edema -...
TRANSCRIPT
EDEMA
II. )mE EFFECTIVENESS OF ULTRAFILTRATION FOR QUANTITATIVELYDETERMININGTHE "FREE WATER" CONTENTOF BLOODPLASMA,AND FOR ESTIMATING PHYSICAL-CHEMICAL CHANGESOF THEPLASMAPROTEINS IN EDEMA
BY MARGARETEMETAKUNDE(From the Otho S. A. Sprague Memorial Institute and the Pathological Laboratory of the
Cook County Hospital)(Received for publication September 14, 1926)
The physicochemical condition of the blood plasma in edema hasbeen the subject of much interest due to the attention which Fisher's(1) writings have drawn to the r6le of the colloids. In connectionwith a discussion of the causes of defective water secretion by nephritickidneys, Fisher suggested that "acid acts upon the tissues of the bodyincluding the blood and lymph. The increased hydration capacityresulting from this makes the tissues hold more water in combinedform (maintenance of body edema) while at the same time it preventswater becoming 'free' in the arterial blood stream." A review of theevidence for and against this theory is entirely outside of the scope ofthis paper. Our interests regarding Fisher's theory are chiefly di-rected towards the researches recently published by Beckmann (2).
By a process of ultrafiltration Beckmann studied the "free" watercon-tent of the blood plasma of seven cases of edema with varyingetiological factors. In some instances he obtained a diminishedamount of ultrafiltrate. Beckmann assumes that the amount ofultrafiltrate obtained from certain of these patients was diminishedbecause the plasma colloids were swollen and held part of the water ofthe blood in combination so that it was not obtained by the process ofultrafiltration. In other instances where he obtained an increasedamount of ultrafiltrate, he assumes that a converse physicochemicalchange had occurred in the plasma proteins, i.e., they were shrunken,thus lessening their ability to combine water, which would result in anincreased amount of free water in the blood plasma.
577
8FREEWATEROF PLASMA
Beckmann used the method of ultrafiltration described by Ellingerand Neuschlosz (3). He also made quantitative determinations of thedry residue of the blood plasma and of the plasma proteins. Heassumes that quantitative differences in the dry residue may causevariation in the amount of ultrafiltrate delivered, independent of theshrunken or swollen condition of the plasma colloids and developed aformula to correct for this variation.
Our purpose in undertaking this research is not to utilize the formuladeveloped by Beckmann, but simply to determine whether measurabledifferences from the normal can be found in the amount of waterobtained by ultrafiltration from the plasma of patients with varyingtypes of nephritis and edema, and to determine whether these differ-ences, if present, bear any relationship to quantitative variation in theplasma proteins. Moreover, if in edema the plasma proteins becomeshrunken or swollen, such changes should be detected by using ultra-filters of graded permeabilities.
METHOD
The details of the technique employed in the process of ultrafiltration and thecontributions of pioneer workers have been discussed by Bechhold (4); more re-cently Zilva and Muira (5) have contributed to the method by a different processfor standardizing membranes. Cushny (6) in his researches directed towarddetermining whether or not the crystalloids of blood plasma exist in combinationwith colloids, reduced the high pressure advocated by Bechhold (1 to 20 atmos-pheres) to 21 cm. of Hg.
Weemployed the essential principles of the technique advocated by Beckmann,i.e., a pressure equal to 77 mm. Hg, and the gel for making membranes which was aglacial acetic acid solution of parlodion. Figure 1 illustrates the simple apparatuswhich proved entirely effective for producing and maintaining, at a constantlevel, low hydrostatic pressures. The pressure is raised by establishing a siphonwhich is not interrupted during the experiment, so that if a lowering of pressureoccurs in the system by a slight leak of air or the filtering through of a few cubiccentimeters of fluid, it is immediately restored with water from the reservoir.
