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EFFECTS OF SULPHONAMIDES ON SERUM-PROTEIN, PLASMA VISCOSITY, AND ERYTHROCYTE-SEDIMENTATION RATE
298
reported with adrenocorticotropic hormone. Our experience with progesterone confirms this. Venning (1946) reported a considerable rise in excretion of corticoids in pregnancy. In the light of recent reports
that
on
the beneficial effect of cortisone it therefore
seems
likely that remissions of rheumatoid arthritis occurring during pregnancy are due to increased production not of progesterone but of steroids with a cortisone-like action. It is of interest, however, to note that the metabolism of progesterone in rheumatoid arthritis appears to be abnormal. After the injection of progesterone the quantity of pregnanediol recoverable from the urine is greater in patients with rheumatoid arthritis than in normal persons (Sommerville et al. 1950). SUMMARY
doses of progesterone were administered to 5 with rheumatoid arthritis. No evidence of therapeutic benefit was obtained. No fall in the number of circulating eosinophils was noted after the injection of 100 mg. of progesterone. It is suggested that the remissions of rheumatoid arthritis which occur during pregnancy are not due to progesterone but may b’e attributable to increased production of other steroids by the adrenal cortex.
Large patients
We are indebted to Prof. L. S. P. Davidson for his criticism and advice in the preparation of this report.
helpful
REFERENCES
Browne, J. S. L., Henry, J. S., Venning, E. H. (1937) J. clin. Invest. 16, 678. Hench, P. S. (1938) Proc. Mayo Clin. 13, 161. Kendall, E. C., Slocumb, C. H., Polley, H. F. (1949) Ibid, 24, 181. Reich, N. E. (1949) Amer. Practitioner, 4, 1. Sommerville, I. F., Marrian, G. F., Duthie, J. J. R., Sinclair, R. J. G. (1950) Lancet, Jan. 21, p. 116. Spies, T. D., Stone, R. E. (1949) Ibid, ii, 890. Thorn, G. W., Bayles, T. B. (1949) Practitioner, 163, 365. Forsham, P. H., Prunty, F. T. G., Hills, A. G. (1948) J. Amer. med. Ass. 137, 1005 Touw, J. F., Kuipers, R. K. W. (1938) Acta med. scand. 96, 501. Venning, E. H. (1946) Endocrinology, 39, 203. Browne, J. S. L. (1936) Proc. Soc. exp. Biol., N.Y. 34, 792. Henry, J. S., Browne, J. S. L. (1937) Canad. med. Ass. J. 36, 83. —
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EFFECTS OF SULPHONAMIDES ON SERUMPROTEIN, PLASMA VISCOSITY, AND
ERYTHROCYTE-SEDIMENTATION RATE JOHN HARKNESS M.B., B.Sc. Glasg. CHEMICAL
PATHOLOGIST, CENTRAL LABORATORY, PORTSMOUTH
THE effects of ordinary therapeutic doses of sulphanilamide and sulphapyridine on the serum-protein levels, plasma viscosity, and erythrocyte-sedimentation rate (E.s.R.) have been investigated in normal persons. MATERIAL AND METHODS
The subjects of the experiments were apparently normal men, aged 20-63, but they were not physically examined. Their lives were restricted to a routine of so as to reduce other work, exercise, food, sleep, &c.. a stimuli to minimum and to allow the physiological successive blood samples to be removed under almost identical conditions. To reduce the emotional stimuli, the men were not told on which days blood would be removed, and they were not likely to be mentally disturbed by taking tablets. Six men were given 2 g. of sulphanilamide by mouth, followed in 4 hours by another 2 g., and thereafter by 1 g. 4-hourly until a total of 20 g. had been administered.
With another six men the same procedure was carried out with sulphapyridine. Venous blood was drawn, with minimal stasis, from the fasting men at the same hour of the day. Part of the blood was allowed to clot to produce serum, which was later separated by centrifuging ; and part was used to make an accurate 4 : 1 mixture with 3-8% sodium citrate solution. Clinical pathological estimations were made of (1) serum total protein, by the falling-drop specific-gravity technique of Barbour and Hamilton (1926) ; (2) plasma viscosity by the technique of Houston et al. (1945); (3) serum viscosity by the same technique ; (4) packedcell-volume (P.c.v.) by centrifuging citrated blood in capillary hæmatocrit tubes for 1 hour at 3000 r.p.m. ; and (5) maximal corrected E.s.R. by the method described by Houston et al. (1945). RESULTS
The were
sulphanilamide and sulphapyridine only the sulphanilamide results are Figs. 1 and 2 show these results in graphical
effects
identical,
presented.
