1358
ANALYTICAL CHEMISTRY
varied from about 0.3 to 25 and [H+] is varied from 1 tu 4. With increasing chloride concentration the value of K ' decreases in accordance with the greater transformation of F e + + + into FeCl++, FeC12+, FeC13, and possibly other species (2). A plot of log K' versus hydrochloric acid concentration appears to give a straight line in the range 1 to 4 AI (Figure 1). No great accuracy is claimed for the values of K' reported, and the clow conformity to linearity may be fortuitous.
9.0
L
8.5
(3
s
8.0
7.5 ~~~
1
P
3
4
HYDROCHLORIC ACID CONCENTRATION, M
Figure 1. Variation of K' for Ferric Cupferrate in Chloroform with Hydrochloric Acid Concentration A few determinations of K' in 1 AI perchloric acid were made The value 7 X 108 was obtained (average of 5 values ranging from 3.6 X 108 to 1 x 10*0), but this figure must be regarded as ver: tentative because of the unfavorable experimental conditions. When the concentration of free nitrosophenylhydroxyIaminein the chloroform phase is 0.1 M the ratio [FeCf3],/Z[Fe], for 1 IM
hjdrochloric acid medium is 6 X lo5, on the basis of the above value of K'. A double extraction under these conditions (final concentration of excess reagent 0.1 AI in the chloroform) should therefore reduce the ferric iron concentration of the aqueous solution to a very small value. The extent of extraction of such metals as aluminum, which do not readily give a cupferrate precipitate in mineral acid medium, cannot be stated at present, although a rough estimate of k" for aluminum can be based on the reported solubility products of ferric and aluminum cupferrates, which are, respectively, 1 X and 2 X according t o PyatnitskiI. If the solubilities of ferric and aluminum cupferrate in chloroform are roughly the same, the value of K' for aluminum rupferrate should be of the order 3 X lo2 in 1 Af hydroehlori( acid. The magnitude of this value indicates that the hydrogen ion and cupferron concentration may have to be controlled with somp care for a satisfactory separation when much iron is present. AP a matter of fact, aluminum cupferrate tends to be extracted from sulfuric acid solutions less acid than 4 Ar (3). If the extraction equilibrium is reversible (the reversibility does not seem to have been tested), a better separation can be achieved by applying the principle of retrograde extraction-i.e., the chloroform solution of ferric cupferrate containing a little aluminum cupferrate and excess cupferron is shaken with a hydrochloric acid solution of suiiable concentration containing added cupferron. By proper choiw of conditions, the major part of the aluminum in the chloroform phasp can be removed without transfer of any significant amniini of iron. LITERATURE CITED
(1) Pyatnitskii, I. V.,Zhur. Anal. Khim., 1, 57 (1946). (2) Rabinowitch, E., a n d Stockmayer, W. H., J . Am. Chem Soc 64 335 (1942). (3) Strafford, N., a n d Wyatt, P. F., Analyet, 72, 54 (1947).
R E C E I V ~June D 13, 1949.
Quantitative Study of Reaction between Beryllium and Quinizarin-2-sulfonic Acid MYRON W. CLCCI, W . F. NEUMAN, AND B. J. RIULRYAIN Department of Radiation Biology, University of Rochester, Rochester,
F
AIRHALL (1) has reported a colorimetric method for the determination of beryllium based on its reaction with an anthraquinone dye, quinizarin-2-sulfonic acid. An attempt to duplicate Fairhall's results led to a critical study of the factors involved in the reaction. The data obtained served as the basis for a revised procedure which permits the analysis of beryllium in quant,ities from 1 to 20 micrograms with a probable error of 3.1 %. EXPERIMENTAL
The procedure described by Fairhall (1) served as a point of departure for these studies. In investigating various anthraquinone derivatives as color reagents for beryllium, 1,4-dihydroxyanthraquinone-2-sulfonir acid (quinizarin-2-sulfonic acid), buffered a t pH 7.0 Kith ammonium acetate, was found to give a red color which is proportional to the amount of beryllium. The color develops rapidlv rprtrhps a maximum in 5 minutes, and does not fade for several
