Spectrophotometric Determination of Magnesium with Sodiw m 1-Azo-2-hydroxy-3-(2,4-dimethylcarboxanilido)naphthalene-I ’-(2-hydroxy benzene-5-sulfonate) CHARLES K. MA“’
and
JOHN H. YOE
Pratt Trace Analysis Laboratory, Department o f Chemistry, University of Virginia, Charlotterville, V a .
A new colorimetric method for the determination of trace concentrations of magnesium is described. The reagent is sodium l-azo-2-hydroxy-3-(2,4-dimethylcarboxanilido) naphthalene 1’- (2 hydroxybenzene-5sulfonate). The method has a spectrophotometric sensitivity of 0.0008 y of magnesium per sq. cm., and a practical sensitivity of 1 part of magnesium in 50,000,000 parts of solution. I t is applicable to the determination of 0.5 to 10 y of magnesium. Interferences by diverse ions have been studied and methods of separation, by extraction and by ion exchange fractionation, have been developed for use in the determination. Magnesium in “synthetic blood ash” solutions and in limestones has been determined with good precision and accuracy.
-
-
-
X 6 mm. in inside diameter with a reservoir and ball and socket connections for application of pressure was used for the ion exchange separation. A reservoir for backwashing was attached to the bottom by a t-cvo-way stopcock. Standard Magnesium Solution. The standard magnesium solution was prepared from reagent grade magnesium oxide, A sample of the ignited oxide (800’ to 820”)weighing 1.658 grams was dissolved in 1 liter of 0.1-Vhydrochloric acid to give a solution containing 1 mg. of magnesium ion per nil. This solution v a s standardized gravimetrically by the ovine method and was stored in a polyethylene bottle. Buffer Solution. The buffer was prepared by dissolving reagent grade borax in water to give an 0.08M solution. Reagent Solution. The reagent solution was prepared by dissolving sodium l-azo-2-hydroxy-3-(2,4dirnethylcarboxanilido)naphthalene-l’-(2-hydroxybenzene-5-sulfonate) in 95% ethyl alcohol to give a solution containing 0.15 mg. of reagent per ml. The compound may be obtained from the LaMotte Chemical Products Co., Towson, Baltimore, .Md. Resin. D o m v SOX12 (200 to 400 mesh) was used after wash1) until the ing it repeatedly s i t h dilute hydrochloric acid (1 washings a e r e colorless, Oxine Solution. The oxine solution was prepared by dissolving 5 grams of Eastman Khite Label 8-quinolinol in 100 ml. of chloroform. A fresh solution must be prepared each day. Other Reagents. All other reagents were analytical grade and were used without further purification.
+
C
OLORIMETRIC methods in general use for small quantities of magnesium are based on indirect determinations after the formation of insoluble precipitates and on the formation of colored lakes. In one indirect method magnesium ammonium phosphate is precipitated, followed by the colorimetric determination of phosphate ( 7 ) . A similar procedure involves the precipitation of magnesium 8-quinolinolate, with solution of the precipitate and colorimetric estimation of the oxine ( 8 ) . -4 number of direct colorimetric methods utilizing the formation of color lakes have been advanced, Titan Yellow having received the most attention (6). Recently, procedures involving the formation of a colored complex of magnesium with Eriochrome Black T have been described ( 3 , 5, IO) This paper describes a method for the determination of trace quantities of magnesium based on the formation of a colored complex of magnesium with sodium l-azo-2-hydrosy-3-( 2,4-
PROCEDURE
