FORMATION AND DESTRUCTION OF URIC ACID
The chemical structure and sources of uric acid having been dealt with,
we are now in a position to resume our narrative, and to take up the
thread at the point when Horbaczewski revealed the derivation of uric
acid from nucleic acid. It now devolves upon us to scrutinise more
narrowly the process by which the formation of _uric acid_ from _nucleic
acid_ is achieved. Incidentally, it will not be unprofitable to note, if
only briefly, the steps by which the necessary expansion of our chemical
and physiological knowledge of nucleic acids has been acquired.
As may be imagined, the primary difficulty was to prepare nucleic acids
of such purity as admitted of their elementary chemical analysis. The
necessary researches were to a large extent confined to two types of
nucleic acid, one derived from _yeast_, and the other from the _thymus
gland_; in other words, to representatives of the only two nucleic acids
in nature, one derived from the nuclei of _animal_ cells, the other from
the nuclei of _vegetable_ cells.
A feature common to nucleic acids of animal and vegetable origin is
that, on hydrolysis with boiling mineral acid, they yield two purin
derivatives, _guanine_ and _adenine_, and a pyrimidin derivative,
_cytosine_. From thence as regards their remaining constituent elements
they display distinctions. Thus animal nucleic acids yield _thymine_, and
contain a _hexose_ group in their molecule. On the other hand, vegetable
nucleic acids give forth _uracil_ and possess a _hexose_ group.
To sum up, _nucleic_ acid is a chemical complex, made up of phosphoric
acid with purin bases, pyrimidin bases and carbohydrate radicles.
Moreover, nucleic acids, whatever their source, show a striking
similarity in structure, containing always two amino-purins (adenine
and guanine), two pyrimidines (either cytosine and uracil, or cytosine
and thymine), and a carbohydrate. Now, while _purin bases_ are always
present, yet, in respect of their _carbohydrate_ group, nucleic acids
display variations; this, according as they are of animal or vegetable
origin. If the former, the carbohydrate group is a _hexose_ (contains six
carbon atoms) with _thymine_. If the latter, it contains _pentose_ (five
carbon atoms) with _uracil_.
The constancy in the content of the various nucleic acids is such that
Levene and Jacobs have felt justified in putting forward the following
provisional formula as to the constitution of a _nucleic acid_ of animal
origin.
[Illustration: STRUCTURAL FORMULA OF NUCLEIC ACID]
DISTRIBUTION OF THE ENZYMES
The enzymes responsible for the disruption of the _nucleic acid_ complex
are not to be found in all the body tissues. Moreover, the distribution
of the enzymes in the various organs and tissues varies in different
species of animals. Of the various organs the _liver_, _spleen_,
_thymus_, and _pancreas_ more particularly contain enzymes in abundance.
As to their varied location in different animals, it may be noted that
the enzyme responsible for the oxidation of _xanthine_ into _uric acid_,
viz., _xanthine-oxidase_, is found in man only in the liver. In other
animals, also, it is of localised distribution, being as a rule only
found in the liver or in the liver and kidney. The dog, however, appears
to be an exception, _xanthine-oxidase_ being found in a variety of its
tissues.
_Adenase_, the deaminising enzyme, is not to be found in any organs
in man. Neither does it exist in any of the tissues of the rat.
Consequently, if _adenine_ be injected subcutaneously in rats, it
undergoes oxidation, without abstraction of its amino group.
On the other hand, _guanase_, also a deaminising enzyme, is in man to be
detected in the _kidney_, _lung_, and _liver_, but not in the pancreas or
spleen. In the pig, however, guanase is lacking, and its absence no doubt
explains why deposits of guanine may occur in the muscles constituting
the so-called _guanine gout_ met with in swine. It is worthy of note also
that in pigs’ urine the content of purin bases exceeds that of uric acid.
To sum up, in man the enzyme, _xanthine-oxidase_, which forms uric acid
from xanthine, is located chiefly or exclusively in the _liver_. This,
of course, represents the _final_ stage of purin metabolism, but the
antecedent chemical processes involved in the disruption of nucleic acids
are initiated by the action of enzymes in the _intestinal juices_ and
_wall_, and to a consideration _seriatim_ of these changes we now proceed.
