CALCIUM DISODIUM EDTA IN TRANSFUSION
HEMOSIDEROSIS
TIBOR J. GREENWALT, M.D., AND V. ELIZABETH AYERS
Laboratories of the Milwaukee Blood Center, Inc., Milwaukee, Wisconsin
Venesection has been used for reducing the iron-load in hemochromatosis. 3 ' 4
In this condition there is an unusual ability to regenerate hemoglobin. The patient with chronic anemia not associated with blood loss who receives many
transfusions presents a different problem. There is no physiologic mechanism
for disposing of the approximately 250 mg. of iron introduced with each pint of
blood. Phlebotomy is not possible in the face of deficient red cell production or
an accelerated rate of destruction. Chemical compounds which form stable, nonionizable complexes with iron may be of value in enhancing excretion of iron.
British Anti-Lewisite (BAL, 2,3-dimercaptopropanol) is known to have a beneficial effect on intoxication with certain heavy metals.12 Ohlsson and his associates8 have reported an encouraging outcome following its use in a case of
transfusion hemosiderosis. Apresoline (Hydralazine hydrochloride) combines
with ferric ions 9 ' 10 but its use to increase iron output has not been reported.
Ethylenediaminetetraacetic acid forms soluble metal chelates in which the
polyvalent metal ion is bound in non-ionic form. It has been used successfully
in treating lead poisoning. 1 ' 2 Recently Wishinsky and co-workers16 reported
their experiences with the intravenous administration of the calcium disodium
salt of EDTA in a case of hemochromatosis. We are reporting our results with
the oral and intravenous administration of calcium disodium EDTA in 4 patients with chronic aregenerative anemia who had received many blood transfusions.
M A T E R I A L S AND METHODS
The patients employed in this study were selected because they had aregenerative anemias requiring transfusion of large quantities of blood. The presence of
some degree of exogenous hemosiderosis was assumed. Iron studies to establish
the presence of an excessive iron-load were made in 2 patients.
The calcium disodium salt of ethylenediaminetetraacetic acid was used in all
the studies and will be referred to as EDTA.* One patient received EDTA dissolved in 250-500 ml. of 0.9 per cent sodium chloride by slow intravenous drip.
The oral doses were administered in hard gelatin capsules. The child was given
an aqueous solution of the drug in milk. All the patients were requested to take
a glass of milk with each dose and were advised to increase their daily intake
of dairy products.
Received for publication October 18, 1954; accepted, December 2, 1954.
Dr. Greenwalt is Medical Director.
* We are indebted to Dr. Norman W. Karr, Director of Clinical Research, Riker Laboratories, Inc., Los Angeles, California, for the initial supplies of calcium disodium EDTA
for intravenous use.
The EDTA powder used for oral therapy was kindly supplied by the Alrose Chemical
Company, Providence, Rhode Island.
266
March 1955
TRANSFUSION HEMOSIDEROSIS
267
All glassware used in these experiments was placed in dichromate cleaning
solution overnight. After thorough rinsing in distilled water it was immersed
in 5 N hydrochloric acid. Finally it was rinsed 5 times in iron-free distilled water
obtained by redistilling distilled water in an all-glass system.
Twenty-four-hour urine specimens were collected and refrigerated until the
time of analysis. The daily urinary output of iron of each patient was determined for 3 to 4 days prior to the administration of EDTA.
Serum iron was determined by the method of Ramsay.11 Iron-binding capacity of serum was measured as described by Rath and Finch.13
TABLE 1
E F F E C T O F E D T A ON U R I N A R Y I R O N E X C R E T I O N
Patient and Dates
Total Dose of
CaNaj EDTA
Diagnosis
Urine Iron
Excretion in
Gm.
Range in
Av.
24-hr.
Excretion of
Urine
Iron
mg.
m%.
tug.
