SULFURMETABOLISMIN CYSTINURIA1

By JAMESC. ANDREWSANDALEXANDERRANDALL(Fronm the Departments of Physiological Chemistry and Urology, School of Medicine, University of Pennsylvania,

Philadelphia)

(Received for publication April 10, 1935)

Although the literature contains numerous stud-ies of cystinuric cases, further data on the metab-olism of various sulfur compounds in the cysti-nuric individual is desirable, not only for the lightit may throw on an interesting anomaly but alsobecause the cystinuric may be valuable for theinvestigation of the course of sulfur, metabolismin the normal individual. Among all the elementsconcerned in animal metabolism, sulfur is uniquein that it undergoes the largest change of valence,the maximum valence change of -2 to + 6 beingaccomplished when hydrogen sulfide is oxidized tosulfate, and a change only slightly smaller beinginvolved in the normal metabolism of cystine.Since the magnitude of this change is not paral-leled by any other element it hardly seems sur-prising that some individuals fail to accomplishcomplete oxidation of ingested sulfur. Indeedwe might reasonably expect cystinuria to be ofmuch more common occurrence than it is. Thefollowing report deals with the results of adminis-tering certain sulfur compounds to a cystintiric.

EXPERIMENTAL

The subject, a 14 year old boy, had a history ofkidney involvement with a right nephrectomy formultiple kidney stones at age 12. The surgicalaspects of the case have been commented on re-cently by Herman and Lee (1). The subject isCase II in their report. Cystine has been con-stantly present in the subject's urine, quantita-tive determinations showing excretion of 0.4 to0.5 gram cystine per day. However, examinationof the urines of both parents gave negative results.At the time of our experiments the subjectweighed 35 kilos and was otherwise in normalphysical condition.

1A preliminary abstract of this paper was presentedat the Pittsburgh meeting of the American Associationfor the Advancement of Science, December 1934.

A preliminary abstract of the work was published inthe Proceedings of the Society of Experimental Biologyand Medicine 1934, 32, 455.

Total sulfur was determined by the Benedict-Denis procedure and total sulfate-sulfur afterhvdrolysis with HC1. In all cases the determina-tion was made gravimetrically as BaSO,. Cystinewas determined by the Sullivan colorimetric pro-cedure as modified by Brand et al. (2).

Table I shows the sulfur distribution in 24-hoururine samples of the subject at various periodsover several months. With the exception of theexperiments of September 23 and September 24,the subject was on normal miscellaneous diet witha somewhat higher sulfur intake than would befurnished by the two quarts of milk ingested perday (and for one day previous) during the experi-ment of September 23 and 24. On May 24, in-gestion of 10 grams sodium bicarbonate daily wasbegun. Although direct cystine determinationswere not made during the April period, they weremade for some days before May 24 (not recordedon the table) and the results indicate that the in-gestion of sodium bicarbonate has no effect on thedegree of sulfur oxidation nor on the cystine out-put. In this we are in agreement with the resultsof Lewis and Lough (3) and Robson (4), as op-posed to those of Looney, Berglund and Graves(5). The lack of effect of either sodium bicar-bonate or sodium citrate in decreasing the cystineoutput of the cystinuric has been confirmed bylater experiments. During the latter part ofMay, the sodium bicarbonate was replaced by anequivalent amount of sodium citrate which wasingested daily until the September experiments.No effect on the cystine output or on the sulfurdistribution was observed. We feel, therefore,that the only object to be gained by the feedingof base to the cystinuric patient is that of keepingthe cystine more completely in solution and of pre-venting formation of calculi. In this, we aremerely increasing the natural tendency of cys-tinuric urines to be more alkaline than those ofnormal subjects because of the usual restrictionof protein in the diet and also because of the ex-

517

JAMES C. ANDREWSAND ALEXANDERRANDALL

TABLE I

Oxidation of sulfur by a cystinuric subject

CystineNitro- Total Sulfate Unoxi- Percen- Cys- S per

Date ume gen S dized tage oxi- tine cent of RemarksN S dation S unoxi-

dized SI

grams grams grams grams per cent grams per cen

April 23 3400 7.58 0.632 0.357 0.275 56.5 Normal hospital dietApril 24 3700 7.30 0.589 0.370 0.219 62.8April 25 3100 6.22 0.614 0.330 0.284 53.8April 26 2670 6.80 0.542 0.356 0.186 65.7April 27 1810 7.02 0.669 0.423 0.246 63.2 .. .. .

