represent the molecular granulations which the blood without the globules will have furnished.
In the "Memoire sur les matieres albuminoides," I still considered the molecular granulations of which I was treating as being microzymas such as they existed in the blood, and I showed that, like the fibrinous microzymas, they liquified fecula-starch and decomposed oxygenated water. In fact, 1 c.c. of humid molecular granulations of the entire blood, disengaged, in 12 hours, 26 c.c. of oxygen from 2 c.c. of water oxygenated to 15 volumes of oxygen; and 1 c.c. of humid molecular granulations of defibrinated blood, disengaged, in 12 hours, 23 c.c. of oxygen from 2 c.c. of the same oxygenated water.
These granulations possess then the properties of the fibrin and of the fibrinous microzymas. But are they really isolated microzymas? Are they not precisely the fibrin, such as it exists in the blood? That which led me to put this question was, first, the observation of molecular granulations of the fibrin spontaneously changed and, second, that the weight of these granulations is often greater than the weight of the fibrin which the same volume of blood was able to furnish.
I then treated granulations obtained from the blood, diluted in alcohol as I had treated those of the altered fibrin, or the fibrin itself, to isolate from it the microzymas.
Treatment of the haematic molecular granulations with very dilute hydrochloric acid. The humid deposit of the isolated granulations is treated with very dilute hydrochloric acid; in contrast with what happens to fibrin they disappear almost instantaneously at the ordinary temperature, even when the acid is diluted to one in a thousand. In some min­utes ten grammes were dissolved in a cloudy liquid from which microzymas, as small as those of the fibrin from whipping, were slowly deposited.
The limpid hydrochloric solution, separated from the microzymas, exactly saturated by carbonate of ammonia, furnished a precipitate of fibrinine, which when well washed does not decompose oxygenated water, while the microzymas, isolated and washed, decompose it, etc.
The rotatory power of the materials dissolved by the hydrochloric acid was (a) j = —74°.
As to the rotatory power of fibrinine, it has been found in acetic solution (a) j = —68°.9.
That is to say, perceptibly those of the fibrin and of fibrinine of fibrin under the same conditions.
The very great difference between fibrin in the state of molecular granulations and fibrin obtained by whipping resides then essentially in the manner of reacting with regard to very dilute hydrochloric acid, the solution of ordinary fibrin, being a function of time and of temperature.
The blood of the ox and that of the rabbit behaved exactly as did that of the sheep. But the blood of the duck offered interesting peculiarities, as we shall perceive presently.
The only condition for the success of the experiment, but rigorously indispensable in every case, is that the shed blood shall flow directly from the vessel into alcohol at 35-40 per cent., the volume of which should be twice that of the blood to be collected. The least interval, as, for instance, receiving the blood into a porcelain capsule or one of glass, from which to pour it into the diluted alcohol is sufficient to compromise the result. Under circumstances similar to the last the deposit seemed to be made more rapidly and instead of being pulverulent, it was flocculent. The deposit, collected as usual, 24 hours
end page 115 missing
which (when poured directly into the alcohol) it was effected at once. Hence it results that the matter of the molecular granulations of the deposit formed by the instantaneous mixture of the blood, as it issues from the vessels, with the dilute alcohol, is in a state, chemical, physiological and an­atomical nearest to that which it assumes in the circulating blood. It is, therefore, necessary to obtain the most exact idea of the physical constitution of the molecular granulations in that condition, in order to understand their constitution in the blood at the moment that this is held by its flowing into the alcohol.
It is an invariable fact that the molecular granulations of the deposit formed by the instantaneous mixture of the blood, at the time of the bleeding, with twice its volume of alcohol at 35-40 per cent, are immediately dissolved by very dilute hydrochloric acid to a cloudy liquid containing the microzymas, and that at low temperature. And it is a no less invariable fact that at the moment of the mixture with alcohol the blood appears to be dissolved, so much so that a thin layer of the liquid is almost transparent; under the micro­scope no globules are seen, and it is with difficulty that one can perceive certain particles which are themselves trans­lucent. The deposit, which is then made very slowly, is not the result of a formation first of all of a precipitate due to some reaction or coagulation of some dissolved matter; it is quite otherwise, since the globules which swim insoluble in the circulating blood are themselves dissolved, while being des­troyed. It is further evident that if the deposit had been pro­duced by coagulation, by the alcohol, of a substance which existed dissolved in the blood and which became insoluble in water from the fact of this coagulation, but became soluble in very dilute hydrochloric acid, the whole should be dissolved by the latter! But the microzymas are the persisting insoluble residue of the deposit as they are of the fibrin obtained by whipping. But, further, this important fact must be borne in mind to which I have already called attention that it is precisely when an interval, even a very short one elapses between the venesection and the mixture of the blood with the alcohol, that the molecular granulations o the deposit are not immediately dissolved by the dilute hydrochloric acid; that is to say, that a certain coagulation has occurred.
