Anal. Chem. 1986, 58, 2161-2166
2161
Metal Chelate Fluorescence Enhancement by Nonionic Micelles: Surfactant and Auxiliary Ligand Nature Influence on the Niobium-Lumogallion Complex Alfred0 Sanz-Medel* and M. M. Fernandez Perez Department of Analytical Chemistry, Faculty of Chemistry, University of Oviedo, Ouiedo, Spain Miguel De La Guardia Cirugeda and J. L. Carrion Dominguez Department of Analytical Chemistry, Faculty of Chemistry, University of Valencia, Valencia, Spain
The effect of the chemical nature and degree of condensation of nonionic micelle-forming agents on the fluorescence Intensities of the chelate nlobium-lwnogalllon are Investigated. The observed micellar enhancement factor (MEF) of the fluorescence depends on the chemical type of the surfactant (an MEF of 19.3-25.5 was observed for Nemols, 14.7-19.5 for Tritons, and 9.5-5.6 for Genapols). The surfactant chain length did not influence the MEF. The influence on the observed MEF of the micellar charge and of the nature of the auxlllary ligand used to malntaln Nb(V) In solution Is studled as well, from parallel absorptlometrlc and fluorometric measurements. A new analytical method for the fluorometrlc determination of Nb(V) Is proposed (detectlon timlt 0.007 ppm) based on the ternary complex Nb(V)-tartaric acld-lumogaUion formed at pH 1 in micelles of Triton X-100. A dlscussion on the role of surfactants In the mlcellar fluorescent reaction is presented.
It is well established today that the local microenvironment in a micellar aggregate is dramatically different from that in the homogeneous aqueous bulk solution. This microenvironment in micellar media may lead to significant increases in photoluminescence ( I ) and in chemiluminescence ( 2 ) quantum yields. Therefore it is clear now that better limits of detection can be achieved in the fluorometric (3, 4 ) , phosphorimetric (5), and chemiluminescent ( 2 ) methods by analytically exploiting photophysical and/or photochemical phenomena taking place in micellar systems. By now it appears that the enhanced fluorescence method is more effective and convenient than the current phosphorescence method (6). Since the early work by Ishibashi and Kina (7-9) at the beginning of the last decade, interest on the fluorescence enhancement by surfactants is steadily increasing over the last few years (4, 10, and references therein) in order to establish fluorescent determinations of metal ions more sensitive and convenient. At present, a lot more of fundamental work is needed in order to be able to predict which type of surfactant will enhance the fluorescence of a particular metalchelate system. Were such basic knowledge established, an intelligent search for new micellar fluorometricmethods would be encouraged. We have reported previously on some trends observed in the fluorescence enhancement of metal chelates in cationic micelles ( 3 , 1 0 , I I ) :first of all a lumophoric organic reagent should be selected to produce a highly fluorescent binary compound able to interact strongly with the micellar aggregates. Unfortunately, very few reagents have been studied so far in this context (4, IO). Once a given metal-chelate compound has been selected, the most important factor governing the observed fluorescence enhancements is the nature of the surfactant used: charge type and chemical 0003-2700/86/0358-2 161$01.50/0
structure of the surfactant, besides formation of true homomicelles ( l o ) ,are the leading properties determing the final degree of interaction lumophor-micelles. We have shown how in the “organized” medium, Le., the micellar medium, hydrophobic solvatation of the chelate and stabilization of its singlet excited state would account for many of the interactions and effects observed. Most of our studies, however, have concentrated on positively charged micelles: in such cases it seems clear that the electrostatic interactions are the driving force that facilitates the closer approaches of the negative complex to the positively charged micellar surface. There, even very close to the surface (11,121,hydrophobic interactions between hydrocarbon parts of the complex and of the micelle would be operative. In other words, if a charge type effect can combine with the more typical hydrophobic interactions at the micellar level, both kinds of interactions seem to act concurrently bringing about the largest enhancements observed of the fluorescence intensity. Nonionic micelles are also known to produce fluorescence enhancements of certain metal-chelates (4, 7 , 8 , 1 3 ) .In fact, the practical importance of these reactions is rocketing today due to their enormous potential in fluoroimmunoassay (14). The use of europium labels and time-resolved fluorescence is becoming today a new powerful tool not only in research but in routine clinical analysis. The principle of such technique is based on the use of nonionic micelles to improve the fluorometric determination of rare-earth metals with ,!3-diketones and tri-n-octylphosphine oxide as a mixed