May 24 1320 6.82 0.604 0.398 0.206 65.9 0.099 48.0 10 grams NaHCO3per dayMay 25 1760 6.86 0.566 0.332 0.234 58.7 0.134 57.2 . . .May 26 1300 7.06 0.551 0.314 0.237 57.0 0.107 45.1

May 27 1280 8.02 0.583 0.368 0.215 63.1 0.112 52.1May 28 780 5.83 0.456 0.269 0.187 59.0 0.087 46.5 5 grams glutamic acid + 2.55 grams gly-May 30 980 7.19 0.533 0.346 0.187 64.9 0.093 49.7 cine daily in addition to NaHCO3May 31 1060 6.90 0.485 0.305 0.180 62.9 0.105 58.3June 1 950 6.69 0.496 0.302 0.194 60.9 0.108 55.7

September 23 1245 6.77 0.434 0.225 0.209 51.8 0.100 47.8 Milk (2 quarts) + crackers and fruit

September 24 800 7.28 0.471 0.274 0.197 58.2 0.080 40.6 .. " . " . ..

October 27 1230 7.78 0.679 0.422 0.257 62.2 0.094 36.7 Egg diet (55 grams egg protein)October 28 740 8.13 0.902 0.607 0.295 67.3 0.078 26.3 .. . . I I ..November 5 1140 4.88 0.528 0.268 0.260 50.7 0.080 30.8 Low protein (30 grams protein)November 6 960 4.60 0.513 0.257 0.256 50.1 0.081 31.5 .. . .A ..

September 23 715 5.87 0.684 0.567 0.117 82.9 Normal subject-same age. Milk (2 quarts)September 24 1065 9.15 0.711 1 0.590 0.121 83.0 + crackers and fruit

cretion of an appreciable portion of the sulfuras cystine instead of as sulfuric acid. The factthat in cystinuria one of the principal sources ofurinary acidity (sulfuric acid) is diminished inquantity furnishes, to a partial degree, the alka-linity favorable to keeping the cystine in solution.If we compare the sulfur oxidation in normal withthat in cystinuric subjects and calculate the effectof this deficiency in sulfuric acid in the cystinuricon the primary-secondary phosphate ratio (thechief buffer system of normal urine), we find thatthe cystinuric urine should average from 0.4 to0.8 pH higher than the normal depending on vari-ations in sulfur and phosphate output. The uri-nary acidity of the present cystinuric has beencompared with that of a normal subject of thesame age and sex, both being on identical diets forseveral days. Complete urine samples collected ineach case over a period of two and one-half daysgave average pH values of 6.5 for the normaland 7.1 for the cystinuric.

The solubility curve of cystine with change inpH demonstrates the possibility of increasing its

solubility at higher pH values and makes it diffi-cult to evaluate the conclusions of Patch (6) asto the lack of efficacy of alkali. In the case ofour present subject, x-ray examination has indi-cated a definite improvement resulting from ad-ministration of alkali. It is obvious that if asubject has already deposited small calculi or"gravel," administration of sufficient alkali tomarkedly increase the solubility of the cystine andcause it to be excreted in solution will producea temporary rise in the total cystine excretion.

The suggestion has sometimes been advancedthat the metabolism of cystine is so bound upwith the formation of glutathione that administra-tion of glycine and glutamic acid might decreasecystine excretion. While there is little to supportsuch a view we thought it worth while to feedthese two amino acids in equimolar amounts for aperiod of some days. As indicated in Table I,5.0 grams glutamic acid + 2.55 grams glycine(supplying a total of 0.95 gram nitrogen) werefed daily for approximately one week. Neitherduring that period nor for some days following

518

CYSTINURIA

was there any noticeable effect on the amount ofcystine excreted. In this, our results are as com-pletely negative as those of Robson (4) whenglutamic acid alone was fed.