N. Y .
hours. The most satisfactory range for colorimetric comparison in a visual colorimeter is 1 to 20 micrograms of beryllium. KOcolor developed when beryllium and the dye were mixed under the conditions given by Fairhall (1). Heat (100" C. for 10 minutes) was necessary to develop the color. Because ammonium aretate is not an effective buffer in the region of pH 7.0, a search for a more suitable buffer was instituted. Histidine monohydrochloride was found to be a fairly effective buffer a t pH 6.5 and it did not interfere with the color reaction as did phosphate, bisulfite, borate, and maleate. It was also observed that the colored lake formed between beryllium and the dye could be easily salted out, but that this separation was prevented by the addition of gum arabic as a color stabilizer. With these preliminary changes in the Fairhall procedure, the reaction wm studied in more detail
Instruments. Measurements of absorption spectra were madr with a Model DU Beckman quartz spectrophotometer, using the
V O L U M E 21, N O , 11, N O V E M B E R 1 9 4 9
1359
A critical study has been made of factors affecting the reaction between beryllium and quinizarin-2-sulfonic acid, an anthraquinone dye suggested by Fairhall as a color agent for beryllium. Under optimal conditions of pH, temperature, etc., quinizarin-2-sulfonic acid may be used satisfactorily for the determination of microquantities of beryllium (1 to 20 micrograms) with a standard deviation of 4.65%. However, the conditions of operation must be rigorously controlled and numerous substances interfere markedly with color formation.
'blue-sensitive" phototube and 1-cm. Corex absorption cells. t'he colorimetric readings \$-ere made with a Klett-Summerson photoelectric colorimeter using the appropriate filter (interference type, Farrand Optical Company, New York, S.Y,). A Beckman pH meter, laboratory model G, wasused for all pH determinations. Reagents. Technical grade lI4-dihydroxyanthra uinone-2,ulfonic acid obtained from the Eastman Kodak %ompany, Rochester, N. Y., was purified by dissolving the sodium salt in ivater, acidifying with hydrochloric acid, pouring into 95% ethyl alcohol, and filtering the precipitate which formed. The reagent was prepared as a 0.l6y0 aqueous solution. Histidine monohydrochloride monohydrate was obtained from Sutritional Research Biochemicals, Cleveland, Ohio. The buffer nas prepared as 0.1 M solution containing ly0 gum arabic as a colloid stabilizer and adjusted to the proper pH by additions of 2.0 AVsodium hydroxide, A crystal of thymol was added to retard bacterial decomposition of the reagent. The standard beryllium solution was prepared by dissolving 1.OOO gram of spectrographically pure beryllium in dilute hydroPhloric acid, The solution was transferred to a 1-liter volumetric
flask and made up to volume with distilled water. The final acid concentration was approximately 0.1 AT. Portions of this stock solution w?reevaporated to dryness, and the residues were ignited to beryllium oxide and weighed; the average value of 1.00 mg. of beryllium per ml. \vas found. For analysis, appropriate dilutions of this stock solution were made. 5001
z
,400W
(r
LT
300 -
U
t
%zooCC
2 Ll
2
I /*..
/=16MG
a
loo-
I
1.6t
20MG
0
J
ol/'
I
I
0 IO 20 MICROGRAMS OF BERYLLIUM
I
30
Figure 3. Colorimetric Response of BerylliumDye Complex as Function of Dye Concentration
goo"
"
"
'
,
--..*.
