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Alkaloidal Products of Cellular Life.

Before entering upon the study of the true toxins, which are products of an alkaloidal nature and of unknown composition, it is necessary to say a few words regarding the most definite of the toxic alkaloidal principles that are frequently encountered under various conditions, conjointly with the true toxins, particularly in venoms, and which, furthermore, are closely allied to these albuminoid toxins.

These principles are formed in essentially reducing media, whether it be within the body of the organism, and by the simple exercise of its normal function, in which case the principles bear the generic name leucomaines1; or whether due to the action of anaerobic microbes, when they are designated as ptomaines.2 These basic principles, which are essentially the products of cellular secretion, are usually toxic, and sometimes even extremely so.

As we shall presently see, ptomaines are essentially products formed during putrefactive fermentation. The toxic properties of extracts from the cadaveric fluids have long been known. Already in 1838 Panum3 had met with these products in snake venoms. Bergmann and Schmiedberg4 in 1868 isolated from septic pus a toxic substance which they named sepsin; and almost at the same time Zuelzer and Sonnenschein5 reported having isolated from anatomical preparations an alkaloid possessing mydriatic properties. It is, however, due particularly to the researches of Selmi and Armand Gautier that we are now so well informed regarding these toxic principles.

The labors of Armand Gautier were first published in his Traité de Chimie Appliquée à la Physiologie; those of Selmi in the Actes de l'Académie de Bologne.

At first sight, there appears to be a great difference between these alkaloidal bases, the ptomaines and leucomaines, and the albuminoid toxins proper. The toxic bases of the first two groups are quite definite chemical products which can be generally obtained quite pure, and frequently in crystalline form. The toxins proper, on the other hand, are highly complex albuminoid substances which greatly resemble the true diastases in all their properties.

Nevertheless, between the toxic alkaloids, ptomaines and leucomaines, and the toxic albuminoids, or more properly toxins, there exists no absolutely sharp line of demarcation, but there is a gradual passage from the one to the other by every intermediary grade, as a result of the breaking down of the albuminoid molecule.

We shall see, moreover, as we proceed, that these substances are formed under coexistent circumstances, and that they are, hence, found together, whether it be in virus or in snake venom.

We will first consider the ptomaines, and then the leucomaines.

Ptomaines.

This name is more specially reserved to designate those alkaloidal substances, generally highly hydrogenized, that are formed outside the organism, from the fermentative action of anaerobic microbes on albuminoid substances.

These bases are generally volatile, with an intense and tenacious purulent odor; often, however, they possess a floral odor (aubépine, syringa), and even like that of musk. They combine readily with acids and with the chlorides of the heavy metals, yielding crystallizable salts.

The ptomaines afford no specific reaction whereby they may be readily identified; and their identification is effected only after a painstaking analysis.

We must here call attention, however, to several of their more common properties, beginning with their basic character, their oxidizability by the air and consequently their well-defined reducing power—a property that led Selmi to propose a mixture of ferric chloride and potassium ferricyanide as a reagent for their detection.6 They are precipitated by all the general reagents for the vegetable alkaloids. Selmi has given several reactions, such as those afforded by sulphuric, hydrochloric, and nitric acids, which appear, however, to apply much more to the impurities present than to the bases themselves.

The physiological action of these bases varies greatly; in some the action is an extremely toxic one, as in the case of neurine and muscarine, which are true ptomaines; there are others, such as cadaverine and putrescine, which are quite innocuous. The physiological action of these bases, like that of the true toxins, is studied by making hypodermic injections of solutions of the bases in healthy animals, such as guinea-pigs, rabbits, and dogs.

In animals, the principal phenomena observed by Selmi to follow the injection of the substances are the following: At first dilatation of the pupil, then constriction; tetanic convulsions, soon followed by muscular relaxation, and retardation, rarely acceleration, of heart-beat; absolute loss of cutaneous sensibility; loss of muscular contractility; paralysis of the vasomotors; greatly retarded respiration; stupor, followed by death with the heart in systole.

It must be observed that in a number of cases where toxic researches had been made in the past, these bases had been mistaken for poisons which were believed to have been introduced into the organism with criminal intent. No one will ever know how many have fallen victims in the past to ignorance regarding the cellular mechanism!

