SOME OBSERVATIONS ON THE BEHAVIOR OF THE ENDOGENOUS PURIN ... · 232 Endogenous Purin Excretion In the course of these investigations we have also studied the effect of the administration

SOME OBSERVATIONS ON THE BEHAVIOR OF THE ENDOGENOUS PURIN EXCRETION IN MAN.

BY J. J. K. MACLEOD AND H. D. HASKINS.

(From ihe Physiological Laboratory, Western Reserve University.)

(Received for publication, June I 7, 1906.)

Since the publication, in 1900, of Burian and Schur’s first paper on the metabolism of the purins,2 a considerable amount of interest in this subjeyt has been awakened in the physiological world. The derivation of the urinary purins from two sources- from the food (exogenous) and from the tissues (endogenous)-- and the variability in the amount of the endogenous moiety in different individuals but its constancy in any one individual while living under normal conditions and on a purin-free diet, are among the most important points established by these workers. Siv&n,s quite independently of Burian and Schur, simultaneously demonstrated the dual origin of the urinary purins and the constancy of the endogenous moiety, and during the past five years these laws; as we may call them, have been confirmed by other workers.

More recently, however, Folin 4 has communicated results which throw doubt on Burian and Schur’s statement that the .endogenous moiety is constant in amount provided that the diet be purin-free (which of course implies that changes in the amount of purin-free food ingested will be without effect on the purin .excretion) and, in view of the fact that this point is of funda- mental importance in the study of the whole question of purin metabolism, we have thought it advisable to place on record certain results bearing on the subject which we have obtained .during the past year.

I H. M. Hanna Research Fellow, Western Reserve University. a R&an and ScBur, Arch. f. d. ges. Physiol., lxxx, p. 241, 1900; lxxxvii,

p. 2.39, 1901; and XCiV, p. 273, 1903.

8 Siv*n, Skand. Arch. f. Physiol., xi, p. 123, 1901.

4 Folin, Amer. Journ. of Physiol., xiii, pp. 45, 66, and 117, 1905.

231

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232 Endogenous Purin Excretion

In the course of these investigations we have also studied the effect of the administration of citrates, and of alkalies, on the endogenous pm-in excretion, for, so far as we are aware, no work on this aspect of the question has been recorded since the publication of Burian and Schur’s paper.

The present investigations were carried out on three pesfectly healthy young men of whom one was one of the writers of this paper (H. D. H.) and the other two, students in this college. A moderate amount of work, mental and physical, was practised during the investigation by all three. The behavior of the urinary nitrogen, urea, and ammonia was investigated by one of us, the results being recorded in a separate paper in this Journal, where detailed accounts of the diet, body weight, etc., will also be found.

CONSIDERATION OF KESULTS.

Expeknent I. (TABLE I., H. D. H.)

This observation was divided into three periods; during the first of these, the diet consisted largely of tapioca, bread, milk, etc. ; it contained 3.5 grams of nitrogen. During the second period, eggs and milk were freely taken, but no meat or other purin-containing food; total nitrogen, 15 grams. During the third period, meat was taken in place of the eggs and milk of the previous diet: total nitrogen, I 6.5 grams. Since Folin has found potatoes to influence the purin excretion, these were excluded from the diet throughout.

About ten hours of laboratory work was performed daily. The urine collected during the first day of the low diet was dis- carded. The Camerer-Arnstein method was that employed for estimating the purins, the average of the duplicates being taken.

On the last two days of the first two periods, citrate of sodium was taken in five-gram doses two or three times a day. This rendered the urine strongly alkaline to litmus and caused it to effervesce briskly on the addition of acids.

The excretion of purin-nitrogen, exclusive of the citrate days, was fairly constant, the average during the second period (0.146


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J, J. R. Macleod and H. D. Haskins 233

gram)’ being slightly higher than that during the first period (0.137 gram).

TABLE x.-H. D. H .-BODY WEIGHT, 61-63 KILOS.

Nature of Diet.

Starch and cream, containing 3.5

Eggs, milk,. etc., contam ing 15 gm F. g;z; win

&VS. i

Diet include< 390 gm. lean steak. Total liitrogen 16. ! grams.

I.

I

s

r=

1

5

965 1025

Iii 880 830

1165 910 770 825

Excreted per diem in Urine

‘itrogel irams.

6.94 6.71

K 6:OS

PurinG~;~gen.

-._ ---~ -.__

Average

0.186 0.222 0.273 0.247 0.246

Body weight 63.6 Kg.

Citrate of sodium in 6 gm. doses 3-4 times a day,

z----

Day missed.

