Spectrofluorimetric Determination of Traces of Fluoride Ion by Ternary Complex Formation with Zirconium and Calcein Blue

Tan Lay Har and T. S. West Chemistry Department, Imperial College of Science and Technology, London, S. W.7., England

ALL SPECTROFLUORIMETRIC and, with two exceptions, all spectrophotometric procedures for the determination of fluoride ion are based on the ability of fluoride to abstract cations such as aluminium or zirconium from strongly ab- sorbing or fluorescing complexes, thus liberating the free reagent. Such methods tend to be unselective. The two spectrophotometric methods which differ are kinetochromic spectrophotometry ( I ) , which depends on the catalytic action of fluoride ion on the zirconium-xylenol orange reaction, and that based on the formation of a strongly absorbing 1 :1:1 ternary complex with cerium(II1) or lanthanum, alizarin complexan, and fluoride (2) . The kinetochromic method allows fluoride to be determined down to 0.005 ,ug/ml with an “effective” molar absorptivity of 2 X 105 but must be operated under rigidly controlled conditions. The alizarin complexan ternary complex method is easy to apply, how- ever, and in view of its high molar absorptivity, 3 X lo4, its stability, and the fact that the reaction is not interfered with by other anions, it seemed logical to look for other ternary systems for the determination of fluoride. It was considered that the sensitivity could be improved even further by a fluorimetric method and this led us to an investigation of the effect of fluoride on the fluorescence intensity of the zirco- nium-Calcein Blue complex (3).

Since its almost simultaneous introduction by Eggers ( 4 ) and Wilkins (5), Calcein Blue has been used extensively as a metallofluorescent indicator for the complexometric titration of several metal ions with EDTA. Only recently (3 ) has it been used as a direct fluorimetric reagent for the determination of zirconium at pH 5 . 5 . It was found that the addition of fluoride to the zirconium-Calcein Blue complex resulted in an enhancement of fluorescence intensity. In this paper we describe an investigation of this effect and the development of a method for the determination of fluoride based on this intensification of fluorescence

EXPERIMENTAL

Apparatus. Fluorescence measurements were made with a double monochromating spectrofluorimeter (Farrand Opti- cal Co. No. 10244) fitted with a 150-W Xenon arc lamp (Hanovia Division No. 901 C-1) and RCA IP 28 photomulti- plier and equipped with a Honeywell-Brown Recorder. Fused quartz cells (10 X 20 x 50 mm) were used throughout.

Reagents. SODIUM FLUORIDE SOLUTION 1O-jM. Prepare a lO-,M stock solution by dissolving 41.99 mg of analytical reagent grade sodium fluoride in a liter of distilled water. Ten milliliters of the stock solution were diluted to 1 liter to obtain a 10-6M solution.

1 ml of 10-5M F- E 0.1899 pg F

(1) M. L. Cabello-Tomas and T. S . West, Talanta, 16 781 (1969). (2) M. A. Leonard and T. S. West, J. Chem. SOC., 4477 (1960). (3) R. V. Hems, G. F. Kirkbright, and T. S. West, ANAL. CHEM.,

(4) J. H. Eggers, Talanta, 4 38 (1960). (5) D. H. Wilkins, ibid., p 182.

42,784 (1970).

CALCEIN BLUE lO-4M. Prepare a lO-3M solution by dis- solving 160.2 mg of Calcein Blue (Hopkin and Williams) in a few drops of 0.1M potassium hydroxide and diluting to 500 ml with distilled water. Ten milliliters of this solution were diluted to 100 ml with distilled water to give a lO-4M solu- tion. After 2-3 days, all Calcein Blue solutions were dis- carded and fresh solutions were prepared.

Prepare a lO+M solution by dissolving 32.2 mg of ZrOCln in 100 ml of 3 M HCl. It has been shown (6) that zirconium solutions 3 M in HCl do not polymerize on standing. Solutions (10-4M) were prepared by dilution with 3 M HCl.

