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which the acid is actually excreted seems to he less conclusive. It is clear
that the alternating reactions just discussed would go on, whether the salts
Avhich leave the renal tubules are quadriurates or biurates. In the latter case,
the interaction with the phosphates would be the first stage of the process,
and the decomposition of the resulting quadriurates the second. Equations
(1) and (2), above, would occur in reversed order, and the alternation would
then continue as before. The fact that the solid excretion of birds and snakes
consists of quadriurates,^ may be held to support the view that these salts
are the excretory form in man, as also the observation that certain urate con-
cretions found in the kidneys of new-born children approximate in composition
to the quadriurates. 2 But it may be fairly argued that when, as in the
human adult, the mechanism of excretion has become more perfectly suited
to the elimination of a liquid urine, the uric acid will tend to assume the
more soluble form, and all the evidence points to the fact that this form is
the biurate. I have frequently observed that when ammonium urate separates
from a clear acid urine, as an effect of adding neutral ammonium chloride in
excess {vide infra), it is wholly in the form of a biurate. While it is not
inconceivable that a migration of bases occurs under these circumstances, it
is far more likely that the fact points to the pre-existence of biurates in the
urine.

Again, it will be found that many concentrated specimens of urine, when
first passed and while perfectly clear, will, on slight acidification with acetic
acid or with a mineral acid, give an immediate precipitate of quadriurates,
while the same specimen may requ.ire hours before any urate deposit separates
spontaneously. The explanation of this would seem to be that the urine
originally contained the more soluble biurates, and that these are changed
immediately upon artificial acidification, or more slowly by interaction with
phosphates, into less soluble quadriurates.

But whatever may be the primary form of the urates present, it is
in any case important to recall the facts discussed on p. 578. The
reaction between urates and phosphates is a reversible one ; with acid-
phosphates, biurates yield quadriurates ; with basic (monohydrogen)
phosphates, quadriiurates yield biurates. With a certain proportion-
ate mixture of the two types of phosphate the uric acid salts will be
therefore in equilibrium.

In many urines this equilibrium between the phosphates and urates
is established, and the determining reactions described above, therefore,
cease before all the uric acid is liberated. In others, where the
proportion of monohydrogen phosphate is at the outside large, the
equilibrium occurs early, and little or no free uric acid separates. Only
when the original excess of acid phosphate over basic phosphate reaches
an adequate value is the whole of the uric acid set free. In other

1 Roberts, loc. cit.

- Flensbuvg, Jahrcsb. ii. d. Fortschr. d. Tliier-Cliem., Wiesbaden, 1894, Bd. xxiii.
S. 581,



URIC ACID.



591



words, the chief factor which determines the precipitation of uric acid
is the degree of acidity of the urine. Eoberts lias found that two other
agencies exert an influence over this precipitation — the pigmentation
of the urine, and its comparative richness or poverty in sahnes. Other
things l>eing equal, a specimen which is poor in pigments on the one
hand, or in neutral salts on the other, will exhibit a special tendency
to deposit its uric acid in crystals. But while the cj^uestion of acidity
affects that stage of the process which consists in the change from
biurates to quadriurates, the pigmentation and percentage of salts
affect rather the change from quadriurate to free acid. The urinary
pigments and the neutral salts inhibit the decomposition of quadri-
urates by water.




Fig. 52. — Upper half, ammonium urate. Lower half, sodium urate.



UjDoii standing, some specimens of urine deposit urates, not as amorphous
quadriurates, but as crystalline biurates. Ammonium urate is frequently to be
seen in the deposit from alkaline urine in the form of roughly dumb-bell-shaped
masses; and in concentrated specimens sodium urate forms the so-called thorn-
apple crystals (Fig. 52).

Isolation of uric acid from the urine. — If the urine be acidified with
hydrochloric acid, much of its uric acid separates in pigmented crystals,
which tend to adhere to the sides of the vessel. These can be easily
identified by the microscope. But for the purpose of applying the
characteristic tests, a supply of uric acid may be more conveniently and
quickly obtained by adding crystals of ammonium chloride to the
urine till near saturation, and then a few drops of strong ammonia.
The precipitate which falls is at once filtered off, washed from the filter
with a little hot water, and warmed with a few drops of hydrochloric



592 THE CHEMISTRY OF THE URINE.

acid. After cooling, the crystals of uric acid which fall mav he washed
by decaiitation.