Transfer a 5-nil. aliquot of the reagent solution to a 25-ml. volumetric flask; add to this an aliquot containing 0.5 to 10 y of magnesium in not more than 5 ml. of solution which has been adjusted so as to be just acid to phenolphthalein. Add 0.5 ml. of 0.08111borav solution, make up to the mark with 95% ethyl alcohol, and mix. After a t least 30 minutes, meaaure the absorbance of the solution a t 510 mp, using a distilled water blank. Determine the weight or concentration in the unknown from a working curve prepared by this procedure using known amounts of magnesium. dimethylcarbosanilido)-naphthalene-l’-~2-hydro~ybenzene-5-~ul- For best results, it is desirable to prepare a new working curve ionate). with each freshly prepared reagent solution. Maximum precision will not be obtained a t the lower extremity of the working curve; O H hence, it is desirable t o adjust the size of the magnesium aliquot to give 1 to 10 y of magnesium for each determination. DISCUSSION
Color Reaction. Sodium l-azo-2-hydroxy-3-(2,4-dimethylcarbosanilido)-naphthalene-l’-(2-hydroxybenzene-~sulfonate)is a brilliant red solid that is soluble both in water and in 9570 ethyl alcohol to the estent of approximately 0.4 mg. per ml. The compound is stable indefinitely in the solid state, and for at least 4 months when dissolved in 95y0 ethyl alcohol. It is an acid-base indicator with a transition from red to blue-violet a t p H 7 to 8 and another transition from blue-violet to pink a t p H 11 t o 12. In the pH range 8 to 11, magnesium ion causes a color change from blue-violet to red, yielding solutions with absorbance peaks of 555 (reagent) and 540 (complex) mp. Hbwever, when solutions are prepared in %yoethyl alcohol instead of water, the color of the reagent is blue instead of violet; that of the complex is salmon pink, rather than red. Absorption spectra of solutions prepared under these’conditions are shown in Figure 1. The absorbance peak of the reagent is shifted from 555 to 615 mp; the comples then shows two peaks, one at 510 mw and another a t 540 mp. .4pplication of the method of continuous variations t o this system indicates the presence of reagent-magnesium ratios of 1 to 1 and 2 to 1.
I
SOoNa Sodium l-azo-2-hydroxy-3-(2,4-diniethylcarbosanilido)naphthalene-1’-(2-hydroxybenzene-5-sulfonate) The color reaction has been studied in detail t o determine optimum conditions for its use. Data on the seneitivity, precision, accuracy, and interferences by diverse ions are presented. APPARATUS AKD REAGENTS
Spectrophotometer. A Beckman spectrophotometer, Model DU, with matched 1-cm. Corex cells was used, p H Meter. p H measurements were made with a Beckman Model G p H meter, using shielded glass electrodes for external operation. Ion Exchange Column. A column consisting of a tube 20 cm. 1 Present address, Department of Chemistry, Pnirersity of Texas, Austin, Tex.
202
203
V O L U M E 2 8 , NO. 2, F E B R U A R Y 1 9 5 6 Optimum Conditions. The addition of ethyl alcohol to the system causes a desirable divergent shift of the absorbance maxima of the reagent and complex. Experiments involving a change of the alcohol concentration indicate that it should be maintained at about SOT0or higher. Several buffer solutions were tried: a boras buffer (0.08M), pH 8.95, was found to be satisfactory. For accurate measurements, it is necessary to hold the pH fairly constant. This may be done by making the magnesium solutions, from which aliquots are to be taken, just acid to phenolphthalein (which does not interfere with the determination) and then following the above procedure. The reagent concentration may vary from 20 to 40 p . p . m , but it is necessary to use the same concentration for standards and unknowns.