STAGES IN DISRUPTION OF NUCLEIC ACID
As might be expected from the complex structure of the nucleic acid
molecule, a number of ferments are concerned in its disruption. The
gastric and pancreatic juices contain not a trace of any enzymes. Thus,
when _nucleo-protein_ is subjected to the gastric juice a moiety of
protein is readily split off and hydrolysed to peptone and other products
of proteolysis.
But the nuclein element remains unacted upon until it comes under the
action of the pancreatic juice. Hydrolysis then ensues, and the ingested
nuclein is broken down into nucleic acid and protein. The _nucleic acid_
remains unaffected by the pancreatic juice, but, coming in contact with
the _succus entericus_, it undergoes partial decomposition through the
action of a ferment called nuclease or _nucleic_-acidase. Under its
disruptive effect the nucleic acids or _poly-nucleotides_ are further
split up into groups known as _nucleotides_. The two _pyrimidine_
nucleotides split off and undergo no further change. But, through the
action of another ferment, _nucleotidase_, the _purin_ nucleotides are
further decomposed to yield _nucleosides_ (substances of the glucoside
class made up of a combination of a purin base with a carbohydrate group
of the nucleic acid with which also phosphoric acid is linked).
No further stage in hydrolysis of nucleic acid occurs in the intestine,
but the _nucleosides_ are again in turn split up after reaching the
tissues, particularly in the _spleen_, _liver_, and _thymus_. This,
under the action of specific enzymes, _nucleosidases_, which succeed in
breaking the nucleosides down into the so-called “building stones” of
the _nucleic acid molecule_, phosphoric acid group, carbohydrate group,
pyrimidine and purin bases, especially adenine and guanine. The adenine
and guanine thus formed are, by the action of the ferments _adenase_
and _guanase_, converted and, by the removal of their amino group,
transformed, adenine into hypoxanthine, and guanine into xanthine, thus:—
C₅H₅N₅ + H₂O = C₅H₄N₄O + NH₃; C₅H₅N₅O + H₂O = C₅H₄N₄O₂ + NH₃
Adenine. Hypoxanthine. Guanine. Xanthine.
By the action of oxidases also present in the tissues hypoxanthine is
changed into xanthine and xanthine into uric acid (trioxy-purine), this
by a specific ferment xanthine oxidase.
C₅H₄N₄O O—C₅H₅N₄O₂; C₅H₅N₄O₂ O C₅H₄N₄O₃
Hypoxanthine. Xanthine. Xanthine. Uric acid.
[Illustration: SCHEME ILLUSTRATING THE PROBABLE STAGES IN THE PASSAGE OF
PURIN THROUGH THE BODY (WALKER HALL)]
It will be seen that the disintegration of nucleic acid involves many
stages, and its complexity is such that we make no apology for drawing
upon the masterly monograph of Walter Jones for further elucidation
of this intricate question. In relating the history of nucleic acid
in the animal body Jones has found it convenient to introduce certain
terms wherewith to designate the various elements of the _nucleic
acid molecule_. Thus, the molecule in its entirety is termed a
_tetra-nucleotide_. The cleavage of this complex is initiated by the
action of two specific enzymes. Through their agency the tetra-nucleotide
is first cloven into two _di-nucleotides_, which immediately divide up
into four _mono-nucleotides_. These ferments are:—
(1) _Phospho-nuclease_ (which splits off the phosphoric acid radicle,
leaving a nucleoside, guanosine or adenosine).
H₂PO₄—C₅H₈O₃—C₅H₄N₅O + H₂O----->H₃PO₄ + C₅H₉O₄—C₅H₄N₅O
Guanylic acid. Phospho-nuclease. Guanosine.
(2) _Purin-nuclease_ (which splits off the purin radicle, viz., separates
out both phosphoric acid and carbohydrate groups, leaving free purin
bases).
H₂PO₄—C₅H₈O₃—C₅H₄N₅O + H₂O----->H₂PO₄—C₅H₉O₄ + C₅H₅N₅O
Guanylic acid. Purin-nuclease. Guanine.
Now, in sequence to either of the foregoing cleavages by the phospho- or
purin-nucleases another set of enzymes come into the field. Under their
_deaminising_ effect the amino group is abstracted, with the formation
of either free _oxy-purins_ or oxy-purins still bound in glucoside-like
combination with sugar.