In investigating the metabolism of sulfur com-pounds administered by mouth we have also useda procedure in which urine collections were madeby two hour periods during the day and for thefollowing 12 hour period overnight. The subject,having been kept on a nonprotein diet the day be-fore the experiment, voided and discarded hisurine at 6 a.m. Samples were then collected at7, 9, arid 11 a.m., etc. until 7 p.m. The following12 hour period (7 p.m. to 7 a.m.) was also col-lected. Since the compound to be studied was in-gested at 7 a.m. on the day of the experiment, 24hours were allowed for its elimination, a periodinsufficient in most cases, for complete eliminationof the sulfur. On the day of the experiment nofood was ingested until the evening meal andpractically no protein was allowed until the follow-ing day. Constant quantities of water were takenhourly but in spite of this the bi-hourly outputoften varied considerably. The sulfur compoundwas ingested by mouth in quantity amounting to0.40 gram sulfur.

0

I-5x0

0 481216 2024 -TIME [ ROUR

FIG. 1. PERCENTAGEOXIDATION OF SULFUR RESULT-ING FROM ADMINISTRATION OF VARIOUS SULFUR COM-POUNDSIN AMOUNTSEQUIVALENTTO 0.4 GRAMSULFUL

I = 1-cystineII = dl-cystine

III = dl-methionineIV = blankV = cysteic acid

Arrow indicates time of administration of compound

Since the chief variation in the effects of dif-ferent sulfur compounds is to be found in the pro-portion of the sulfur oxidized and excreted assulfate, the results of a number of such experi-ments are summarized in Figure 1 by plottingpercentage oxidation against time in hours. Thepercentage oxidation represents the ratio of totalsulfate sulfur to total sulfur in the urine specimen.This percentage is higher in the samples collectedearlier in the day but as the experiment morenearly approaches a fasting basis, this figure inblank experiments becomes approximately 30 percent. Under the subject's normal dietary condi-tions this percentage is roughly doubled. Thelater rise in the curve is the result of the resump-tion of food intake on the sulfur partition in thesample collected the following morning.

In agreement with the classical picture of cys-tinuria, the present subject is able to oxidize freecystine, administered by mouth, as well as a nor-mal individual. This is indicated by the rise inthe curves for both 1- and dl-cystine during theearly part of the 24 hour period.

The comparative feeding of 1- and racemic cys-tine was prompted by the remarkable results re-ported by Loewy and Neuberg (7) who found acystinuric subject capable of oxidizing " stone cys-tine" to sulfate but incapable of so oxidizing"protein cystine" (from hair). The suspicionthat such results might be caused by difference inoptical configuration (the " protein cystine " hav-ing been racemized during preparation) is in-creased by the findings of duVigneaud, Craft andLoring (8). These investigators obtained, withrabbits, a percentage oxidation for d-cystine ofapproximately half the magnitude of that for the1-isomer. While it is true that our figures showsomewhat slower oxidation of racemic cystineduring the first 12 hours, the differences are notgreat and the results, as a whole, more nearly ap-proach those of Hele and Pirie (9) on dogs andLawrie (10) on rats. The figures for the excesssulfate excreted as a result of cystine ingestion,when corrected for the average fasting level, indi-cate a somewhat higher recovery of the sulfur for1- than for racemic cystine, but here again thedifferences are not highly significant, consideringthe difficulty of assigning a definite value for theaverage fasting level.

519

4

i4

JAMES C. ANDREWSAND ALEXANDERRANDALL

In the case of the 1-cystine experiment, about75 to 80 per cent of the sulfur administered, was

recovered as sulfate-sulfur in 24 hours, whereasthe same calculation applied to the experimentwith racemic cystine indicates a recovery of 60to 65 per cent. In neither case was any signifi-cant rise in unoxidized sulfur noted. In studyingthe absorption of cystine from dogs with isolatedintestinal loops, Andrews and Johnston (11) ob-tained results indicating slightly more rapid ab-sorption of racemic cystine. This, however, byno means implies a more rapid or complete meta-bolic oxidation.