[email protected] of so- p~‘3npm

times a day.

Body weight ,61.6 Kg.

During the third period, a daily ration of 390 grams of lean steak was taken, but no other pm-in-yielding food stuff. Allow- ing that lean steak contains o. 06 per cent. of purin nitrogen (Burian and Schur) and that 50 per cent. of this (0. I I 7 gram) appears in the urine as exogenous purins, we have for &he endogenous excretion during this period, o. 130 gram of purin nitrogen,

Regarding the effect of citrate administration, it will be noted that during Period I a distinct increase in purin nitrogen

1 In this average the purin value for Day VIII is omitted on account of errors in technique.


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occurred on the first day that citrate was taken, but a diminution during the second day. The average excretion for the two days was 0,151 gram of purin nitrogen. During Period II the increase, thus induced, was very marked on the first day and it was also very distinct on the second day.

Experiment II. (TABLE II, L.)

Throughout this observation a purin- and potato-free diet was taken, For the first eight days, this contained 5.5 grams of nitrogen and during the next nine days 23.2 grams. The energy value of the diet was carefully maintained at about 3000 c.

TABLE II.--L.-AVERAGE BODYWEIOHT, 56.7 KIIOS.

N o.

:I III IV

VVI VII

VIII --

IX

11 XII

5::’ xv

XVI

XVII

Nai%* Of

Purin-free diet with 5.5 gm. nitrogen

Ci t ra ie day

Purin-free Siet with 23.2 grams nitrogen.

I Citrate

days.

-

vol:F Urine.

c. c.

1122 995

1200 1318 810

1850 1200

Jitit~~ Purb&rogen.

--__

9.99 0.269 rerages 7.27 0.152 6.35 5.90 :470 3.86 E 3 0:093

0.138

6.16 5.17 t:xi

960 5.05 0.131

:zx

E 975

1420 1840

7.47 0.133 12.05 14.83 it :xz

0: 125 With

:4”:2: XVII

0.123 1 0.126 14.87 0.256 16.23 0.288

1285

1230 -__

15.61 0.248

15.43 0.136

-+

- I Excreted per diem in Urine

- I 1

-

Remarks.

N’gt. 56.4Kg.

N’gt. 56 Kg.

Sodium citrate taken till tmimi;/ust al-

W ‘gt.‘57 Kg.

Sodium citrate till urine jistinctly alkaline. Urine acid. ammonia normal. Y’g1.67.5 Kg. ---___


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J. J. R. Macleod and H. D. Haskins 235

Omitting Days IV and V when the results were out of line,’ it will be seen that break down of body proteid occurred during Period I and retention of proteid during Period II.

The average daily excretion of purin-nitrogen (omitting the first and the citrate days in each period) was, for the first period, o. 138 gram, and for the second, o. 126 gram.

Citrate administration did not cause an increase in purins during Period I-although it affected the ammonia excretion (p. 228)-but it had a very distinct influence on this during Period II. In neither period did it affect the excretion of nitrogen,

Experimelzt III. (TABLE III., E.)

In this case also a purin- and potato-free diet was taken, For the first nine days of the observation this contained a daily average of 5.2 grams of nitrogen: for the remaining twenty days the average was 15 grams, The energy value was about 3000 C throughout. The subject was on diet for three days before the urine was analyzed.

Sodium bicarbonate was taken on the last two days of the low- diet period, and again on the fourth day of the liberal diet. Sodium citrate was taken on the fourteenth and again on the twenty-first, twenty-second, and twenty-third days, i.e. during the liberal diet.

In this case estimation was made not only of the total purin- nitrogen but also of the uric acid (Folin and Shaffer’s modification of Hopkin’s method).

The nitrogen excretion behaved as in the previous cases, Disregarding, for the present, the effect of citrate and alkali, it will be noted that the daily average excretion of total purins was the same on the low as on the liberal diet, viz., o . I 74 gram of purin-nitrogen for the low, and o. 168 gram for the liberal diet. This is in accord with Burian and Schur’s law. On the other hand, a comparison of the uric acid excretions during these diets reveals a somewhat higher average daily excretion on the liberal than on the low diet, viz., o . I 43 gram of uric acid nitrogen for the latter, and o. I 54 gram for the former. This difference is,

1We have, however, recorded the values obtained on these days, since we cannot account for the irregularity.


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TABL

E 1x

-E

.-AVE

BAG

B BO

DY

WEI

GH

T.

71.3

KI

LOS.

T-

Excr

eted

pe

r di

em

in Ur

ine.

7-

Rem

arks

. - n.

I NO

. Na

ture

of

V’$‘

!=z

of

Diet

. c.

