artificial proteolysis than those connected with the possible
absorption of the more diffusible products of digestion. Thus, in the
living stomach there is an ever-increasing secretion of hydrochloric
acid, and perhaps also of pepsin, more or less proportional to the
extent of proteolysis. On this point Brandl’s experiments again give
us some light. Thus, it was found that the introduction of an aqueous
solution of peptone into the empty stomach led to the secretion of an
acid fluid containing on an average 0.24 per cent. HCl, while, under
similar conditions, the introduction of sugar or potassium iodide
was followed by the secretion of a fluid containing on an average
only 0.13 per cent. HCl. Further, the absolute amount of acid found
after the introduction of peptone was far greater than when sugar or
iodide was introduced, since peptone led to an increase of at least
fifty per cent. in the volume of fluid secreted. Hence, proteolysis in
the living stomach may give rise to such an increased production and
secretion of hydrochloric acid that formation of the terminal products
of gastric digestion may be greatly accelerated. That such in fact is
the case, I have no manner of doubt, but that it may result in the
complete conversion of the so-called primary and secondary proteoses
into peptone I very much question. In fact, such examinations as I have
made of the stomach-contents after a suitable test-meal have always
resulted in the finding of a relatively large amount of proteoses. To
be sure, true peptone may be detected and in fairly large amounts,
but whenever a quantitative determination of the relative proportion
of the two has been made, the proteoses have always been in excess. I
have already reported elsewhere the results of some experiments in this
direction made on a healthy young man, where the stomach-contents were
withdrawn at varying periods after the ingestion of weighed amounts
of coagulated egg-albumin. Thus, in one experiment[146] the stomach
was thoroughly rinsed with water, after which 138 grammes of finely
divided coagulated-albumin, equal to 16 grammes of dry albumin, were
ingested. Three-quarters of an hour thereafter, the stomach-contents
were withdrawn by lavage and analyzed. As a result, 1.41 grammes of
albumoses were separated and weighed, and 0.84 gramme of peptones,
the relative proportion being expressed by sixty-two per cent. of
albumoses and thirty-seven per cent. of peptones, calculated on the
2.25 grammes of soluble products recovered. This expresses the general
character of the results obtained in experiments of this nature, and in
my opinion adds emphasis to the statement already made, that complete
peptonization is not a feature of pepsin-proteolysis, either in the
artificial or in the natural process as it takes place in the living
stomach.
[146] Journal of Physiology, vol. xiv., p. 501.
Gastric digestion is to be considered rather as a preliminary step in
proteolysis, preparatory to the more profound changes characteristic
of pancreatic digestion, in which the ferment trypsin is the important
factor. We can thus see how, as in the case of Czerny’s dogs, an
animal may be perfectly nourished without a stomach, digestive
proteolysis being carried on solely by the pancreatic fluid. You will
remember that two of the dogs operated on by Czerny and his pupils
lived between four and five years after the operation, with the stomach
completely removed, and yet during this period they were well nourished
and ate all varieties of food with apparently a normal appetite.[147]
Evidently, then, in some cases at least, digestive proteolysis
can be carried on without this preliminary action of the gastric
juice. Ogata[148] arrived at essentially the same conclusion by the
establishment of a duodenal fistula, shutting off the stomach from the
intestine by means of a small rubber ball which could be inflated with
water. On then introducing coagulated egg-albumin and other forms of
proteid matter into the duodenum, he found that digestion was at least
sufficiently complete to satisfy all the demands of the system. The
only unsatisfactory result was with collagenous foods, which plainly
showed the need of a preliminary acid digestion. More recently still,
Cawallo and Pachon,[149] working in Richet’s laboratory, have studied
the digestibility of different kinds of proteid foods in a dog, upon
which they had performed a gastrectomy; the entire fundus, as well as
the pyloric portion, of the stomach having been removed. In an animal
so operated upon, after recovery was complete, solid food, as meat,
was completely digested when taken in small quantities at a time. Raw
meat, however, was less completely utilized, the fæces showing portions
of undigested fibres. Still, it was apparent that intestinal digestion
alone was capable of accomplishing all that was necessary for the
complete nourishment of the animal, when it had once become accustomed
to the changed condition of its alimentary tract.
[147] Bunge’s Physiologische und Pathologische Chemie, p. 152.
[148] Ueber die Verdauung nach der Ausschaltung des Magens. Du
Bois-Reymond’s Archiv f. Physiol, 1883, p. 89.
[149] Une observation de chien sans estomac. Comptes Rendus hebd. de la
Société de Biologie, December 1, 1893.