ligand ( 14-1 6 ) . In spite of such importance, the absence of any charge-type effect in nonionic micelles and the lack of knowledge on the effect of chemical nature of these surfactants on fluorescence enhancement make it virtually impossible at present to explain why some nonionic surfactants increase the fluorescence while others, very similar, are quenchers for the same fluorescent complex (13). In the course of study of the trends existing in the fluorescence enhancement of metal-chelates by micelles (IO), the present communication describes some fundamental studies on the effect of the chemical nature and degree of condensation of several nonionic micelle forming agents on the fluorescence of the complex niobium-lumogallion (IO). At the same time the effect of the micellar charge and especially the nature of the auxiliary or “mixed” ligand used in this fluorescent system are also discussed. EXPERIMENTAL SECTION Reagents. A 100 ppm stock solution of Nb(V) was prepared from Nb205(Scharlau) as described in the literature (17). A 1.016 X M solution of lumogallion (ICN Pharmaceuticals) was prepared fresh in water. Stock solutions (15% w/v) of the following nonionic surfactants were prepared by dissolving the surfactant in water: (a) Nonylphenol-ethylene oxide condensates. Nemol K-36, K-1032, K-1035, K-2030, and K-3030 (Masso and Carol) with an 0 1986 American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 58, NO. 11, SEPTEMBER 1986
average number of ethylene oxide moles of 7.2, 13.4, 17.8, 25.8, and 36.4, respectively, were used. (b) tert-Octylphenol-ethyleneoxide condensates. Triton X-114, X-100, and X-405 (Fluka) with an average condensation degree of 7.8, 10.3, and 44.2, respectively, were essayed. (c) Ethylene oxide-propylene oxide condensates. Genapol PF-10, PF-20, and PF-40 (Hoechst), with average molecular weights of 2080, 2040, and 2840, respectively, and with ethylene oxide-propylene oxide mole ratios of 0.23, 0.57, and 1.01, respectively,were studied. Genapol PF-80, with an 80% of ethylene oxide and average molecular weight of 6600-9300 according to manufacturer specifications, was also tested. (d) Poly(ethy1ene glycols) (ICI) with average molecular weights of 190-210,380-420,570-630, and 5800-6800, respectively, according to manufacturer specifications, were tested. Those surfactants were characterized by NMR (18), UV (19), and IR (20) spectroscopies. In the case of the ethylene oxidepropylene oxide condensates, the IS0 norme was used for the determination of the molecular weight from the hydroxyl group (21) and NMR spectroscopy was used for the determination of the ethylene oxidelpropylene oxide mole ratio (22). Stock solutions of the “auxiliary ligands” were prepared 1 M in all cases. Apparatus. Fluorescence intensity measurements were initially made on a Perkin-Elmer MPF-44 spectrofluorometer,with a high-pressurexenon tube and 1.0-cm quartz cells. A Shimadzu difference spectrofluorometer, Model RF-520, equipped with a xenon lamp and a U-135 s recorder, was eventually used for the nonionic surfactant chemical nature studies. Absorbance measurements were made on a Perkin-Elmer, Model 124, spectrophotometer. A Julabo (Paratherm 111) thermostat with temperature control to f0.2 “C was employed to hold constant the sample compartment temperature; pH measurements were made with a WTW pH meter, Model 139. A simple, conventional, stalagmometer was used for surface tension measurements. General Procedure. Aliquots (0.5 mL) of a 10 ppm Nb(V) solution are transferred to a 25-mL volumetric flask. Then the following reagents are added: tartrate solution (0.25 mL of M), HCI solution to fix the acidity (1mL of 3.6 M), lumogallion M), and finally 2.5 mL of a 15% solution solution (1mL of of Triton X-100. Alternatively, for study of the influence of the chemical nature and condensation degree of other nonionic surfactants on the fluorescence of the complex, 2.5 mL of a 15% (w/v) solution of each surfactant to be tested is added at optimum experimental conditions to form the Nb(V) complex in Triton X-100 (between brackets). For every set of experiments the apparatus is adjusted with a ternary complex solution containing 200 ppb of Nb(V) and 1.5% of Triton X-100. The fluorescence measurements were obtained 4 h after the preparation of solutions containing Triton, Nemol, and poly(ethylene glycol) and 5 h later for solutions containing, Genapol (using in all cases water as a blank) to ensure reaching of the equilibrium. In order to determine the break point or critical micelle concentrations (cmc),aqueous solutions of the different surfactants assayed were prepared at concentrations ranging from 4 x to 1% (w/v) and their surface tension was measured with the stalagmometer. All the surface tension measurements in each series are carried out in a single session, in order to prevent changes in the experimental conditions, and 24 h after the preparation of the solutions so that the dynamic monomer-micelle equilibrium has been totally reached.