N M
Normal control
0
1.5 (oral)
1.05 in 3 days
3.39 in 3 days
.15-.54
.88-1.31
0.35
1.13
TG
Normal control
0
1.5 (oral)
.36 in 3 days
2.40 in 3 days
0-.240
.55-1.12
0.12
O.SO
1.27 in 4 days
32.35 in 9 days
0-.57
1.97-5.3S
0.32
3.51
.72 per d a y ;
daily spot
checks
0.02-1.52
0.72
0
4.26 in 3 days
.93-1.74
1.42
0
2.47
1 clay
after
transfusion
5.15 in 5 days
.5S-1.S7
1.03
.69 in 3 days
.06-.55
0.23
1.89 in 6 clays
.02-.64
0.31
.43 in 3 days
.09-.22
0.14
1.39 in 13 days
.00-.19
0.10
G F
Sept. 9, 1953
Sept. 10
Sept. 18
Dec. 19
Jim. 18, 1954
Aplastic
anemia
N A
Dec. 11, 1953
Dec. 13
Dec. 14
"Refractory"
anemia
22.5 (oral)
6
3.5 (oral)
Dec. 15
0 McG
Myelosclerosis
Feb. 9, 1954
Feb.11
Feb. 12
F e b . 17
LJ
Feb. 22, 1954
Feb. 23
Mar. 7
0
18 (intravenous)
0
4.5 (oral)
Congenital
hypoplastic anemia
0
1.8 (oral)
268
GREENWALT AND AYEES
Vol. 25
Urinary iron was determined in duplicate on aliquots of the 24-hour collections. The method recommended by Sandell14 was used. Recovery procedures
with known amounts of iron added to normal urine with and without EDTA
gave an average recovery of 99.0 per cent (range 88.5-106.7 per cent). The average error of duplicate determinations was ±1.9 per cent. All the readings were
made with a Beckman model " B " spectrophotometer. A wavelength of 508 m/x
was used for urine iron and 520 m/i for the serum analyses.
RESULTS
Prior to starting the clinical experiments the excretion of urinary iron of 4
rabbits averaging 3.5 Kg. in weight was studied. Precautions were taken to
avoid contamination of the urine collected with extraneous iron and feces. The
average of 18 24-hour urinary iron determinations during the control period was
45 tig. Each rabbit was given 100 mg. of EDTA intravenously for 2 days and
200 mg. for 2 more days. The average of 16 24-hour urinary iron values during
this period was 108 MgThe effect of small oral doses of EDTA on the urinary iron output of 2 normal
adult persons is shown in Table 1.
The increase in urinary iron excretion in these control studies appeared to be
significant and therefore the 4 selected patients were subjected to similar investigations. The results obtained in these cases are given in Table 1.
REPORT OF CASES
Case 1. G. F., a 35-year-old white man with aplastic anemia, was under observation for
37 months and received 103 pints of blood. He gave no history of loss of blood. His skin
had a light gray cast.
The results of the urinary iron excretion studies are given in Table 1. During the 4-day
control period 1.27 mg. of iron were excreted in the urine. With the administration of EDTA
intravenously for 5 days the urinary iron output rose and remained elevated for 4 days
after discontinuation of therapy. The total iron output for this 9-day period was 32.35
mg., whereas the expected total based on the control studies was 2.S5 mg. The patient was
given EDTA orally for 31 consecutive days. Twenty-four-hour urine studies were done
only at intervals during this time. The iron output was inconstant but at times amounted
to 2-4 times the baseline values. Serum iron and unsaturated iron-binding capacity (UIBC)
values were as follows:
Sept. 9 to Sept. 14, 1953
Dec. 17, 1953
Jan. 11, 1954
Jan. IS .
June 21
Serum iron (pg/100 ml.)
UIBC
297, 267, 397,
480, 410, 449
440
406
278
444
—
0
0
—
—
Prothrombin, serum protein, calcium and phosphorus levels revealed no significant
alterations.
Case Z. N. A. was a 62-year-old white woman with "refractory" anemia. She was followed clinically from April, 1952 up to the time of her death in May, 1954. During this
period she was given 62 pints of blood. Her skin gradually acquired a slate-gray hue. There
were no hemorrhagic complications.
March 1955
TRANSFUSION HEMOSIDEROSIS
269
The studies in this patient were incomplete. Her baseline urinary iron excretion was
high. Hemosiderin granules were demonstrated in the urinary sediment by the Prussian
blue technic. It is interesting to note the sharp rise in iron output following blood transfusion. Daily oral administration of 0.5 to 1.0 Gm. of EDTA for short periods did not result
in increased output of urinary iron (Table 1).