The extraction of these bases is a tedious and difficult operation. The materials must first be exhausted with water slightly acidulated; then, after precipitating the albuminoids by boiling and defecating by adding lead acetate, the liquid is evaporated to one-half its volume and dialyzed in a vacuum.7

Phosphomolybdate is then added to the dialyzed liquid, and the precipitate formed, which now contains all the bases, decomposed by boiling with lead acetate. After removing the excess of lead, there is thus obtained a limpid solution of all the alkaloidal bases in the form of acetates. These are separated by alcohol and by means of fractional precipitations with various metallic salts, depending upon the known properties of the bases.

In order to facilitate their study, the ptomaines have been grouped under two distinct classes, the one embracing the cadaveric or putrefactive ptomaines, of undetermined microbial origin, the other containing the ptomaines formed by microbes of known character. Each of these two groups is itself divided into subgroups, as shown in the following table:

GROUP I.

CADAVERIC PTOMAINES OF UNDETERMINED MICROBIAL ORIGIN.

a. Amines. b. Guanidines. c. Oxamines (fatty or aromatic). d. Amido Acids. e. Carbopyridic Acids and analogues. f. Undetermined Ptomaines.

GROUP II.

PTOMAINES OF KNOWN MICROBIAL ORIGIN.

a. Ptomaines extracted from microbial cells.

b. Ptomaines from pathological urines.

We will not here enter upon a detailed study of the bases belonging to each of these groups. This subject is a vast one, requiring for its treatment a volume devoted to it alone. We will here simply touch upon the principal properties of several of the bases of each of the subgroups named.

Bases of Group I.

a. Amines.—Among these we find nearly all the fatty amines, such as the methylamines and the cyclic alkaloids such as pyridine. They are formed particularly by the putrefaction of fish.

Certain of these bases are very toxic, for instance trimethylene diamine, the collidines, and the parvolines.

b. Guanidines.—Among the products of ordinary putrefaction there has been found so far only methylguanidine, C₂H₇N₃. This is a highly toxic base of which 0.2 Gm. is fatal to a guinea-pig.

c. Oxamines.—Under this designation the following bases are comprised: 1. Neurine bases; 2. oxygenized aromatic bases; 3. bases of unknown constitution. Amongst them we find neurine and choline, which are toxic, and betaine, which is innocuous. They are found particularly in putrid fish.

d. Amido Acids.—These ptomaines, which are usually innocuous in small quantities, are particularly the products of the decomposition of albuminoid substances. Among them we find glycocoll, leucine, and tyrosine, as members of this group.

e. Carbopyridic and Carboquinoleic Acids.—So far only one base is known belonging to this group, and that is morrhuic acid, which is found in the decomposed livers of codfish, and which is a powerful appetizer and stimulant in disassimilation.

f. Undetermined Ptomaines.—Under this heading are classed certain undetermined bases, such as those found in normal urines, and in spoiled meats and bread.

Bases of Group II.

a. Ptomaines Isolated from Cultures of Pathogenic Bacteria.—Bacterial cultures contain, besides the true toxins, a certain number of alkaloidal bases which sometimes possess considerable toxicity.

In the cultures of streptococcus pyogenes there are found trimethylamine and xanthic bases; in those of staphylococcus pyogenes aureus are found xanthic bases and creatinine; while pyocyanine and pyoxanthine are found in the cultures of bacillus pyocyaneus, etc.

b. Ptomaines Isolated from Pathological Urines.—Toxic ptomaine bases have been found in the urines of a large number of diseases.8 It is quite probable that these bases are the results of a general pathological condition due to some bacterial disease, the toxic products of which are eliminated by the kidneys.

From the urines of epileptics Griffiths9 isolated a colorless base crystallizing in prisms having the formula C₁₂H₁₅N₅O₇, and which was found to be exceedingly toxic; the same investigator isolated from the urines of eczematous subjects a ptomaine which he named eczemine,10 and which is also highly toxic.

In certain cases of cystinuria there are found in the urine sulphurized ptomaines, and in measles the urine contains an undetermined ptomaine, rubedine, which is very poisonous. Typhotoxine, a very toxic ptomaine, has been isolated from the urine of typhoid patients; erysipeline, a hardly less toxic base, exists in the urine of erysipelatic subjects; while spasmotoxine, tetanotoxine, and tetanine, exceedingly active alkaloids, are found in the urines of tetanus patients.11

As a general rule, all abnormal urines contain toxic bases; the kidneys appear, in fact, to serve as a means of eliminating the toxic products that form in large quantity whenever, and for whatever cause, the organism ceases to functionate normally, whether it be as a whole, or in any one of its parts.12