These facts are cited not to belittle the importance of gastric
digestion in the nutrition of the body, but rather to emphasize the
probability that pepsin-proteolysis is simply a preliminary step in
digestion; that its function is not in the direction of a complete
peptonization of the proteid foods ingested, but that its action is
especially directed to the production of soluble products, proteoses,
which can be further digested in the small intestine, or perhaps
directly absorbed after they have passed through the pylorus, or even
from the stomach itself to a certain extent.
SOME PHYSIOLOGICAL PROPERTIES OF PROTEOSES AND PEPTONES.
It is very evident from what has been said that all forms of proteid
matter, _i.e._, all the members of the three main groups spoken
of in our classification of the proteids, excepting only nuclein,
reticulin, and the keratins, are capable of undergoing proteolysis with
pepsin-hydrochloric acid. Further, in every case the main products
of the transformation are proteoses; viz., albumoses, caseoses,
gelatoses, vitelloses, myosinoses, etc., according to the nature of
the proteid undergoing proteolysis; true peptones being formed in
less abundance. Corresponding to each of these groups are primary and
secondary proteoses, all possessed of many points in common, both
chemical and physiological, yet differing from each other in many minor
respects. These are the important products of gastric digestion, of
pepsin-proteolysis, and it may be well to consider for a moment some
of the physiological properties of the proteoses and of peptones as
well, in order that we may the better comprehend the general nature of
these substances with reference to their possible action in the economy.
As far back as 1880, Schmidt-Mülheim[150] discovered that the injection
of aqueous solutions of peptone into the blood-vessels of living dogs
was attended by a series of remarkable phenomena. Thus, the animal
passed at once into a condition of narcosis resembling that produced by
chloroform, accompanied by a fall of general blood-pressure so great
that the animal was liable to die, as from asphyxia. Further, there
was evidence of some marked change in the condition of the blood, as
indicated by loss of the power of spontaneous coagulation, while the
peptone itself evidently underwent some alteration, or else was rapidly
eliminated, since it could not be detected in the blood a short time
after its introduction. These experiments, however, were not conducted
with true peptone but with Witte’s “peptonum siccum,” which at that
time, at least, was composed in great part of proteoses. The general
character of these interesting results was confirmed by Fano,[151] who
found that the injection of so-called peptone in the proportion of 0.3
gramme per kilo. of body-weight was sufficient to bring about complete
narcosis, together with loss of coagulability on the part of the blood.
Very suggestive, however, was the fact that Fano, on trying similar
experiments with the peptone formed by pancreatic digestion, viz.,
with antipeptone, which presumably contained a far smaller proportion
of proteoses, failed to obtain like results; the tryptone, so-called,
being exceedingly irregular in its action, in many cases producing no
effect whatever.
[150] Beiträge zur Kenntniss des Peptons und seiner physiologischen
Bedeutung. Du Bois-Reymond’s Archiv f. Physiol., 1880, p. 33.
[151] Das Verhalten des Peptons und Tryptons gegen Blut und Lymphe.
Ibid., 1881, p. 277.
The discovery at this date of the several albumoses, and their presence
in large amounts in all so-called peptones, led to a study of their
physiological action with special reference to the observations
of Schmidt-Mülheim and Fano. Politzer,[152] working under Kühne’s
guidance, was the first to experiment in this direction, and his
results are full of interest as throwing light on the action of the
individual albumoses. Thus proto, hetero, and deuteroalbumose are all
active physiologically, giving rise when injected into the veins of
dogs and cats to strong narcotic action, varying somewhat in intensity
in different individuals. There is also produced a marked fall in
blood-pressure, due apparently to vaso-motor paralysis, the action
being manifested chiefly, if not wholly, on the splanchnic region.
Thus, after an injection of one of these albumoses, the mesenteric
vessels are always strongly congested, accompanied frequently by the
appearance of a bloody serum in the peritoneal cavity. Narcotic action
is manifested only so long as the blood-pressure remains sub-normal,
and is due presumably to this marked accumulation of blood in the
large abdominal veins, thus leading to anæmia of the brain. Albumoses
and peptones injected into the jugular vein likewise produce fever,
presumably through some action on the nervous system by which the
equilibrium of tissue-metamorphosis is interfered with.[153]
[152] On the Physiological Action of Peptones and Albumoses. Journal of
Physiol., vol. 7, p. 283.
[153] Ott and Collmar: Pyrexial Agents, Albumose, Peptone, and Neurin.
Journal of Physiol., vol. 8, p. 218.