400 500 WAVE LENGTH (MILLIMICRONS)
600
Figure 1. Absorption Spectra of Beryllium-Dye Complex 2;s. Reagent Blank A . Beryllium-dle complex B. Reagent blank
0
I
I
1
2
4
6
8 MG O F ADDED DYE
I
IO
Figure 2. Colorimetric Response of Beryllium-Dye Complex us. Reagent Blank Measured ageinst function of dye concentration
Effect of pH on Absorption Spectra. The color of the beryllium-dye complex was markedly affected by variations in pH. Accordingly, absorption spectra of the blank and the beryllium-dyc, complex were studied over the pH range from 4.0 to 9.0. The curves were obtained with solutions containing 6.0 microgranis of beryllium and 0.1- ml. of 0.5yo aqueous 1,4-dihydroxyanthraquinone-2-sulfonic acid, buffered with 2 ml. of 0.1 111 hist,idine monohydrochloride in 1% gum arabic solution, all in a totai volumc of 20 ml. Blanks identical in every way, but lacking tieryllium, were also prepared. The solutions were hea.ted at 100" C. for 10 minutes and cooled to room temperature under tap water prior to reading. I t was observed that, at acid pII, beryllium affected the alsorption of the dye only slightly. A4talkaline pH (8.8), beryllium increased absorption of the dye a t ivave lengths from 500 to 600 mH, but the blank itself absorbed strongly. Between pH 6 and 7 the beryllium-dye complex showed some absorption from 550 to 600 r n N where the absorption of the blank was negligible. I t was evident that, for color contrast (beryllium complex us. blank), a wave length of 575 mp was optimal and the reaction should 1 ) ~ buffered at pH 6.5 (cf. Figure 1). Effect of Dye Concentration. . 4 series of solutions wa., I J ~ V pared containing 4.0 micrograms of beryllium, 2 ml. of buffer solution (pH 6.5), varying amounts of dye solution, and distilled water to give a total volume of 20 ml. Blanks (no beryllium) \ : e ~ prepared for each dye level. The plot of colorimetric response VL d>-econcentration, shown in Figure 2, indicates that a dye coticei~:ration of about 2 mg. in a total volume of 20 ml. is optimal Le., there was a large net color difference with a low blank. Because previous studies ( 2 ) have shown that, under certait, 4v.mstances, the o2tirnnl quaiitit- of dye varies as a function (J{ ~
ANALYTICAL CHEMISTRY
1360
the amount of beryllium being analyzed, a series of standard curves was obtained employing varying quantities of dye. In these experiments, the v o h m e was 25 ml., pH was 6.5, buffer concentration was 0.01 N , and the beryllium content varied from 1.0 to 30 micrograms. The results are presented in Figure 3. I t is apparent that the color-beryllium relation does not conform to Beer's law. A series of curves was obtsined, the curvature of which varied with the quantity of dye added. An almost h e a r curve was observed with 1.6 mg. of dye. This quantity was considered optimal and was employed in most subsequent atudies. Effect of BufEer Concentration. A series of standard curves was obtained with varying concentrations of histidine buffer. [n this series, the following conditions were employed: pH 6.5, volume 25 ml., amount of dye 2 mg., beryllium content 2 to 16 micrograms. Three concentrations of buffer were studied: 0.1, 0 01. and 0.005 M . The results are presented in Figure 4.
.-
joOt
Table I.
Recovery of Known Quantities of Beryllium
Added
Found
Added
Y
Y
Y
0.50 1.0 2.0 3.0 4.0 5.0
0.55 0 . 9 5 , l .o, 1 , o 2.2 2.0 3 . 2 :3 . 1 , 3 . 2 4.3,4.2 5 . 1 ,5 . 2 , 5 . 3 6.2 6.1 6.8: 7 . 3 , 7 . 4
6.0 7.0
Table 11. Foreign Ion
8.0 9.0 10.0 11.0
13.0 15.0 17.0 19.0
Found 7
7.9,8.4 8.7,9.5,9.4 10.0,10.3 11.3 13.6 15.4 17.4 18.6
Effect of Interfering Substances Amount R e uired to Give Interjerence Equivalent to 0.5 y of Be IMQ.