C.

*

---

$1

;p:r

::’

tai&

g 4.8

15

Gm

. N.

V

Sodiu

m

( :i%

VI

bl.

carL

?o-

) 10

10

nate

days

---

-.

Body

we

ight

, 71

.2 Kg

.

urine

alk

aline

.

-

Two

days

m

issed

.

: I Twod

aysm

ised.

I

Ur$e

ac

id.

a.

. .

. .

A=W

0.

143

0.15

4

8.63

10.70

‘Z:E

10.91

11

.62

E:E

11.38

11

.27

11.58

13

.12

:Ef

:%

10.74

14.42

:::iZ

1 I

VII

, Pm

-in-fr

ee

760

[O.ll

l)

0 17

8 I

0.17

4 i

0.16

3

0.19

1 0.

189

8%i

p;

1

:::z

0:

178

I

gg

oh36

8%

0:141

ET;

0.131

0.13

8

0.16

s

0.15

4

0.174

diet

con

- I

%I1

SN!5

/

X so

dim

/ i3k

arbo

-

%I

n&e

day.

1 I

0.176

0.187

0.208

XIII

5:”

Sod.

citra

td

ET:1

XE

TI

f F

XXI

Sod&

m

XXII

Cifn

zft?

xx11

4 da

ys.

I I- ET

XX

VI


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J. J. R. Macleod and H. D. Haskins 237

however, only noticeable after the liberal diet had been taken for some time and is so small as to be negligible. It is scarcely allowable to infer anything regarding the behavior of the purin bases, since the only values we have for these are calculated from the difference between the purin and uric acid figures. These values for the periods bracketed in the table are as follows: 0.033, 0.048, 0.038, 0.019, 0.014, and 0.034 grams of purin nitrogen.

The administration of alkali, during both the low and the liberal diets, produced a distinct increase in the total purin and uric acid excretions. During the latter diet, this effect did not appear on the day of taking the alkali but was well marked for four days following this, the urine reacting acid towards litmus on these latter days. At any rate, we are not able to explain the high purin excretion on Days XI, XII, and XIII in any other way, there having been no irregularity in diet or other unusual condition to explain it.

Citrate of sodium was taken on Day XIV, during the liberal diet. On the first day, it did not influence the previously high purin excretion but it will be noticed that on the two succeeding days a somewhat smaller amount than normal both of purin and uric acid was excreted, It was given later in the experiment (Days XXI, XXII, XXIII), producing no effect until the third day of its administration but being followed by a high excretion lasting several days.

DISCUSSION OF RESULTS.

The consta,ncy of the purin excretion notwithstalzdiug considerable variations in the diet is clearly demonstrated by our results. It is true that in the case of H. D. H. a slight average increase was Jbtained during the more liberal diet, but, since the excretions of only three days are included in the average, no weight can be given to so slight a variation. This result, like those of Siv&n,r Rockwood, Walker Hall.,3 and others, would therefore corroborate Burian and Schur’s conclusion that the endogenous

1 SivPn, lot. cit. 1 Kockwood, E. W., Amer. Joum. of Physiol., xii, p. 38, 1904.

3 Walker Hall, The Purin Bodies of Food Stuffs, Manchester, 1902; also, Brit. Med. Journ., Sept. 24, 1904.


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moiety is uninfluenced by the nature and amount of the diet, provided that this is purin-free and sufficient in amount [and of proper composition] 1 to prevent starvation. Folin has, as remarked above, doubted the truth of this law. In two respects, however, Fofin’s experiments differ from those of Burian and Schur: firstly, in the method used for analysis,2 and secondly, in that the low diet in Folin’s observations contained very much less nitrogen than did that of Burian and Schur.

It is on account of the first of these points that, in the case of our third observation (on E.), we carried out parallel estimations by the two methods employed by these workers.’ It will be noted that the average results for uric acid rare slightly higher on the liberal than on the low diet (viz., o. 143 gram uric acid nitrogen on the low diet as against o. 154 gram on the liberal diet), but the difference is too small to be considered of any account. We must conclude, therefore, that the apparent dis- crepancy between the conclusions of the two groups of workers is due to differences in experimental conditions, viz., to Folin’s low diet containing much less proteid than did that employed by Burian and Schur.

Polin’s low diet in the two cases in which he noted a very distinct diminution in uric acid contained an average of about one gram of nitrogen and consisted of starch and cream. Rurian and Schur’s low diet contained over nine grams of nitrogen and was varied in nature: milk and eggs for four days, and vegetables for four other days. The distinct falls in the excretion of uric acid, observed by Folin, occurred when this starch and cream diet was taken. In two of Folin’s experiments in which the low diet was a mixed one (vi& Tables VI and VII of Folin’s paper, pp. 106-109 and also p, 90) the endogenous purins were not diminished, even although there was a fall in the total nitrogen excretion of seven and ten grams respectively.