RESULTS AND DISCUSSION Previous work (10) demonstrated that the addition of micelles of Triton X-100 to the fluorescent complex of Nb(V) with the azo dye lumogallion produced a dramatic enhancement of the fluorescence (specially at pH 1). It is also known that the fluorescence of ‘this complex is very sensitive to the presence of “auxiliary” ligands (23), used to prevent Nb(V) hydrolysis and precipitation via mixed ligand complex formation, and this behavior could be modified by a micellar medium. For both reasons, this system appears as an adequate probe to study fluorescent nonionic micellar effects and the
A
0.2
0.1
\ BIN
TERN
SLS
CPI
TRlT
Flguure 1. Effect on absorbance of Nb(V-lumogallion complex (BIN) of mixed Ngand natue (TERN) and micelle charge type: (1) tartaric acid, (2)flucfide, (3)oxalic acid, (4) ckrii acid, (5)hydrogen peroxide; [Nb] = 1.07 X M [Lum] = 5.14 X lob M; [oxalic acid] = 0.5 X M; [fluoride] = [tartaric] = 10 X M; [hydogen peroxide] = [citric acid] = 5 X M; [surfactant] > cmc and maximum wavelength
used in each case.
influence of mixed ligand formation on the measured fluorescence intensity in a micellar medium. In order to get a deeper insight into the natme of the micellar reactions, pardel absorptiometric measurements were carried out. Mixed-Ligand Nature and Micelle Charge Influence. Spectrophotometric measurements of solutions of the Nb(V)-lumogallion complex, carried out to study the influence of common Nb(V) auxiliary ligands (hydrogen peroxide, fluoride, tartaric, oxalic or citric acids), demonstrated that “ternary (mixed ligands)” complex formation seems to take place a t any pH (from 1 to 6 were tested) in the aqueous solution. The change in absorbance depends on the nature of the ligand and on the pH but is most notorious when hydrogen peroxide is added (see Figure 1 “bin” and “tern”, where only the extreme pH values studied are illustrated). The addition of tensoactives of different charge at pH 3 did not alter significantly the spectra (a slight red shift of 5-15 nm in the presence of the surfactants) or the sensitivity of the corresponding ternary complexes. At higher pH (Figure lb) the positively charged micelles seem to “sensitize” slightly the absorbance of the ternary complexes while the nonionic micelles produced the reverse effect. In any case it is clear from the absorptometric results that the ternary (or mixed ligand) complexes do interact with the micellar media. This interaction depends on the nature of the auxiliary ligand (H202 always showed an anomalous behavior) and on the charge type of the micelle (anionic micelles of SLS hardly modified the spectral characteristics of any ternary complex at any pH). Spectrofluorometric results (Figure 2) showed that the slight fluorescence of the Nb-lumogallion complex increases due to ternary complex formation with any of the studied auxiliary ligands, provided that pH >2. The enhancement due just to auxiliary ligand addition is more noticiable the higher the pH, being maximum at pH 6 in the sequence: hydrogen peroxide > fluoride > oxalic > tartaric > citric. The addition of different charge type micelles to these ternary complexes at different pH values produced dramatic changes on their fluorescence intensity (IF)depending on the charge of the micelle; while the anionic SLS only brings about very small increases in I F a t any pH, the cationic surfactant BCP originates dramatic enhancements of IF at any pH and for any ternary complex. Maximum IF occurs a t pH 6 using tartaric acid as auxiliary ligand as shown by Figure 2b. It is interesting to note how a t low pH (pH l),the nonionic surfactant Triton X-100 produces an enhancement of IF even
ANALYTICAL CHEMISTRY, VOL. 58, NO. 11, SEPTEMBER 1986
If
I
2163
I,
.I pn 1
10
8
6
I
4
. I BIN
TERN
LL8
CPI
TRIT
BIN
TERN
8LI
CPI
TRIT
Flguro 2. Effect of mixed ligand nature and micelle charge type on
2
fluorescence intensity (at maximum emission wavelength for each system and ail conditions as in Figure 1). Table I. Excitation and Emission Maxima of the Ternary Complexes at pH 1 in the Absence and Presence of Surfactants of Different Charge system
surfactant
micelle
Triton
nonionic positive negative
Nb-lum-tartaric acid
BCP SLS Nb-lum-oxalic acid Triton
BCP SLS
nonionic positive negative
Nb-lum-citric acid Triton
BCP SLS
nonionic positive negative
Nb-lum-H,O,
Triton
BCP SLS
nonionic positive negative
Nb-lum-fluoride Triton
BCP SLS
nonionic positive negative
A,,
A,
490 495 515 495 490 495 505 490 490 495 495 490 495 495 519 490 490 495 505 490
630 610 615 618 630 610 620 615 630 610 620 620 620 600 625 615 630 610 615 615
higher than that produced by BCP. For higher pH values, this nonionic micellar effect decreases very sharply as illustrated comparing Figure 2, parts a and b. In fact all the graphs at intermediate pH were less pronounced but similar in shape to those at pH 6. Table I shows the effect upon other relevant spectral characteristics at pH 1 (excitation and fluorescence maxima of the corresponding ternary complexes) of the addition of different charge-type micelles. A comparison of the spectrophotometric and the corresponding fluorometric results (compare Figure l a and 2a) shows how to observe micellar fluorometric enhancement; it is not necessary that micellar spectrophotometric sensitization occurs; Le., the nature of the fluorescence enhancement by micelles appears different from the enhancement in "sensitized" spectrophotometricreactions. Rather than an increase in the apparent probability of the electronic transition involved (molar extinction coefficient), a sheltering or protection of the lowest singlet excited state by the micelles to avoid its radiationless deactivation (increase of quantum efficiency) seems to be the most probable mechanism in micellar fluorescence.
A m Figure 3. Excitation and emission spectra at pH 1 of: (A) the ternary Nb(V)-iu~all!m-tartrate system, [Nb] = 1.07X lo-' M, [Lum] = 5.14 X 10- M, [tartaric acid] = lo-' M; (6) the above system with 0.5% of Triton X-100; (C) the above solution without niobium (blank).
