V O L U M E 26, N O . 3, M A R C H 1 9 5 4
553
Table 1. Rj Values of Substituted Quinones and Colors Produced by Chromogenic Spray, 5 % Aqueous Sodium Hydroxide Compound 1.4-NaphthoquinoneUnsubstituted 2-Methyl2-Methoxy2,3-Dimethoxy2-Methox~-3-h~drosv2-HydroxyZ-Methy1-3-acetyl3-Methyl-6-succino2 3-Dichloro2k!hloro-3-methoxy-
*
2-Chloro-3-ethoxy-
Fa+ +.- salt of 2-chloro-3-hydroxy.
2-Amino2-Amino-3-chloro2-Acetylamino2-Dimethylamino2-Chloro-3-dimethylamino2-Chloro-3-ethylamino-
Rf
(One-dimensional chromatograms) Colorn
0.85 0.87 0.81 0.84 0.37 0.47 0.83 0.53 0.41 0.33 0.86 0.27 0.89 0.42 0.82 0.83 0.83 0.83 0.87 0.88
OB OB 0 PUP PUP 0 PaB P Y 0 Pa0 PaRO RO Op TB
0 POB
0
OP PO
RI
Compound
1,4-Saphthoquinones 2-Chloro-3-n-decylaminoZ-(S-Acetanilido)-3-chloro2-Methylmercapto- . 2-Mercapto-3-chloro-
Z-Methyliminonaphtho(2,3)1,3-Dithiole-4,9-dione-methochloride monohydrate 1,2-Naphthoquinone Benzoquinones p-Benzoquinone p-Toluquinone p-Xyloquinone 2.5-Dichloro-p-benzoquinone
2.5-Dichloro-3,6-dicarbethoxy-a-benzoquinone 2.3,5,6-Tetrachloro-p-benzoquinone 3,4,5,6-Tetrachloro-o-benzoquinone Others 2,2,3,4.2-Pentachloro-l-ketotetrahydronaphthalene
Color PaY PaOB
0.91 0.89 0.84 0.43 0.95
Y Y
0.92 0.46 0.84
PaYB P PaY
0.87 0.85 0.97 0.28 0.93
PaR YR PaB PaT PaY
0.94 0.22 0.90
PaY PaY PaYB
0.90
PaPR
PaY
P a , pale; P, pink: B, brown: Y ,yellow: 0, orange; R , red: P u . purple. Impure compounds producing t a o spots on chromatographing.
running in the standard solvent, but before spray development. These eluates and the corresponding solutions prior to running on paper were compared on a Beekman Model DU spectrophotometer. The absorption curves produced mere superimposable in the four cases tried, indicating that no change occurred during the solvent run. This method would be of great aid in the isolation and identification of naturally occurring naphthoquinones. In working with the above procedure, it was found that flavonoid and phenolic substances were also chromatographed and developed. However, other solvents and chromogenic sprays are better suited for the isolation and identification of such compounds (1, 2 ) . Spectrophotometric absorption curves from eluate samples of chromatographic spots will serve as a means for preliminary classification of the compound producing the spot.
ACKNOWLEDGMENT
The authors are grateful to the Naugatuck Chemical Division, U. S. Rubber Co., for supplying the majority of the quinones studied. This research was supported in part by a grant from the Sational Science Foundation, Washington, D. C. LITERATURE CITED
(1) Bate-Smith, E. C., and Westall, R. G., Biochem. et Biophys. Acta, 4, 427 (1950). ( 2 ) Gage, D., and Wender, S. H., ASIL. CHEY.,23, 1582 (1951). (3) Jeanes, .4llene, Wise, C. S., and Dimler, R. J., Ibid., 23, 415
(1951). (4) LIcSew, G. L., and Burchfield, H. P., Contrib. Boyce Thompson Inst., 16, 357 (1951). RECEITE D for review J u n e 11, 1953.
Accepted September 17, 1953.
