Am peromet ric Determination of Ethy Iened iaminetet raacet ic Acid with Zinc Ions W. S. WISE and N. 0 . SCHMIDT Department
o f Sugar
Chemistry and Technology, Imperial College o f Tropical Agriculture, Trinidad, 8. W. 1.
A method for the determination of total and available ethylenediaminetetraacetic acid in solutions containing the calcium or magnesium chelates is described. The solution is titrated amperometrically with zinc sulfate at a low pH for total ethylenediaminetetraacetic acid. The available ethylenediaminetetraacetic acid is determined at a higher pH when calcium is not displaced from its chelate. If magnesium is present, glycine is added to prevent the zinc displacing the magnesium from its chelate with ethylenediaminetetraacetic acid.
T
HE sodium salts of et'hylenedianiinetetraacetic acid (EDT-4) recently have become prominent owing to t,heir ability to form soluble chelate compounds with many meta1,ions. The chelate formed wit,h calcium ions is of special interest, and because the formation of this particular chelate ethylenediaminetetraacetic acid is applicable in many industrial applications. I n investigating the w e of ethylenedi:tminctet,raacetic acid for dissolving scale from factory evaporator tubes, determination of both the free et,hylenediaminetetraacetic acid and the calcium chelate in the aqueous solutions used was required. These solutions acquired a deep reddish-brown color during their use. The available methods (1-4) for the determination of ethylenediaminet,etraacetic acid and its chelates were found to be in this case unsatisfactory.
I
0
i
indicated well defined polarographic \yams. The addition of zinc ions to a solution containing ethylenediaminetetraacetic acid results in no polarographic wave until free zinc ions are in excess of those required to chelate all of the ethylenediaminetetraacetic acid. DETERMI3 &TION OF TOTAL ETHY LENEDI AMINETETRAACETIC ACID
Experimental. A sample of pure ethylenediaminetetraacetic acid 1% as prepared by precipitation from a solution of analytical grade disodium salt of ethylenediaminetetraacetic acid with hydrorhloric acid. The acid 1% a8 recrystallized from water and dried. A standard solution of ethylenediaminetetraacetic acid as prepared by dissolving a weighed amount of this acid in sufficient potassium hydroxide to form the dipotassium salt. A standard zinc solution 1%as prepared by dissolving (analytical reagent grade) zinc in dilute sulfuric acid and partly neutralizing with sodium hydroxide. A known amount of the ethylenediaminetetraacetic acid solution nas pipetted into a polarographic cell containing 25 ml. of a huffer, 0.5-Vin both acetic acid and sodium acetate. The dissolved oxygen was then removed by gassing out with nitrogen; the applied potential TTas kept constant a t -1.3 volts (21s. saturated calomel electrode) and the solution was titrated with 0.0515Jf zinc sulfate solution added from a microburet. Results. A typical titration curve shown in Figure 1, A , is the expected form. Such curves were obtained in solutions containing the ethylenediaminetetraacetic acid chelates of calcium or magnesium and the current readings became constant almost immediately, indicating that the displacement of calcium and magnesium ions by zinc ions was very rapid under these conditions. Some of the results obtained by this method are given in Table I.
Table I. Determination of Total Ethylenediaminetetraacetic Acid by Titration in Acetate Buffer
I-
z W
Substance Added
a a
3 V
Moles of Added Substance X 105
n
0.00
0 0 0 0
0 35 1 17 2.37 1.72 1 73 1.74 1.74 1.73
80 72 2100 -0
0
0 5
100
10
I 5
M L . OF
2.0
ZINC
2 5
3.0
3 5
L O
SULFATE
Figure 1. Amperometric titration curves 10 n i l . of 0.0171.1.1 EDTA in 25 nil. of acetate buffer, titrated w i t h 0.0493.1.1 ZnGOr 8. 15 ml. of 0.0171.W E D T A . 5 nil. of 0.0199.M CaClz, in 25 ml. of 0.1,V KOH, titrated with 0.0515M ZnSO. -4.