Interference' Effect
f
/ 101 N
MICROGRAMS OF BERYLLIUM Figure 4. Colorimetric Response of Beryllium-Dye Complex as Function of Salt Concentration
Increasing the concentration of buffer increased the color intensity. Xot only did beryllium samples show greater color intensity; the blank was also increased. For this reason, a compromise was chosen as optimal: 0.01 M buffer was employed in Rubsequent work. Similar effects were noted when varying quantities of sodium chloride were added to the solutions being analyzed. It appears that the color reaction is sensitive to variations in the ionic strength of the solution and not to a specific interaction with the buffer. Effect of Time on Stability. A series of beryllium standards in the concentration range 1 to 20 micrograms per 20 ml. of solution was prepared together with blanks, heated a t 100" C. for 10 minutes, cooled to room temperature under tap water, and read in a Klett-Summerson photoelectric colorimeter equipped with a Farrand interference filter a t 575 mp. Readings taken over a 2-hour time interval to determine whether there was any appreciable drift with respect to time showed that the solutions were relatively stable for a t least 2 hours. In fact, there mas a slight increase (3 to 4%) in color intensity after standing. The time of hrating will vary somewhat with the diameter of the tubes employed by the analyst. Wide-diameter tubes may require as long as 30 minutes to reach full color development. Because overheating tends to reduce the color density, optimal conditions camlot be stated cateporicrJly but must be arrived a t by experimentatiori. PROCEDURE
Based on the conditions outlined above, the following procedure was adopted. Blanks and standards are analyzed with each group of unknouns.
a L reading lower and H reading higher than solutions containing beryl lium standard. CC, color change; P p t , precipitation.
One to 15 ml. of the solution to be analyzed were taken, the pH was adjusted to approximately 6, and the solution was transferred to a tube graduated a t 20 mi. Two milliliters of histidine buffer and 1.0 ml. of a 0.16% aqueous solution of quinizarin-2 sulfonic acid were added and the mixture was diluted to a volume of 20 ml. The tube was immersed in a boiling water bath for 10 minutes and cooled in tap water and the optical density was d e termined by means of a Beckman spectrophotometer (at 575 mp) or a Klett-Summerson colorimeter equipped with a filter (interference type, 575 mp, Farrand Optical Comprtny). Optical measurements were converted to micrograms of beryllium by means of a standard calibration curve which is essentially a straight line. Accuracy. A series of unknown solutions of pure beryllium chloride was prepared by a disinterested person and analyzed by the procedure given above. The results are presented in Table I. In view of the small amounts of beryllium analyzed, the accuracy was considered satisfactory. Interfering Substances. The influence of foreign ions was investigated. The experiments were made on solutions containing 10 micrograms of beryllium in 20-ml. volume. The values OD Table I1 indicate the quantities of interfering ions which cause an error equivalent to 0.5 microgram of beryllium. The common anions which might be used in the preparation of samples (sulfate and chloride) fortunately do not materially affect the color reaction, if the total molarity does not exceed 0.005 Many common cations, however, and fluoride and phosphate interfere markedly and thereby limit the applicability of the method. Discussion. Quinizaiin-2-sulfonic acid, suggested by Fairhall ( 1 ) as a colorimetric agent for bervilium, can be used successfully for the accurate analysis of beryllium only if the conditions of color development are rigorously controlled. Necessary precautions must be taken to exclude interfering substances, in particular, phosphate, fluoride, and aluminum. Because the method requires the use of pure solutions, its applicability is a t present restricted to analyses of beryllium dusts as used in inhalation studies and analyses of air samples around beryllium plants. The method may be applied to other substances if the beryllium is first isolated. LITERATURE CITED
(1) Fairhall,
L. T..Natl. Inst. Health, U.S. Public Health Service.
Bull. 181. 10-14 (1943). ( 2 ) Underwood, A , , Neuman, W-.F., and Carlson, A . B., University of Rochester, Atomic Energy Rept. M-1951 (1947).
RECEIVED June 9, 1919. Based on work performed under contract with the Atomic Energy Commission a t the University of Rochester Atomic Energs Project, Rochester, PIT. Y.