Flame Photometric Determination of Phosphate WILLIAM A. DIPPEL, CLARK E. BRICKER, and N. HOWELL FURMAN Department o f Chemistry, Princeton University, Princeton,
EVERAL
extensive discussions of flame photometric inter-
S ferences have appeared in the literature 8). Although cations, anions, and organic substances have been investigated, (1-6,
the cationic interferences have been studied most thoroughly. Probably the only useful generalizations regarding these interferences are that both their directions and magnitudes are dependent upon the instrument used and that the interferences are minimized a t high dilutions. In making a more extensive study of anion interferences, it was found, in agreement Kith the work of Parks et al. ( 6 ) , that phosphate had a pronounced inhibiting effect on the flame intensity of potassium and a lesser inhibiting effect on sodium. Its effects on calcium or magnesium are most striking, however, since a plot of emission intensity against concentration of phosphate a t a constant calcium or magnesium concentration exhibits a reversal (Figures 1 and 2). &4tthe lower phosphate concentrations, the emission intensity of calcium varies linearly and inversely with the phosphate Concentration. At higher phosphate con-
N. J. centrations, the emission intensity passes through a minimum and then increases until finally enhancement occurs a t phosphate concentrations exceeding approximately 1X for magnesium or 1.5.11for calcium. The linear relationship between phosphate concentration and emission intensity has been made the basis for the quantitative measure of phosphate in the concentration range between 0.005 and 0.012.11 phosphoric acid. Rieman and Helrich ( 7 ) employed an ion exchange column to remove interfering cations prior to a pH titration of the phosphoric acid present in a sample of phosphate rock. Likewise, the procedure described here employs a cationic exchange resin on the hydrogen cycle to remove all calcium and other cations present in the sample. The phosphate passes through the column and is collected in the effluent solution. After a known amount of a standard calcium solution is added, the intensity of the calcium flame is measured on the flame photometer. The weight of phosphorus pentoxide present is obtained from a calibration curve previously prepared from a series
554
ANALYTICAL CHEMISTRY
of solutions containing the same amount of calcium that was added to the unknown and varying known amounts of phosphoric acid. REAGENTS AND APPARATUS
All reagents used were C.P. grade chemicals. The 10,000 p,p.ni, calcium solution was made bv TTeighing out 2.4973 grams of calcium carbonate, treating with a slight excess of concentrated hydrochloric acid, evaporating to dryness, taking up in distilled water, and diluting to 100-ml. volume. This solution was stored in a polyethylene bottle. The cation exchanger used in this work was 20- to 50-mesh Dowex 50 resin. This nuclear sulfonic acid resin n-as chosen because i t vas available in the laboratory and preliminary tests showed that it removed all cations quantitatively from the solutions under the conditions required for dissolving the samples. Possibly some other resin might have been a better choice, and many other resins would undoubtedly work as well, The ion exchange columns n ere made by cutting off the tops of broken 50-ml. burets so that a column of resin 8 to 9 inches long could be supported in the buret. Each of these tubes contained 10 grams of air-dry resin supported on a plug of glass wool placed above the stopcock to prevent the resin from washing out of the column. The photometer used was the Beckman DU equipped with a standard photomultiplier detector and with a Model 9200 oxyacetylene burner attachment.
flame photometric readings was aln ays within f1%T. This fluctuation in the intensity of the flame corresponds to a variation of rt0.0002M in the phosphate concentration. Taking known phosphate samples through the exchange resin, the recovery was found to be as good as the reproducibility of the flame. Thus, the accuracy of the procedure is determined by the reproducibility of the photometer and not by the remainder of the procedure. When samples from two naturally occurring phosphate rocks were analyzed by the recommended procedure, the results shown in Table I were obtained.