Although administration of cysteic acid to thecystinuric individual would not be expected toproduce results different from those of the nor-

mal, such an experiment appears not to be re-

corded in the literature and we therefore includedin Figure 1, the curve of one of several such ex-

periments, conducted in the same way as describedabove. Since the cysteic acid is not converted tosulfate by the organism, the output of sulfate-sulfur remains relatively constant while the un-

oxidized portion increases moderately and the oxi-dation percentage drops. In another cysteic acidexperiment, the percentage of sulfur oxidationreached the low level of 17.4 per cent. However,the absolute increase in sulfur elimination was in

all cysteic acid experiments so slight that the sul-fur recovery amounted to only 30 to 40 per centof that fed. This is all the more surprising inview of the very rapid absorption of cysteic acidfrom intestinal loops of dogs observed by An-drews and Johnston (11), but accords with thelow recoveries of sulfur observed by Schmidtand Clark (12) after feeding cysteic acid to dogs.Further investigation of the cause of these lowrecoveries of cysteic acid, both in the normal andthe cystinuric organism is in progress.

The administration of methionine to the cysti-nuric individual presents several features of muchinterest. Not the least of these is the suggestionof Brand, Cahill and Harris (13) that cystinuriamay be essentially a disturbance in methioninemetabolism as indicated by their finding that theoutput of cystine by a cystinuric individual was

much increased (nearly doubled) by administra-tion of large doses of methionine. The methi-onine curve in Figure 1, obtained by feeding anamount of methionine equivalent to 0.40 gramsulfur, demonstrates that this amino acid is notnearly so readily oxidized to sulfate as is cystine.

In Table II are presented the results of a longerexperiment in which methionine equivalent to 0.50gram sulfur was fed after a preliminary period,and twelve hour urine samples were collected for

TABLE II

Administration of di-methionine(Urine sampks colkcted in five 12-hour periods)

Period Volume Total Total Sulfate Unoxidized Percentage Cystine Methionine a X cc.N S S S oxidation S S (2 dm.)

cc. grams grams grams grams grams grams degrees

Cystinuric subject

I 640 4.63 0.294 0.180 0.114 61.2 0.058 0.014 -115II* 690 e 3.92 0.453 0.243 0.210 53.6 0.062 0.084 -138

III 350 4.23 0.364 0.249 0.115 68.4 0.052 0.020 -70IV 490 3.72 0.276 0.152 0.124 55.1 0.042 0.018 -113V 710 4.19 0.310 0.173 0.137 55.8 0.071 0.017 -135

Normal subject

I 350 5.34 0.383 0.327 0.056 85.4 0.019 -39II* 1160 6.83 0.772 0.633 0.139 82.0 0.069 -116

III 560 6.42 0.457 0.393 0.064 86.0 0.028 -45IV 800 6.28 0.511 0.440 0.071 86.1 0.023 -64V 310 4.10 0.262 0.211 0.051 80.5 0.014 -43

* 2.33 grams. dl-methionine (-0.50 gram S) administered at the beginning of Period II.

520

CYSTINURIA

48 hours after its administration. In this case,the cystinuric was compared with a normal sub-ject of the same age and sex. Both subjects werekept on the standard milk-cracker diet used for theexperiments of September 23 and September 24in Table I during the day preceding the first periodof urine collection as well as during the entire ex-periment. The methionine was administered bymouth at the beginning of Period II.

Examination of Table II shows that here again,administration of methionine has little influenceon the percentage oxidation of the sulfur witheither subject since it produces proportional in-creases in both oxidized and unoxidized fractions.The figures for cystine sulfur excreted by thecystinuric demonstrate that this increase is notto be accounted for by an increased cystine out-put. The sharp rise in unoxidized sulfur notedin Period II is not accompanied by a significantrise in cystine sulfur while the rise in the latterin Period V is evidently the result of an increasedurine volume at that time (see below). Onestriking difference to be noted between the twosubjects is the promptness with which the normalsubject excreted the extra sulfur administered ascontrasted with the slower and somewhat erraticexcretion by the cystinuric patient.

The methionine administered was the dl-formwhereas the natural isomer is the 1 form ([a]D=- 7.2). On the supposition that the organismmost readily oxidizes the natural isomer and dis-cards all or part of the d-form, we have soughtto account for the fact that dl-methionine is lesscompletely oxidized than cystine and gives riseto increases in the unoxidized S fraction. Forthis reason methionine was determined directlyin these urines.

