STUDIES ON PROTEINOGENOUS AMINES. Work on the proteinogenous amines thus far reported ...

16 Pages·2003·0.99 MB·English

STUDIES ON PROTEINOGENOUS AMINES.
XXI. THE INTESTINAL ABSORPTION AND DETOXICATION OF
HISTAMINE IN THE MAMMALIAN ORGANISM.
BY KARL K. KOESSLER AiVD MILTON T. HANKE.
(From the Otho S. A. Sprague Memorial Institute and the Department of
Pathology, University of Chicago, Chicago.)
(Recei: ed for publication, January 26, 1924.)
Work on the proteinogenous amines thus far reported shows that
the human intestinal tract is normally inhabited by microorgan-
isms which have the faculty of decarboxylating histidine to
histamine and tyrosine to tyramine. We were able to show that
this amine production (decarboxylase activity) is a specific
function of certain species of bacteria and of certain strains within
the species; microorganisms which decarboxylate histidine do not
decarboxylate tyramine and vice versa. The decarboxylation of
the amino acids, histidine and tyrosine, and probably of other
amino acids resulting in amine production, is thus a normal path-
way in the catabolism of amino acids in the intestinal tract which
occurs side by side with deamination. Histamine (and probably
also tyramine) is a normal constituent of the cecal and fecal matter
of man; i,?., of the large intestines.
This last point, the actual demonstration that relatively large
amounts of histamine are present in the intestinal tract of healthy
men, invited further investigation into the fate of this amine in
the organism. For it deserves the meditation of the physiologist.
as well as of the clinician, how it is possible that such quantities
of a highly poisonous substance could be harbored within the
organism without producing symptoms of intoxication. There are
several explanations which present themselves.
1. The histamine is detoxicated within the intestinal lumen.
The contents of the intestine, by means of their bacterial and
cellular enzymes, might produce rapid changes in the structure
and constitution of the amine of such nature that it is deprived of
889
This is an Open Access article under the CC BY license.
890 Studies on Proteinogenous Amines. XXI
its toxicity. On this assumption the histamine found by us in
the intestinal contents would represent only a transitional stage
in protein metabolism and would not be an end-product.
2. Histamine is not absorbed. It might, for example, be so
firmly adsorbed by the solid constituents of the intestinal contents
that it is not free to pass through the intestinal wall.
3. Histamine is absorbed, but during absorption is at once
detoxicated by the animal tissues (e.g. by the liver or by the cells
of the intestinal wall itself).
Certain phases of this problem of the intermediary metabolism,
fate, mode of absorption, and dctoxication of imidazole ethylamine,
have been approached by previous investigators and before we
report our own attempts directed toward solution of these
questions it might be well to review briefly those papers which
have a direct bearing on our subject.
Dale and Laidlaw,l investigating the fate of histamine by
perfusion experiments through the liver, obtained some evidence
of its disappearance, but the limit of the destructive power of the
liver appeared to be reached very quickly. Neither by adding
one large dose (200 mg. or more) nor by successive small addi-
tions could they get evidence of a total destruction amounting to
more than 10 mg.
In a paper on the mode of action of histamine on intravenous
injection Oehme2 showed that many multiples of the acute lethal
dose of a dilute histamine solution might be injected provided
the injection is done slowly. He showed that a very rapid
detoxication must be taking place in the animal organism and
demonstrated further that the difference between peripheral
venous injection and’ that through the portal way disappears
completely on slow injection. All the previously conducted
comparison experiments have, as far as we know, been carried
out in this way; i.e., a peripheral venous injection has been
compared with a portal injection. Dr. H. G. Wells has called
our attention to the fact that these two routes are hardly com-
parable because in one case only one capillary system is traversed
by the poison; namely, the pulmonary; in the other instance,
1 Dale, H. H., and Laidlaw, P. P., J. Physiol., 1919, xli, 318.
* Oehme, C., Arch. exp. Path. u. Pharmakol., 1913, Ixxii, 76.
K. K. Koessler and M. T. Hanke 891
however, two are traversed, the hepatic and pulmonary. It should
be more correct to compare the injection into the femoral artery
with one in the portal system for in this way two capillary syst.ems
have to be traversed in both instances. We have adopted this
mode of procedure in our work.
A very interesting and important piece of work was reported
by Guggenheim and Liiffler in 1916.3 These investigators
showed-extending the previous work of Ewins and Laidlaw on
p-hydroxyphenylethylamine-that most proteinogenous amines,
such as isoamylamine, phenylethylamine, p-hydroxyphenyl-
ethylamine, indolethylamine, and imidazole ethylamine, are de-
toxicated in the animal organism. The detoxication proceeds
through deamination and oxidation by way of the alcohol to
the aldehyde and finally results in the formation of fatty acids
of the same number of C atoms as the amines. Only the catabo-
lism of histamine (p-imidazole ethylamine) to imidazolacetic acid
could not be proved. If 1 gm. of histamine in solution was
perfused for several hours through a rabbit liver, no considerable
destruction of the amine could be demonstrated by biological
methods.