Table I. Sample Bureau of Standards phosphate rock s o . 56 (31 5.5% PzOa)
Analyses of Phosphate Rocks Plot Found, %
Sample Taken, Gram 0.1320
32.20
0.2413 0.0954
0.1390 0 1150 0 1208
31.18 31.99 31.16 3 1 48
Average Average error Average deviation
RESULTS
Using known amounts of phosphoric acid and not taking the samples through the exchange resin, the reproducibility of the
f O . 17 0.45
41.99
40.56 41.31 40.45 41.02
PROCEDURE
A sample of dried, poffdered phosphate rock estimated to contain between 0.0036 and 0.085 gram of phosphorus pentoxide is weighed into a 50-ml. beaker to which 10 ml. of concentrated hydrochloric acid and 5 ml. of concentrated nitric acid are added. This is heated to gentle boiling on a medium hot plate for 1 hour, then evaporated to dryness, and baked for about 15 minutes to dehydrate the silica. The dry residue is then taken up in 2 drops of concentrated hydrochloric acid and 20 ml. of vater. This mixture is poured into the top of the column through a funnel containing a filter paper to prevent the insoluble silica from entering the column. The filter paper and the column are washed with distilled water until a total of about 85 ml. of effluent solution is collected in a 100-ml. volumetric flask. About 10 minutes are required to collect this quantity of effluent if the stopcock on the column is wide open. After all cations present in the original sample have been replaced by protons. 10 ml. of the standard 10,000 p.p.m. calrium solution is pipetted into the volumetric flask and the resulting solution is diluted to volume with distilled water. The intensity imparted to the flame by this solution, now containing 1000. p,p.m, of calcium and an unknown amount of phosphoric acid, is measured on the flame photometer a t 422.7 mp. The slitnidth and sensitivity settings are so chosen to give a dial reading of 100% T for a solution containing 1000 p.p.m. of calcium and no phosphoric acid. The calibration curve is previously constructed by adding known amounts of standardized phosphoric acid to solutions containing 1000 p.p.m. of calcium. The same sensitivity and slit-LYidth settings are maintained for the unknowns and the standards and all measurements are made a t a wave length of 422.7 mp. .A convenient method of operating the ion evchange columns employs three identical columns packed with Donex 50. Two of these columns are connected in series with a rubber stopper, so that the flow rate is the same in both columns and can be controlled by a single stopcock. Jl'hile these two columns are being used for removing cations from samples, the third column is regenerated by slowly passing 3N hydrochloric acid through i t from a large reservoir. The regenerated column is washed with distilled water before i t is used for removal of cations. After four samples have been run through the series combination, the top column is placed on the regeneration cycle, the lower column is placed on top, and the newly regenerated column is glared on the bottom. By rotating columns in this manner, a fresh column is always being prepared and the series arrangement provides extra exchanging capacity to ensure against any possible break-through by calcium or other cations which might affect the emission intensitv.
32.32 31.72
d 2 71
Average Average error Average deviation
ii 34
+O 14 0 67
DISCL5SION
Procedure. The procedure finally adopted was shown to be more convenient for the treatment of representative phosphate rocks than two alternative procedures which were also tried. All procedures used the treatment nith exchange resins described in the recommended procedure. First, a hydrofluoric acidperchloric acid attack which required the use of platinum crucibles was tried. Bumping was a constant source of trouble even when a sand bath n-as used and the time allon-ed to take the samples to dryness n-as excessive. Both precision and accuracy were poorer by this method than one would expect from the reproducibility of the burner alone. However, it is possible that this attack could be used if a more refractory sample is encountered. Secondly, the procedure of Rieman and Helrich (7) employing pure 12.V hydrochloric acid did not completely dissolve the Bureau of Standards sample. Therefore, the results were always IOIY and the precision was extremely poor; the values for the phosphorus pentoxide content varied between 21.3 and 28.8%. With more easily soluble samples this procedure may be satisfactory. Effect of Phosphoric Acid on Flame Intensities. Figure 1 shows the effect of various concentrations of phosphoric acid on the flame emission of 50 p.p.m. of sodium, 50 p.p.m. of potassium, 100 p.p.m. of calcium, and 500 p.p.m. of magnesium. It is obvious that the phosphoric acid causes a reversal in the emission intensities of magnesium and calcium, but i t only tends to decrease the intensity of the sodium or potassium flame. The effect of phosphoric acid on sodium is much less than with any of the other cations and, furthermore, is nearly independent of concentration beyond 0.4M phosphoric acid. Phosphoric acid itself shows an almost continuous emission between 320 and 700 mp, which increases linearly with phosphoric acid concentration a t any given wave length. This emission of phosphoric acid may very well explain the horizontal portions of the curves for sodium and potassium shown in Figure 1that is, the inhibition of the emission intensities of sodium or potassium might continue to increase with increasing phosphoric acid concentrations, but this inhibition is cancelled by the selfemission of the phosphoric acid. On the other hand, the emis-
555
V O L U M E 2 6 , NO. 3, M A R C H 1 9 5 4
I01V-. ,
1.0
9
'
"
"
"
Figure 1.