Optimization of the System Nb(V)-LumogallionTriton X-100. From all the above results, it is clear that at pH 1 the fluorescence of the ternary systems Nb(V)-Y-lumogallion (Y being fluoride or oxalic, tartaric, or citric acid) is strongly enhanced by nonionic micelles of Triton X-100 (Figure 2a). The low pH and the high fluorescence of these reactions could offer the basis for a selective and sensitive determination of the metal, apart from constituting an adequate probe to study micellar enhancements of the metal chelates. Therefore, all the reaction variables were carefully optimized. Figure 3 shows the uncorrected excitation and fluorescence spectra of the ternary complex Nb-lumogallion-tartaric acid alone and in the presence of Triton X-100 at pH 1 (as obtained in the MPF-44A). Tartaric acid was eventually selected as the mixed ligand, Y, because it is the most common auxiliary agent used for dissolution and stabilization of Nb(V) solutions and because a relatively high concentration of tartaric acid (Q3%)can be tolerated without virtual loss in the analytical signal. The rest of the auxiliary ligands studied produced sharp decreases in the fluorescence observed even when small excesses of the ligand, over the molar Nb(V) content, were added. Addition of Triton X-100 micelles to this ternary system causes not only a dramatic increase in fluorescence intensity but also a 5-nm red shift in the excitation and a 20-nm blue shift in the emission maximum (see Table I). The influence of the pH on the fluorescence intensity (IF) of the ternary complex alone and in the presence of nonionic micelles is given in Figure 4. As can be seen, the ternary complex reaches its maximum fluorescence between pH 4 and 6, while the micellar medium shifts the optimum pH for complex formation to pH 1. At higher acidities the observed IF decreases. HCl acid turned out to produce higher fluorescent signals than HNOBor H2SO4 acids. Although temperature of the measurement in the cuvette was not very critical around 20 "C, some photodecomposition
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ANALYTICAL CHEMISTRY, VOL. 58, NO. 1 1 , SEPTEMBER 1986
’f
20
,\ \3 15
\
\
10
? 5
r
C.I.C.
5
)i/ L
I
0.1
0.2
0.3
0.4
0.6
o (~um].3.67*1Q‘Y 1
3
5
PH
Flgure 4. Influence of pH on fluorescence intensity of (A) the ternary Nb(V)-lu~Nlon-tartratesystem, [Nb] = 1.07 X M; [Lum] = 7.34 X 10 M; [tartaric acid] = 2 X lo-* M; (9) the above system
in the presence of Triton X-100. and kinetic effects were noticed when studying the influence of standing time. The observed IF increases steadily with standing time of solutions in the flasks at room temperature. If the ternary complex is heated for 20 min a t 40-50 “C in a water bath and allowed to cool before final addition of the surfactant, a stable signal is obtained after about 1 h of standing time (otherwise4 h are necessary to obtain the same signal without heating). The influences of surfactant and lumogallion concentrations have been plotted in Figure 5. The observed effect of increasing concentration of Triton X-100 (curve 5B) shows clearly that fluorescence enhancement starts only after the cmc of this surfactant has been reached (0.014% w/v final concentration, in 0.1 M HC1 acid). The fluorescence only levels off after surfactant concentration is high enough (>0.5%). These facts demonstrate the micellar nature of the observed fluorescence enhancement: only when the “organized” medium is established can the ternary complex be accommodated in the surface of the micelles and so fluorescence is favored. The effect of the lumophoric reagent is illustrated in Figure 5A. The intensity increases with concentrationof Lumogallion until a plateau is reached at around lo4 M final concentration of the reagent (above 7-fold molar excess over the Nb(V) concentration tested). The detection limit of the fluorometric niobium determination (3ab criterium, ab being the standard deviation of the blank) was 0.007 ppm and the calibration graph was linear up to 0.6 ppm of the metal. Effect of Chemical Nature and Degree of Condensation of the Nonionic Surfactant. In order to investigate the influence of the nature of the nonionic micelle, we measured the fluorescence enhancement in solutions of the ternary complex of the same concentration and at pH 1prepared as described for Triton X-100, but adding in each case a final concentration of 1.5% (w/v) of the different nonionic surfactants assayed. At the same time, we have tested the effect of the addition of poly(ethy1ene glycols) of different molecular weight (i.e., condensation degree) for comparative purposes, as these reagents do not exhibit amphiphilic character. For every surfactant we obtained the corresponding calibration graph, in the linear response interval, in order to be able to compare the sensitivity in each case. The standard deviation of the corresponding blanks, ab, was also worked out from ten independently prepared blank solutions without niobium.