4 0
500
540
580
preparing working curves, when the correction is made, a working curve is obtained which passes through the origin. Also, it is then not necessary t o prepare a new working curve for each reagent solution as suggested above because changes of reagent concentration within the range of 20 to 40 p.p.m. have no effect. The absorbance of the complex does not attain its maximum value instantly; it increases about 1% during the first 30 minutes after preparation of the colored solutions. Then it remains constant for more than an hour. It is necessary, therefore, to allow a t least 30 minutes for the full development of the color. Absorbance measurements of similar solutions that were placed in thermostats a t 15' and 30'. respectively, showed no variation in absorbance. Hence, normal temperature changes in the laboratory introduce no error. The reagent was tested with 71 ions on the spot plate to determine the effect of diverse ions. Thirty ions were found t o cause interference. For quantitative evaluation, an ion concentration that causes a deviation of more than 0.010 unit in the absorbance of 0.12 p,p.m. of magnesium as the complex was arbitrarily taken as an interference. Table I summarizes the tolerances of some of the more common cations. I n addition to the ions listed in Table I, the following ions are also known to interfere: As++* Be++, C e + + + ,Crf-', D y + + + ,E u + + + ,Gd+'+ Au+++, MoOr--, iYdLLL, Kb'+, Pd+-, Pr+++, S m + + + , TI+++,T m + + + , and Yb'+". I t is necessary, therefore, either to remove or t o mask most of the metal ions before a magnesium determination can be made. The common anions, halides, nitrate, sulfate, phosphate, acetate, and silicate, interfere only when they form salts that are insoluble in ethyl alcohol. This depends on the type and concentration of cation present. The limiting concentration of the alkalies is approximately 0.01.l'.
Table I. Interfering Ions .Idde cl Concentration, Limiting Ion AI Ba'cd-+ Ca'+
620
cut-
0.03 0.08
FeTFe++Pb Rln Si GO:- Sr-
Figure 1. Absorption spectra of reagent and its magnesium complex
0.02
0.1
++*&
0.01 0.01 1000
-
14 p.p.rn. of reagent 14 p.p.m. of reagent plus 0.4 p.p.m. of magnesium
As seen in Figure 1, the reagent absorbs a t the nave lengths of maximum complex absorption (510 and 540 mp): however, the complex peaks do not overlap the reagent peak. -4s the reagent absorbs much less a t 510 mp than at 540, the former is better for analytical use. Obviously, the absorbance at 510 mp is the sum of the absorbances of the complex and the unreacted reagent. The absorbance due to unreacted reagent may be calculated and deducted from the total, giving the absorbance due t o the magnesium complex. This may be done because a proportional relationship exists between the reagent absorbance a t 510 and a t 615 mp. Since the absorbance of the complex at 615 mp is almost nil, the total absorbance of the solution a t that wave length may be taken as a measure of the amount of unreacted reagent. Hence, the portion of the absorbance a t 510 m p due t o unreacted reagent may be calculated from this measurement. I n practice, it has been found convenient to prepare a calibration curve by measuring the absorbances of solutions of varying reagent concentration at the two wave lengths (510 and 615 mp) and plotting one against the other ( 4 ) . The absorbance at 510 mpwhich is due t o unreacted reagent may then be read from the calibration curve. Although equally precise results are obtained by either method of
0.01 0.01 0.01
co++
jVAV E L E h G T t -
1. 2.
P.P.AI. 4 2
as
+
1 0.03
Zn--
Precision, Sensitivity, and Range. I n order to measure the precision, five series of 11 to 13 replicate determinations were made. The precision, expressed as the standard deviation from the mean for each of the five series, is shown in Table 11.
Table 11. Precision of Determination Standard Deviation from Mean Rlagnesiuin Concentration, P.P.M.
a
.Ihsnrbance" 510 nip
Absorbance unit
% of
magnesium present
These values are the arerages of 11 t o 13 replicate determinations.