The ultrafiltration sacs employed in the experiments of table 1, were made ac-cording to the technique used by Beckmann, i.e., paper Soxhlet extraction thim-bles were impregnated with a glacial acetic acid solution of parlodion (DuPont)by suspending each with a loop of thread sewed through the upper part 1 cm. fromthe top and filling with the gel. The gel was of such consistency that completeimpregnation (indicated by drops of gel falling from the outer surface of the sac)occurred in 3 to 5 minutes. The excess of parlodion was allowed to drain off,
578
MARGARETEMETA KUNDE
then the sacs were fixed in distiUed water and washed in running water for 12 hours.Whennot in use they were kept under distilled water in the ice chest. The mem-branes were brought into the pressure system by mounting each on a perforatedrubber stopper of slightly less diameter than the membranes. Short glass tubing,one end of which was connected to the pressure system, the other end passing
I.~ ~~Z
FIG.'1.PSESMMNj
FIG. 1. PREssuRE SYSTEmwrmH MimBANs ATTACHED
4
1, cistern; 2, rubber tubing; 3, pressure bottle; 4, collecting bottles; 5, metaltop; 6, "T" tubes; 7, "T" tube for manometer; 8, membranes; 9, rubber stoppers.
through the perforated stoppers, admits the pressure from the system into themembranes. Above the filtration sac, the tubing passes through a large rubberstopper which loosely covers the receiving bottle. This prevents excessive evapo-ration of the ultrafiltrate, but admits atmospheric pressure to the outside of themembranes. The amount of blood plasma placed in each sac in all experimentswas 5 cc. The duration of the experiment was 3 hours unless otherwise stated.
579
TABLE 1
Showing variations in amount of ultrafiltrates obtained when unstandardized membranesare used
Five cubic centimeters of blood plasma was placed in each ultrafiltration sac. Timeof each experiment = 3 hours. Pressure 77 mm. Hg. (M) = male, (F) = female, (E) =
palpable edema.
Sac Ultra- Protein in CCase number number filtrate ultrafiltrate jClinical diagnosis
CC.
2 0.8 0 Acute nephritis (E) (M)1(Su) 4 2 .6 0 Acute nephritis (E)1S5 1 .9 0 Acute nephritis (E)
6 2.2 0 Acute nephritis (E)lO(G. T.) 17 2.3 ++ Acute nephritis (F)19 (C. N.) 30 2.45 0 Acute nephritis (E) (F)
31 1.9 0 Acute nephritis (E) (F)32 2.0 0 Acute nephritis33 1.4 0 Acute nephritis
20 (C. N.) 34 1.35 0 Acute nephritis35 1.3 0 Acute nephritis36 2.0 0 Acute nephritis37 1.7 0 Acute nephritis (E) (M)
l 38 1 .8 0 Acute nephritis (E)21 (J. C.) 39 2.2 0 Acute nephritis (E)
40 2.0 0 Acute nephritis41 2.0 0 Acute nephritis
3 (S. W.) 7 2.0 0 Normal (M)4 (A. J.) { 8 1.1 0 Normal (M)
5 (Y. W.) 10 2.7 0 Normal (F)3 (F. W.) 5 2.4 0 Normal (M)
2 (B. F.) { 6 216 0 Normal (M)14 (C. J.) 22 1.6 0 Normal (F)16 (A. J.) 24 1.5 0 Normal (F)
5 (Y. W.) 11 3.4 + Normal (F)15 (F. H.) 23 1.4 0 Nutritional edema (E) (M)11 (N. N.) { 18 2.0 0 Diabetes (M)
12 (N. A.) 2 2.0 0 Postpartum 8th day13 (V. C.) 21 1.5 0 Postpartum 9th day
6 (H. S.) lOa 3.8 ++ Postpartum 7th day25 1.8 + Toxemia of pregnancy
17 (R. B.) 26 1.3 0 Toxemia of pregnancy27 1.3 0 Toxemia of pregnancy
18 (R. B.) 28 1.3 0 Toxemia of pregnancy29 1 .8 0 Toxemia of pregnancy
6 (H. S.) 12 3.4 + Postpartum 8th day13 2.8 0 Hypertension (F)
8 (E. D.) 14 2.8 0 Hypertension
9. 16 2.6 0 Hypertension9 (D. M. C.) 16 2.8 0 Chronic nephritis (M)
580
MARGARETEMETA KUNDE
RESULTS
The data presented in table 1 show the amount of ultrafiltrateobtained from the plasma of normal individuals, patients withtoxemias of pregnancy, normal postpartum patients, and patientswith- chronic.and acute nephritis with and -wit-hout edema of cardiac orrenal origin.' The only group of this series that shows any degree ofconstancy in the amount of ultrafiltrate obtained is the toxemia ofpregnancy group. The most extreme degree of variation occursamong members of the other groups. Obviously, attempts to -estab-
TABLE 2
Variations in amount of ultrafiltrate obtained from the blood plasma of a dog, usingdiferent sacs
The duration of the experiment in each case is 3 hours. Amount of plasma placed ineach sac is 5 cc. Pressure 77 mm. Hg. The ultrafilters are not standardized.