Eight milli- liters of concentrated ammonia (18M) were diluted to 100 ml with distilled water.

Procedure. CALIBRATION CURVE. Transfer by pipet 5-30 ml (at suitable volume intervals) of 10-5M sodium fluoride into 100-ml flasks, Add 2 ml of 10-4M Calcein Blue and just sufficient NH, solution (ca. 3-4 ml of an approximately 1.5M NH3 solution) to adjust the pH of the final solution to 2.5. Mix the contents of the flask and dilute to volume.

The fluorescence of the solutions was measured after 20-30 min at 410 nm with the excitation wavelength set at 350 nm.

Unknown solutions containing 0.9-6 pg F (or 20.01 ppm in concentration) were analyzed in a manner similar to that used to set up the calibration curves.

Because of day-to-day variation in lamp intensity, it is necessary to run two or three standards through with each batch of unknowns, as in all single beam fluorimetric pro- cedures, to establish the position and slope of the calibra- tion curve.

ZIRCONYL CHLORIDE 10e4M.

AMMONIA SOLUTION (APPROXIMATELY 1 S M ) .

Finally, add 2 ml of lO-4M Zr in 3 M HCl.

RESULTS AND DISCUSSION

Spectral Characteristics. Figure 1 shows the excitation and fluorescence emission spectra of a 2 x 10-6M solution of Calcein Blue at p H 2.5, (a) and (b), respectively, a 1 :1 zirco- nium-Calcein Blue solution (c) and (d), respectively, and a 1 : 1 : 1 zirconium-calcein Blue-fluoride solution (e) and cf), respectively, at the same concentration and pH, and obtained with identical instrumental settings of slit and gain, etc. These spectra are uncorrected for variations in the emission, characteristics of the 150-W Xenon arc lamp, the response characteristics of the IP 28 photomultiplier, and the diffraction gratings.

It will be seen that at pH 2.5 the addition of zirconium depresses the fluorescence originating from Calcein Blue at 450 nm and produces a new emission maximum of lower in- tensity at 410 nm. Similarly, the excitation maxima are changed from 325 nm for Calcein Blue to 350 nm for its zirconium complex. The addition of fluoride to the zir- conium-Calcein Blue complex increases the fluorescence in- tensity, but the same maximum excitation and emission wave- lengths are retained as for Zr:CB. This indicates that a ternary complex is formed and eliminates the possibility that

(6) B. C. Sinha and S . Das Gupta, Analyst (London), 92, 558 (1967).

136 ANALYTICAL CHEMISTRY, VOL. 43, NO. 1, JANUARY 1971

EXCITATION 100 1

E MISSION

Figure 1. Fluorescence excitation and emission spectra for Calcein Blue, its 1 : l binary complex with Zr(1V) and its 1 : l : l ternary complex with fluoride and ZrW) (a) CB excitation (fluorescence at 450 nm) (b) CB fluorescence (excitation at 410 nm) (c) CB-Zr excitation (fluorescence at 325 nm) (d) CB-Zr fluorescence (excitation at 350 nm) (e) CB-Zr-F excitation (fluorescence at 410 nm) (f) CB-Zr-F fluorescence (excitation at 350 nm). Concentrations of CB/Zr/F at 2 X 10-6M. pH 2.5

34

L I 2 3 4 5

h O U R S

Figure 2. Fluorescence stability plots us. time a t specified p H values

Solid lines Zr :CB Dotted lines Zr:CB:F All other conditions as recommended in Procedure

the fluoride is simply abstracting zirconium from the 1 : 1 Zr :CB complex and releasing the more strongly fluorescent Calcein Blue. This was further confirmed by adding a 500- fold excess of fluoride to the Zr : CB complex which resulted in a decrease in fluorescence intensity at the excitation and emis- sion maxima of the zirconium-Calcein Blue complex and an increase in fluorescence intensity at the emission and excitation maxima of the reagent itself.

