LITERATURE CITED
(4) Jones, H. B., Baum, H., Ibid., 27,
( 1 ) Atack, F. W.,J . SOC. Chott. Ind. Japan 34,936 (1915). (2) Barton, C . J., ANAL. CHEW 20,
99 (1955). (5) Luke, C. L., Braun, IC. C., Ibid., 24, 1120 (1952). (6) Mellan, I., “Organic Reagents in Inorganic Analysis,” pp. 227-50, Blakiaton, Philadelphia, Pa., 1941.
1068 (1948). (3) Blacdcl, W.J., Lcwip, \V. B.,Thomas, J. W.,Ibid., 24,50‘3 (1952).
(7) Snyder, L. J., IND.ENQ. CHEM., ANAL. ED. 17, 37 (1945). (8) Welcher
F. J v “Organic Analytical Reagcnta,” Vol. IV, p. 355, Van Nostrand, New York, 1948. RECEIVED for review October 18, 1980. Accepted January 3, 1961.
Ion Exchange Separation of Calcium and Strontium Application to Determination of Total Strontium in Bone MARVEN A. WADE and H. J. SElM Universify of Nevada, Reno, Nevada
b The difference in the stability constants of the EDTA complexes of strontium and calcium i s utilized to effect their separation by ion exchange. The sample is dissolved in EDTA and, after the pH has been adjusted to 4.80, the solution is passed through a Dowex 50-X8 column. The remaining calcium is then eluted with EDTA at a pH of 5.30. After removal of alkali metal and ammonium ions with 0.75N HCI, the strontium is eluted with 3N HCI and determined flame photometrically at 460.7 mp using an oxygen-hydrogen flame. The method has been used for the routine determination of strontium in bone ash and is applicable to any type of sample. The procedure has been applied successfully to Ca-Sr ratios as great as 20,000 to 1 .
has bccii applied by Roberts (4), who reported the strontium content of two different adult femurs to be 281 and 480 pg. per gram of ash. These methods of detcrmining strontium all require elaborate and expensive instrumentation Strontium can readily be determined in bone ash flame photometrically once i t is separated from the large amount of calrium present. I n bone ash, the calcium-strontium ratio is of the order of 4000 to 1. A method is described for the ion exvhange separation of calcium and strontium utilizing (rthylenedinitri1o)tctraacetic arid (EDTA). The Ca+l ions are complrxed with EDTA and the CaEDTA+ complex and Sr+l ions are separated using Dowex W X 8 resin. THEORETICAL
R
on thc metabolisin of strontium and its distribution in the body has been difficult because of t h r lack of an analytical method suitable for routine analysis Research in this direction is important because the radioactivr species, Sr”, is considered t o be the most dangerous fission product resulting from nuclear explosions and reactors. Strontium is present in only trace amounts in body tissues, and the main difficulty involved in its determination in samples such as milk and bone is separation from the large amount of calcium present. Neutron activation has been applied by Sowdrn and Stitch (7’) t o the determination of strontirini in bone. Thcse investigators found the concentration to be of the order of 100 pg. per gram of ashed tissue. Thurbcr el al. (B), using emission spcctroscopy, reported thc world-wide average of strontium in human bone to be 172 pg. per gram of ash. X-ray fluorescence ESEARCH
The reaction of a metal ion with EDTA is represented by Equation 1, where Y represents EDTA. M+a
+ HZY-1
=
MY-’+ 2 H C
the elution of the calcium through the resin. This technique of cation separation has bcen applied by Fritz and Umbreit (3) t o the separation of several binary mixtures, and by Davis (1) t o the separation of calcium and strontium when present in equal amounts. Farabee (9)utilized a similar procedure for the determination of radiostrontium in urine. APPARATUS
All flame photometric determinations were made with a Beckman Rlodel D U spectrophotometer equipped with a 9220 flame attachment and an osygenhydrogen burner. All p H mcasurements were made with a Beckman Model G pII meter. The ion rxchrtnge columns used were similar t o those described by Seim, Morris, and Frew (61, but were made from borosilicate glass 14 nim. in outside diametcr and held a column of resin 27.6 cm. in height. The reservoir had a 250-ml. capacity, and the column and reservoir were connectcd with a 19/22 standardtaper ground-glass joint.
(1)
This equation shows that the system is dependent on the H + concentration, and increasing the acidity of the solution will decrease the concentration of the metaI-EDTA complex. The concentration of the complex also depends on the stability constant; the larger the log K z , the lower the p H at which the metal-EDTA complex will exist. Since the logs Kz for the calcium and strontium complexes with EDTA are 10.59 and 8.63, .respectively (6),the calcium mill complex at a lower p H than the strontium. A second factor favoring the separation by ion exchange is that the resin affinity is greater for strontium than for calcium. Both of these factors favor the retention of the strontium and
REAGENTS
Ion Exchange Resin. Dowex 50X8, 50-100 mesh, analytical grade, hydrogen form (Bio-Rad Laboratories, 32nd and Griffin Ave., Richmond, Calif.), is thoroughly washed with water and 30 ml. arc transferred to a n ion exchange column. After elution with 250 ml. of 3 N HCl and 50 ml. of H20 t o remove impurities, 150 ml. of 10% NH,Cl are added to convert the resin to the ammonium form. After equilibration with 50 ml. of 2% EDTA“4, the column is ready for use. EDTA. The EDTA used in this study was supplied by Geigy Industrial Chemicals, Saw Mill Rd., Ardsley, N. Y. The EDTA is purified before use by dissolving in NHIOH, filtering, VOL. 33, NO. 6, M A Y 1961
793
and precipitating a t a pH of 1.6. A 2% solution of the ammonium salt of is prepared by EDTA, EDTA-"4, dissolving 80 r a m of the purified EDTA in NH48H, diluting t o 4 liters, and adjusting the pH to 6.30. All other reagenta used were analytical grade and all solutions were prepared using deionized distilled water and stored in polyethylene bottlea.