The spectrophotometric sensitivity is 0.0008 y of magnesium per sq. cm. For comparison the recorded sensitivities of other methods are: Titan Yellow, 0.006 7 ; quinalizarin, 0.017 y: and
A N A L Y T I C A L CHEMISTRY
204
Brilliant Yellow, 0.02 y of magnesium per sq. cm. (8, p. 649). The practical sensitivity of the new reagent may be taken as 1 part of magnesium in 50,000,000 parts of solution. Solutions containing this concentration will have absorbances that are about 0.015 unit greater than those of solutions which contain no magnesium. Beer’s law is obeyed up t o 0.2 p.p.m. of magnesium; in addition, absorbances are sufficiently reproducible to allow the range of the determination to be extended to 0.4 p.p.m. The concentration range for optimum precision is 0.12 t o 0.4 p.p.m. of magnesium. Compensation for Calcium Interference. Interference by calcium is of special importance because it frequently occurs with magnesium and because of the difficulty in separating small quantities of these ions. In the determination of magnesium in the presence of calcium, the absorbance a t 510 m r is greater than when the calcium is absent. Immediately on preparation of the colored solutions, a red color forms, but i t fades almost completely within an hour, leaving a solution with an absorbance only slightly higher than if calcium had been absent. The enhancement of color due t o the presence of calcium is not entirely a function of the calcium concentration. When the concentration of calcium is increased, the absorbance increases up to 0.4 p.p.m. and remains constant up to 12 p.p.m., above which a red precipitate is formed. Hence, if the calcium concentration in the colored solution is between 0.4 and 12 p.p.m., or is adjusted to this concentration range, it is possible to compensate for the interference. The procedure for the determination of magnesium in the presence of calcium is identical with that already given, except that it is necessary to make certain that the calcium concentration will fall within the specified limits (0.4 t o 12 p.p.m.) when a determination is performed and when the working curve is prepared. I n addition, it is necessary to allow sufficient time for the initial color due to calcium to fade. After 1 hour this fading is nearly complete; the average change in absorbance during the second and third hours is about 3%. Because the fading of solutions that contain varying amounts of magnesium proceeds a t about the same rate, it is possible t o achieve good precision by using a waiting period of 1 5 minutes. The precision and sensitivity of the method modified to compensate for the interference by calcium are about the same as those of the unmodified method. A series of 14 replicate determinations that contained 0.08 p.p.m. of magnesium and 8 p.p.m. of calcium showed a standard deviation of 5.2% of the magnesium present. The spectrophotometric sensitivity of the modified method is 0.0005 y of magnesium per sq. cm., compared with 0.0008 y of magnesium per sq. cm. for the unmodified method. SEPARATION O F INTERFERING METAL IONS
Extraction with 8-Quinolinol. 8-Quinolinol (8-hydroxyquinoline) reacts with 23 metal ions in neutral or weakly acid solution to give quinolinates which are soluble in chloroform (8, p. 115); under these conditions it does not react with magnesium. To show that the proposed method of separation is compatible with the determination, experiments were performed involving the separation of the following metal ions from magnesium: aluminum, cadmium, cobalt, copper(II), iron(III), manganese(II), nickel, and zinc. For this purpose, solutions containing 50 y of magnesium and 200 y of each of the other ions in approximately 10 ml. of dilute hydrochloric acid ( 1 9 ) Rere used. The following procedure gives satisfactory separations.
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The p H of the solution is adjusted to approximately 7 with 5 % sodium bicarbonate solution. The solution is then extracted five times with 5-ml. portions of 5% 8-quinolinol in chloroform and twice with 5-ml. portions of chloroform. Magnesium may then be determined in an aliquot of the aqueous phase without further treatment. When iron is present in quantities exceeding 100 y, it is desirable to adjust the pH initially to approximately 3, then
extract with two or three portions of 8-quinolinol solution, neutralize the solution, and proceed as above. In this way, excessive hydrolysis of iron salts is avoided; otherwise incomplete separation might result. When only 1 or 2 mg. of metal ions are present it is not necessary to use buffer solutions to control the pH, because the amount of hydrogen ion liberated is not sufficient to affect the acidity appreciably a t p H 3; and the buffer capacity of the system produced by the addition of the bicarbonate is sufficient to prevent large changes a t p H 7 . When more than 1 or 2 mg. of metal ions are present, as in the precipitation of major constituents of the sample, it is necessary t o add a buffer solution.
Table 111. Recovery of Magnesium from Resin Column after Separation of 100 y of Magnesium from 1 l f g . of Calcium Per Cent 104.8 91.4 88.2 94 0 97.4
IV.