Influence of p H on Stability of the Complex. Figure 2 shows the fluorescence intensity of a solution 2 x 10-BM with respect to both zirconium and Calcein Blue, at various pH values, plotted against time.

The dotted curves were obtained similarly, but with fluoride added in 1 :1 proportion to the Zr:CB complex. The fluo- rescence of the solutions developed to a maximum value within 10 minutes. The Zr:CB binary complex shows maxi- mum intensity at higher pH values than the ternary Z r : F :CB

Figure 3. Continuous variations plot

F us. Zr:CB All other conditions as recommended in Procedure

complex as shown previously (3). The drop in intensity with time at p H >3 is probably due to progressive hydrolysis of the zirconium complexes. At pH 2.5, all solutions showed no significant change in fluorescence intensities within 4 hours, and hence all experiments were carried out at this pH.

Nature of the Complex. Job's method of continuous varia- tions was applied between the 1 : 1 zirconium-Calcein Blue complex and the fluoride ion (see Figure 3). This gave a binary complex to fluoride ratio of 1.2 : 1.

Mole ratio plots with varying concentrations of Calcein Blue against a fixed amount of a 1 : 1 zirconium fluoride solu- tion and of zirconium solution showed that in excess Calcein Blue, the fluorescence due to the ternary complex is con- siderably diminished (see Figure 4). The decrease in the

ANALYTICAL CHEMISTRY, VOL. 43, NO. 1, JANUARY 1971 137

Figure 4. Effect of excess of Calcein Blue (relative to Zr) on the fluorescence of the ternary complex with fluoride

Table I. Study of Interferences 4p of F by Recommended Procedure

Ion added, mole ratio

Ion/F C1- (500) Nos- (500) S04'- (500) s04'- (50) sod2- (5)

POa3- (500) POa3- (50) B407'- (500) B407'- (50) Mood2- (500) M004'- (50) Mood2- (5) WO4'- (500) W0h2- (50) w04'-(5) VOz- (500) VOs- (50) Oxalate (500) Oxalate (50) Oxalate (5) Tartrate (500) Tartrate (50) Tartrate (5) Acetate

Interference, z 0 0

+33 +11

0

Quenched - 67

20 0

- 42 -9

0 Quenched - 87 - 21 -15

0 - 89 - 80 - 55 - 12 - 66 - 45

0

Ion added mole ratio

Ion/F Be2+ (50) Be2+ (5) Ca2+ (500) Mg2+ (50) Mn2+ (50)

Mn2+(5) Ag+ (50) Ag+ (5) ~ 1 3 + (50) ~ 1 s t (5) Cd2+ (50) Ni2+ (50) Zn2+ (50) Co(I1) (50) corn (5) Pbz+ (50) Sb(II1) (50) Sb(5) AsV (50) AsV (5)

Interference, z - 33

0 0 0

Quenched (brown solution) - 13

White ppte + 10 - 39 - 16

0 0 0

- 22 0 0

- 9 0

- 16 0

presence of excess Calcein Blue (relative to Zr) may be ex- plained by the breakdown of the ternary fluoride complex in favor of a polynuclear zirconium-Calcein Blue complex. Previous studies of the binary complex at p H 5.5 showed that excess CB had a depressant effect on the fluorescence also, although it was not so marked as for the ternary complex at p H 2.5. This depressing action of excess CB is safely ac- counted for, however, by formulating a 1 : 1 reaction with the fluoride ion.

From both the continuous variation and mole ratio plots, it was concluded that a 1 :1:1 ternary complex was formed. The slightly high figure obtained for the Calcein Blue in the ratio was attributed to its partial decomposition when dis- solved in alkali and its low assay because it is manufactured for use as a metallo-fluorescent indicator and could not be obtained 100% pure. Attempts at purification were not successful.