3
B
PROCEDURE
One gram of bone ash is dissolved in HCI, filtered, evaporated to dryness, and redissolved in 200 ml. of 2% EDTA-NH4. After the pH has been adjusted to 4.80, the solution is passed through the ion exchange column prepared as described above. The flow rate for this and all succeeding steps is maintained at 1 to 2 ml. per minute. The column is then eluted with 250 ml. of 2% EDTA-"( a t a pH of 5.30 which removes the calcium, followed by 50 ml. of water to remove the excess EDTA-NH,. Ammonium and residual alkali metal and magnesium ions are removed ' by elution with 250 ml. of 0.75N HCI. The strontium is then eluted with 250 ml. of 3N HC1. After evaporating to dryness, the strontium chloride is diaolved in water, diluted to 25 ml. in a volumetric flask, and determined flame photometrically, by measuring the emission a t 460.7 mp. The readings are compared against a standard curve prepared by measuring the emission produced by solutions containing 5, 10, 15, and 20 pg. of strontium per ml. The resolution of calcium and strontium ions achieved by this separation procedure is represented in Figure 1. EXPERIMENTAL
Elucidation of the p H range in which the resolution of Cay-* and Sr+9 ions would be possible became the first objective of this invcstigation. A sample of Baker's analyzed calcium carbonate was analyzed for strontium a t eight p H values ranging from 4.50 to 5.80.
k
3
N HCI
--a
OF SOLUTION
Figure 1. Separation of calcium and strontium on Dowex 50-X8 resin using EDTA
Strontium-89 tracer was added to each of the 1-gram samples of calcium,carbonate. Figure 2 shows the percentage of calcium and strontium retained by the resin us. pH. The calcium was d e termined flame photometrically and the strontium radiometrically after elution with HCl. Figure 2 shows that at pH 5.2 to 5.8 all the calcium is eluted from the resin by the EDTA-NH, and at pH 4.5 to 5.3 all the strontium is retained by the resin. These data illustrate that in the p H range of 5.2 to 5.3, the separation is quantitative. The elution pattern of calcium in the EDTA-"( waa followed by collecting fractions and analyzing each for calcium. Figure 3 shows the elution curves for calcium at p H 4.5, 5.0, and 6.6. These curves are representative of the eight pH values checked and illustrate that the elution patterns are very similar. The maximum amount of calcium retained was 12% at a pH of 4.5. These results indicate that the p H of the EDTA-NH4 solution containing the sample is not critical. Subsequent work showed that the pH of the sample solution could vary from 4.6 to 5.2, with successful quantitativc separation
of calcium and strontium in calcium carbonate, if the pH of the EDTA-N& solution used in the elution step were maintained at 5.30. Table I shows that added strontium was not quantitatively recovered above the bmg. level when the sample and elution EDTA-NH, solutions were both maintained at pH 5.30. Adjusting the p H of the sample solution to 4.8 and subsequently eluting with EDTA-N& at pH 5.3 increased the recovery of 125 mg. of added strontium from 69.2 to 95.6%. The procedure adopted takes advantage of the additional resolution made possible by using a lower pH for the sample solution. This should also compensate for small changes in resin characteristics and fluctuations in pH. This separation procedure results in the resolution of strontium from all the elements expected to be present in bone except barium. The magnesium is eluted from the resin with the calcium as the MgY-* complex. The phosphate
Table 1. Effect of pH of Sample Solution on Recovery of Strontium (pH of EDTA-NH, elution maintained at 6.3)
Sr
H of
Recovery of Sr, %
6.3 4.8 6.3 4.8 6.3 4.8 6.3 4.8
100.4 100.0 96.8 99.2 77.1 95.2 69.2 95.6
Added,a Sr Found, {ample Mg. Mg. Soh. 5 26 SO
5,
76
J
pM OF SOLUTION
126
Figure 2. Effect of pH on retention of calcium and strontium by Dowex 50-X8 resin eluted with EDTA
794
ANALYTICAL CHEMISTRY
0
5.02 6.00 24.20 24.80 67.80 71.40 86.60 119.60
All eamplee contained 400 mg. calcium.
Figure 3. Elution of Cay'' at different pH values
Table 11. Determination of Strontium in Various Types of Materials
Strontium Found, ag.lG.
Type of Sample
Limcstone, Sample
NBS
l a (Sr 0.19,%)
reptd.: CaCOa, Bakers analyzed (Sr reptd.: .eo