Tests and reactions — («) The murexicle test. — If a small quantity of
uric acid be placed upon a watch glass, a little strong nitric acid, or a
few drops of bromine water added, and the whole taken to dryness upon
the water-bath, an orange-red residue is obtained which, if touched with
a drop of ammonia, yields a fine purple colour. If a minute quantity of
sodic-hydrate solution be subsequently added, the purple colour changes
to blue ; while, on warming the alkaline solution, all colour is discharged.
The water-bath should always be used for evaporation in applying this test,
and if the watch glass be allowed to remain on the bath for a considerable
time, after evaporation is complete, a red colour will develop without
further treatment, and the residue will dissolve to a purple solution in
distilled water. This is the most delicate method of applying the test.

The residue left by the action of the nitric acid or bromine water consists
of various oxidation products of uric acid, amongst which is alloxantin
(CgHgN^Og or CsH^N^Og.H^O). This substance yields, with ammonia,
ammonium purpurate, which is the purple product of the test.

(b) If uric acid be dissolved in a little caustic soda, a few drops of
Fehling's solution added, and the solution boiled, a yellowish precipitate
of cuprous oxide is obtained (cf. p^ 608).

(c) An alkaline solution of uric acid gives, on the addition of a few
drops of a solution of pliosijliomolyldic acid, a dark blue precipitate with
a metallic lustre, which under the microscope is seen to consist of small
six-sided prisms.^

Estimation. — The methods now used for the estimation of uric acid depend
either upon the insolubility of its silver compound in ammoniacal solutions, or
upon the depression in solubility which ammonium urate undergoes in the
presence of other ammonium salts. Of the silver processes the Salkowski- ^
Ludwig ^ method is the most accurate. In this the phosphates of the urine are
first precipitated by the addition of an ammoniacal solution of magnesium
chloride, containing ammonium chloride (magnesia mixture). Without filter-
in" off the phosphates, a solution of ammoniacal silver nitrate is next added,
which gives a further precipitate of silver-magnesium urate. After standing,
the mixed precipitates are filtered off, washed, and treated with a solution of
potassium-hydrogen sulphide, which decomposes the silver compound, forming
silver sulphide and potassium urate. The black precipitate of the former is
filtered off, and the uric acid liberated in the filtrate by the addition of hydro-
chloric acid. It is finally separated by filtration and weighed.

The writer ^ has modified the previous methods employed for the separation
of uric acid as ammonium urate in such a way that the precipitation is absol-
utely complete, and the results are as accurate as those of the foregoing method,
while much more easy to obtain.^ The urine (100 c.c.) is saturated with
chloride of ammonium, and alloAved to stand for two hours, when the resulting
ammonium urate precipitate is filtered off", washed from the filter with hot
water, and the uric acid liberated by warming with hydrochloric acid.
After standing it is filtered off, washed, and weighed.

1 Offer, Centralhl. f. Physiol., Leipzig u. Wien, 1894, Bd. viii. S. 801.

- Arch. f. d. gcs. Physiol., Bonn, 1872, Bd. v. S. 210.

^ ZiHchr.f. anal. Chem., Wiesbaden, 1885, Bd. xxiv. S. 637.

'' Hopkins, Journ. Path, and Bacteriol., Edin. and London, 1893, vol. i. p. 450.

5Cf. V. Jakscb, " Klinische Diagnostik," 1896, 4th edition, S. 428, 431; Ritter,
Ztschr. f. physiol. Chem., Strasburg, 1895, Bd. xxi. S. 288; Luff, Goulstonian Lectures,
1897, Lect. i.



URIC ACID. 593

Variations in the, amount. — (a) The, relation to urea; the effects
of diet. — Variations in the quantity of uric acid have been considered
from two different points of view. By some, these variations have
been expressed always in relation to the quantity of urea excreted
simultaneously. Such observers have felt that an increase or decrease
in uric acid, which merely accompanies a corresponding change in the
general nitrogenous metabolism, is of less physiological significance than
a variation which occurs independently of (or out of proportion to) the
latter; and since the urea excretion is a measure of this general
metabolism, the uric has been, by such writers, referred to the urea
output as a standard. Other and more recent authorities, seeing the
origin of uric acid in an entirely distinct series of events within the
body, and observing that the urea : uric acid ratio has no stable
value, have recommended the entire neglect of this relation, prefer-
ring to express the uric acid output always in terms of its absolute
amount.