Case S. 0. McG., a 57-year-old white man, received 33 pints of blood during the 12month period since the diagnosis of myelosclerosis was established. He had no hemorrhagic
phenomena. His serum iron level was 304 ng. on Feb. 10, 1954 and no TJ.I.B.C. was demonstrable. The iron excretion data presented in Table 1 indicate that following the oral ingestion of 0.5 to 1.0 Gm. of EDTA daily there was a slight rise in urinary iron.
Case 4- L. J., a 4-year-old white girl, developed pallor at the age of 2 months. Subsequent
studies established the presence of erythroid aplasia of the bone marrow. She has been
under our observation for almost 4 years and has received the equivalent of 25 pints of
blood. Daily oral administration of 0.10-.20 Gm. of EDTA in solution did not result in
any increase in excretion of iron (Table 1).
DISCUSSION
The urinary iron output of patient G. F. was increased ten-fold by the intravenous administration of 18 Gm. of EDTA over a 5-day period. Wishinsky
and associates16 and Figueroa and his co-workers6 reported similar results. I t is
evident that this form of therapy is not practicable for extended use to remove
significant quantities of iron.
Oral doses of 0.5 to 1.0 Gm. of EDTA daily resulted in 3- to 6-fold increases
of urine iron in 2 normal persons. This led us to think that protracted oral administration might have some merit. The results given in Table 1 indicate that
a slight increase in output of urinary iron occurred in 2 of the 4 patients. The
dose used was small but the recent report of Figueroa and associates.6 Indicates
that oral dosage up to 8.0 Gm. per day is not more effective.
Foreman and co-workers6 have shown that in rats the fecal as well as the
urinary excretion of intravenously administered Fe 59 is increased after intraperitoneal injection of "Fe-3 Specific,"* a chemical homolog of EDTA. They
found oral administration to have a similar effect. Over 95 per cent of the activity appears in the urine when C14-labeled EDTA is given intravenously to
normal adult persons.7 Less than 5 per cent of the labeled material is absorbed
from the intestinal tract when given by the oral route. The iron from ferric
sodium ethylenediaminetetraacetate is absorbed from the intestinal tract and
utilized for hemoglobin regeneration at the same rate as ferrous sulfate in anemic rats.16
It would be hazardous to draw any definite conclusions from the data which
have been presented. Intravenously injected EDTA has been shown to increase
the urinary iron output. The increment in urinary iron following oral administration of EDTA may not represent iron withdrawn from the tissues. It is possible that the chelating agent can render dietary iron more readily available for
absorption. In that case the increase in urinary output of iron after oral inges* A chelating agent manufactured by Bersworth Chemical Co., Framingham, Massachusetts. Its structure was not given.
270
GREENWALT AND AYERS
Vol. 25
tion of EDTA may at least in part represent renal clearance of iron absorbed.
In fact, Foreman and associates6 have shown that, when Fe 69 and "Fe-3 Specific" are fed simultaneously to rats, a marked increase in urinary excretion of
the tracer occurs while the fecal excretion is essentially unaffected. Likewise,
Figueroa and co-workers5 report preliminary observations that the oral administration of EDTA to patients does not affect fecal output of iron. Careful studies
on iron balance will be necessary before further interpretation is possible.
No toxic manifestations were encountered in the patients in this study. The
possibility of producing hypocalcemia is largely circumvented by using the calcium salt of EDTA. Studies with C14-labeled Ca-EDTA indicate that it passes
through the body unaltered.7 I t is rapidly and completely eliminated and therefore long-term toxicity is unlikely. The gradual depletion of essential ions as a
result of prolonged administration is possible.
The use of chelating agents to reduce overload of iron deserves more investigation. The results with oral administration have been disappointing and prolonged intravenous therapy is not practical. The intravenous use of Ca-EDTA
with all transfusions given to patients who are likely to be exposed to the dangers
of exogenous hemosiderosis may be of value. Perhaps a "depot" preparation
can be developed for intramuscular injection. Some of the newer aminopolycarboxylic acid derivatives may prove to be of greater value in enhancing loss
of iron than those now available for clinical use.