Leucomaines.13

The leucomaines are basic substances, nearly allied to the ptomaines, but still more closely related to the ureides. They are formed directly or indirectly by the breaking down of protoplasmal albuminoids. The agents that effect the breaking down are the hydrolyzing ferments of the economy. It is well to recall here that these phenomena of hydrolyzation occur within the cell itself and in a practically reducing medium, as we have already stated. The inmost mechanism of these phenomena cannot here be detailed; it will be found described by Armand Gautier in the Chimie Biologique, and in his work Chimie de la Cellule Vivante.14

The extraction of these bases is an extremely delicate operation. It is necessary to operate with a large quantity of substance, say 50 kilos. The substance is finely chopped, then exhausted with twice its weight of water acidulated with acetic acid (0.2 Cc. per liter) and containing a trace of oil of mustard, which is intended to act as an antiseptic. The albuminoids are precipitated by boiling, the solution then filtered, evaporated in a vacuum at 60° C., and the bases extracted with 95-per cent. alcohol.

The alkaloidal bases obtained in this manner are separated by crystallization from alcohol or by various other chemical methods, the description of which we will not enter upon here.

In order to facilitate the study of the leucomaines they are classed under three groups, according to their chemical affinities. These groups are as follows:

1. Xanthic Leucomaines.—The bases of this group appear to have a composition resembling that of uric acid. When hydrolyzed, they yield urea and guanidine. They are weak bases, and exhibit both basic and weakly acid properties. They all possess the common characteristic of being precipitated by copper acetate in acid solution with heat, and by ammoniacal silver nitrate in the cold.

According to Kossel, these bases are derived from the nucleo-albumins which are found in the cell nuclei, and which are, as we know, substances rich in nitrogen and phosphorus.

Among the bases of this group may be mentioned adenine, C₅H₅N₅, which is obtained from infusions of tea.15 This base is non-toxic; it was discovered by Kossel,16 and it crystallizes easily.

Some others of this group are:

Guanine, C₅H₅N₅O, non-toxic, discovered by Unger; pseudo-xanthine, obtained from muscular tissues; sarcine, C₅H₄N₄O, also but slightly toxic, discovered by Scherer; xanthine, C₅H₄N₄O₂, which is found in many urines, and which acts as a stimulant on the cardiac muscles; paraxanthine, C₇H₈N₄O₂, a toxic base found in certain pathological urines; caffeine and theobromine, powerful diuretic bases; and carnine, C₇H₈N₄O₃, from meat, a muscular stimulant like caffeine.

2. Creatinic Leucomaines.—These have for their type guanidine; they differ from the xanthic bases in that they are not precipitated by copper acetate, but frequently are by ammoniacal silver nitrate. They yield double salts with the chlorides of zinc and cadmium. To this group belong glycocyanine, C₃H₇N₃O₂, and glycocyanidine, C₃H₇N₃O, both very toxic; creatine, C₄H₉N₃O₂, only slightly toxic; creatinine, C₄H₇N₃O; lysatine, which very easily decomposes to form urea; lysatinine, xanthocreatine; arginine, a vegetable base, etc.

3. Neurinic Leucomaines.—These have none of the characteristics of the preceding bases; their type is neurine, a highly toxic base found in the brain, nerves, and certain fish ova. These bases are sometimes normally produced by the animal economy, and are also frequently the result of microbic action. They are the result of the simple phenomena of fermentative hydrolyzation of protagons and lecithins. Among these bases are choline, a weak alkaloid, and betaine, which appears to be non-toxic.

The former has the formula C₅H₁₅NO₂; it was discovered by Stocker. Wurtz synthesized it by combining trimethylamine and glycol-monochlorhydrine, and treating the resulting hydrochloride with silver oxide. Betaine, C₅H₁₁NO₂, is found in beets; it was discovered by Scheibler. Neurine is, chemically, trimethylvinylammonium hydrate.

4. Undetermined Leucomaines.—Among these bases several are important in more than one respect. For instance spermine, which is found in the sperm, is a strong base possessing a powerfully dynamic and tonic action on the nerves. It acts as an oxidizer. Spermine was first obtained by Schreiner17 from the sperm of mammifers in which it occurs as a phosphate. It has the formula C₅H₁₄N₂. It was physiologically studied by Poehl, Tarchanoff, Weljaminoff, and Joffroy.18 Plasmaine, a toxic base found in the blood and discovered by R. Wurtz,19 has the formula C₅H₁₅N₅; protamine, from fish milt, was discovered by Micocher.20