The figures were obtained by the method ofBaernstein (14). For each determination a 50 cc.sample of the urine was evaporated to drynessbelow the boiling point and the determination wasmade on this dried residue. It was demonstratedthat methionine, added to the urine before evap-oration, could be 94 to 96 per cent recovered.Normal urines give a small titration by thismethod, which, expressed in terms of methioninesulfur, amounts to about 0.02 to 0.04 gram per24 hours. The table shows that both normal andcystinuric subjects exhibited 4 to 6 fold increases

in this value during the period immediately fol-lowing methionine ingestion. However, the abso-lute amount of the increase accounts for not morethan 10 to 15 per cent of the methionine sulfuringested although it does account for 60 to 70 percent of the increase observed in the unoxidizedsulfur fraction. The fact that the methionine wasin the dl form lends some support to the view thatin these experiments oxidation may be less com-plete than would have been the case had the nat-urally occurring isomer been administered.

The last column of the table shows the productobtained by multiplying the optical activity ob-served with D-light in a 2 dm. tube at 250 C. bythe volume of the sample for that period. Theresulting figure is therefore an arbitrary measureof the excretion of some levorotatory constituentsand is obviously higher with the cystinuric be-cause of the presence of 1-cystine. With moder-ately constant urinary volumes, marked increasesin cystine output should be evident in these figures.However, the only case in which such an increaseoccurs is in Period II of the normal subject whenthe methionine ingestion was followed by markeddiuresis. That this increase in the optical figureis not due to excretion of cystine or homocystineis evidenced by the fact that all samples from thenormal subject gave negative cyanide-nitroprussidetests. In this, we fail to confirm the results ofVirtue and Lewis (15) and Vars (16) with rab-bits and dogs respectively but it should be notedthat our dose, per kilo of body weight, is far lessthan that used by them. The possibility of forma-tion of homocystine from the methionine in thecystinuric is practically excluded by the fact thatFolin-Marenzi determinations (which respond toboth cystine and homocystine) have given valuesfor cystine which were not appreciably higher thanthose obtained by the more specific Sullivanmethod.

In order to investigate the effect of still largerdoses of methionine on the cystinuric organism,a similar experiment was run on the same subjectover a period of fifteen days under carefully con-trolled dietary conditions. The diet used con-tained 300, 50 and 100 grams respectively of car-bohydrate, protein, and fat daily. Twentv-fouwrhour urine collections were made. After estab-lishing this metabolic level for several days, the

521

JAMES C. ANDREWSAND ALEXANDERRANDALL

subject was given a 5.0 gram dose of cystine inwater suspension. On the 4th, 5th and 6th daysafter cystine ingestion large doses of dl-methioninewere given. The results are summarized in TableIII.2

The results of this experiment confirm ourprevious conclusions: that with the present cysti-nuric subject, no very marked increase in cystine

may therefore conclude that there is no evidenceof excretion of homocystine. The figures formethionine sulfur again show an increase follow-ing methionine ingestion although the amount issmall as compared with the amount of methionineingested. The column showing optical activity(Table III) confirms the previous conclusion asto the constancy of cystine output.

TABLE III

Administration of di-methionine(Twenty-four hour urine samples)

Day |Volume | N | Creatinine Total Sulfate Unoxidized Cystine Methionine a X cc.DayVolume N 5~~~~~s S xiato 5 5 (2 dm.)

1

2345*6789t

lot11$12131415

cc.

184015201580200017901680174015401490162020002000196017601480

grams

7.277.527.317.717.937.117.026.587.036.927.517.167.016.836.87

grams

0.780.670.750.820.760.580.510.810.730.680.510.660.690.520.80

grams

0.7120.6850.7560.6161.4400.4250.4520.5180.8240.7880.9520.4960.5760.4040.572

grams

0.2360.3200.2970.3621.1260.1920.2360.2670.4730.4610.6450.2380.3130.2170.276

grams0.4760.3650.4590.2540.3140.2330.2160.2510.3510.3270.3070.2580.2630.1870.296

per cent33.646.739.358.878.245.352.251.557.458.567.748.054.353.748.3

grams

0.1470.1260.1240.1350.0910.0880.0900.0870.1010.1020.1080.1110.10o8i0.099-0.095'

grams0.0380.0410.0390.0280.0220.0210.0250.0220.0510.0440.0770.0210.0190.0230.020

* 5 grams 1-cystine (= 1.333 gram S) administered at the beginning of the 5th day.t 2 grams dl-methionine (=0.43 gram S) administered at the beginning of both 9th and 10th days.$ 5 grams dl-methionine (= 1.075 gram S) administered at the beginning of the 11th day.