Further, Politzer found that all of the albumoses either delayed or
prevented altogether the coagulation of the blood, in conformity with
the observations of Schmidt-Mülheim and Fano. In all of these actions
the primary albumoses appeared most effective, deuteroalbumose least
so. Heteroalbumose, however, was constantly most active, especially in
delaying the coagulation of the blood. With amphopeptone, there was
far less narcosis and less diminution of blood-pressure, while the
effect on the coagulability of the blood was more or less variable,
frequently being entirely negative. Antipeptone, on the other hand, was
found almost wholly wanting in any constant effects, although in one
instance deep narcosis was produced. Thus, from Politzer’s experiments,
it was made clear that the albumoses, when introduced directly into
the blood-current, possess a far greater toxic action than either
amphopeptone or antipeptone. Albumoses, in sufficiently large doses,
were invariably fatal, while peptones never produced fatal results
so long as the kidneys of the animal remained intact. The extreme
solubility and diffusibility of peptones, coupled perhaps with their
marked diuretic action, lead to rapid elimination through the kidneys,
and their consequent removal from the system.
Many of these observations made with the albumoses I have repeated
with several of the proteoses and peptones more recently studied, as
protocaseose, protoelastose, the globuloses, and others. The results
may be taken as practically confirmatory of the older observations,
and I make mention of them in this general way simply to emphasize
the fact that all of the proteoses, though perhaps showing individual
peculiarities, are possessed of marked physiological properties, which
plainly testify to their toxic nature, when introduced directly into
the blood-current.
Young animals are particularly sensitive to the injection of
proteoses into the blood, even when the introduction takes place very
gradually.[154] Thus, a young, healthy dog of 2 kilos. body-weight,
eight weeks old, died in one hour after the injection into the jugular
vein of 1 gramme of protoalbumose in 20 c.c. of water, thus affording
a good illustration of the extreme toxicity of this albumose when
introduced directly into the blood.
[154] Neumeister: Ueber die Einführung der Albumosen und Peptone in den
Organismus. Zeitschr. f. Biol., Band 24, p. 284.
Of greater interest, physiologically, are the changes the individual
proteoses undergo after their injection into the blood. As already
stated, peptone so injected may appear in the urine wholly unaltered.
Thus, Neumeister[155] has made injections of both amphopeptone
and antipeptone in the case of dogs, and was able to detect the
peptone very quickly in the urine. I have made like experiments with
other forms of peptone and obtained similar results; thus, a pure
amphopeptone formed from casein by pepsin-proteolysis (2 grammes in
15 c.c. water) was injected into the jugular vein of a dog weighing 5
kilos. The urine collected during several hours after the injection
was heated to boiling, and saturated while hot with ammonium sulphate.
The filtrate, on being tested with cupric sulphate and potassium
hydroxide, gave a fairly strong biuret reaction for peptone. Another
similar experiment made with antipeptone, formed from the myosin of
muscle-tissue, gave like results.
[155] Zeitschr. f. Biol., Band 24, p. 287.
With proteoses, however, different results are obtained, as
Neumeister[156] first pointed out. These bodies introduced into the
blood undergo more or less of a change prior to their excretion in
the urine, the change partaking of the character of a hydrolytic
cleavage in which the primary proteoses are transformed into secondary
proteoses, while deuteroproteoses are changed into peptones. This
is not necessarily to be interpreted as meaning that the full
equivalent of the proteose injected appears in the urine, but that the
portion which is eliminated through the kidneys tends to undergo a
transformation somewhere _en route_, akin to the change produced in
pepsin-proteolysis. As to how common or complete this transformation
is under the above circumstances, we have no positive knowledge. Such
a hydrolytic change certainly occurs in the case of the dog, and the
experimental evidence is in favor of the view that the transformation
is effected in the kidneys by the pepsin secreted through the urinary
tubules, where there is momentarily a formation of free acid. In the
rabbit, on the other hand, no such change occurs; the urine from this
animal contains practically no pepsin, and consequently the proteoses
eliminated through the kidneys are excreted unaltered. As, however, the
experiments of Stadelmann[157] and others have shown that the urine of
all carnivora, and of man as well, contains a ferment which, on the
addition of a suitable amount of hydrochloric acid, will digest fibrin
with formation of the ordinary products of pepsin-proteolysis, it is to
be presumed that all proteoses passing through the kidneys will undergo
at least some change prior to their excretion in the urine.
[156] _Ibid._, p. 284.
[157] Untersuchungen über den Pepsin-fermentgehalt des normalen und
pathologischen Harnes. Zeitschr. f. Biol., Band 25, p. 208.