95 2
Table IV. Concentration of 3fetals in Whole Human Blood Constituent Aluminum Boron Calcium Chromium Copper Iron Lead Magnesium Manganese Nickel Phosphorus Potassium Sodium Zinc
Concentration
P.P.M. 0.1 1.0 50 1 1.2 400
0.6
50 0.1
0.1 100 2000 2000 15
Separation of Calcium and Magnesium. When small quantities of calcium and magnesium are to be separated, the usual methods (precipitation of calcium oxalate or calcium sulfate) are unsatisfactory, owing to solubility and coprecipitation losses. A procedure has been devised, based on an ion exchange frsctionation, which permits the separation of microgram quantities of magnesium from calcium. During the development of a procedure for the separation of milligram amounts of the alkalies, Sutton and Almy (9) noted that calcium and magnesium were also fractionated. A similar procedure has proved effective for the present purpose.
PROCEDURE. Wash the resin column with 25 ml. of 2.0K hydrochloric acid and use the washings as a blank. Then place on the column the sample containing magnesium in a weakly acid solution, the volume of solution corresponding to the sample volume being discarded. Pass 30-ml. portion of the 2.ON hydrochloric acid elutriant through the column at a flow rate of 1 ml. per minute (20 to 50 em. of mercury pressure), discarding the first 5 ml. and reserving the remainder for the magnesium determination. Evaporate both the blank and the solution containing magnesium to dryness in silica dishes and take up in 10 to 15 ml. of 0.002N hydrochloric acid. Dilute this solution to a volume which permits withdrawal of 1 to 10 y of magnesium in a 3- to 5-ml. aliquot. Analyze identical aliquots of the solutlon containing magnesium and of the blank by the procedure for the determination when calcium is not present. Wash the column \?ith 20 ml. of dilute hydrochloric acid (1 f 1) to remove calcium and then back-wash with approvimately 50 ml. of water. The column is then ready for use again. DISCUSSION.The new procedure has been used in the separation of 25 to 100 y of magnesium from 1 to 2 mg. of calcium. The data in Table I11 illustrate the efficiency of the separation.
205
V O L U M E 28, NO. 2, F E B R U A R Y 1 9 5 6 The ion exchange fractionation can be combined with the 8-quinolinol extraction by placing the aqueous phase remaining after the extraction on the column and eluting it according t o the recommended procedure. Two dilute hydrochloric acid solutions that contained 50 y of magnesium, 200 y each of aluminum, cadmium, cobalt, copper, iron, manganese, nickel, and zinc, and 1 mg. of calcium were subjected t o the combined separation procedure. Determination of magnesium in aliquots of the resulting solutions showed 100% recovery from each. The improvements in these recoveries over those listed in Table I11 may be due t o compensating errors. Starting with a sample in solution, the combined separation and determination requires from 3 t o 4 hours. APPLIC4TION TO COMPLEX MATERIALS
For the purpose of demonstrating the applicability of these procedures to the separation and determination of magnesium, two types of samples were chosen. The fimt a a s a “sbnthetic blood ash” solution containing many of the metallic elements found in human blood. This was analyzed using the extraction and ion exchange techniques to remove interfering ions. The second type of sample was a group of low-magnesium limestones, chosen because their high calcium content permitted a test of the modified procedure. Preparation of “Synthetic Blood Ash” Solution. Values for the metals present in whole human blood given by A41britton ( I ) , together with values made available by the Pratt Trace Analysis Laboratory, meie used in the preparation of the solutions for this part of the a-orb. These values are given in Table IV.