If the oxy-purins are free, the following is the reaction:—
C₅H₅N₅O + H₂O------>C₅H₄N₄O₂ + NH₃
Guanine. Guanase. Xanthine.
Should, however, the guanine glucoside be present:—
C₅H₉O₄—C₅H₄N₅O + H₂O------>C₅H₉O₄—C₅H₃N₄O₂ + NH₃
Guanosine. Guanosine-deaminase. Xanthosine.
In the latter instance a hydrolysing enzyme, _xanthosine-hydrolase_, by
its action, splits off xanthine. We see, therefore, that by either route
the end-product is the same. Following a like series of changes, the
adenine radicle is transmuted into hypoxanthine. This either directly by
the action of adenase:—
C₅H₅N₅ + H₂O------>C₅H₄N₄O + NH₃
Adenine. Adenase. Hypoxanthine.
or indirectly through the agency of adenosine-deaminase, the
hypoxanthine-glucoside (inosine) is formed, and subsequently the
hypoxanthine is split off.
Xanthine and hypoxanthine are, therefore, now to hand, and given the
presence of oxygen, their oxidation to _uric acid_ ensues:—
C₅H₄N₄O + O---------------------> C₅H₄N₄O₂
Hypoxanthine. Hypoxanthine-oxidase. Xanthine.
C₅H₄N₄O₂ + O----------------->C₅H₄N₄O₃
Xanthine. Xanthine-oxidase. Uric acid.
Now, in man and the anthropoid apes, _uric acid_ is the end-product of
_purin_ catabolism. In contrast therewith in most mammals only a minimal
amount of the exogenous or endogenous purins escapes in the urine as uric
acid. Most of it undergoes further oxidation into _allantoin_,[15] this
change taking place in most mammals chiefly in the _liver_. According to
Schittenhelm, if nucleic acid be given to dogs, pigs or rabbits, from
93-95 per cent. thereof appears in the urine as allantoin, and only 3-6
per cent. as uric acid, and 1-2 per cent. as purin bases.
[Illustration: DISRUPTION OF NUCLEIC ACID MOLECULE (AMBERG AND JONES).]
In man, as in most mammals, uric acid is formed chiefly in the _liver_
from purins, and in the preceding table Amberg and Walter Jones
schematically represent the various steps by which disruption of the
nucleic acid molecule is attained, and uric acid formed.
DESTRUCTION OF URIC ACID
_Uricolysis_, or the destruction of uric acid, is, in most mammals,
achieved through the agency of the oxidising enzyme _uricase_, which
oxidises uric acid to _allantoin_. Consequently, in their instance,
purin bases, ingested as such or set free in the tissues, appear in the
urine, not as uric acid, but in the form of _allantoin_. On the other
hand, both in man and in the anthropoid apes, this particular enzyme
uricase is absent. In accordance therewith, only a trace of allantoin is
to be found in the urine of man and the higher apes, while in the lower
animals, _e.g._, dogs, pigs, and rabbits, a large proportion of the purin
excretion assumes this form.
Now, the absence of _uricase_, in _man_, is held to be proved by the
fact established by Wiechowski and others, viz., that uric acid, if
_injected subcutaneously_, may be almost wholly recovered in the urine,
and moreover, unchanged. On the other hand, the total excretion of uric
acid and the other purin bodies by no means tallies exactly with the
amount of the uric acid ingested as purin bases in the food and that
produced from the tissues; in other words, it has been found that,
when given by the mouth, _nucleic acid_ or _purins_ are by no means
_quantitatively_ excreted in the urine, even though not only _uric acid_,
but also _allantoin_ and the _purin bases_, are included within the
estimate. According to most experiments, a considerable proportion of the
purin-nitrogen intake, about 50 per cent., is excreted as _urea_.
The question then arises as to what becomes of that moiety of the _food
purins_ which fails to appear in the urine as _uric acid_. Now the amount
of allantoin that appears in the urine is negligible. Moreover, Ackroyd,
having shown that the organism cannot destroy allantoin, it is possible
that the minimal amounts excreted thereof in the urine are all derived
from the food.