However this may be, it is very evident that the proteoses formed
in gastric digestion cannot be absorbed as such directly into the
blood-current. Introduced into the blood, they behave in such a manner
as to warrant the conclusion that they are truly foreign substances,
and the system makes a brave endeavor to remove them as speedily as
possible. The same may be said of amphopeptones, from which we may
conclude that all of these products of pepsin-proteolysis undergo
some transformation during the process of absorption, by which their
toxicity is destroyed and their nutritive qualities rendered fully
available for the needs of the body. Discussion of this question,
however, will be left until the next lecture.
In view of these pronounced physiological properties of the proteoses,
it is interesting to recall the now well-known fact that many of
the chemical poisons produced by bacteria are proteose-like bodies,
chemically, at least, closely akin to the proteoses resulting from
pepsin-proteolysis. Thus, Wooldridge[158] as early as 1888 pointed out
that an alkaline solution of tissue-fibrinogen exposed to the action
of anthrax-bacilli suffered some change, so that when introduced into
the blood it possessed the power of producing immunity to anthrax.
This observation was verified by Hankin,[159] who further showed that
the substance formed by the anthrax-bacilli was a veritable albumose,
and that it truly possessed the power of producing immunity. Sidney
Martin[160] carried the matter still further, and by growing the
anthrax-bacilli in a pure solution of alkali-albuminate prepared
from blood-serum, proved the formation of both primary and secondary
albumoses, as well as of peptone, leucin, tyrosin, and a peculiar
alkaloidal substance of pronounced toxic properties. Martin finds that
the albumoses are not as poisonous as the alkaloid, and surmises that
the alkaloid is contained in the albumose molecule in the nascent
state; further, he suggests that the albumoses in small doses may exert
some protective influence, while in larger doses they act as vigorous
poisons. How true this may be I cannot say, but my own experience
convinces me that the anthrax-bacilli grown in a culture medium
composed of alkali-albuminate, prepared from egg-albumin, to which
the necessary inorganic salts and some glycerin have been added, do
give rise to albumoses and peptones which are truly endowed with toxic
properties.
[158] Versuche über Schutzimpfung auf chemischem Wege. Du
Bois-Reymond’s Archiv f. Physiol., 1888, p. 527.
[159] British Med. Journal, October, 1889.
[160] Proceed. Royal Society, 1890, vol. 48, p. 78.
Albumose-like bodies have also been obtained by Brieger and
Fränkel[161] with the bacillus of diphtheria. These, too, were endowed
with powerful poisonous properties, and when introduced into the
tissues of the body gave rise to reactions resembling those produced
by the Löffler bacillus. In my own laboratory, recent experiments
made with the bacillus of glanders have shown that when grown in a
slightly acid medium containing alkali-albuminate, albumoses, peptones,
and crystalline bodies such as leucin and tyrosin are formed in
considerable quantities. Kresling[162] has reported similar results.
With the tubercle-bacilli, many like results have been recorded. Thus,
among others, Crookshank and Herroun[163] have reported the finding of
albumoses, peptone, and a ptomaine when the bacilli have been grown in
glycerin agar-agar, and also in fluid media.
[161] Untersuchungen über Bacteriengifte. Berlin, klin. Wochenschrift,
1890, p. 241 and 268.
[162] Ueber die Bereitung des Malleins und seine Bestandtheile.
Abstract in Jahresbericht f. Thierchemie, Band 22, p. 634.
[163] Journal of Physiology, vol. 12, p. 9.
Koch[164] has made a special study of the albumose which he considers
as the specific toxic agent of the so-called tuberculin. This albumose
was found by Brieger and Proskauer[165] to have a somewhat peculiar
composition, inasmuch as it contains forty-seven to forty-eight per
cent. of carbon and only 14.73 per cent. of nitrogen, agreeing,
however, in this respect very closely with the peptone formed from
egg-albumin by the action of bromelin.[166] Still more recently,
Kühne[167] has made a thorough study of this albumose, as well as of
the other products elaborated by the growth of the tubercle-bacillus.
He designates all of the peculiar albumoses formed by these bacilli
as _acrooalbumoses_. They are endowed with marked chemical and
physiological properties, causing a rise of temperature when injected
into the blood, as well as other phenomena more or less pronounced.
It is thus evident there is ample ground for the statement that all
nutritive media in which pathogenic bacteria have been planted are
liable to contain, sooner or later, toxic substances, many of which at
least are closely related to, if not identical with, the albumoses. It
is not my purpose, however, to consider these points in detail, nor to
quote the many results obtained by other workers in this direction.
[164] Deutsche med. Wochenschrift, 1891, p. 1180.