The most recent work on this subject is reported by NIeakins
and Harington.4 These investigators, studying the absorption
of histamine from the intestine in the cat, found that histamine
introduced as hydrochloride, was absorbed from all parts of the
intestinal tract. The rate of absorption was most rapid from the
ileum, somewhat less from the duodenum, and very much less,
but still quite definite, from the cecum and stomach. Regarding
the part which the liver played in the destruction of histamine
after absorption these authors, on the basis of the work with
animals in which Eck fistulas were made, feel that the liver
exercises a protective function, which is ascribed to the cushioning
effect of its extensive capillary network, which prevents a sudden
flood of the substance from reaching the general blood stream
and thus is regarded as more mechanical than chemical.
Our own experiments directed toward the solution of these
problems were of two kinds:
3 Guggenheim, M., and Liieffler, W., Biochem. Z., 1915-16, lxxii, 325.
4 Meakins, J., and Harington, C. B., J. Pharmacol. and Exp. Therap.,
1921, xviii, 455; 1923, xx, 45.
892 Studies on Proteinogenous Amines. XXI
1. Dogs and guinea pigs were fed known amounts of histamine
dichloride. After the animals had been kept under observation
for some time, they were killed. The intestinal contents, the
intestinal tract, and the liver were then examined for histamine.
In this way we were able to ascertain the approximate rate at
which the amine disappeared from the intestinal tract.
2. Large dogs were anesthetized and prepared so that blood
pressure and respiratory tracings could be obtained. Dilute
solutions of histamine dichloride were then injected, at definite
rates, into the saphenous vein, the femoral art,ery, the mesenteric
vein, the splenic vein, and the duodenum. It was possible for
us, in this way, to determine the minimum effective dose of hista-
mine for the venous system, the buffer effect of the capillary net-
work of the dog’s leg, and the effectiveness of the liver as a
detoxicating organ.
I. On the Absorption and Disappearance of Histamine from the
Alimentary Tract.
Mock of Procedure.
A gelatin capsule containing the finely powdered histamine
dichloride was placed in the posterior pharynx of guinea pigs in
such a manner that it passed into the esophagus intact. The
animals were carefully watched to make certain that they had
swallowed the capsule. Histamine has thus to be introduced
directly into the stomach, for comparatively small doses will
cause death when the amine comes in contact with the mucous
membrane of the mouth, through which it is rapidly absorbed.
In another set of experiments a watery solution of histamine
dichloride was introduced directly into the stomach of young dogs
by means of a stomach tube.
The analytical procedure used in determining histamine was
that described in the preceding paper.
Experiment i.-Approximately 80 per cent of the histamine present in the
intestinal contents of a guinea pig can be recovered by our method.
Two large guinea pigs, weighing each approximately 800 gm., which had
been fed exclusively on carrots, were asphyxiated. The entire alimentary
tract was removed and the contents of the stomach and intestines washed
out with 0.85 per cent salt solution.
K. K. Koessler and M. T. Hanke
To the contents of the alimentary tract of one animal were added 100 mg.
of hist.amine dichloride. The contents from the other guinea pig were
treated with 20 mg. of histamine dichloride. The two mixtures were then
separately hydrolyzed and analyzed for histamine.
Histamine added. Histamine recovered. Efficiency of method.
mg. %7. per cent
100 80 80
20 16.8 84.3
The methods seems, therefore, to recover approximately 80 per cent of
the histamine present irrespective of the initial concentration.
Experiment S.-Histamine is not changed by a 24 hour tncubation with
intestinal contents.
100 mg. of histamine dichloride in solution were intimately mixed with
the contents of the stomach and intestines of a guinea pig which weighed
about 800 gm. and the mixture was then incubated for 24 hours at 37°C.
At the end of this time it was hydrolyzed and analyzed for histamine.
Exactly 60 mg. of histamine dichloride were recovered, 80 per cent of the
amount originally added.
\Ve proved, in Experiment I, that our method is 80 per cent efficient when
applied to this kind of material; hence we are justified in concluding that
the constituents of the intestinal contents do not modify or destroy hista-
mine so that the amine is unrecognizable. The first of our speculative
possibilities to account for the disappearance of histamine seems thus to be
eliminated by this experiment.
E.cperiment 3.-100 my. of histamine, when fed to a guinea pig, does not
cause death. The amine disappears from the alimentary tract within $4
hours.