Figure 5. Structure of Calcein Blue at various pH values related to its fluorescence and suggested structure for the weekly fluorescent binary Zr : CB and moderately fluorescent Zr : CB: F ternary complexes

Order of Addition of Reagents. The effect of of the order of mixing of the solutions was investigated. I t was found that the zirconium must be added after Calcein Blue; other- wise the fluoride produces no effect. The fluoride, however, could be added at any stage of the mixing.

Sensitivity of Method. The development of the method was carried out with test solutions 10-jM in fluoride. However, the maximum instrumental sensitivity was not used and it was possible to determine down to 10-7M concentrations in the final solution.

Interference Studies. The influence of up to 500-fold ex- cess of a selection of 11 other anions likely to interfere by forming complexes or precipitates with zirconium was ex- amined as were the effects of 13 cations likely to form pref- erential complexes with Calcein Blue or fluoride ion. The results of this study, which are summarized in Table I show that at a fivefold excess, tungstate, acetate, tartrate, and phosphate cause low results while, at 50-fold excess, molybdate also yields low recoveries and sulfate high recoveries. Among the cations, aluminum, manganese, and silver caused slight interference at fivefold excess, while at 50-fold excess, arsenic, antimony, beryllium, and cobalt caused low recoveries.

Nature of Complex Formation. The fluorescence of Calcein Blue at intermediate pH's has been attributed ( 4 ) to the formation of a hydrogen bridge between the phenolic oxygen and the nitrogen from the iminodiacetic acid group, Figure 5 (a), thus preventing torsion of the group about the C-CH2 bond and minimizing nonradiative dissipation of energy. At high pH, deprotonation occurs, Figure 5 (b) , and at low pH the N is protonated, Figure 5 (c), both resulting in breakage of the hydrogen bridge and freeing of the side chain, thus weakening the fluorescence yield by rotational dissipation of energy, etc.

Little is known about the chelate chemistry of zirconium, but it will be seen from the spectra presented in Figure 1 that at pH 2.5 the Zr:CB complex is less fluorescent than the reagent. This can be explained by the breakage of the H bridge on formation of a chelate complex with Zr, see Figure 5 (4. Thus, because of the largely ionic nature of the 0-Zr bond the Zr:CB complex yields less fluorescence than the Calcein Blue alone at the same pH. The enhancement of fluorescence on addition of fluoride, Figure 5 (e ) , can be explained by the incorporation of fluoride ion in the CO-

ordination sphere of the zirconium, i .e . , replacement of CO-

ordinated HzO molecules by the small fluoride ion, which has a

138 ANALYTICAL CHEMISTRY, VOL. 43, NO. 1, JANUARY 1971

very high affinity for zirconium. The electrophilic effect of fluoride may cause withdrawal of electrons resulting in the formation of a more covalent 0-Zr bond. This strength- ening of the degree of covalency should favor stabilization of the excimer and, hence, increase the quantum efficiency of the fluorescence.

The above explanation is consistent with the observation that at higher pH’s the binary Zr-Calcein Blue complex is more fluorescent than the reagent and addition of fluoride then has little or no effect.

amounts of fluoride in aqueous solution down to 0.01 ppm. Of the common anions, only phosphate interferes seriously at fivefold excesses. A 1 : 1 :1 ternary complex formation is responsible for fluorescence production. The increased fluorescence yield is attributed to conformational stabilization of the excimer of the fluoro- phore in the ternary complex. The method is rapid and the procedure is reproducible. Analyses show a precision of i 3 z near the middle range of calibration curves.

Very few cations interfere.

RECEIVED for review May 12, 1970. 1970.

for a grant to purchase the spectrofluorimeter.