Many workers have studied the polarography of the metal chelates of ethylenediaminetetraacetic acid, and, therefore, it should be possible to titrate amperometrically the ethylenediaminetetraacetic acid with a metal ion, the chelate of which was not reduced a t the dropping mercury electrode. The zinc chelate waa found to give no reduction wave, a t least in the buffers considered, while reduction of free zinc or zincate ions
Titration, M1. of 0.0515.M Zinc Sulfate
Total E D T A , Moles X 106 Added Found0 1.8
6.1 12.2 9.0 9.0 9.0 9.0 9.0
0
1.8 6.0 12.2 8.9
9 .o 9.0 9.0 9.0
All determinations were carried out in acetate buffer of pH 4.6, but further experiments showed that the method is valid a t least over the range from p H 3.5 to 5.5. The accuracy of the determination depends upon the accuracy of reading the end point of the titration curve (Figure 1). This accuracy was found to be within 0.02 ml., or within 5 1% 1%hen approximately 2 ml. of the zinc solution are required for the titration. The experiments were completely reproducible within the accuracy of the determination. . Among other uses of the method, are the determination of ethylenediaminetetraacetic acid content of commercial samples (giving very satisfactory results), the standardizing of solutions made from analytical grade sodium salts of ethylenediaminetetraacetic acid, and a convenient way of standardizing zinc 1469
ANALYTICAL CHEMISTRY
1470 solutions by a volumetric method. The use of an amperometric titration to determine zinc has been published previously (6, 7 ) . DETERMINATION OF AVAILABLE
glycine the magnesium was displaced from its chelate, but as the glycine concentration was increased an end point was observed a t the titration corresponding to the amount of free ethvlenediaminetetraacetic acid (Figure 2, B ) .
ETHYLEYEDIARIINETETRAACETIC ACID
Although zinc is able to displace calcium from its ethvlenediaminetetraacetic acid chelate under acid or neutral conditions, it seemed possible that conditions could be found where zinc is not able to displace calcium. If this were so, the available pthylenediaminetetraacetic acid-Le., that not chelated by cdciuni-could be determined in solutions containing both ethylenediaminetetraacetic acid and its calcium chelate. If the amperometric titration was carried out in an alkaline medium then zinc could not displace calcium from its chelate, but calcium cqould displace zinc from the zinc chelate with ethylenediaminetetraacetic acid. The same effectwas described by Laitinen and SI-mpson ( 6 ) in a method of titrating calcium amperometrically. The results obtained for the determination of free ethylenediaminetetraacetic acid in solutions containing calcium are given in Table 11. I n this method 0.1N potassium hydroxide solution was substituted for the acetate buffer and the applied potential as -1.7 volts (US. S.C.E.). A typical titration curve is shown in Figure 1, B. The potassium hydroxide concentration must range from 0.05 to 0.5Y in order to obtain satisfactory titration.
M L . OF Z I N C SULFATE Figure 2. Amperometric titration curves of EDTA solutions containing magnesium -4. 5 ml. of 0.0171M E D T A , 2 ml. of 0.0204M MgSOa in 20 rnl. of tartrate buffer (10% ootassium hvdroeen tartrate in 2iV NaOH) B, C , D . 5 ml. of 0.0171M E D T A , 2 ml. of 0.0204M hfgSO4, titrated with 0.0493M ZnSO4, in presence o f : B. 20 ml. of glycine buffer (0.5M in K O H a n d 0.5M in glycine) C . 5 ml. of glycine buffer and 15 ml. of water D. 2 ml. of glycine a n d 18 ml. of water .%I. Theoretical end point for available E D T A 2'. Theoretical end point for total E D T A -
Table 11. Determination of Available Ethylenediaminetetraacetic Acid in Solutions Titration, Available E D T A , Total EDTA, Calcium, 1\11. 0.0515M lo' Theoretical FoundMoles x 106 hloles X 10s Zinc Sulfate 3.4 0 0.66 3.4 3.4 8.5 1.64 8.6 0 8.6 17.2 3.33 17.1 0 17.1 15.7 15.8 9.9 3.05 25.7 7.1 1.37 7.2 9.9 17.1 4 5 4.6 4.0 0.87 8.6 0 5 0.10 0.6 8.0 8.6