Histamine dichloride, 100 mg. enclosed in a gelatin capsule, was fed to an
800 gm. guinea pig. During t.he 1st hour the animal showed some signs of
discomfort. It seemed depressed, sneezed occasionally, scratched its nose,
and salivated profusely. These symptoms had disappeared completely
at the end of 2 hours. The animal’s normal appetite was reestablished.
The following morning, 24 hours after the histamine had been ingested, the
guinea pig seemed quite normal. It was killed by asphyxiation; the entire
alimentary tract was removed, free from adhering fat and omentum, slit
open, and the contents carefully removed by washing with physiological
salt solution. The aliment.nry tract and the contents were separately
hydrolyzed and analyzed for histamine. The alimentary tract did not con-
tain histamine. The contents contained only 1.6 mg. In some uay, 98.4 mg.
of histamine disappeared from the alimentary tract without causing death and
without eliciting marked symptoms.
Experiment 4.-100 mg. of histamine, when fed to a guinea pig, have dis-
appeared from the alimentary tract to the extent of 61.6 per cent within 2 hours.
894 Studies on Proteinogenous Amines. XXI
Experiment 3 was repeated excepting that the animal was killed 2 hours
after the histamine had been injected and that the liver was removed as
well as the alimentary tract, and analyzed for histamine. This animal
neither vomited nor defecated during this time interval. The stomach
was found to be acutely dilated with fluid. Solid matter was almost absent.
Efficiency Histamine
of method. present.
cent
m7. per WT.
Intestinal contents.. 26.8 50 33.5
Alimentary tract.. . . . . 4.7 lOO(?) 4.7
Liver . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.5 lOO(?) 4.5
Total.................................................... I 42.7
In this case, 66.5 mg. of histamine dichloride disappeared from the
intestinal contents; 4.7 mg. of this were surely in the tissue of the alimentary
tract and 4.5 mg. had been transported to the liver. The remaining 57.3
mg. were not accounted for. If we assume that the histamine was ab-
sorbed, as such, to pass into the portal blood stream or into the lymphatic
circulation and thence to the systemic circulation, the body of this guinea
pig was being injected with histamine at the rate of 0.5 mg: per minute. It
is well known that one intravenous injection of 0.5 mg. of histamine dichlo-
ride will kill a guinea pig of this size. We are, therefore, forced to conclude
that the histamine, as such, could not have been thrown into the general
circulation at any such rate of speed.
The experiment indicates that the histamine passed into the
intestinal wall as histamine, for we found some of it there. It, is
also ,clear that some of the histamine reached the liver as histamine,
because we found it in this organ. This experiment tells us
nothing, however, about what happens to the major part of the
amine after it leaves the lumen of the intestine. Two possi-
bilities present themselves :
1. The amine may be absorbed and pass into the liver as such.
In this case the liver must possess a highly developed faculty for
depriving histamine of its toxicity, because, if such large doses
of amine were to pass into the systemic circulation, death would
be the inevitable outcome.
2. Histamine may be detoxicated in its passage through the
intestinal wall. In t.his case we should be ascribing to the thin
intestinal wall a function that seems almost prodigious and we
recall hesitatingly how other functions that have been ascribed
K. K. Koessler and M. T. Hanke
to this membrane from time to time have all proved untenable
(e.g. the synthesis of proteins).
But in this instance it is possible that we may have to deal with
the phylogenetic development of a protective mechanism, for
histamine exerts a very stimulating effect upon the musculature
of the intestines eliciting violent contractions. The ability of
the intestinal mucosa to detoxicate amines might have had to
be acquired in the phylogenesis of man.
Before we turn to a consideration of the above problems, we
shall describe a typical example of the conditions prevailing when
histamine is fed to a dog, from which it becomes clear that the
results obtained with the guinea pig are more general in their
application.
Experiment 5.--500 mg. of histamine when fed to a 5 lcilo dog have disap-
peared from the alimentary tract to the extent of 53 per cent within 2 hours.
After convincing ourselves that puppies show no signs of discomfort after
the ingestion of 500 mg. of histamine dikhloride, we introduced 500 mg.,
dissolved in 200 cc. of water, by stomach tube into a puppy weighing 5.2
kilos. There was no reaction of any kind. The animal did not vomit.
The dog was killed with illuminating gas 2 hours after the amine
was ingested.
The entire alimentary tract was removed and freed from fat and omen-
turn. A ligature was placed tightly at the cardiac and pyloric ends of the
stomach to prevent loss of material. The intestine was cut off just below
the stomach. The material was, in this way, divided into two fractions;
namely, the stomach and the intestines. Each of these was separately
hydrolyzed and analyzed for histamine.
The stomach and contents weighed 500 gm. This organ was acutely
dilated with fluid. Solid matter was practically absent. The contents
were not separated from the stomach wall; the analysis was conducted on
the entire material.