Sac number Ultrafiltrate Protein in ultrafiltrate Percentile increase over
cc. per cent
la 3.5 ++ 412a 3.1 Very slight 253a 2.8 None 164a 3.3 None 375a 3.6 ++ 506a 2.4 None 07a 2.9 None 208a 3.0 Slight 259a 2.4 None 0
lOa 3.0 None 25Ila 3.2 None 3312a 3.6 + + 50
lish the amount of free water available from the blood plasma of normalindividuals, or to classify the findings from pathological sera in clinicalgroups on the basis of these results is entirely futile. Either theavailable water of the blood in conditions of health and in these
'The clinical cases studied in this report were all patients at the Cook CountyHospital, Chicago, Illinois. Wegratefully acknowledge our indebtedness to theinternes and resident physicians of this hospital. Thanks are especially due to Dr.H. A. Singer, whose cooperation made the clinical material used in this researchavailable.
581
5FREEWATEROF PLASMA
TABLE 3
The influence of adsorption by the membranes in reducing the amount of ultrafiltrates obtainedfrom membranes standardized but once, and used repeatedly
The amount of plasma used, pressure and time, are the same as in tables 1 and 2.All studies excepting those marked * were on blood plasma obtained from patients of thepathological obstetrical ward.
Date, 1925 -b w Case number Clinical diagnosis Cniino lrfle
~~~~~~I~~~~~~~~~
Toxemia of pregnancy.
Slight palpable edema
No edemaNo edemaSlight edemaSlight edemaNo edemaNo edemaNo edemaNo edemaNo edemaToxemia of pregnancy.
No edemaToxemia of pregnancy.
No edemaToxemia of pregnancy.
No edemaToxemia of pregnancy.
No edemaToxemia of pregnancy.
No edemaToxemia of pregnancy.
No edemaToxemia of pregnancy.
Marked edemaToxemia of pregnancy.
Marked edemaToxemia of pregnancy.
Marked edemaToxemia of pregnancy.
Marked edemaToxemia of pregnancy.
No edema
Used several times
Used several timesUsed several timesUsed once
Used once
Used several timesUsed several timesUsed several timesUsed several timesUsed several titesUsed several timesUsed several timesUsed several times
Used several times
Used several times
Used several times
Used several times
Used several times
Used once
Used once
Used several times
Used once
Used several times
March 31
March 31March 31April 8April 8April 23April 23May 2May 8May 8May 8May 15April 3
April 3
April 3
April 21
April 24
April 24
August 22
August 22
August 22
August 22
March 25
cc.
1.3
1.31.32.11.251.251.31.21.251.341.41.41.15
1.2
1.1
1.5
1.5
1.45
2.8
2.8
1.6
2.8
1.3
mgm.per cc.
55
73
88
89
7167
82
68
56
1 (R. B.)
1 (R. B.)1 (R. B.)1 (R.B.)1 (R. B.)1 (R. B.)1 (R.B.)1 (R. B.)1 (R. B.)1 (R. B.)1 (R. B.)1 (R. B.)2 (0. N.)
2 (0. N.)
2 (0. N.)
2 (0. N.)
2 (0. N.)
2 (0. N.)
3 (E. D.)
3 (E. D.)
3 (E. D.)
3 (E. D.)
4 (A. B.)