Accepted September 22, We thank the Agricultural Research Council for a

The method suggested provides a positive fluorimetric grant-in-aid of this work and the Science Research Council

CONCLUSIONS

method of very high sensitivity for the determination of trace

Fluorometric Methods for Europium and Terbium

Dora E. Williams and John C. Guyon Unicersity of Missouri, Columbia, Mo. 65201

Determination of

A VARIETY OF P-diketones have been used to form tetrakis che- lates with europium and terbium for analytical purposes and as potential laser materials. Their utility as laser materials is limited by their high ultraviolet extinction coefficients, which would indicate good sensitivity in methods for the determina- tion of europium and terbium. Since these chelates involve charge transfer from the chelating agent to the metal ion, the fluorescence wavelength is characteristic of the metal ion and provides a method for the determination of europium and ter- bium in the presence of other rare earth ions. Among those chelating agents previously tried are thenoyltrifluoroacetone ( I ) and trifluoroacetylacetone (2) . Since trifluoro-P-dike- tones had been utilized, it was decided to try a hexafluoro-P- diketone, in the hope that more sensitive methods could be developed. This paper is a summary of the studies made upon the 1,1,1,5,5,5-hexafluoro-2,4-pentanedione chelates of europium and terbium.

EXPERIMENTAL

Apparatus. All fluorescence measurements were made on a Perkin-Elmer Fluorescence Spectrophotometer MPF-2A.

Reagents. The stock chelating agent solution was prepared by diluting 1 ml of Eastman 1,1,1,5,5,5-hexafluoro-2,4-pen- tanedione to 100 ml with 95 z ethanol. A stock 100 ppm of europium solution was prepared by dissolving 11.6 mg of K & K europium oxide in 0.5 ml of 1.25M HCI, then diluting to 100 ml with 95 z ethanol. A stock 100 ppm of terbium so- lution was prepared by dissolving l l .8 mg of K & K Tb407 in 5 ml of concd HCI and dilution to 100 ml with 95z ethanol. A 1-ppm solution of each lanthanide was prepared by dilution of 1 ml of each of the previous solutions to 100 ml with 9 5 z ethanol, neutralizing the solution with alcoholic K O H prior to complete dilution. Finally, a 50-ppb working solution of

(1) E. C. Stanley, B. I. Kinneberger, and L. P. Varga, ANAL.

(2) D. L. Ross and J. Blanc, Aduan. Chem. Ser., 71, American CHEM., 38, 1362 (1966).

Chemical Society, Washington, D. C., 1967, Chap. 12.

Table I. Effect of Rare Earth Ions Tolerable level, ppm

Europium Terbium Rare earth Added as method method

Ce (NH4hCe(N0d4 1 1 DY DY C13 1 0 . 5 Er ErCh 0 .1 0 . 5 E U EunOs . . . 1 Gd GdC13 0.1 1 Ho HoCIs 0.1 1 La LaCI3 1 1 Nd NdCI3 1 1 Pr PrCI3 0.06 1 Sm SmC13 0.06 1 Tb TbC13 0 .1 . . . Tm TmCI3 1 1 Yb YbC13 1 0 . 5

-~

each one was prepared by dilution of 25 ml of the 1-ppm so- lutions to 500 ml with 95% ethanol. Other rare earth solu- tions used in foreign ion study were rare earth chlorides sup- plied by K & K. The remainder of the foreign ion solutions reported were prepared from analytical reagent grade salts.

EUROPIUM METHOD

Procedure. Pipet an aliquot containing between 50 and 4000 ng of europium into a 50-ml volumetric flask. Neutra- lize the solution before the addition of chelating agent. If there is any deviation from neutrality at the time the chelating agent is added, the fluorescence intensity is reduced to that of the ethanol blank. Add 0.2 ml of the 1% chelating agent solution to the flask. This results in a final chelating agent concentration which gives the calibration curve the largest linear range.

Allow the solution to stand a t room temperature for 10 minutes prior to dilution with 95% ethanol and 30 minutes after dilution to obtain maximum fluorescence intensity.

ANALYTICAL CHEMISTRY, VOL. 43, NO. 1, JANUARY 1971 139