0.6
1
1.5
2 0
[ralron x-loo].%
Figure 5. Lumophor reagent (A, where [Triton] = 0.5%) and Triton (B, [Lum] = 7.34 X lo-‘ M) Influence on the fluorescence. Metal and tartaric acid concentrations are given in Figure 4.
X-100
The instrumental sensitivity was adjusted in all cases by using a reference solution and fluorescence measurements were made consistent every day by comparing with the observed IFof the freshly prepared ternary complex in Triton X-100. The kinetic character of these reactions was again evidenced because a stable signal was only obtained after 4 h of standing time for nonylphenols, tert-octylphenols, and poly(ethy1ene glycols); 5 h had to be allowed when using ethylene oxidepropylene oxide condensates to get a reproducible value of IF.
The results obtained have been summarized in Table 11 in terms of equations of the linear calibration graphs (with the correlation coefficients observed), detection limits, and observed micellar “enhancement factor”, MEF (ratio between the slope observed in a given surfactant and that characteristic of the ternary system alone). It is evident from these results that all the amphiphilic surfactants (nonylphenols, tertoctylphenols, and propylene oxide-ethylene oxide condensates) enhanced the fluorescence intensity much more than the nonsurfactant poly(ethy1ene glycols). Moreover, for each family of nonionic surfactants, a certain homogeneous “enhancement factor” is noticed: Nemols producing the highest enhancement, followed by Tritons and Genapols. In other words, the chemical structure of the monomers of the tensoactive, which will eventually form the nonionic micelle, really matters on the final fluorescence observed. Within a given family of surfactants, however, our results do not indicate any significant trend of IF enhancement with molecular weight (degree of condensation). It has to be pointed out here that although we used in all cases a 1.5% concentration of the surfactants, regardless of its chemical nature or molecular weight, such concentration is well above the cmc of any of the surfactants studied (see Figures 6 and 7). For ethylene oxide-propylene oxide condensates, a much higher surfactant concentration is needed to get saturation of IF; in any case, with a saturation concentration (15%) the value of IF is only twice that observed for 1.5% as shown by Figure 7. In other words, these kinds of compounds should still exhibit much worse “enhancement factor” even at high concentrations. As the detection limit depends also on the absolute value of the fluorescence of the blanks, given by the first term of the analytical curves, it is worth noting that with some exceptions the order of magnitude of the intercept with the IF axis, depends again mainly on the surfactant chemical family considered. Surfactants for which the emission of the complex was noticed slightly more red shifted than usual (at 605 nm
ANALYTICAL CHEMISTRY, VOL. 58, NO. 11, SEPTEMBER 1986
2165
Table 11. Characteristics of the Fluorescence of the Niobium-Lumogallion-Tartrate System Enhanced by Nonionic Surfactans surfactant
A,,
Nemol K-36 Nemol K-1032 Nemol K-1035 Nemol K-2030 Nemol K-3030 Genapol PF-10 Genapol PF-20 Genapol PF-40 Genapol PF-80 PEG 200 PEG 400 PEG 600 PEG 6000
0
linear calibration equation
DL,ppb
MEF
IF = 0.912 + (6.68 X 10-3)c r2 = 0,999
23.15 7.77 7.41 6.16 16.7 2.4 2.1 2.17 2.06 13.68 22.73 4.09 5.27 14.21 18.19 16.59 11.39
1 16.0 19.5 14.7 19.3 19.3 19.2 25.5 24.1 4.9 3.9 5.5 3.3 1.4 1.2 1.4 2.2
nm
, , ,X
595 605 600 595 605 600 595 590 595 605 605 590 590 590 590 590 590
490 490 495 505 490 490 490 495 495 485 490 490 490 490 485 485 490
Triton X-114 Triton X-100 Triton X-405
It1
nm
IF = 45.77 + 0.107~ r2 = 0.99 IF = 25.16 + 0.130~ r2 = 0,999 IF = 29.03 + 0.098~ r2 = 0.99 IF = 354.9 + 0.129~ r2 = 0.9999 IF = 25.72 + 0.129~ r2 = 0.999 IF = 17.73 + 0.128~ r2 = 0.9999 IF = 18.92 + 0.170~ r2 = 0.9999 IF = 16.02 + 0.161~ r2 = 0.999 IF = 10.88 + 0.033~ r2 = 0.99 IF = 18.33 + 0.026~ r2 = 0.99 IF = 2.53 + 0.037~ r2 = 0.999 IF = 1.96 + 0.022~ r2 = 0.999 IF = 0.38 + 0.0095~ r2 = 0.9 IF = 1.84 0.0081~ r2 = 0.999 IF = 1.49 + 0.0094~ r2 = 0.999 ZP = 1.63 + 0.015~ r2 = 0.999
+
I r f
0.2 ,
0.4 ,
0.6 l
0.0 l
,1.0
,1.2
l1.1
l1.8
L1.6
,2.0
;
INEMOL K-20301,k
Figure 8. Effect of the concentration of Nemol K-2030 on the fluorescence intensity of the niobium-lumogallion-tartrate system at pH 1. 0.01 0.014 0.02 '1
0.03 0.04 0.05 0.06 0.01 [OENAPoL PF-'dl4
f
4 0.85 o,75[ 0.115 PENAPOL PF-44,).