582
MARGARETEMETA KUNDE
TABLE 3-Continmcd
.10
Date, 1925 2 Case number Clinical diagnosis Condition of ultrafilter- 0AWQZ _ _ _ _ _ _
Toxemia of pregnancy.No edema
Toxemia of pregnancy.No edema
Slight edemaNo edemaNo edemaNo edemaToxemia of pregnancyToxemia of pregnancyToxemia of pregnancyToxemia of pregnancyHypertensionNormalNormalNormalEclampsiaEclampsiaEclampsia. No edemaEclampsia. No edemaEclampsia. No edemaEclampsia. No edemaNormal pregnancyNormal pregnancyNormal pregnancyNormal pregnancyNormal pregnancyNormal pregnancyNormal pregnancyNormal pregnancyNormalNormalNormalNormalNormalNormalNormalNormalNormal
Used several times
Used several times
Used several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several timesUsed several times
mgm.per cc.
7074
596162
72
56
65
67
98
68
March 25
March 25
April 2April 6April 6April 6May 12May 26May 26June 6June 2July 10July 10July 10July 8July 8April 28April 28April 28May 6May 1May 1May 11May 1
April 13April 16April 18August 25May 20May 27May 28May 29June 30June 30June 30May 10June 1
cc.
1.3
1.3
1.31.01.11.21.51.01.51.0'1.02.52.62.82.32.151.21.11.11.11.11.31.71.11.61.61.31.81.41.51.01.31.21.21.21.51.6
4 (A. B.)
4 (A. B.)
4 (A. B.)4 (A. B.)4(A. B.)4 (A. B.)5 (B. M.)6 (L. W.)7 (E. G.)7 (L. G.)8 (L. B.)9 (L. C.)9 (L. C.)9 (L. C.)
10 (A. B.)10 (A. B.)10 (V. L.)10 (V. L.)10 (V. L.)11 (D. E.)12 (N. N.)*12 (N. N.)*13 (A. A.)*14 (P. B.)*15 (N.P.)*15 (N. P.)*16 (F. F.)*17 (F. U.)*18 (C. J.)*18(C. C.)*19 (A. R.)*20 (M. B.)*21 (B. E.)*21 (B. E.)*21 (B. E.)*22 (E. G.)*23 (L. B.)*
-
583
FREE WATEROF PLASMA
diseases is exceedingly variable, or sufficient uniformity in the per-meability of the membranes made according to the method describedabove, had not been attained.
This second possibility was investigated, and the results of anexperiment planned to test the uniformity of the ultrafiltration sacsare given in table 2. In this test 12 sacs were used; 5 cc. of bloodplasma from the same dog was placed in each sac and the sac fitted tothe pressure apparatus. At the end of 3 hours a 37 per cent variationwas found in the total amount of protein-free ultrafiltrate deliveredfrom the several sacs. Obviously this difference was due, not to achange in the "free" water content of the blood plasma of the dog, butto differences in the ultrafiltration membranes.
Recognizing the need of a more constant degree of permeability inour membranes, we attempted to eliminate variations in the thicknessby using a gel (7 per cent glacial acetic acid solution of parlodion) ofsuch consistency that the paper Soxhlet thimbles were completelyimpregnated in 2.5 to 3 minutes. They were then drained anddried for 8 minutes, immersed in the gel, drained again for 5 minutes,fixed in distilled water and washed as already described. The mem-branes were then standarized by placing 5 cc. of distilled water in eachand fitting into the pressure apparatus. Only those were chosen foruse that delivered 5 cc. of distilled water in 75 to 90 minutes, at apressure of 77 mg. Hg. Such thimbles delivered 2.8 to 3.0 cc. ofprotein-free filtrate from 5 cc. of oxalated plasma from normalindividuals or dogs in 165 to 180 minutes.
Table 3 shows the amount of ultrafiltrate obtained from a series ofwomen, some of whom were in normal state of health, others weresuffering from toxemia of pregnancy, or were normal postpartumpatients. The membranes used in the experiments of this series wereall standardized with distilled water and then tested with normalserum as mentioned above. After this, in most instances, they wereused to test the amount of ultrafiltrate that could be obtained from theblood plasma of nephritics or patients with deferred diagnosis. Theywere then used in the series of cases reported in table 3. These dataare important only in showing how consistently false results may beobtained because of adsorption by the gel of colloidal substances inthe plasma.
584
MARGARETEMETA KUNDE
TABLE 4
Data selected from experiments in which standardized membranes are usedRecalibration shows little or no change in the membranes during the experiment.* (M)
= male; (F) = female. Time, pressure and amount of plasma are the same as in table 1.