of
so
form. The serum-protein, plasma viscosity, serum viscosity, and E.s.R. had all increased within 48 hours of the administration of the sulphanilamide. (The sulphapyridine tests were done at different time intervals, and tne
increases
evident within 24 hoursof giving that drug.) That these increases are not due to hæmoconcentration can be ascertained from the P.c.v. values, in which the variation is smaller and were
inconstant, and by making allowance for this variation in calculating the changes in these other values (as has been done for the serum-
protein only in fig. 1). No work has previously been published on the daily variations in
the plasma and viscosities in response
serum
to
ordinary physiological stimuli, but my
experi-
indicates that such variations should Fig. I-Variations in plasma viscosity, serum visbe less than cosity, packed cell volume, and serum-protein of sulphanilamide. Results are on exhibition of the as expressed percentages of values before original value. administration of sulphanilamide. The lines join The changes the means of the experimental results. ence
1%
299 in every instance in this series were greater than 1% ; and in several instances the change was sufficiently great to raise the plasma viscosity out of the range usually described as normal into the range which one associates with the reaction to organic changes in the body. DISCUSSION
The present series of liave not had any further
12
tests
is
opportunity
small, but I
for
experiments
with
drugs
these suit-
on
able subjects under suitable basal conditions. The results are offered to draw attention to an effect of the
sulphonamides which has not Fig. 2—Variation in erythrocyte-sedimentation previously been rate on exhibition of sulphanilamide. Results described but are expressed as percentages of value before administration of sulphanilamide. The line seems worthy of further investijoins the means of the experimental results. gation. The findings can be discussed from two aspects.
(1) Effects of Sulphonamides
Plasma and Serum The interaction of the sulphonamides with the bloodprotein has been described by Davis (1942), but there has been no mention of any action exerted by them on the amount of the proteins. The increase in serum-protein can be detected within 24 hours of giving sulphonamides and disappears within 3 days of stopping them. From fig. 1 it seems that the effect on the second day was greater than on the third day-i.e., the protein was beginning to fall though sulphanilamide was still being given. This ’opinion is supported by previous work (Harkness, unpublished) in which the sulphanilamide was administered in similar doses to ten normal persons for 10 days and the E.s.R. (Westergren) increased slightly and then returned to normal while the drug was still being exhibited ; no such changes occurred when the same routine was carried through without giving on
sulphanilamide. The effect on the protein must be regarded as the primary change in the blood constituents, of which the plasma viscosity, the serum viscosity, and the The relationship of the E.s.R. are but reflections. viscosity to the protein has been discussed elsewhere (Houston et al. 1945, Harkness et al. 1946, Harkness and -Whittington 1947), as also has the relationship of the E.S.R. and protein (Bendien and Snapper 1931, Fraser and Rennie 1941). Further metabolic studies should show whether the protein increase is due to increased protein formation or decreased utilisation.
(2) Significance of Clinical Pathological Tests These results also indicate that there is a danger in accepting the changes in the laboratory tests during an illness as being due entirely to changes in the disease since sulphonamides alone can produce significant alterations in plasma viscosity, serum viscosity, and E.s.R. Thus, in a case of pyrexia of unknown origin treated empirically with sulphonamides (admittedly bad therapeutics) the findings after 24 hours might lead to a long search for an organic lesion. Furthermore, the fall in these values after the second or third day, whether the sulphonamides were stopped or continued could be interpreted as an improvement in a non-existent disease. Similarly there has been a division of opinion over the action of salicylates in rheumatic diseases. While
some workers-e.g., Reid (1948)-claim that the E.S.R. is so reliable a test that they are willing to use it as the criterion of amelioration in the rheumatic process, others claim that the salicylates have a direct depressing action on the E.s.R., but no mechanism for effecting the depression has yet been demonstrated. Before the results of clinical pathological tests can be used for ascribing certain actions to drugs in disease,. the effect of the drugs on these tests should be fully studied in normal persons. Nevertheless, there is no guarantee that, in the presence of a disease for which the drug is given, reactions identical with the " normal ones will be educed.