It seems to us that, in the sense in which it is expressed by

iThe words in brackets are not included in Burian and Schur’s statement of the law.

2Folin estimated uric acid alon;! by his and Schaffer’s modification of Hopkin’s method,--Zeitschr. f. ~hysid. Chem., xxxii, p. 552, 1901.

Burian and Schur estimated not only uric acid but also total purins

and purin bases @tie Burian and Schur, LX. cit.). The total purin nitrogen results are those from which the authors drew their main conclusions.


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J. J. R. Macleod and H. D. Haskins

them Burian and Schur’s law * holds good notwithstanding Folin’s results, for such an unusual diet as this observer employed can scarcely be considered as normal. The extreme importance of the constancy in endogenous purin excretion under normal conditions lies, we think, largely in the fact that it gives us a rational basis from which further work on this intricate subject can be done, and we consider that Burian and Schur, and subsequent workers, have amply justified this view of the case. We ought to take Burian and Schur’s endogenous excretion as our working standard, and if we do so then we must conclude that Folin has succeeded in producing variations in this standard by unusual conditions of diet just as we, and others, have done by the administration of drugs, and Rurian has done by muscular exercise. There can be no doubt that, at no very distant date, many other conditions will be discovered which influence the excretion. It should not be lost sight of that the two groups of workers were studying the question from different standpoints: Folin to show that with the greatest possible variations in purin-free diet certain changes take place in the composition of the urine; Rurian and Schur, to show that with distinct, but not excessive, changes in the purin-free diet the purin excretion is unchanged and could therefore be considered as the normal endogenous excretion for that individual.

The i+zdividual variation in the endoge+zous moiety is very noticeable in our results: for H. D. H. it is o. 138 gram purin- nitrogen, for I,., 0. I33 gram, and for E., 0.173 gram. It is interesting to note that H. D. H. and 1‘. were much smaller men than E., so that Walker Hall’s assertion2 that the endogenous moiety bears a direct relationship to the body weight would seem to be confirmed by our results. The only worker, so far as we are aware, who has questioned this individual factor in determining the amount of the endogenous purin excretion is Loewi.3 As

1 Rurian and Schur state this part of their law as follows: “Es ist nur dafur zu sorgen, dass die Nahrung das Nahrungsbediirfnis in1 gauzen deckt ” (Zoc. cit., p. 303).

* Walker Hall, Zoc. cit. 8 Loewi, Arch. f. exp. Path. u. Pharm., xliv, p. I; also Arch. f. d. gcs.

Physiol., lxxxviii p. 296.


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pointed out by Burian,’ however, the observations on which Loewi bases his conclusions are inadequate to prove the point.

The Effect of Citrate Ad&&ration.-Citrate of sodium was taken by each of the observed persons at two periods during the observation. In the cases of H. D. H. and I,. it was taken during the low, and again during the liberal diet. In the case of E. it was taken at two periods during the liberal diet. The effects of the administration on the endogenous purin excretion may be summarized as follows:

During the low diet, in the case of H. D. H., an increase amounting to 29 per cent. of the previous average excretion was caused on the first day of administration, but a decrease on the second day. In the case of L. no change was produced by the administration while on the low diet. During the liberal diet, the citrate caused a marked increase in all the cases,except on the first occasion on which it was given to E. when the previously high purin excretion, following alkali administration, probably masked its effects. The exact manner of increase in the three remaining observations on the liberal diet was different in each case: in the case of H. D. H., the increase on the first day was about I 12 per cent. of the previous average amount, but only about 20 per cent. on the second day. In the case of L. the increase on the first day was about I go per cent., on the second day about I 27 per cent. and on the day after the administration had been stopped there was still an increased excretion amounting to about g6 per cent. In the case of E., there was no distinctly increased purin excretion on the first two days of citrate administration, but on the third day an increase amounting to about 40 per cent. was noted. Two days then elapsed before the urine was again examined, but on the next two days (i, e., third and fourth after discontinuing the drug) an increase amounting to about 34 per cent. was present. These averages are calculated from the total purin values, but it Gill be seen that those for the uric acid agree closely with them.

A similar increase in the excretion of endogenous purins has been noted to follow the taking of salicylate of soda (Walker

1 Rurian, ibid, xciv, p. 273, 1902. See also article “The Metabolism

of the Pm-ins” in Recent Advances in Physiology and Bio-chemistry, edited by 1,. Hill, Longmans, Green, & Co., 1906.