Figure 7. Effect of the concentration of Genapol PF-40 (A) and poly(ethy1ene glycol) (PEG6000) (B) on the fluorescence intensity of the niobium-lumogallion-tartrate system at pH 1.
Table 11) showed anomalously high values of the blanks. This could be attributed to the presence of impurities in the commercial surfactants used. Such impurities would not virtually disturb the MEF but would increase the blank fluorescence (see the data for the case of Nemol K-36). In conclusion, the addition of a 1.5% (w/v) final concentration of nonionic surfactant to the ternary complex Nb(V)-lumogallion-tartaric acid, at pH 1, produced an enhancement of the IF that depends on the chemical type of surfactant used: an MEF of 19.3-25.5 for the condensates of ethylene oxide and nonylphenol, from 14.7 to 19.5 for the
condensates of ethylene oxide and tert-octylphenol, and finally an enhancement of 3.3-5.6 times for the condensates of ethylene oxide and propylene oxide (better enhancements can be obtained in the last case by increasing ten times the concentration of the tensoactive agent). Finally, the poly(ethy1ene glycols) (PEG), which are lacking the tensoactive character, can modify the IFobserved, but only slightly when compared with those having surfactant properties (Figure 7). Role of the Surfactant in the Micellar Fluorescent Reaction. With surfactant "sensitized" spectrophotometric determinations, at least two different basic metal chelatesurfactant interactions have been demonstrated: formation of unusual ion-association complexes (11,12,24) or formation of true micelles where the chromophoric metal-chelate is solubilized and "organized" (5). In both cases, the microenvironment experienced by the chromophore is different from that for bulk water. All the information available so far on metal chelate fluorescence enhancement by surfactants indicates (4,10,12) that the phenomenon of enhancement of fluorescence is not observed unless a minimum concentration of true micelles (charged or uncharged) is secured in the solution. In other words, with surfactant-enhanced fluorometric determinations, only the solubilizing-organizing mechanism, characteristic of true micelles, seems to be operative. Of course this claim should be specially true with nonionic surfactants where the absence of charge prevents any ion-association or electrostatic interaction with the chelate. On the other hand, it has been stated (10) that one of the most important factors, governing the -metal chelate fluorescence enhancement by surfactants, is the charge type of the micelle. Present work results (Figure 2) confirm how the decisive factor is the charge of the micelles but relative to the actual charge of the metal chelate. Let us consider first our results at low pH. At pH 1 the ternary complex could be thought of as consisting of two moieties, a more hydrophobic one (the phenolic ring with the -OH group protonated a t such pH) and the hydrophilic one having the dissociated sulfonic group (see Figure 8). Then, this ternary complex could be solubilized-organizedby the Triton X-100 nonionic micelles in the following manner: the -OH group of the complex would form hydrogen bonds with oxygen of the ethylene oxide chains (Figure 8) or/and van der Waals attractions would take place between the hydrophobic benzene ring of the complex and those rings of the surfactant chains exposed, through dynamic bending and disorder (25), to external water and counterions existing in the micellar surface. A more rigid and ordered environment results in this way where the fluorophor would be sheltered from collisional deactivations. As shown by Figure 8, however, even at pH
2188
ANALYTICAL CHEMISTRY, VOL. 58, NO. 11, SEPTEMBER 1986 TYPICAL
CH3-(CH4
-;HaO{CHiCH~O);H CH3
TRITON
C M C , I w VI’.