Dis-Total tilled
Ultra-protein ~~~~~waterCasenjmber Date,1925 Uiltra-e prtin Clinical diagnosis deliv-Casenumber Date, filtrate sin eredaftertest
CC.
2.1
2.1
2.8
2.8
2.62.8
2.3
2.15
2.62.6
2.82.7
3.0
3.1
2.5
2.83.02.72.5
3.0
3.2
mgm.per cc.
73
68
72
56
63
88
100
83
525160
62
Toxemia of pregnancy.Slight palpable edema
Toxemia of pregnancy.Slight palpable edema
Toxemia of pregnancy
Toxemia of pregnancy
Acute nephritis.edema (M)
Marked
Acute nephritis
Chronic nephritis andnicious anemia (M)
CC.
4.7
4.7
5
55
4.8
4.7
4.94.9
55
per- 5
5
Acute nephritis. Edema (M)j 4.8
Acute nephritis. Edema (M)
Chronic nephritis (F)
554.44.4
5
August 19
1 (R.B.) {
2 (E.D.)
3(L.C.) {
4 (A. B.)
April 8
April 8
August 22
August 22
July 10July 10
July 8
July 14
July 14June 18
June 8June 8
August 19
August 11
March 14
May 18August 22September 29September 29
August 20
5 (Su)
6 (G. T.)
7 (Mi)
8 (C. N.)
l
9 (J. C.)
10 (L. D.)
11 (N. N.)
585
i
I(M) 5
FREE WATEROF PLASMA
TABLE 4-Concluded
Case number
12 (C. T.)
I13 (C. B.)
14 (Y. W.)
Date, 1925
August 24August 24August 24
September 15September 15
August 26
August 26
15 (H. S.) August 31
16 (A. J.)
17 (D. H.)
August 25August 25
f April 6l April 6
Ultra-filtrate
cC.
3.13.23.1
2.52.5
2.73.0
3.4
2.82.8
2.32.35
Totalprotein
inplasma
mgm.per cc.
68
81
Clinical diagnosis
Acute nephritis. Edema
Hypertension (F)
Normal (M)
Normal. Postpartum
Normal
Normal (M)
* Protein was present in the ultrafiltrates of cases 10 (L. D.) and 15 (H.S.).
Critical examination of the technique and further analysis andexperimentation showed that the low values were due to lessenedpermeability of the membranes, and that these ultrafilters had changedduring the process of using, and would no longer deliver 5 cc. of dis-tilled water in 90 minutes. This tendency to change in permeabilitymakes it necessary to know the degree of permeability of the mem-
branes at the end of each experiment as well as at the beginning. We-find that the membranes usually become changed as a result of the
first ultrafiltration of blood plasma, so that instead of delivering 5 cc.
of distilled water in 90 minutes, they deliver 5 cc. of distilled water intwo hours; therefore two hours becomes the standard for subsequentrecalibration tests. The membranes may become "set" at this point;if so they may be repeatedly used without a measurable reduction inthe permeability. But they must be tested after each using withdistilled water, since changes may occur after the second or subseqiient
586
Dis-tilled
waterdeliv-eredaftertest
cc.
555
55
55
5
55
4.74.7
MARGARETEMETA KUNDE
using. When sufficient adsorption has occurred to reduce the per-meability of the membranes about one-half they may remain remark-ably constant for a number of experiments, or the permeability mayslowly continue to decrease or suddenly increase. A calibration testmade with distilled water reveals their condition.
Table 4 gives the results obtained from a series of 33 ultrafiltrationexperiments made with the blood plasma of 17 subjects, includingnormal men and women, patients with nephritis with and withoutedema and hypertension, and patients suffering from toxemias ofpregnancy. These results are chosen from experiments where re-calibration of the membranes with distilled water showed that thepermeability of the membranes had remained relatively constantduring the experiment. The data show that there is no significantdifference in the quantity of ultrafiltrate obtained from these patientsor normal individuals, regardless of the amount of edema or nephritispresent.
DISCUSSION
The amount of ultrafiltrate obtained by filtering serum throughmembranes made according to the technique advocated by Beckmann,is of no significance unless the limits of effectiveness of the ultrafiltersare definitely known. The permeability of the membranes dependslargely upon the amount of gel entering into the composition and atpresent no method for exactly controlling this suggests itself.