We reach then the conclusion that the granulations o the deposit formed instantaneously in the mixture represent the nearest state to that which they have in the blood at the precise moment of the bleeding.
But what is the relation between the part soluble in dilute hydrochloric acid of these molecular granulation: and the microzymas which remain undissolved?
It is the same which I have pointed out in the molecular granulations which exist in the deposit from the spontaneous alteration of the fibrin in carbolated water, WHERE EACH GRANULATION IS ASPHERICAL MASS OF ALBUMINOID MATTER HVING A MICROZYMA FOR ITS CENTRE. In fact, the molecular granulations of the deposit formed by the alcoholized blood are like that, round, spherical, motile; that is to say, animated by the brownian movement, representing an exceedingly minute mass of albuminoid matter having a microzyma for centre. The very dilute hydrochloric acid dissolves the enveloping albuminoid matter, leaving the central microzyma undisolved.
A microzyma for nucleus, enveloped as by an atmosphere by a mass of albuminoid matter, insoluble in water, but which very dilute hydrochloric acid dissolves, such then is the physical constitution of a molecular granulation formed by blood diluted in two volumes of alcohol at 35-40 per cent. It may be called a microzymian molecular granulation.
But do molecular granulations thus constituted exist anatomically? and do any such exist in the blood? Yes, and the example is not solitary,1 but the haematic microzymian molecular granulation, with its special albuminoid atmosphere, is a prime example of this kind. It only remains to represent that state of this atmosphere in the blood.
I revert to the remark already made, that the deposit in the alcoholized blood is not the result of the precipitation of some dissolved substance contained in the blood, according to the plasma hypothesis. But direct observation had already permitted Estor and me to declare that the blood, as it issues from the vessels before the commencement of the formation of the clot, contains, around the globules, an innumerable number of microzymas (that which we took for microzymas) most readily to be seen in the blood of very young animals— for example, of kittens from three to forty days old; and these microzymas we found to resemble those of the liver, but more transparent; and we did not fail to add that the reason they had escaped the attention of histologists was because of their minuteness and transparency.2 It was in reality a diligent idea which enabled us to find them where they had not before been seen. In the defibrinated blood they cannot be found.
1. Before I discovered the microzymian molecular granulations with an albuminoid atmosphere. I had already observed some of them of another kind. In isolating the microzymas of the pancreas, as Estor and I had isolated those of the liver, I found them enveloped with an atmosphere of complex matter. Treatment with alcoholized ether and with water dissolved the matters of the enveloping atmosphere and the naked pancreatic microzymas became visible with their special minuteness and color. The vitellin microzymas of the yolk of the eggs of birds are also enveloped with a complex matter. When the sort of tissue which constitutes the yolk of these eggs is steeped in a good deal of distilled water the intergranular materials dissolve and the vitellin molecular granulations are deposited, spherical, sometimes mixed with vitellin globules. Washing with water removes everything which can be dissolved in it; then treatment with ether and with alcoholized ether dissolves the enveloping matter, a son of alloy, an amalgam of fatty bodies and of lecithin. Finally, washing with water and again with ether yields the vitellin microzymas of a perfectly white color. The vitellin microzymas and the pancreatic exist then well enveloped, in the condition of microzymian molecular granulations.2. C. R-.VoI. LXIX, p. 713.
That which we took for transparent microzymas, visible with difficulty, were the haematic microzymian molecular granulations, the same as those of the deposit in the alcoholized blood, except that the albuminoid atmosphere of the latter is a condensed atmosphere, contracted, become opaque, while in the blood it is inflated, soft and mucuous, hyaline, and can again, as will be presently shown, become inflated in blood, to which water had been added. The theory of the phenomenon presented by the blood diluted in the alcohol is as follows:
When the blood is directly received into the alcohol under the specified conditions, its anatomical elements are rudely placed in new conditions of existence; while the globules are destroyed and their coloring matter (the haemoglobin) dissolves, the soft and mucoid atmosphere of the insoluble microzymian molecular granulations condenses by degrees and hardens around each central microzyma; then, becoming more dense, the microzymian molecular granulations are deposited.