SUMMARY
The exhibition of sulphanilamide and sulphapyridine to normal persons increased the serum total protein, the plasma viscosity, the serum viscosity, and the E.S.R. This effect requires further study, but it should be taken into consideration in assessing the significance of changes in these tests in patients under treatment with.
sulphonamides. REFERENCES
Barbour, H. G., Hamilton, W. F. (1926) J. biol. Chem. 69, 625. Bendien, W. M., Snapper, I. (1931) Biochem. Z. 235, 14. Davis, B. D. (1942) Science, 95, 78. Fraser, T. N., Rennie, J. B. (1941) Brit. J. exp. Path. 22, 81. Harkness, J., Houston, J., Whittington, R. B. (1946) Brit. med. J. i, 268. Whittington, R. B. (1947) Biochim. Biophys. Acta, 1, 487. Houston, J., Harkness, J., Whittington, R. B. (1945) Acta tuberc. scand. 19, 153. Reid, J. (1948) Quart. J. Med. 17, 139. -
THE MALE FROG (Rana esculenta) PREGNANCY TEST AND ITS CLINICAL
APPLICATION J. BIENIARZ M.D. Gdansk ASSISTANT,
OBSTETRICS
AND
GYNÆCOLOGY
ACADEMY, GDANSK,
CLINIC,
MEDICAL
POLAND
THERE is a considerable difference of opinion about the value of the male frog, Rana esculenta, in the Galli Nlainini (1948) test. Hinglais and Hinglais (1948), Biernacka and Pigon (1948), Fotin et al. (1948), Hernandez Andueza (1948), Aznar Ferreres (1948), and Bach et al. (1949) viewed it enthusiastically, whereas Schockaert et al. (1948), and Hinglais and Hinglais (1949) thought poorly of it. To assess its value 312 tests were made between December, 1948, and March, 1949-200 with pregnancy urine (114 from normal pregnancies, and 86 from abnormal pregnancies) and 112 controls with urin0 from non-pregnant persons, all patients in our obstetrics and gynaecology clinic. The pregnancy tests were classified into (1). those done in early (first sixteen weeks) normal pregnancy ; (2) those done in advanced normal pregnancy ; (3) those done in early (first sixteen weeks) abnormal pregnancy ; and those done in advanced abnormal pregnancy. In this way we could compare results in our groups not only with one another (to see whether they were influenced by the duration of pregnancy or by its abnormal character) but also with those of other workers, who had almost all worked with urine from cases of early normal pregnancy, when the need for a laboratory diagnosis is most imperative. ’
METHOD
We followed Galli Mainini’s (1948) usual technique, an amount of urine equal to a twentieth of the frog’s body-weight was found sufficient. In this. way positiveresults of varied intensity were obtained, which could be estimated by an approximate count-of the spermatozoa in a of cloacal urine under
except that
droplet
reported with adrenocorticotropic hormone. Our experience with progesterone confirms this. Venning (1946) reported a considerable rise in excretion of corticoids in pregnancy. In the light of recent reports
that
on
the beneficial effect of cortisone it therefore
seems
likely that remissions of rheumatoid arthritis occurring during pregnancy are due to increased production not of progesterone but of steroids with a cortisone-like action. It is of interest, however, to note that the metabolism of progesterone in rheumatoid arthritis appears to be abnormal. After the injection of progesterone the quantity of pregnanediol recoverable from the urine is greater in patients with rheumatoid arthritis than in normal persons (Sommerville et al. 1950). SUMMARY
doses of progesterone were administered to 5 with rheumatoid arthritis. No evidence of therapeutic benefit was obtained. No fall in the number of circulating eosinophils was noted after the injection of 100 mg. of progesterone. It is suggested that the remissions of rheumatoid arthritis which occur during pregnancy are not due to progesterone but may b’e attributable to increased production of other steroids by the adrenal cortex.
Large patients
We are indebted to Prof. L. S. P. Davidson for his criticism and advice in the preparation of this report.