The results of Table I1 show that in solutions containing both free ethylenediaminetetraacetic acid and ethglenediaminetetraacetic acid combined with calcium, the free ethylenediamine tetraacetic acid can be determined by amperometric titration in alkaline solution with zinc sulfate. However, when the same method was applied to solutions containing magnesium, the zinc could displace magnesium from its chelate with ethylenediaminetetraacetic acid when the titration was carried out in a potassium hydroxide solution. .4 titration curve similar to Figure 2, D,was obtained. Unlike the previous titration curves obtained, no definite end point was observed; but, when a large amount of zinc was added the titration curve tended toward a straight line which extrapolated to the point corresponding to the total ethylenediaminetetraacetic acid in solution. This behavior shows that in the presence of a large hydroxyl ion concentration zinc is just able to displace magnesium from its chelate and does so completely only when in excess. I n order, therefore, to prevent zinc from displacing magnesium a substance must be added t o the solution which sets up another equilibrium with the zinc ions present and further reduces the amount of free zinc ions available for ethylenediarninetetraacetic acid chelation. Alkaline tartrate was first tried, but this chelate was so strong that the zinc could not chelate with ethylenediaminetetraacetic acid to any extent. Therefore, no titration curve was obtained (Figure 2, A ) . I n an ammonia-ammonium chloride buffer, zin'c r a s still able to displace magnesium. However, glycine was found to possess the desired characteristics. Titration curves obtained with two concentrations of glycine are shown in Figure 2, B and C. With a buffer containing small concentrations of
I
When the experimental end points were compared with the theoretical end points there was a small constant error. I n using an ethylenediaminetetraacetic acid solution without added magnesium, the error was proportional to the glycine concentration, and an impurity in the glvcine apparently was responsible for the error. This was confirmed by the fact that the ash from glycine was found to chelate ethylenediaminetetraacetic acid. A correction for the impurity in the glycine was determined by titrating a known volume of a standard ethylenediaminetetraacetic acid solution in the glycine buffer.
Table 111. Determination of Available Ethylenediaminetetraacetic Acid in Solutions Containing Magnesium Using Glycine Buffer Titration hI1. of MFg0.0?93M Total E D T A , nesium, zinc Corrected Moles X 106 Moles X 105 sulfate 0.65 5.2 2.0 0.50 1.32 8.6 2.0 1.17 2.0 1.86 2.01 12.1 8.1 2.39 2.54 20.6 4.1 1.46 1.61 12.1
Available E D T A , hloles X I O 5 Theoretical Found 3.2 3.2 6.6 6.5 10.1 9.9 12.5 12.5 8.0 7.9
-4s a result of this work the following procedure is established for the determination of available ethylenediaminetetraacetic acid in solutions which may contain magnesium. A known amount of the solution t o be analyzed is added to 20 ml. of a buffer, 0.5-44 for glycine and 0.5N for potassium hydroxide, giving p H 11.1. The solution is gassed out with nitrogen and titrated amperometrically with zinc sulfate solution a t a potential of -1.7 volts (21s. S.C.E.). Purification of the glycine is not necessary before use, provided the amount of impurity is determined and the appropriate correction applied.
1471
V O L U M E 2 7 , N O . 9, S E P T E M B E R 1 9 5 5 The results obtained using this procedure are given in Table 111. Allowing for the error due to the impurity in the glycine, good results were obtained. Since this method can be used when calcium is present, the use of the glycine buffer is preferable for routine determinations of available ethylenediaminetetraacetic acid: if a sample contains both magnesium and calcium, erratic results are obtained only rvhen potassium hydrovide is used.
ACKNOWLEDGMENT
The work described in this paper is part of a program of research carried out under the aegis of the British R e s t Indies
Sugar Research Scheme. The authors wish t o thank L. F. Wiggins for his interest in this work. LITERATURE CITED
(1) Bersworth Chemical Co., Tech. Bull. 2, (1953). ( 2 ) Darbey, 9., ANAL.CHEM.,24, 373 (1952). (3) Furness, W., Crawshaw, P.. and Davies, W. C., Analyst, 74,629 (1949). (4) Kerkow. F. W,, 2. anal. Chem., 133, 281 (1951). (5) Laitinen, H. A , , and Synipson. 1%. F., ~ N A L .CHEM.,26, 556 (1964). ( 6 ) Pribil, R., and hlatyska, B., Cheni. L i s t y , 44, 305 (1950). (7) Pribil. R., and Vicenova, E., I h i d . . 45, 177 (1951) RKEIVF.Dfor review l l a r c l i 80, 1B.54.