excretion follows the ingestion of large amountsof methionine. It should also be noted that whatincrease in cystine output was observed accom-

panied the larger urine volumes. Wehave noted,on several other occasions, that diuresis is accom-

panied by an increased cystine output. The di-vergence between our results in this particular andthose of Brand and coworkers (13), suggests thatthere exists considerable variation between cysti-nuric subjects, and points to the desirability ofexperimentation with as wide a range of subjectsand conditions as possible. The figures for cys-tine sulfur were obtained, as before, by the Sul-livan method but no appreciable increase was ob-tained by the use of the Folin procedure. We

2We wish to acknowledge the kindness of Dr. ErwinBrand of the New York State Psychiatric Institute infurnishing the dl-methionine used in this experiment.Wewish also to acknowledge the assistance of KathleenC. Andrews in controlling the dietary conditions of theexperiment and in carrying out numerous analyses.

The percentage recovery of the sulfur is, as

would be expected, somewhat low. One couldhardly expect to administer doses of cystine andmethionine of the size used here without havingconsiderable loss in the feces.

In spite of the constancy of the total nitrogenthere is obviously considerable variation in thecreatinine figures, and it is somewhat suggestivethat the most marked variations date from theday on which the large dose of cystine was given.The variability of creatinine output in cystinuricsubjects has been commented on both by Alsbergand Folin (17) and by Brand, Harris and Biloon(2). These authors ascribe a rise in creatininewhich they observed to an unusual amount of ex-

ercise on the part of their subjects and since our

present subject was leading a normal school lifeduring the experiment with the daily amount ofexercise varying considerably this may account forour variations. The extremely cooperative atti-tude of the subject as well as the constancy of the

degrees

-340-420~360t

_300-320

-280 --260-460-440-460-420-340

522

CYSTINURIA

total nitrogen figures negates any assumption ofincomplete collections.

TABLE IV

Increase in free cystine content of cystinuric urines onstanding

Free cystine content of sample

At once After 10 days After 100 days

grams grams grams

0.034 0.044 0.0440.023 0.0360.050 0.0720.034 0.043 0.0440.036 0.0520.030 0.038 0.0360.056 0.056 0.0410.039 0.038 0.0370.050 0.057 0.0610.039 0.041

Table IV illustrates an interesting quality ex-hibited by some cystinuric urines. Brand, Har-ris and Biloon (2) have pointed out that the Sul-livan colorimetric method for cystine when ap-plied to very freshly voided cystinuric urines,sometimes gives values which are decidedly lowerthan those obtained when the same method is ap-plied later to the same samples. Because of thewell known specificity of the Sullivan procedureand its usual failure to respond to cystine com-plexes, this increase was explained by assumingthe excretion of an unstable complex which quicklydecomposed. The figures recorded in Table IVshow that in most cases we substantiate these find-ings as far as our subject is concerned. It willbe noted that this increase, when it is observed,is maintained on standing for over 3 months (at00 and preserved with chloroform). We havealso noted that cystinuric urines over eight monthsold show practically no further change in cystineconcentration. The statement made by Magnus-Levy (18) that such urines lose their cystine aftersome months is hard to explain except on the as-sumption that slow racemization occurred and thatthe higher solubility of the racemized cystine ad-versely affected the precipitation method he used.

The hypothetical cystine complex is, however,far less stable than the data in Table IV imply.Wehave observed an increase of practically thesame magnitude after keeping the fresh samplefor 24 hours at-0° C. The instability of the com-

plex is further emphasized by the fact that ourpresent subject, while giving evidence of its ex-cretion, has also formed deposits of practicallypure cystine. As far as we are aware, no cysti-nuric urine on record has ever given a negativeSullivan test when freshly voided, and certainlysuch a finding could hardly be expected in anysubject with even small deposits of free cystine inkidney or bladder.