helpful
REFERENCES
Browne, J. S. L., Henry, J. S., Venning, E. H. (1937) J. clin. Invest. 16, 678. Hench, P. S. (1938) Proc. Mayo Clin. 13, 161. Kendall, E. C., Slocumb, C. H., Polley, H. F. (1949) Ibid, 24, 181. Reich, N. E. (1949) Amer. Practitioner, 4, 1. Sommerville, I. F., Marrian, G. F., Duthie, J. J. R., Sinclair, R. J. G. (1950) Lancet, Jan. 21, p. 116. Spies, T. D., Stone, R. E. (1949) Ibid, ii, 890. Thorn, G. W., Bayles, T. B. (1949) Practitioner, 163, 365. Forsham, P. H., Prunty, F. T. G., Hills, A. G. (1948) J. Amer. med. Ass. 137, 1005 Touw, J. F., Kuipers, R. K. W. (1938) Acta med. scand. 96, 501. Venning, E. H. (1946) Endocrinology, 39, 203. Browne, J. S. L. (1936) Proc. Soc. exp. Biol., N.Y. 34, 792. Henry, J. S., Browne, J. S. L. (1937) Canad. med. Ass. J. 36, 83. —
—
-
-
EFFECTS OF SULPHONAMIDES ON SERUMPROTEIN, PLASMA VISCOSITY, AND
ERYTHROCYTE-SEDIMENTATION RATE JOHN HARKNESS M.B., B.Sc. Glasg. CHEMICAL
PATHOLOGIST, CENTRAL LABORATORY, PORTSMOUTH
THE effects of ordinary therapeutic doses of sulphanilamide and sulphapyridine on the serum-protein levels, plasma viscosity, and erythrocyte-sedimentation rate (E.s.R.) have been investigated in normal persons. MATERIAL AND METHODS
The subjects of the experiments were apparently normal men, aged 20-63, but they were not physically examined. Their lives were restricted to a routine of so as to reduce other work, exercise, food, sleep, &c.. a stimuli to minimum and to allow the physiological successive blood samples to be removed under almost identical conditions. To reduce the emotional stimuli, the men were not told on which days blood would be removed, and they were not likely to be mentally disturbed by taking tablets. Six men were given 2 g. of sulphanilamide by mouth, followed in 4 hours by another 2 g., and thereafter by 1 g. 4-hourly until a total of 20 g. had been administered.
With another six men the same procedure was carried out with sulphapyridine. Venous blood was drawn, with minimal stasis, from the fasting men at the same hour of the day. Part of the blood was allowed to clot to produce serum, which was later separated by centrifuging ; and part was used to make an accurate 4 : 1 mixture with 3-8% sodium citrate solution. Clinical pathological estimations were made of (1) serum total protein, by the falling-drop specific-gravity technique of Barbour and Hamilton (1926) ; (2) plasma viscosity by the technique of Houston et al. (1945); (3) serum viscosity by the same technique ; (4) packedcell-volume (P.c.v.) by centrifuging citrated blood in capillary hæmatocrit tubes for 1 hour at 3000 r.p.m. ; and (5) maximal corrected E.s.R. by the method described by Houston et al. (1945). RESULTS
The were
sulphanilamide and sulphapyridine only the sulphanilamide results are Figs. 1 and 2 show these results in graphical
effects
identical,
presented.
of
so
form. The serum-protein, plasma viscosity, serum viscosity, and E.s.R. had all increased within 48 hours of the administration of the sulphanilamide. (The sulphapyridine tests were done at different time intervals, and tne
increases
evident within 24 hoursof giving that drug.) That these increases are not due to hæmoconcentration can be ascertained from the P.c.v. values, in which the variation is smaller and were
inconstant, and by making allowance for this variation in calculating the changes in these other values (as has been done for the serum-
protein only in fig. 1). No work has previously been published on the daily variations in
the plasma and viscosities in response
serum
to
ordinary physiological stimuli, but my
experi-
indicates that such variations should Fig. I-Variations in plasma viscosity, serum visbe less than cosity, packed cell volume, and serum-protein of sulphanilamide. Results are on exhibition of the as expressed percentages of values before original value. administration of sulphanilamide. The lines join The changes the means of the experimental results. ence
1%
299 in every instance in this series were greater than 1% ; and in several instances the change was sufficiently great to raise the plasma viscosity out of the range usually described as normal into the range which one associates with the reaction to organic changes in the body. DISCUSSION
The present series of liave not had any further
12
tests
is
opportunity
small, but I
for
experiments
with
drugs
these suit-
on
able subjects under suitable basal conditions. The results are offered to draw attention to an effect of the
sulphonamides which has not Fig. 2—Variation in erythrocyte-sedimentation previously been rate on exhibition of sulphanilamide. Results described but are expressed as percentages of value before administration of sulphanilamide. The line seems worthy of further investijoins the means of the experimental results. gation. The findings can be discussed from two aspects.
(1) Effects of Sulphonamides
Plasma and Serum The interaction of the sulphonamides with the bloodprotein has been described by Davis (1942), but there has been no mention of any action exerted by them on the amount of the proteins. The increase in serum-protein can be detected within 24 hours of giving sulphonamides and disappears within 3 days of stopping them. From fig. 1 it seems that the effect on the second day was greater than on the third day-i.e., the protein was beginning to fall though sulphanilamide was still being given. This ’opinion is supported by previous work (Harkness, unpublished) in which the sulphanilamide was administered in similar doses to ten normal persons for 10 days and the E.s.R. (Westergren) increased slightly and then returned to normal while the drug was still being exhibited ; no such changes occurred when the same routine was carried through without giving on
sulphanilamide. The effect on the protein must be regarded as the primary change in the blood constituents, of which the plasma viscosity, the serum viscosity, and the The relationship of the E.s.R. are but reflections. viscosity to the protein has been discussed elsewhere (Houston et al. 1945, Harkness et al. 1946, Harkness and -Whittington 1947), as also has the relationship of the E.S.R. and protein (Bendien and Snapper 1931, Fraser and Rennie 1941). Further metabolic studies should show whether the protein increase is due to increased protein formation or decreased utilisation.