"
2.0 MOLRRITY
"
3.0
'
I
1
'
4.0
H,PQ
Effect of Phosphoric .Acid on Flame Intensities
A . 50 p.p.m. of sodium, wave length 589.3 mp, 0.02-mm. slit width 50 p.p.m. of potassium, wave length 767 mp. 0.10-mm. slit width C. 100 o.n.m. of calcium, wave length 422.7 mp. 0.025-mm. slit widihD . 500 p.p.m- of magnesium, wave length 370.8 mp, 0.06-mm. slit width
B.
sion intensity of phosphoric acid is much too weak to account for thp enhancement observed for both calcium and magnesium. Interferences. Since phosphoric acid has such a significant effect on the emission of calcium and magnesium, it is important to know the effects of other anions. Much additional work is required in order to draw quantitative conclusions but it can be said that the effect of phosphate on the emission intensity of the calcium flame is greater a t low concentrations of calcium than a t higher concentrations. Perhaps this same conclusion cannot be drawn for all anions. A summary of the results from a preliminary examination of the effects of various anions on the intensity of the calcium flame is shown in Table I1 The extent of the individual interferences depends upon the concentration of calcium as well as that of the interfering substance. For example, riith 100 p p m of calcium, 0.06M perchloric acid showed a 4OY0 enhancement 7%-hichdecreased nearly linearly until, a t 3.231, only S70 enhancement vias observed. With 1000 p.p.m. of calcium. 0.06.11 perchloric acid caused'only 17% enhancement. Since a 0.04-mm. slit width was used with 100 p.p.m. of calcium and a 0.02-mm. slit width n-ith 1000 p.p.m., the per cent of enhancement in these tn-o caqei: mny not be as divergent as the numerical figures indicate.
a t 0.3631 hydrochloric acid. As the concentration of the hydrochloric acid is increased, the enhancement slowly decreases until the flame appears to be unaffected a t 4.5M hydrochloric acid. Blthough the mechanism of flame photometric interferences of the type mentioned is not fully understood, i t can probably be said that these are chemical or photochemical effects, that they are not purely a result of instrumental factors, and that the viscosity is an insignificant variable. I t has been shown that the rate of sample aspiration is inversely proportional to the specific viscosity. If the logical assumption be made that the flame intensity is directly proportional to the rate of aspiration, the flame should vary inversely with phosphate concentration if a visccsity effect is solely responsible for the inhibition. That this is not so is obvious when one considers the very strong inhibitions shown in Figure 1. It is suggested that a process which robs the flame of some of its excitation energy is responsible for the extreme inhibition a t low phosphate concentrations. This process could be anhydride formation or merely a heat capacity effect. Because the energy necessary for dehydration of phosphoric acid is relatively large compared to other acids, it would be expected to exert the greatest cooling effect and therefore the greatest inhibition. S o good explanation for the enhancement of the calcium and magnesium flames is known. Calibration Curves. The effects of various concentrations of phosphoric acid on the intensities of the flames from three different calcium concentrations are shown in Figure 2. I t is Yignificant that for each calcium concentration there is a range of phosphate concentrations over which the inhibition of the calcium flame increases nearly linearly. Although the range of phosphoric acid concentrations over n-hich these curves are linear is very small for the low calcium concentrations, this method for detecting phosphate is most sensitive a t low calcium concentrations. S o effort has been made to determine the limit of detection of phosphate by the inhibition of a very weak calcium flame.