.-\ccepted February 24, 19.5.i.
Paper Chromatography of Alkali and Alkaline Earth Cations H. T. GORDON and C. A. HEWEL Department of Entomology and Parasitology, University of California, Berkeley, Calif.
This method separates and specifically detects 0.1 to 10 y of the common cations potassium, sodium, calcium, and magnesium on one-dimensional paper chromatograms run for 2 hours. Complete separation of barium, strontium, calcium, magnesium, and beryllium is also possible. Alany other inorganic and organic cations and anions can be separated and identified. Polyvalent anions may interfere with cation analysis, hut can usually be removed by precipitation as insoluble lead salts on the paper strip. The method is of blood and other usually applicable to 1 - ~ 1volumes . biological fluids without preliminary ashing.
Solution ID-1-B. Prepare by dissolving 145 mg. of 8-quinolinol and 360 mg. of 2-aminoethanol (Eastman 1597) in 50 ml. of C.P. acetone. Solution ID-2. Dissolve 100 mg. of bromocresol purple indicator (Eastman 745) in 10 ml. of absolute ethyl alcohol and dilute to 100 ml. with C.P. acetone. Add one or two drops of 28% ammonia in a a t e r to give the solution a red-yellow color, Solution ID-3 consists of 170uranyl zinc acetate in absolute ethyl alcohol containing 5y0 by volume of acetic acid (glacial, reagent). Heating on a m t e r bath is necessary to obtain complete solution. Uranyl zinc acetate is obtainable from Fisher Scientific (Catalog KO. V-11).
P
APER partition chromatography makes possible the rapid separation and detection of microgram quantities of inorganic ions such as potassium, sodium, calcium, magnesium, and chloride, which are widely distributed in minerals, soil, water, and living organisms. Flame photometry has been perfected to measure less than 0.1 y of sodium, potassium, and calcium ( 7 , 1 4 ) ,but paper chromatography may prove to be equally useful. I t is a semiquantitative technique in its own right, and, because it does not destroy the analytical sample, it can be used for separations preliminary to final analysis by colorimetry. fluorimetry, or flame photometry. The objective of the research reported in this paper was to obtain caomplete separation of potassium, sodium, calcium, and magnesium on chromatograms spotted with 1 pl. or less of blood drawn from small organisms such as insects, without preliminarv ashing to destroy organic matter. While many solvent systems have been used for inorganic ion separations on paper (4,Q), a new four-component system was devised for this work, especially for ascending one-dimensional chroniatography of the alkali and alkaline earth cations. The chromatographic technique has also been slightly modified to improve reproducibility of R , values. REAGENTS
Paper-Washing Solvent. Mix distilled water, pyridine (spectrochemical grade), and acetic acid (glacial, conforming to ACS qualifications) in the volume ratio 80 to 15 to 5. Developing Solvent. &fix isopropyl alcohol (98 to 99T0,such as Eastman 212), pyridine (spectrochemical grade), acetic acid (glacial, reagent), and distilled water in the volume ratio 8 to 8 to 1 t o 4. Lead Acetate Solution. Prepare 0.1M basic lead acetate in dilute acetic acid, by dissolving 580 mg. of basic lead acetate in 9.5 ml. of water plus 0.5 ml. of glacial acetic acid. Solution ID-I-A consists of 0.1% of reagent 8-quinolinol in C.P. or spectrochemical grade acetone.
Figure 1. Simplified apparatus for ascending one-dimensional paper chromatography Inset, upper right-hand corner, illustrates use of concentrating strip for solutions more dilute than 0.001M
Traces of inorganic ions are common contaminants in chenlicals, and it is desirable to use the purest obtainable reagents in this work. Samples of pyridine and bromocresol purple from some sources have given very poor results. APPARATUS
The chromatographic apparatus consists of borosilicate glass tubes, 51 mm. in diameter and 380 mm. long, open a t both ends (Figure 1). The ends are fitted with 90.26 corks through which