Walpole (7) showed, by chemical analysis, that the amount of gel inadjacent 5 cm. strips of membrane varies appreciably even where themembranes are made after the most accurate and painstakng method,and no paper support is introduced. Furthermore, micrometermeasurements show considerable differences in the thicknesses ofsuch strips. In the method used by Beckmann, no factor in thetechnique is controlled excepting the length of time taken to im-pregnate the paper Soxhlet thimbles, and that is given a leeway of 3 to5 minutes. The paper thimbles vary at different points. This givesa base irregularly impregnated with gel and hence with differencesin porosity at different points. Other conditions which we have foundto cause changes which reduce the amount of ultrafiltrate are: first,the degree of drying of the membranes before fixing in water; second,
58i7
FREE WATEROF PLASMA
swelling of the gel which occurs in some cases, after 12 hours of washingin water, so that membranes used only for experiments with distilledwater deliver lessened amounts of ultrafiltrate after several days;third, exposure to drying during the preparation for an experiment;fourth, the adsorption of colloidal substances by the gel.
The data submitted in table 2 show that uniform results cannot beobtained from blood plasma of the same subject where the abovefactors are operating in an unknown way. The 12 membranes used inthe series of table 2 had not been previously used, so the 50 per centdifference found in the amounts of ultrafiltrate obtained from the sameblood is due either to changes in the permeability that occurred duringthe process of making, or to differences in the amounts of adsorptionwhich took place during the experiment.
Proper emphasis must be placed upon the important factor ofadsorption. If our work had been interrupted at the point where theinvestigations of table 3 were completed, we could have advocatedthe false conclusion that the water available, by methods of ultra-filtration, from the blood plasma of normal women, postpartumpatients and patients with toxemias of pregnancy, is 40 to 50 per centless than that available from normal men. The results were obtainedfrom membranes used several times without recalibration.
Comparable amounts of ultrafiltrate may be obtained from mem-branes of the same calibration, but recalibration is necessary aftereach experiment in order to make sure that the permeability has notchanged during the experiment.
Standardized membranes, recalibrated, were used for the experi-ments given in table 4. These results indicate that the quantity ofultrafiltrate obtained in three hours with pressure of 77 mm. Hg fromblood plasma of normal individuals, postpartum patients (both un-complicated and of the toxemia group), chronic and acute nephritiswith and without palpable edema, does not vary to any significantdegree. Further evidence supporting these results was obtained fromthe following experiments which differs from those previously reported,only in that the time for the ultrafiltration was extended until no morefluid was delivered from the membranes (9 to 15 hours). The bloodplasma from 10 subjects was tested for "free" water content accordingto the process of ultrafiltration. The subjects included 3 patients with
588
MARGARETEMETA KUNDE
chronic nephritis (2 had moderate edema), 1 with acute nephritisaccompanied by marked edema, 1 with essential hypertension and 5normal individuals. Twenty ultrafiltration experiments were made inthis series, using standardized and recalibrated membranes. Thetotal amounts of ultrafiltrates delivered from the 5 cc. portions ofplasma varied from 4.3 to 4.7 cc. Only one sac delivered as small anamount as 4.3 cc. and on recalibration with distilled water, the per-meability of this sac was found to be markedly reduced. Webelievethe slight differences in the amounts of ultrafiltrate delivered by theother sacs to be due to undetectable changes in the permeability ofthe membranes. Wedo not offer these results as evidence that theavailable water of the blood plasma may not vary under some condi-tions. Webelieve, however, that such changes cannot be detectedby this method.
Ultrafiltration, according to Bechhold, is not a method for detectingthe amount of water available, but for separating colloidal particles ofdifferent sizes. The two processes are no doubt closely related, but notidentical. For example, membranes of a given porosity will hold backall colloidal particles above a certain size regardless of the amountof fluid filtered through; the lower limit of permeability for a givencolloidal partide is well 'defined. In measuring water the degree ofrefinement in the technique is infinitely more difficult, because mole-cules of water filter through when the porosity is entirely too small topass other substances found in the blood plasma. If 5 cc. of distilledwater is placed in a membrane of diminished porosity, only a smallamount of ultrafiltrate is obtained at 77 mm. Hg and 3 hours time.If the same amount of water is placed in a membrane of slightlygreater porosity, it will completely filter through (length of time andpressure as above). Using the same membranes and reducing thepressure to 33 mm. Hg (approximately the hydrostatic pressure of thecapillaries) only 80 per cent of the distilled water passed through atthe end of 9 to 15 hours. Obviously this is not a test of the amount offree water in the contents of the filters, but a failure in some cases of thegel membranes to give rapid passage to free water under a given hydrostaticpressure because of the fineness of the pores. The same condition holdsto a great degree in experiments with blood plasma.