And it is because the mucoid albuminoid atmosphere, insoluble in water, is condensed and hardened, laid hold of as it were, by the alcohol before being coagulated, that it is dissolved immediately in very dilute hydrochloric acid, while it is modified by coagulation, and becomes insoluble in it, as it is in the case of fibrin obtained by whipping, if some interval of time separates the venesection from the mixing of the blood with the alcohol. Once more, the condensed atmosphere of the microzymian molecular granulations contains the albuminoid matter in the state nearest to, if not identical with, that which it had in the blood.
The following experiment will instruct us still better as to the nature of the mucoid atmosphere of the haematic microzymian molecular granulations.
Experiment upon blood diluted in a saturated aqueous solution of sulphate of soda.
It is known that blood mixed with several times its volume of a saturated solution of sulphate of soda yields no clot and that the globules are deposited in the mixture without yielding up their coloring matter. Why under these conditions is no clot formed? The following is an attempted explanation of the phenomenon. The experiment ought to be made in winter in freezing temperature.
A volume of sheep's blood is received directly from the jugular vein into four times its volume of a saturated solution of sulphate of soda and the mixture left at rest. Twenty-four hours afterwards the greater part of the globules will be deposited. The supernatant clear liquid is filtered upon a filter lined with sulphate of barium,1 in order to retain the globules and the microzymas which remain in suspension. The filtration is necessarily slow. The filtered liquid, almost colorless, is absolutely clear; mixed with oxygenated water, it slowly sets free a little oxygen.
1. The lining with sulphate of barium is made as follows: The filter should be without a fold; it must be filled with liquid, into which a suitably diluted solution of chloride of barium has been precipitated by a similar solution of sulphate of soda; the filtrate is to be again poured on the filler until that which passes is perfectly clear. It must be so arranged that the bed of sulphate of baryta be at least half a millimetre in thickness. Finally the filter is to be washed with a solution of sulphate of soda.
The liquid which had remained limpid during the whole period of filtration (about 20 hours) furnishes by agitation a small mass of fibrin of a brilliant whiteness, having the membranous appearance of the fibrin obtained by whip­ping. The liquid separated from this matter, again filtered on a filter lined with sulphate of baryta, also sets oxygen free from oxygenated water.
The fibrin separated from the filtered liquid of the mixture of blood and of sulphate of soda, that is to say, a fibrin without microzymas, does not set free oxygen from oxygenated water, but dissolves in it.
A mass of this fibrin without microzymas, about 1 c.c. being placed in 8 c.c. of oxygenated water having six volumes of oxygen, did not set oxygen free even after six days of contact (at least not more than would have been set free from it without the addition), but the fibrin had disappeared; it was dissolved. And the solution was albuminoid, for on treating with Millin's re-agent, a white precipitate appeared which became red on slightly heating it.
As to the clear liquid separated from this fibrin without microzymas, it gave, on acetic acid being carefully added, a slight albuminoid precipitate which has not been further examined. But the liquor separated from this precipitate contained a soluble albuminoid matter precipitated by alco­hol, which, in acetic solution, had a rotatory power: (a) j= — 86°, very different from that of seralbumine.
From this experiment we may conclude that the microzymian molecular granulations are, like the globules, insoluble in blood, where the conditions of their anatomical integrity exist united; but the blood being diluted in the solution of sulphate of soda, although the globules remain insoluble, the albuminoid substance which forms the soft atmosphere of the microzymian granulations is dissolved in the new medium, at least in part, undergoing doubtless some transformation. In fact, while by agitation a part is separated in the condition of an insoluble mass1 having the membran­ous appearance of fibrin, another part remains dissolved and can be separated from it, and has a rotatory power greater than that of seralbumin. The fact of the change is also evident from this; that the insoluble matter of fibrinous appearance is dissolved in oxygenated water without setting oxygen free. And if the limpid liquid of the filtration before and after the separation of this fibrinous substance sets free a little oxygen from the oxygenated water, it is because some of the matter which, in the microzymas, effects this disengagement is diffused in it. Thus the direct experiment demonstrates that that which in the fibrin decomposes the oxygenated water are the microzymas; the intermicrozymian mass or the enveloping albuminoid matter does not decompose it; thus constituting a verification of the facts established in the first chapter; so much so that it is unnecessary to add that the defibrinated blood, treated with sulphate of soda, does not yield any fibrinous substance.