(2) Significance of Clinical Pathological Tests These results also indicate that there is a danger in accepting the changes in the laboratory tests during an illness as being due entirely to changes in the disease since sulphonamides alone can produce significant alterations in plasma viscosity, serum viscosity, and E.s.R. Thus, in a case of pyrexia of unknown origin treated empirically with sulphonamides (admittedly bad therapeutics) the findings after 24 hours might lead to a long search for an organic lesion. Furthermore, the fall in these values after the second or third day, whether the sulphonamides were stopped or continued could be interpreted as an improvement in a non-existent disease. Similarly there has been a division of opinion over the action of salicylates in rheumatic diseases. While
some workers-e.g., Reid (1948)-claim that the E.S.R. is so reliable a test that they are willing to use it as the criterion of amelioration in the rheumatic process, others claim that the salicylates have a direct depressing action on the E.s.R., but no mechanism for effecting the depression has yet been demonstrated. Before the results of clinical pathological tests can be used for ascribing certain actions to drugs in disease,. the effect of the drugs on these tests should be fully studied in normal persons. Nevertheless, there is no guarantee that, in the presence of a disease for which the drug is given, reactions identical with the " normal ones will be educed.
SUMMARY
The exhibition of sulphanilamide and sulphapyridine to normal persons increased the serum total protein, the plasma viscosity, the serum viscosity, and the E.S.R. This effect requires further study, but it should be taken into consideration in assessing the significance of changes in these tests in patients under treatment with.
sulphonamides. REFERENCES
Barbour, H. G., Hamilton, W. F. (1926) J. biol. Chem. 69, 625. Bendien, W. M., Snapper, I. (1931) Biochem. Z. 235, 14. Davis, B. D. (1942) Science, 95, 78. Fraser, T. N., Rennie, J. B. (1941) Brit. J. exp. Path. 22, 81. Harkness, J., Houston, J., Whittington, R. B. (1946) Brit. med. J. i, 268. Whittington, R. B. (1947) Biochim. Biophys. Acta, 1, 487. Houston, J., Harkness, J., Whittington, R. B. (1945) Acta tuberc. scand. 19, 153. Reid, J. (1948) Quart. J. Med. 17, 139. -
THE MALE FROG (Rana esculenta) PREGNANCY TEST AND ITS CLINICAL
APPLICATION J. BIENIARZ M.D. Gdansk ASSISTANT,
OBSTETRICS
AND
GYNÆCOLOGY
ACADEMY, GDANSK,
CLINIC,
MEDICAL
POLAND
THERE is a considerable difference of opinion about the value of the male frog, Rana esculenta, in the Galli Nlainini (1948) test. Hinglais and Hinglais (1948), Biernacka and Pigon (1948), Fotin et al. (1948), Hernandez Andueza (1948), Aznar Ferreres (1948), and Bach et al. (1949) viewed it enthusiastically, whereas Schockaert et al. (1948), and Hinglais and Hinglais (1949) thought poorly of it. To assess its value 312 tests were made between December, 1948, and March, 1949-200 with pregnancy urine (114 from normal pregnancies, and 86 from abnormal pregnancies) and 112 controls with urin0 from non-pregnant persons, all patients in our obstetrics and gynaecology clinic. The pregnancy tests were classified into (1). those done in early (first sixteen weeks) normal pregnancy ; (2) those done in advanced normal pregnancy ; (3) those done in early (first sixteen weeks) abnormal pregnancy ; and those done in advanced abnormal pregnancy. In this way we could compare results in our groups not only with one another (to see whether they were influenced by the duration of pregnancy or by its abnormal character) but also with those of other workers, who had almost all worked with urine from cases of early normal pregnancy, when the need for a laboratory diagnosis is most imperative. ’
METHOD
We followed Galli Mainini’s (1948) usual technique, an amount of urine equal to a twentieth of the frog’s body-weight was found sufficient. In this. way positiveresults of varied intensity were obtained, which could be estimated by an approximate count-of the spermatozoa in a of cloacal urine under
except that
droplet