An attempt has been made to show that urea and uric acid are
always produced in the body, so as to bear a constant and definite ratio
to each other, and that any alterations in this ratio indicate either a
retention of uric acid on the one hand, or a sweeping out of previously
retained acid on the other.^ That this position cannot be maintained in
its entirety is quite certain. Consideration of the effect of varying diet
alone gives sufficient evidence against it. If the two analyses by Bunge,
given in an early section of this article, be examined, we see that upon a
diet of bread, not only is the absolute amount of uric acid less than upon
a diet of beef, but also that the relation to urea is also strikingly less.
On bread the ratio is 1 to 81, on beef it is 1 to 48. Similar results are
obtained, as the writer has found, if the experiments are continued for
many days. If the two substances were always produced in constant ratio
we should have to conclude that a bread diet produces a continuous
storing up of uric acid in the body ; and for this conclusion there is
certainly no evidence.

Again, if we consider the effect of varying the quantity of the
ingested food — its composition being maintained uniform — we find that
on the whole the uric-acid excretion is less affected by such variations
than is the urea, so that we change the value of the ratio merely by
altering the amount of food taken.

It is therefore impossible to look upon the ratio which uric acid
bears to urea as an independent physiological constant, or to conclude
that even wide variations in its value are necessarily pathological.

But some authorities go further than to say that the uric acid
output is more stable than that of the urea, claiming, indeed, that it is
quite unaffected by the absorption of the ordinary proteids of diet — the
albumins and globulins with their derivatives. If this be a fact, and
the production of the acid is independent of variations in these main
nitrogenous constituents of food, we ought certainly, in studying the
quantity in the urine, to neglect its ratio to urea altogether. This ratio
will then be little more, under ordinary circumstances, than an expression
for the urea variations, measured from the more stable uric acid output,
so to speak, as a base line ; while, if we are studying the effect of special
factors upon uric acid production, reference to the urea will be un-
necessary and misleading.

^ Haig, " Uric Acid in Disease."
VOL. I, — 38



594 THE CHEMISTRY OF THE URINE.

Salkowski, in 1889/ was among the first to give prominence to this
view, but the experiments upon which he then based his opinion were not
Avholly calciilated to decide as to what is the effect, if any, of tlie ingestion
of ordinary proteids. They were in the main those of Hirschfeld,- but
the experiments of this investigator were directed to a broader question
than that of uric acid excretion, and the diet for the purposes of his
research was made entirely abnormal, so that definite conckisions on the
point we are discussing cannot be fairly drawn from them. In the experi-
ments of Horbaczewski and Camerer,^ undertaken with the object of ascer-
taining the effect of glycerin, carbohydrates, and fat, respectively, on uric
acid excretion, there Avere certain "normal periods," in which a standard
mixed diet was taken alone. The fact that the diet was carefully maintained
at a uniform level makes these very careful experiments more or less unavailable
for our pur230se. ^Nevertheless, during one of these control periods, Avhich
lasted for many days, the urea excreted fluctuated somewhat widely, pre-
sumably from varying degrees of proteid absorption. Of this period the
author says : " The uric acid eliminated went hand in hand with the nitrogen
excretion. In general, the more the total nitrogen present the more the uric
acid found."

There are but few experiments recorded Avhich bear properly on our
problem, fundamental though it be : that is to say, experiments Avhere the
uric acid and urea (or total nitrogen) have been estimated from day to day
by reliable processes ; while the quantity, but not the quality, of the proteids
ingested has been made to vary widely. Schiiltze ^ found the uric acid rise
Avith increase of flesh diet. Hester and Smith ^ found it raised when the
ingestion of proteids Avas increased, though it was somewhat less affected than
the urea. I myself have repeatedly observed a rise to folloAV an increase in
the diet where the composition of this has been carefully maintained constant.

But these observations are open to one criticism. Whatever the effect of
globulins or albumins, there appears to be no doubt that ingestion of
nucleo-2Jroteids increases the excretion of uric acid ; calves' thymus, with its
abundant nuclein, has been largely used to test this point. Umber ^ and
Weintraud^ have found that Avith thymus the excretion of uric acid may
amount to double that of the same individual upon ordinary proteid (muscle)
diet of equal nitrogenous value.

Is, then, the smaller increase found when ordinary proteid diet is taken,
merely due to any nuclein present and not to the absorption of the ordinary
proteids? We shall be able to add the last Avord to this discussion
immediately.