Wefind no indication of marked physico-chemical changes in the
589
FREE WATEROF PLASMA
plasma proteins in cases of nephritis with or without edema, at least asindicated by the ultrafiltration of the plasma. Results in table 4were obtained by standardized membranes. No protein moleculespass through where the quantity of ultrafiltrate measures 2.8 to 3.0 cc.or less. If the permeability of the membranes is but slightly in-creased so that the amount of fluid delivered is increased by 0.4 cc.the ultrafiltrate gives a slight positive protein test2 in all conditionstested. It does not seem probable that changes in size of the colloidsthrough this small range could account for the binding of water by theblood protein described by Beckmann. However, one must considerthe possibility of various proteins of the blood swelling differently.Hydrostatic pressure equal to 77 mm. Hg used in Beckmann's and myown experiments may tend to overcome the power of the slightlyswollen colloids to hold water, hence at such pressure slight differencesin amount of ultrafiltrate due to the hydration processes of the colloidsmay not be detected.
The nature of the proteins in solution influences the amount ofultrafiltrate delivered much more than do slight quantitative varia-tions found in blood plasma. For example, 5 cc. of a 7 per cent solu-tion of gelatin yields only about 0.7 cc. of ultrafiltrate from thestandard membrane (pressure and time as in table 1), whereas a 7 percent solution of colloids as found in blood plasma yields 2.8 to 3.0 cc.
SUMMARY
1. The amount of ultrafiltrate passing through a gel membranein a given time and at a given pressure is of little significance unlessthe permeability of the membrane is definitely known.
2. Gel membranes should be standardized both before and aftereach experiment to detect changes which may occur during the experi-ment.
3. Ultrafiltration does not measure the free water of the substancefiltered, but the speed with which the membrane can give passage tofree water under a given hydrostatic pressure.
4. When standardized membranes were used in a system with ahydrostatic pressure equal to 77 mm. of Hg the amount of ultrafiltrate
2Nitric acid ring test (Heller).
590
MARGARETEMETA KUNDE
obtained in three hours did not vary significantly whether obtainedfrom the blood plasma of normal men and women or of patientssuffering from nephritis, with or without edema, or toxemias of preg-nancy, or of postpartum patients without complications.
5. Wefailed to find, by this means, evidence either supporting orrefuting the theory that swelling or shrinking of the plasma colloidsmay occur to a degree sufficient to influence the free water content ofthe blood plasma in patients with and without edema.
BIBLIOGRAPHY
1. Fischer, M. H.: Edema and Nephritis. John Wiley and Sons, N. Y., 1921,3rd ed.
2. Bechmann, K.: Deut. Arch. Klil. Med., 1924, cxlv, 22. tJber das BlutodemUltrafiltrationsversuche amBlutserum Odematoser.
3. Ellinger, A., and Neuschlosz, S. M.: Biochem. Zeitsch., 1922, cxxvii; 241.Vergleichende Untersuchungen uber Viscositat und Ultrafiltrationsge-schwindigkeit von Serum.
4. Bechhold, H., and Bullowa, J. G. M.: Colloids in Biology and Medicine. D.Van Nostrand and Co., N. Y., 1919.
5. Zilva, S. S., and Miura, M.: Biochem. Jour., 1921, xv, 422. XLVIII. TheDifferential Dialysis of the Antineuritic and the Antiscorbutic Factors.
6. Cushny, A. R.: Jour. Physiol., 1919, liii, 391. The Colloid-Free Filtrate ofSerum.
7. Walpole, G. S.: Biochem. Jour., 1915, ix, 284. XXVI. Notes on CoilodionMembranes for Ultrafiltration and Pressure Dialysis.
591