1. There is nothing to be surprised at in this spontaneous passage from the dissolved state to an insoluble state. There is a parallel case in a modification of amylaceous matter, which, from a condition of perfect solution, passes by degrees, in the liquor itself, to an insoluble stale.
Such are the facts. But these microzymian granulations, which Estor and I took for microzymas, had they not been already perceived? On this subject the following is the only information I have been able to collect:
"There is to be found, says M. Frey,1 in human blood besides globules agglomerations of small pale granulations 0.00 l mm to 0.002 mm in diameter (Schultze); adding that these granulations, which had been noticed before, showed themselves sometimes with active movements of protoplasm, sometimes with a molecular movement (brownian move­ment). "2
1. Frey, Traite d'histologie et d'histo-chimie; Fr. trans. From the Ger. by P. Spielman, p. 120 (1877). Eng. trans, by A. E. Barker, p. 108. N.Y., Appleton. 1875. Note'p- sup. 2. Frey. loc. cit., p. 121.
Further these molecular granulations had been observed in other humors and animal tissues and many opinions had been expressed as to their role, but it was not known what that role was, nor whether they were organized.
Now, thanks to the anatomical analysis of blood treated with alcohol, the existence of the molecular granulations of the blood is certain, it remains to explain how such an observer as J. Muller did not see them and could have maintained, under microscopic examination, that, except the globules, the whole of the rest of the blood was in a state of perfect solution. To understand this, it is sufficient to take into consideration the anatomical and physical constitution of the haematic microzymian molecular granulations in the blood; a microzyma, (whose diameter at the most is 0.0005 mm) and is enveloped in an atmosphere of a soft substance, mucous and hyaline. But in the blood this mucous atmosphere when much inflated may have the same refractive power as the surrounding liquid, it is not then surprising, the central microzyma being so very small, that it escaped microscopic observation; in fact, it only becomes visible when the albuminoid matter of the enveloping atmosphere, outside of the vessels, begins to undergo the allotropic modifications which cause it to acquire the properties which it possesses in the fibrin.
To convince oneself that this interpretation is a true one, it is sufficient to consider crystallin, the transparency whereof is perfect. Nevertheless, anatomically, crystallin is constituted by two layers of crystalline tubes; further it contains, like other anatomical elements, a crowd of micro-zymas; all this the microscope is incapable of showing directly because all the parts have in the entire organ the same refractive index. But so soon as, by grinding, this organization is destroyed, the conditions of existence of the anatomical elements of the organ being altered, microzymas and crystalline tubes become visible.
To sum up; the facts of this chapter establish definitely that the blood contains a third anatomical element, as constant, as necessary as the globules, consisting of a microzyma enveloped in an atmosphere of a special albuminoid substance, insoluble in the sanguineous medium. This anatomical element, heretofore unrecognized, because of its anatomical constitution, its location and its properties, I have named haematic microzymian molecular granulation.
And now, when one considers that the weight of these microzymian molecular granulations, deduction made of the molecular granulations which are furnished by the same volume of defibrinated blood, represents very nearly the weight of the fibrin obtained from the same volume of blood by whipping, it becomes evident that ordinary fibrin is nothing else than the microzymian granulations, heaped up and soldered together, whose albuminoid atmosphere has undergone, outside of the vessel, an allotropic modification, by virtue of which, from having been directly soluble in very dilute hydrochloric acid, it has become soluble in it, only as a function of time and of temperature.
We shall see how the anatomic constitution of the haematic microzymian granulations and the properties of their enveloping albuminoid atmosphere explain at the same time, mechanically, the phenomenon of the spontan­eous coagulations of the blood and the production of the fibrin by whipping. Meantime let us say that the foregoing demonstrations destroy the hypothesis of the plasma and verify, while completing, the conceptions of Hewson, of Milne-Edwards, of J. B. Dumas, regarding the existence of the fibrin in the condition of fine granulations in the blood.

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