0 "
"
'
"
.005
"
"
.o10
"
MOL R R I T Y
Table 11. Interferences Calcium Concn., P.P.M. 100 100 100 1000 50
1000 1000
Substance -4dded
HC1 H&Oi
HClOh HClOi HCsHiOz HaF2 HsSiOP
Concn. Range, Molarity 0 06 - 4 . 5 0.36 - 2 . 0 0.06 - 3 . 2 0.01- 0.12 0 . 0 4 -5.3 0.002-0.014
... ..
T y p e of Interference Enhancement Inhibition Enhancement Enhancement Enhancement above 0.2.U N o Effect Inhibition
Figure 2. Effect on Phosphate Determination Low High Low
A. B.
"
"
.Ol5
"
"
.020
H3FQ
Calibration Curves for Phosphoric Acid Using Calcium Emission 100 p.p.m. of calcium, 0.04-mm. slit width 500 p.p.m. of calcium, 0.02-mm. slit width
C . 1000 p.p.m. of calcium, 0.02-mm. slit width Wave length of 422.7 mp used for all curves
LOW
Low Sone High
a Effect of this substance was presumed f r o m high results obtained when silica was not dehydrated.
With sulfuric acid, the calcium flame is inhibited about 35% over the entire range from 0.36 to 2.0M sulfuric acid. KO data were taken for lorn sulfuric acid concentrations with calcium but i t is known that this acid shows less pronounced inhibition of the potassium flame than does phosphoric acid. With hydrochloric acid, the calcium flame is gradually enincrease in the emission intensity is observed hanced until a
The most useful calibration curve was that using 1000 p.p.m. of calcium. Points on this curve were checked frequently over a period of 6 months and were always found to agree to within zizl%T. The effect of phosphoric acid on the intensity of the magnesium flame, as shown in Figure 1, is even more pronounced than that of calcium. The dopes of calibration curves using magnesium are accordingly very steep, but suffer from the disadvantage that only a limited range of phosphate concentrations can be analyzed from each curve. Comparison with Other Methods. Classical methods for phosphate involve its separation as the ammonium phosphomolybdate followed by a gravimetric, titrimetric, or colorimetric
556
ANALYTICAL CHEMISTRY
determination. Rieman and Helrich ( 7 ) separate the phosphate by a cation exchange resin and then perform a pH titration. The procedure described in this paper offers no new technique for separating phosphate but provides a new method for its determination. The flame photometric technique is rapid after the separation has been completed. The accuracy that can be expected is of the order of 1% when only a single determination is made. However, if several analyses are averaged, as shown in Table I, the accuracy may be as good as 3 to 5 parts per thousand. High concentrations of various anions mill interfere with the phosphate determination by this method. This makes the accurate determination of low percentages of phosphate in certain mixtures unattractive. Other Applications. It is obvious from the data presented that a small amount of phosphate constitutes a serious interference when calcium, magnesium, or potassium is being determined by the flame photometer. Thus, in order to determine any of these cations accurately in the presence of phosphate, the phosphate should be removed by an anion exchange resin prior to the determination. ..ilthough the determination of phosphate by the flame photom-
eter may have some limitations, it is significant that an inhibiting substance can be determined even though it does not emit in a Lundegardh flame. This principle may have some other useful applications. LITERATURE CITED
Barnes, R . B., Richardson, D., Berry, J. W., and Hood, R. L., IND. ENG.CHEM., ANAL. ED., 17, 605 (1945). Berry, J. W., Chappell, D. G., and Barnes, R. B., Ibid.,18, 19
(1946). . 22,1530 (1950). Conrad, A. L., and Johnson, W.C., A s . 4 ~CHEX., Gilbert, P. T., Hawes, R. C., and Beckman, A. O., Ibid., 22, 772 (1950). Leyton, L., Ann. Repts. Progr. Chem. (Chem. SOC. London), 45, 326 (1948). Parks, T. D., Johnson, H. O., and Lykken, L., ANAL.C m x , 20, 822 (1948). Rieman, W., and Helrich, K., IXD. ESG. CHEM.,ANAL.ED.,19, 651 (1947). \5'est, P. W., Folse, P., and llontgomery, D., ANAL.CHmi., 22, 667 (1950). RECEIVED for review July 25, 1953. Accepted October 24, 1953. T h i s research was supported b y Contract AT(30-1)-937, Scope I of the U. S. lltomic Energy Commission.