x-100
15.10-2
CL
;1
HO-(cHph 0
(CH I - CHdO) - (CHpCHO I )H ;
GENAPOL
V
CH3
PF 40
2 0.10-2
Flgure 8. Chemical formulas of the different families of nonionic surfactants used and speculative formula of the Nb(V)-lumogalliontartrate system.
1there is another charged (much more polar) moiety in the complex which through the -SO3- group holds the posibility of electrostatic interaction with positively charged micelles. This would explain why cationic surfactants also enhance the fluorescence while anionic ones do not (see Figure 2a). At such a low pH, however, the chemical interaction (“ordering or penetration”) of the ternary complex with the positive micelles (CTAB showed analogous behavior to that of CPB) seems to be weaker than the interaction observed with nonionic Triton X-100 micelles. If the negative charge of the chelate is too high (e.g., at pH 6 in Figure 2b) electrostatic interactions should be favored against hydrophobic ones. Then, in such cases, positively charged micella would interact more strongly with the chelate, i.e., cationic surfactants would be more effective than nonionic ones to enhance the fluorescence at such pH (Figure 2b). In any case, it is interesting to note that, for the rest of the conditions being the same, the “micellarenhancement factor” of CPB at pH 1 (MEF = 10)is clearly superior to the MEF of the same surfactant at pH 6 (MEF = 3). We verified that the corresponding MEF a t pH 3 was around 6 for all the ternary complexes. These facta seem to support the general idea that the maximum fluorescence enhancements are observed when electrostatic and hydrophobic interactions can act concurrently (10,12). In the case of nonionic micelles the leading force of the interaction must be hydrophobic attraction. Therefore, the chemical nature of the monomers of the surfactant and their particular affinity to the metal chelate considered should be expected to play the fundamental role. Our results indicate that the micellar enhancement factor for the fluorescence of the ternary system with different nonionic Surfactants actually depends on the chemical nature of the tensoactives. It seems like the presence of an aromatic ring (25)in the surfactant molecule (i.e., nonylphenols and tert-octylphenols) would favor the interaction of the micelles with the more hydrophobic
aromatic ring of the Nb(V) complex (Figure 8). With ethylene oxidepropylene oxide condensate surfactants (Genapols) such aromatic chemical affinity cannot operate. An additional factor to consider is that the presence of branched methyl groups in the chain of genapols could hinder a “good packing” of the molecules of €he ternary Nb(V) complex. It is interesting to note that the reverse behavior was noticed for Al(II1)-morine (13): ethylene oxide-propylene oxide condensate surfactants allowed appreciable fluorescence enhancement, whereas fluorescence quenching occurred in the presence of nonylphenol and tert-octylphenol ethylene oxide condensate surfactants. The effect of increasing the surfactant concentration on the fluorescence of the ternary complex differs from one type of nonionic surfactant to another suggesting the existence of an optimum concentration of micelles in each case to get saturation fluorescence (Figures 6 and 7). Curiously enough we have not experienced the expected influence of the surfactant chain length on the sensitization of the Nb(V) ternary system. In fact, for nonylphenols an increase in the condensation degree from 7 to 37 mol yields only an increase in the MEF from 19.2 to 25.0 whereas for tert-octylphenols an increase in the condensation degree from 8 to 44 mol only gives rise to a slight decrease in the MEF value, having obtained the maximum MEF for a compound with an intermediate condensation degree.
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RECEIVED for review February 11,1986. Accepted May 5,1986.