Further investigation of the chemistry of thiscomplex is highly desirable since some means ofincreasing its stability might have practical bear-ing on the treatment of cystinuria; the excretionof a soluble complex might avoid the dangers ofcalculus formation. At present, however, wemust admit the possibility that this increase incolor may be merely due to the influence of someother urinary constituents on the Sullivan reaction.

SUMMARY

A case of cystinuria accompanied by calculusformation has been investigated with regard tothe metabolism of various sulfur compounds bythe subject.

The cystine output is unchanged by daily ad-ministration of alkali (sodium bicarbonate or so-dium citrate) over a period of several months.Deposition of calculi has, however, evidently beenprevented by this means.

Administration of equivalent amounts of glycineand glutamic acid is without effect on the rate ofcystine excretion.

Administration of 1-cystine by mouth results inpractically complete oxidation of the latter; ad-ministration of dl-cystine is followed by slightlyless efficient oxidation.

Cysteic acid, administered by mouth causes noincrease in sulfate excretion.

Following administration of dl-methionine tothe cystinuric subject we have observed: (1) Nosignificant increase in cystine excretion, (2) noexcretion of homocystine, (3) definite but veryslight excretion of methionine.

The increase reported by Brand and coworkers(2) in the apparent cystine content of these urineson standing has also been observed by us.

BIBLIOGRAPHY1. Herman, L., and Lee, W. E., Cystine nephrolithiasis.

Report of two cases. Ann. Surg., 1935, 101, 746.

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JAMES C. ANDREWSAND ALEXANDERRANDALL

2. Brand, E., Harris, M. M., and Biloon, S., Cystinuria.The excretion of a cystine complex which decom-poses in the urine with the liberation of free cys-tine. J. Biol. Chem., 1930, 86, 315.

3. Lewis, H. B., and Lough, S. A., The metabolism ofsulfur. XIV. A metabolic study of a case ofcystinuria. J. Biol. Chem., 1929, 81, 285.

4. Robson, W., Protein metabolism in cystinuria. Bio-chem. J., 1929, 23, 138.

5. Looney, J. M., Berglund, H., and Graves, R. C., Astudy of several cases of cystinuria. J. Biol.Chem., 1923, 57, 515.

6. Patch, F. S., Cystinuria and cystine lithiasis. Canad.M. A. J., 1934, 31, 250.

7. Loewy, A., and Neuberg, C., tber Cystinurie. Ztschr.f. physiol. Chem., 1904, 43, 338.

8. duVigneaud, V., Craft, H. A., and Loring, H. S.,The oxidation of the stereoisomers of cystine inthe animal body. J. Biol. Chem., 1934, 104, 81.

9. Hele, T. S., and Pirie, N. W., Studies in the sulfurmetabolism of the dog. VIII. The metabolismof glutathione compared with that of other cystinederivatives. Biochem. J., 1931, 25, 1095.

10. Lawrie, N. R., The metabolism of i-cystine in the rat.Biochem. J., 1932, 26, 435.

11. Andrews, J. C., and Johnston, C. G., The absorptionof certain sulfur compounds from intestinal loopsof dogs, J. Biol. Chem., 1933, 101, 635.

12. Schmidt, C. L. A., and Clark, G. W., The fate ofcertain sulfur compounds when fed to the dog.J. Biol. Chem., 1922, 53, 193.

13. Brand, E., Cahill, G. F., and Harris, M. M., Cysti-nuria. II. The metabolism of cystine, cysteine,methionine and glutathione. J. Biol. Chem., 1935,109, 69.

14. Baernstein, H. D., A modification of the method fordetermining methionine in proteins. J. Biol. Chem.,1934, 106, 451.

15. Virtue, R. W., and Lewis, H. B., The metabolism ofsulfur. XXI. Comparative studies of the metab-olism of l-cystine and dl-methionine in the rabbit.J. Biol. Chem., 1934, 104, 59.

16. Vars, H. M., Amino acid metabolism. Fate of dl-methionine in the phlorhizinized dog. Proc. Soc.Exper. Biol. and Med., 1933, 31, 129.

17. Alsberg, C., and Folin, O., Protein metabolism in cys-tinuria. Am. J. Physiol., 1905, 14, 54.

18. Magnus-Levy, A., Kleine Beitrage zur CystinurieBiochem. Ztschr., 1925, 156, 150.

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