Amperometric Titration of Calcium With the Disodium Salt of Ethylenediaminetetraacetic Acid H. A. LAITINEN and R. F. SYMPSON Department o f Chemistry and Chemical Engineering, University o f Illinois, Urbana, HE use of an indicator in amperometric titrations was first described by Ringbom and Wilkman (IO),who added a substance yielding a polarographic current to the titration mixture. The indicator material would not react during the course of the titration, but would react with excess reagent in such a way that a sudden change in polarographic current at some appropriate applied potential occurred a t the end point. The indicator material was added in relatively small concentration, so that the observed change occurred in a narrow interval of the titration. In the present titration the indicator is present in a concentration comparable with that of the sample being determined. The end point is then determined by the traditional extrapolation method, which does not require the addition of small increments of reagent in the vicinity of the end point. The choice of an indicator ion is complicated by the fact that ethylenediaminetetraacetic acid (Versenate) forms more stable complexes with most of the common metallic ions than with calcium. It was desired to find an ion which could be complexed by some other agent to such a degree that the 1-ersenate would react with the calcium until it was titrated and then form the Versenate complex with the indicator ion, decreasing its diffusion current proportional to the decrease in its concentration. I n the ideal case the current would remain constant (except for dilution) until the end point, after which a linear decrease would occur. In selecting such an indicator ion and complexing agent, the equilibrium which exists a t the equivalence point and following the equivalence point must be considered. At the equivalence point of the calcium titration, the following equations describe the equilibria: MX;+p"
+ Cay--=
MYm-4
+ Ca++ + pXn
(1)
where AI" is the indicator metal ion, X" is the competing complexing agent, and Y - - - - is the Versenate ion. +
111.
The equilibrium constant, KA, may be evaluated from K,, K,, and K 8 , which are the dissociation constants of the following complexes:
cay-- $ cat++ y----
'
[ C a ~ + ] [ r ' - - - - ] = Kl [Cay--]
The constant K A should be so small that Reaction 1 does not proceed to the right to any appreciable extent. Beyond the equivalence point, the following equilibrium must be considered: Jfx:+P"
+ Y----
MYm-4 f p-P
(2)
The equilibrium constant is given by
The constant K B must be large enough so that Reaction 2 proceeds quantitatively to the right as soon as an excess of Y----is present beyond the calcium end point. I t is not possible to make accurately qumtitative calculations using these equilibrium constants because of large and unpredictable ionic strength effects. In a qualitative way, however, it is possible to predict a combination of metal ion and complexing agent which might be satisfactory. According to Schwarzenbach and Ackerninnn (1%') the dissociation constant of the calcium Versenate complex is 2.6 X lo-". By an inspection of the magnitudes of the dissociation constants of various combinations of metal ions and complexing agents, it