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J. J. R. Macleod and H. D. Haskins *4*

Hall’) and alcohol (Beebe 2), and from the literature prior to the date of Burian and Schur’s first paper we find that an increased excretion of uric acid has been observed after taking citrate of potash or alkalies (HaigP Gorsky 4). On the other hand, Klemptner s and Burchardd record a diminution in uric acid as a result of the taking of citrates.

The undolrbkd increase in the endogenous purivt excretion which our resulfs show to follow citrate administrations might be accounted for in various ways, viz. :

I. By some of the citric acid, before it had become completely oxidized, being used to bring about a synthesis of two urea molecules into uric acid. Thus, Wiener r has shown that a synthesis of this nature occurs in birds after they have been given such organic acids as lactic, malonic, tartronic, etc., and he further claims that under certain conditions a similar process may take place in mammals.

2. By the citrate raising the alkalinity of the blood and in some way diminishing the destructive power of the organism towards uric acid.

3, By an increase in the amount of blood circulating through the kidneys, whereby, in terms of Luthje’ss and Burian and Schur’ss hypothesis, a larger fraction than normal of the uric acid in the blood would escape through the kidney filter before being destroyed in the liver, etc.

4. For the sake of completeness, we must add to these the hypothesis of Haig lo that the increased alkalinity of the blood causes uric acid, stored away in the liver and spleen, to be re- moved by the urine.

1 Walker Hall, Bvit. Med. Jour., Sept. 1904. 1 Beebe, S’P., Amer. Journ. of Physiol., xii, p. ~3, r9o4. SHaig, A., Joum. of Physiol., viii, p, 211, 1887. 4 Gorsky, Inaugural Dissertation (Russian), St. Petersburg, 1888: from

Atwater and Longworthy’s Digest of Meiabolism Experkenfs. S Klemptner, Inaugural Dissertation, Dorpat, ~889, from Atwater. 6 Burchard, lnaugural Dissertation, Dorpat, 1889, from Atwater. TWiener, Be&. z. them. Physiol. und Path., ii, p. 42, 1902; also Er-

gebnisse der Physiol., i, Biochem. Abth. 8 Liithje, Arch. f. Verduuwgskrunkh., ii, p. 36, 1896. Q Burian and Schur, Arch. f. d. ges. Physiol., Ixxxvii, p. 239, x901.

10 Haig. Ioc. cit.


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Of these possible causes, we may, with certainty, immediately eliminate the third and fourth. The absence of any constant diuresis on the citrate days eliminates the third possibility, as well as the fact that diuresis produced by other causes has no effect on the twenty-four hours’ purin excretion (0. g. the, taking of diuretics, the imbibition of large quantities of water, beer, 1 etc.). The persistence of the increase for some days after discontinuing the drug (cf. Tables II and III) makes Haig’s hypothesis untenable, quite apart from the numerous other known facts which stand against it.

Between the remaining two possibilities there exists this difference that in the one-the synthetic hypothesis-the citric acid acts before it is completely oxidized to carbonate, and in the other that it is first oxidized and so renders the blood and urine more alkaline. To decide between these in our third experiment, we administered sodium bicarbonate instead of citrate, with the result that a similar increase in the purin excretion was obtained, which would point to increased alkalinity as at least one of the causes. Exactly how the increased alkalinity brings about, the increased purin excretion remains an open question, and the possibilities are many: thus, it may diminish the destruction of uric acid by the liver, etc., or it may in some way stimulate the practically dormant synthesis from urea which Wiener believes can under certain conditions come into evidence in the mammal, etc.

CONCLUSIONS.

I. The excretion of endogenous purins is not affected by very considerable variations in diet provided that this contains no purins.

2. The endogenous purin excretion of different individuals is variable.

3. The administration of citrate of sodium until the urine reacts alkaline towards litmus causes an increased excretion of endogenous purins which may continue for some days after discontinuing the drug, even although the reaction of the urine has meanwhile returned to acid.

4, Sodium bicarbonate similarly administered also increases the endogenous purin excretion.

1 Rurian and Schur, lot. cif., p. 347.


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J. J. R. Macleod and H. D. HaskinsPURIN EXCRETION IN MAN

BEHAVIOR OF THE ENDOGENOUS SOME OBSERVATIONS ON THE

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SOME OBSERVATIONS ON THE BEHAVIOR OF THE ENDOGENOUS PURIN ... · 232 Endogenous Purin Excretion In the course of these investigations we have also studied the effect of the administration

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