[165] _Ibid._
[166] Chittenden: On the Proteolytic Action of Bromelin, the Ferment of
Pineapple Juice. Journal of Physiology, vol. 13, p. 303.
[167] Weitere Untersuchungen über die Proteine des Tuberculins.
Zeitschr. f. Biol., Band 30, p. 221.
I wish merely to call attention to the fact that the proteoses, and
likewise the peptones formed by pepsin-proteolysis, are more or less
toxic when introduced directly into the blood, and that they share this
property with the proteoses formed by bacterial organisms, or by the
enzymes which they give rise to. In other words, these primary cleavage
or alteration products of the proteid molecule, however produced,
are more or less poisonous, and if introduced into the blood-current
without undergoing previous change may show marked physiological
action. It is, of course, not to be understood that these bodies are
all alike. They are surely closely related and possess many points in
common, especially so far as their chemical properties are concerned,
but their chemical constitution and their physiological action must
vary more or less with their mode of origin.
In any event, it is very evident that the proteoses and peptones
formed in the alimentary tract by pepsin-proteolysis must undergo some
transformation, before reaching the blood-current, by which their
peculiar physiological properties are modified. This modification may
be associated with a conversion into the serum-albumin, or globulin of
the blood. However this may be, the fact remains that these proteoses
formed so abundantly during digestion can be absorbed and serve as
nutriment for the animal body, but between their formation as a result
of proteolysis and their passage into the blood they are exposed to
some agency, or agencies, doubtless in the very act of absorption, by
which a further transformation is accomplished. With this point we
shall be able to deal more in detail in the next lecture.
LECTURE III.
PROTEOLYSIS BY TRYPSIN--ABSORPTION OF THE MAIN PRODUCTS OF PROTEOLYSIS.
PROTEOLYSIS BY TRYPSIN.
In pancreatic digestion, proteids are exposed to the action of an
enzyme of much greater power than pepsin, one endowed with a far
greater range of activity, and consequently proteolysis as it occurs
in the small intestine becomes a broader and more complicated process.
As you well know, the ferment trypsin manifests its power not only in
a more rapid transformation of insoluble proteids into soluble and
diffusible products, but there is a diversity in the character of the
many products formed which testifies to the profound alterations this
ferment is capable of producing. The primary and secondary products of
pepsin-proteolysis, as well as unaltered proteids, are alike subject
to these changes, and bodies of the simplest constitution may result
in both cases from the series of hydrolytic changes set in motion by
this proteolytic enzyme. The power of the ferment as a contact agent is
astonishing, for in the case of trypsin no accessory body is necessary
to bring out its latent power. Water, proteid, and the enzyme at the
body-temperature are all that is necessary to call forth prompt and
energetic hydrolytic action.
Moreover, hydrolysis does not stop with the mere production of soluble
proteoses and peptones, but the hemi-portion of the latter is quickly
broken down into crystalline bodies, such as leucin, tyrosin, lysin,
lysatin, etc. This special characteristic of the ferment testifies
in no uncertain manner to the existence of inherent qualities in
the inner structure of the enzyme peculiar to the body itself. In
general properties and reactions, pepsin and trypsin may be closely
related; both are products of the katabolic action of specific
protoplasmic cells, but the inner nature or structure of the two must
be quite different. Pepsin, as we have seen, is powerless to produce
any change in proteid bodies unless acids are present to lend their
aid. Furthermore, pepsin is limited in its action to the production
of proteoses and peptones, while trypsin gives rise to a series
of hydrolytic cleavages which result in the ultimate formation of
comparatively simple bodies.
Trypsin, however, in its natural environment is dissolved in an
alkaline medium. Its proteolytic action is therefore carried on, under
normal circumstances, in an alkaline-reacting fluid containing 0.5
to 1 per cent. sodium carbonate, and the proteolytic power of the
ferment is unquestionably manifested to the best advantage in such a
medium. At the same time, it will act, and act vigorously, in a neutral
fluid, and likewise in a fluid having a weak acid reaction, provided
there is little or no free acid present. Thus, in experiments[168] on
blood-fibrin it was found that, while a solution of trypsin containing
0.5 per cent. sodium carbonate, digested or dissolved 89 per cent.
of the proteid in three to four hours at 40° C., a perfectly neutral
solution of the ferment, otherwise under exactly the same conditions,
digested 76 per cent., and a 0.1 per cent. salicylic acid-solution of
the enzyme converted 43 per cent. of the proteid into soluble products.
[168] Chittenden and Cummins: Studies in Physiol. Chem., Yale
University, vol. i., p. 135.