For the analysis of blood ash, a working curve should be prepared by carrying solutions of known magnesium concentration through the combined separation and determination procedure. hnalyses may then be carried out under the conditions used to obtain the data for the working curve, and corrections for ion exchange column blank and for failure t o obtain complete recovery from the separation need not be made. The working curve should cover a range of 25 t o 100 p.p.m. of magnesium in the initial blood sample. Table V shows results of analyses of synthetic blood ash solutions made in this manner. Analysis of Limestone. The analysis of limestone for magnesium was carried out, in part, by the usual procedure. The sample was decomposed with hydrochloric acid and the silica dehydrated by baking on a low temperature hot plate. The hydrous oxides were precipitated with ammonium hydroxide, using bromine water to oxidize iron and any manganese present.
Table 1.1. Determination of Magnesium in Limestone Sample 40.
Magnesium Oxide Presenta,
cc
Sample NO. 1
2
50.0
Magnesium Found, P.P. 11. 23 28 -417. 26 57 47 49 59 47 48 49
47
52
9; 3
.4Y.
50
.4Y.
93 107 100
100
Standard deviation from mean magnesium concentration, 7 67,.
Difference,
5
0.7.5 0.85
Av.
2
0.69 1.04 0.83
-0.02
0.79 0.70 0 69 0.81
.4r.
0.73 0 .7 3
-0.08
AY.
0.9fi 1.06 1.13 1.21 1.10
rO.OO
1.76 1.87 1.71 2 07
Analysis of Synthetic Blood Ash Solutions Magnesium .4dded, P.P. hl. 25.0
%
1
1.10
Table V.
Magnewmi Oxide Found,
1.96 .4r. 1 , 8 5 Standard deviation from certified value 0.12 a
-0.11
Standard Sample Co., dines, Iowa.
Ammonium salts were destroyed by evaporation with concentrated nitric acid and the residue was taken up in dilute hydrochloric acid. Magnesium was determined in an aliquot of this solution by the authors’ procedure modified for the determination of magnesium in the presence of high concentratione of calcium. The ratio of calcium t o magnesium in limestone is sufficiently high to permit the determination to be made without the addition of calcium to the final solution. The results of analyses of four standard samples of low-magnesia limestones are shown in Table VI. LITERATURE CITED
Upon ashing a sample of dried blood in a muffle furnace. the organic matter is burned and driven off, leaving a n ash which consists largely of metallic carbonates and oxides. When this ash is dissolved in hydrochloric or nitric acid, a solution of chlorides or nitrates, is obtained. The synthetic blood ash solution, therefore, was prepared from the chlorides or nitrates of the elements listed in Table IV, except in the case of zinc, which was added as the acetate, and of phosphorus, which was added as sodium dihydrogen phosphate. The xeights of the salts used yield the concentrations of the metals listed, except for sodium, potassium, and boron; the latter elements were not considered in the preparation of the solution, because the amounts that would be introduced by the reagents used in the analysis were large, compared with the quantities originally present in the aliquot of blood ash solution.
Albritton, E. C., “Standard Values in Blood.” pp. 117-19, W. B. Saunders Co., Philadelphia, 1952. Deterding, H. C., and Taylor, R. G.. IND.EN(;.CHEM.,A N A L . ED. 18, 127 (1946).
Dirscherl. W., and Brener, R., Mikrochemie cer. M i k r o c h i n . Acta 40, 3 2 2 (1953).
Harley, J. H., and Wiberly. S. E., “Instrumentnl .\nalysis,” p. 4 2 , Wiley, New York, 1954. Harvey, A. E., Komarmy, J. h l . , and Wyatt. G. >I., A N A L . CHEM.25, 498 (1953).
Kolthoff, I. M., Chem. Weekblad 24, 254 (1927) Marriott, W. M., and Howland. J., J . B i d . C‘hern. 32, 233 (1917).
Sandell, E. B., “Colorimetric Determination of Traces of Metals,’’2nd ed., Interscience, Yew York, 1950. Sutton, W. J. L., and Slmy, E. F., J . Dairy Sci. 36, 1248 (1953). Young, A., Sweet, T. R., and Baker, B. B., . ~ N A L .CHmf. 27,366 (1955). RECEIVED for review- July 27, 1955.
Accepted October 27, 1955.