If the effect of an isolated meal of ordinary mixed diet be studied, it
is found that an increase in the excretion of uric acid occurs ver}'- rapidly
after the food is taken. According to Mares ^ the maximum hourly excretion
occurs at the fifth hour after the meal ; four hours before the urea reaches
its maximum. This observer held, therefore, that it Avas not derived directly
from the ingested proteid, but from cellular activity during digestion.
Horbaczewski confirmed this result, and believed that it Avas due to a digestive
leucocytosis [vide article, " Metabolism "), Avith its consequent liberation of
nucleins in the body. But Camerer^ has recently found that this rise of

^ Virchov/s Archiv, 1889, Bd. cxvii. S. 572 ; comments on a paper by Spilker.
^ Jbid., Bd. cxiv. S. 301.

3 Sitzungsh. d. k. Akad. d. TFissensch., Wien, 1886, Bd. xcviii. Abth. 3, S. 301. '
* Arch.f. d. ges. Physiol., Bonn, 1889, Bd. xv. S. 427.
s New York Med. Jourii., 1892, p. 38.
^ Ztschr.f. klin. Med., Berlin, 1896, Bd. xxix. S. 174.
"> Berl. klin. JVchvschr., 1895, S. 407.
8 Centralbl.f. d. med. JFissensch., Berlin, 1888, S. 2.

^ Zlschr. f. Biol., Mimchen, 1896, Bd. xxxiii. >S. 136 ; also Weintraud, Chem. Centr.-
Bl., Leipzig, 1895, Bd. ii. S. 234.



URIC ACID. 595

uric acid after a meal is by no means marked, unless the food taken contains
nuclein. On a diet composed, for instance, of egg albumin, the rise was very-
small, while during the digestion of non-nitrogenous diet the output of uric
acid Avas even diminished. Camerer holds, therefore, that digestive leuco-
cytosis cannot be the cause of post-prandial increase, but only the actual
ingestion of nucleins, and his results would suggest that we must answer the
question in the previous paragraph in the affirmative, and recognise that
the excretion of uric acid is not increased by the ingestion of ordinary
proteids.

If this be confirmed, we must for the future attach no importance to the
urea : uric acid ratio ; and when we wish to eliminate mere dietetic effects
from our study of other specific variations in the urinary uric acid (when, for
instance, we are endeavouring to ascertain if retention is occurring in disease,
or whether a certain drug is promoting elimination), we must do this by
controlling the ingestion of nucleins during the experiments.

It should be understood, however, that in spite of much labour spent
upon the problem, our knowledge of the relation of urinary uric acid to
diet is scarcely yet upon a firm foundation, and contradictory statements
will be found in the literature. Future investigators may have to face
yet another difficulty, if it be true, as Weintraud ^ affirms, that a true
excretion of uric acid may occur through the walls of the intestine.

One fact is abundantly certain — that great individual differences
exist in uric acid excretion. In spite of all that has been said above, it
is found that, with the ordinary regularity of habits and diet customary
in civilised life, the uric acid output (when the whole twenty-four hours'
excretion is dealt with), and even its relation to urea, will remain fairly
constant in any given individual; whereas, when different individuals
are compared, much greater differences are seen. Before we can say
with certainty what constitutes a pathological or exceptional condition
in any case, we must know the normal behaviour of the particular
organisation in question (Salkowski).

The ratio borne to urea may vary in different healthy individuals
from 1 : 25 to 1:50; the proportion most commonly found being from
about 1:35 or 1 : 40.

(&) Variations apart from diet. — It is a well-established fact that
in newly-born children the uric acid excretion bears a high proportion
to the body weight, and also to the other nitrogenous constituents of
the urine. In the first few days of life 7'8 per cent, of the urinary
nitrogen may be in the form of uric acid.^

The absolute amount is increased by excessive exercise and
diminished by rest. With regard to dmgs, the action of alkalies is
still disputed. It is possible that an isolated dose may temporarily
accelerate excretion ; but, according to Spilker and Salkowski,^ continued
administration diminishes it. There is certainly no foundation for the
statements of Haig, that the excretion of uric acid varies inversely as
the acidity of the urine.* Salicylates undoubtedly increase the amount
in the urine. Pilocarpine produces an increase, possibly from the
leucocytosis which follows its use. Pathologically, there is increase in

1 Chem. Centr.-BL, Leipzig, 1895, Bd. ii. S. 310.
" Hofmeier, Virdwws ArcMv, 1882, Bd. Ixxxix. S. 493.
^ Ibid., 1889, Bd. cxvii. S. 570.

* Cf. Herringliam and Davies, also Herringliam and Groves, Journ. Physiol., Cambridge
and London, 1891, vol. xii. pp. 475 and 478.