The stomach and contents contained i.JY mg. of histamine dichloride.
Since the material was simple in composition, mostly fluid, this figure is
probably not subject to corrections. This is equal to 29.4 per cent of the
amount ingested.
The alimentary tract, below and exclusive of the stomach, weighed 520
gm. This was hydrolyzed and analyzed for histamine. The contents were
not separated fr.om the tract.
The intestines contained 53.2 mg. of histamine dichloride which is 10.64
per cent of the amount originally ingested.
Since this mat.erial was something like human feces in consistency, for
which the efficiency of our method has proved to be 60 per cent, the maxi-
mum amount of histamine that could have been present in the intestine is
88.65 mg., which is 17.73 per cent of the amount originally ingested.
896 Studies on Proteinogenous Amines. XXI
The total recovery was 235.6 mg.; hence 264.4 mg. of histamine dichloride
were absorbed in 2 hours. The average absorption rate was, therefore, 2.2
mg. per minute; but we know that, if histamine were injected into the
systemic circulation at anything approaching this rate of speed, severe
symptoms of intoxication would undoubtedly have developed.
The feeding experiments lead always to the same conclusion.
It is unreasonable to assume that the histamine absorbed passes
into the general circulation as such. The compound must be
removed from the general circulation in some way. We have
already indicated, under Experiment 4, that there are only two
possible explanations. The amine must be detoxicated either in
its passage through the intestinal wall or subsequently by the
liver. The other tissues of the body unquestionably have some
power of detoxicating this amine; but this in itself would not be
sufficient to account for our results.
So much .speculation has surrounded the liver as a detoxicating
organ that, in spite of the .convincing experiments of Meakins
and Harington4 we decided to investigate, in a different way, the
role of the liver as a “detoxicator” of histamine.
II. On Certain Factors Involved in the Extent of Pharmacodynamic
Action of Histamine Following Its Injection into the
Circulation.5
The toxic action of histamine on intravenous injection in the
common laboratory animals is well known since the extensive
studies of Dale and his coworkers. While the pharmacological
effect of the poison is very similar in its symptoms in the different
species, the minimal lethal dose of histamine on intravenous
injection varies considerably in different animals. For guinea
pigs and rabbits 0.6 to 0.8 mg. of histamine per kilo constitutes
the fatal dose.
Working with dogs we found that 30 mg. of histamine per kilo
body weight is insufficient, in unanesthetized animals, to produce
death.‘j
We were, however, more interested to establish the minimal
effective dose of histamine in the dog; i.e., the smallest quantity
5 In the experimental part of this work we were ably assisted by Dr. H. L.
Huber.
6 We were assisted, in these experiments, by Dr. Julian H. Lewis.
K. K. Koessler and 31. T. Hanke 897
of the amine which on intravenous injection would still produce
a definite fall in blood prcssurc.
Large dogs were used in this work (10 to 20 kilos). In our first
experiments ether preceded by an injection of morphine was
used as anest.hetic. We found, however, that the respiration and
blood pressure would remain at a more constant, normal, level
when barbital7 was used as anesthetic. Even with this anesthetic
it is advisable to use a small amount of ether while the intestines
are manipulated.
The dog, after anesthetization, was prepared so that blood
pressure and respiratory tracings could be made simultaneously
on the drum of the same kymograph. The injections were made
with a Woodyatt pump.* The apparatus had to be arranged so
that we could switch from salt to histamine solution, or vice versa,
within a few seconds. To accomplish t,his two pumps were used,
one for histamine and one for salt solution. The solutions were
fed to the machines from burettes so that the rate of injection
could bc easily followed. All solutions were warmed to 40°C.
before they passed into the animal’s circulation. The feed pipes,
from the pump to the animal, were composed of glass. Rubber
connections, where such were necessary, were as short as possible.
Many animals were used in these experiments, but we will
give here only a few graphs which illustrate sufficiently the points
we wish to make.
Fig. 1 shows the effect of injecting histamine into the saphenous
vein. The minimum effective rate was 0.0027 mg. per minute
per kilo body weight. When the inject,ion was made at the rate
of 0.0054 mg. per minute per kilo body weight, there was a pro-
nounced fall in blood pressure, but no respiratory embarrassment.
When the injection rate was increased to 0.0108 mg. per minute,
the animal reacted with a marked fall in blood pressure and
considerable respiratory embarrassment. These experiments were
repeated a large number of times on dogs weighing from I2 to 15
kilos. The results were always the same.
The two graphs listed as Figs. 2 and 3 show the effect of in-
jecting histamine into the saphenous and large mesenteric veins
7 Tstum. A. L., and Parsons, E., J. Lab. and Clin. Med., 1922-23, viii, 64.
8 \vqodyatt, R. T., J. Biol. Chew, 1917, xxix, 35.5.
898