SUMMARY
Calcium disodium ethylenediaminetetraacetate (EDTA) was administered
orally and intravenously to 4 patients with aregenerative anemias who had received many transfusions. The urinary iron output of 1 patient was increased
10-fold by intravenous medication. Oral therapy produced inconstant results.
The problem of mobilizing iron stores with chelating agents is discussed.
•At present no method of administering EDTA has been found which makes
it a valuable agent for the treatment of overload of iron.
Acknowledgment. T h e authors are indebted t o D r . B e r t r a m H . Dessel and D r . N e d G.
Maxwell for their valuable assistance.
REFERENCES
1. B E L K N A P , E . L . : E D T A in t h e t r e a t m e n t of lead poisoning. I n d u s t . Med., 2 1 : 305-306,
1952.
2. BBSSMAN, S. P . , R I E D , H . , AND R U B I N , M . : T r e a t m e n t of lead encephalopathy with calcium disodium versenate; report of case. M . Ann. District of Columbia, 2 1 : 312-315,
1952.
3. D A V I S , W. D . , J R . , AND ARROWSMITH, W. R . : T h e effect of repeated bleeding in hemochromatosis. J . L a b . & Clin. Med., 36: 814-815, 1950.
4. D A V I S , W. D . , J R . , AND ARROWSMITH, W. R . : T h e t r e a t m e n t of hemochromatosis b y
massive venesection. Ann. I n t . Med., 39: 723-734, 1953.
5. F I G U E R O A , W. G., ADAMS, W. S., B A S S E T T , S. H . , R O S O V E , L., AND D A V I S , F . : Effect of
disodium calcium versenate on iron excretion in man. Am. J . Med., 17: 101, 1954.
6. F O R E M A N , H . , H U F F , R . L., O D A , J . M . , AND GARCIA, J . : Use of a c h e l a t i n g a g e n t for
accelerating excretion of radio-iron. Proc. Soc. Exper. Biol. & Med., 79: 520-524,
1952.
7. FOREMAN, H . , AND T R U J I L L O , T . T . : T h e metabolism of C 14 labeled ethylenediaminetetraacetic acid in human beings. J . L a b . & Clin. Med., 43: 566-571,1954.
March 1955
TRANSFUSION HEMOSIDEROSIS
271
8. O H L S S O N , W. T . L., K U L L E N D O R F F , G. T . , AND L J U N G B E K G , L. K . : Transfusion h e m o -
siderosis. Report of a case treated with BAL. Acta med. Scandinav., 145: (Ease. 6 ) :
410-418, 1993.
9. P E R R Y , H . M., J R . , AND SCHROEDER, H . A . : Syndrome simulating collagen disease caused
by hydralazine (Apresoline). J . A. M. A., 154: 670-673, 1954.
10. P E R R Y , H . M . , J R . : A method of quantitating 1-hydrazinophthalizine in body fluids.
J. L a b . & Clin. Med., 41: 566-573, 1953.
11. RAMSAY, W. N . M . : T h e determination of iron in blood plasma or serum. Biochem. J.,
53: 227-231, 1953.
12. RANDALL, R. V., AND S E E L E R , A. 0 . : BAL. New England J. Med., 239: 1004-1009; 1040-
1046, 1948.
13. R A T H , C. E . , AND F I N C H , C. A.: Serum iron t r a n s p o r t : Measure of iron-binding capacity
of serum in man. J . Clin. Investigation, 28: 79-85, 1949.
14. SANDELL, E . B . : Colorimetric Determination of Traces of Metals. New York: Interscience, p p . 382-385, 1950.
15. SEEBERG, V. P . , HIDALGO, J., AND W I L K E N , W.: Hemoglobin regeneration following oral
administration of chelated iron. Science, 119: 608-609, 1954.
16. W I S H I N S K Y , H . , W E I N B E R G , T . , P R E V O S T , E . M . , B U R G I N , B . , AND M I L L E R , M .
J.:
Ethylenediaminetetraacetic acid in the mobilization and removal of iron in a case of
hemochromatosis. J . L a b . & Clin. Med., 42: 550-554, 1953.