596 THE CHEMISTR Y OF THE URINE.

conditions of leukaemia, and this may be said to be the only well-
established fact as to the effect of disease on uric acid excretion. In
gout, although the urate deposits form so prominent a factor, the question
of the amount excreted in the urine is still unsettled. In this country,
at any rate, many cases occur in which, as originally observed by Sir A.
Garrod, the excretion during the chronic condition is greatly diminished,
whereas, in relation to the acute attack, increased elimination may
occur.^ Pyrexia alone does not produce any marked increase ; but
in certain specific fevers with a definite crisis, a large temporary
increase may occur, depending, according to Horbaczewski, upon the
associated leucocytosis.

Uric acid is one of the commonest constituents of urinary calculi.

(e) The xanthin bases. — Several members of this chemical group
are found in urine, in variable but always small amount. Xanthin
itself was discovered by Marcet in 1819 as a constituent of a urinary
calculus, and its presence in urine was first demonstrated by Strecker in
1857. In addition to xanthin, the following members of the group
may be present — heteroxanthin, 2Jct'i''ci'Xanth'in, hypoxanthin (sarkin),
guanin, adenin, and carnin.

All these substances are closely related to each other and to uric
acid ; and the chemical group to which they belong also contains
certain important vegetable bases.

The relation of xanthin to uric acid is best understood by a com-
parison of the structural formulse ; our knowledge of the constitution of
the base being due to E. Fischer.

NH— C— NH NH— C = ]S\

I II >co I I )co

CO C— NH/ CO C— NH/

IsTH— CO NH— CH

(uric acid) (xanthin)

Xanthin contains one atom less oxygen than uric acid, while hypo-
xanthin contains one less than xanthin.

C,H,N,03 C^H^N^O^ C^H^N.O

(uric acid) (xanthin) (hypoxanthin)

In the laboratory means have not been found to pass from one of
these three compounds to another by oxidation or reduction ; but in the
body the steps involving oxidation can certainly occur.

Heteroxanthin and paraxanthin are homologues of xanthin, the
former being its methyl- and the latter its dimethyl-derivative ;
paraxanthin is therefore an isomer of the vegetable bases, theobromin
and theophyllin.

Guanin in an imido-xanthin ; that is to say, it is xanthin with an
oyygen atom replaced by an NH group ; and adenin bears the same
relation to hypoxanthin.

C5H4N4O.O C5H4N,O.NH C,H4N4.0 C.H^N^.NH

(xanthin) (guanin) (hypoxanthin) (adenin)

Uric acid and the xanthin bases are grouped together by recent

^ Cf. Fawcett, Guy's IIosp. liep., London, 1895 ; Luff, Goulstonian Lectures, Lect. i.



THE XANTHIN BASES. 597

German writers ^ under the term, " alloxuric substances," a name meant
to show then- relation on the one hand to alloxan,^ and on the other to
urea ; the bases themselves may be designated the " alloxuric bases."

The amount of the xanthin bases in the urine has been generally
understated until lately ; they amount collectively to something like
one-tenth of the uric acid present ; that is to say, an average of Ol to 0'07
grm. of the combined bases is excreted per diem (Camerer, Salkowski).
Of xanthin itself some 0'02 to 0"03 grm. is found, upon a mixed
diet.

General properties. — That the xanthin compounds, unlike uric acid,
are basic in character, is probably due to the fact that the CO group is
absent from the central carbon chain. (Of. graphic formulee above.)

Their basicity is, however, very feeble, and many of their compounds
with acids are decomposed by water — ^just as is the " sulphate " of uric
acid ; while, on the other hand, they are all capable of forming metallic
derivatives and compounds with other bases. They contrast sharply
with uric acid in their easy solubility in mineral acids. In ammonia
they are also soluble (with the exception of guanin). Xanthin itself
dissolves to a very slight extent in water, but the other bases are more
soluble.

They are precipitated from urine — (1) By the addition of phospho-
tungstic or phosphomolybdic acids in acid solution ; (2) by silver nitrate
in ammoniacal solution ; and (3) by copper salts ; especially in the
presence of thiosulphates. When precipitated by any of these methods,
they are accompanied out of solution by uric acid {vide infixt).

Isolation and estimation. — It is beyond the scope of this article to describe
in detail the separation of the xanthin bases individually. Very large quanti-
ties of urine (100 litres and upwards) are required for the purpose. If the
precipitate obtained by adding ammonia, and afterwards ammoniacal nitrate



Online LibraryE. A. (Edward Albert) Sharpey-SchäferText-book of physiology; (Volume v.1) → online text (page 84 of 147)