Accordingly, if, as experimental feeding with _purins_ or _nucleic acid_
appears to indicate, purins are destroyed in the body they “pass through
some other route than allantoin, and possibly, that part of the purin
which is destroyed does not pass through the stage of uric acid.” Such is
Wells’ opinion, and he reminds us that _in vitro_ the destruction of uric
acid can be attained by other routes than through allantoin. Thus, it can
be broken down into glycocoll, ammonia, and CO₂, or by another method of
disintegration it furnishes first alloxan (C₄H₂N₂O₄), then parabanic acid
(C₃H₂N₂O₃), which in turn yields oxalic acid and urea.
But while it is probable that there is more than one way in which uric
acid can be decomposed in the body, nevertheless there is, according to
Wells, no evidence that either of the alternative routes above suggested
is ever affected in the animal body. In this _impasse_ Siven suggests the
further possibility, viz., that the moiety of the food-purins which fail
of recovery from the urine undergo partial destruction in the intestine
by _bacteria_.
Stewart, however, in his “Physiology,” discussing _uricolysis_, maintains
that a considerable destruction of uric acid and other purin bodies
goes on in the body and mainly in the _liver_. He reminds us that when
uric acid is heated in a sealed tube with strong hydrochloric acid, it
breaks down into glycin, carbon-dioxide and ammonia, and he maintains
that “there are grounds for believing that a similar decomposition takes
place in the body, and that the products are then transformed into urea
in the _liver_”; this, through the agency of a special ferment called the
_uricolytic enzyme_.
Also, Flack and Hill, discussing the metabolism of _nuclein_, hold that
some of the uric acid thus formed may be transmuted into urea by an
uricolytic ferment present in the _liver_, muscles, and _kidneys_. This
same agent they consider “probably destroys a considerable amount of the
uric acid formed in the body. Indeed, uric acid, even when given in the
food, owing to the presence of this enzyme, causes no increase in the
uric acid output of the body.”
On the other hand, Wells, discussing the destruction of uric acid,
observes that repeated investigations show “that the tissues of man have
no power whatever to destroy uric acid _in vitro_; the earlier reports of
positive uricolysis undoubtedly being erroneous.” His final conclusion,
after weighing all available evidence, is that it is highly probable that
in man “most of the purin absorbed from the food, and practically all
the purin from cell metabolism, is converted into uric acid and excreted
as such.” MacLeod, however, reflecting on the fact that uric acid is not
destroyed when extracts of the organs are incubated at body temperature
with uric acid or its precursors, bids us bear in mind that, “although
the uric acid is thus shown not to be destroyed _in vitro_, it may
nevertheless be destroyed in the living animal.”
We see, therefore, that the question, Whether uric acid can undergo
destruction in the human body? is still a matter of dispute, and must,
pending further investigation, remain _sub judice_. Still, despite the
conflict of evidence, clinicians have felt justified in assuming that one
of the factors in the genesis of gout may be an entire _absence_ or a
_diminution_ in the amount or activity of this _uricolytic ferment_.
But the awkward fact remains that all researches up to date have failed
to establish the presence in the human body of any enzyme which can
decompose uric acid. Should, therefore, future investigators place beyond
the reach of cavil the claim that no _uric-acid-destroying enzyme exists
in the body_, it would seem that, _ipso facto_, man, through lack of this
capacity for rapid oxidation of uric acid, is, by this same disability,
rendered a potential victim of _uric acid retention_ and _deposition_.
Elucidation of this vexed point seems more probable in view of the
striking discovery recently made by R. Benedict, viz., that in one
particular breed of dog, the Dalmatian, _uricase_ is wholly absent. In
respect of this _lack of a uric-acid-destroying ferment_, the Dalmatian
breed of dog has a _purin_ metabolism apparently identical with that of
man.[16] Thus, if fed on a purin-free diet, he passes large quantities of
uric acid, and if the latter be injected subcutaneously, elimination in
quantity as such ensues; this, in striking contrast to what obtains in
all other animals in whom, as before noted, uric acid is mostly oxidised
to _allantoin_ before excretion. Now, as MacLeod observes, investigation
into the metabolism of nucleic acid has, in man, been hampered greatly,
in that the absence of uricase from his tissues, prior to Benedict’s
discovery, rendered experimental researches on the lower animals
valueless. But, in light of the above revelation later by R. Benedict, it
may reasonably be hoped that in the near future our knowledge as to the
location and nature of the intermediary chemical processes occurring in
the metabolism of nucleic acids may be materially clarified.