ANALYST, JANUARY 1992, VOL. 117 57 Characterization Lipophilized Ana of an Optode Membrane for Zinc(ii) Incorporating a logue of the Dye 4-(2-Pyridylazo)resorcinol Kemin Wang," Kurt Seiler,t Bruno Rusterholz and Wilhelm Simon Swiss Federal institute of Technology (ETHJ, Department of Organic Chemistry, Universitatstrasse 16, CH-8092 Zii ric h, Switzerland A chemical optical sensing system (optode) for Zn" based on a lipophilized metal ion indicator dye [ 1 -octadecyloxy-4-(2-pyridylazo)resorcinol; ETH 24641 dissolved in a plasticized poly(viny1 chloride) membrane has been developed. In the measuring range from 1 x 10-6to 3 x 10-3 mol dm-3Zn2+ (pH 4.8), the absorbance response shows a good correlation with the theoretically derived formulae. The optode membranes were found to reach 95% of the final signal within 5 min (&).When exposed to a continuous sample flow (1 ml min-I), the optode membrane did not lose a perceptible amount of the indicator over a period of 10 h. The selectivity of the optode membrane for the response towards Zn2+ ions in pH-buffered solutions is described. Keywords: Optode membrane; indicator; plasticized poly(vin yl chloride) membrane; spectrophotometry; zinc(//) ion Organic indicator dyes, which are normally used in the spectrophotometric determination of various metal ions, play a central role in the design of chemical optical sensors (optodes). I-8 Many optical sensing layers with immobilized classical indicators," which exhibit a marked change in their absorption spectrum on complexation of the metal ion, have been described.Most frequently, the immobilization of the indicator is achieved by covalent binding or by electrostatic attraction to a resin. Unfortunately, the group responsible for the complexation is often inaccessible to the metal ion after the immobilization process. In addition, very strong binding of the metal ion by the indicator often prohibits the reversible response of the sensor. Saari and SeitzJ.i have developed a fluorescence sensor for and Be" by binding the ligand morin with cyanuric chloride on a cellulose membrane. Zhujun and Seitz6 have described a fluorescence sensor for the detection of Allt1, Mg", Zn" and Cd", which is based on 8-hydroxyquinoline-5-sulfonic acid attached electrostatically to an ion-exchange resin. Chau and Porter7 have utilized a similar procedure for the immobil- ization of calcichrome on an anionic polymer film.An attempt to construct a sensor for transition metal ions with the fluorogenic indicator calcein was not successful, because of its lack of reversibi1ity.x The performance of these sensors is usually given in terms of dynamic measuring range. response time and selcctivity, whereas data on lifetime and signal recovery are often absent. A different approach has recently been proposed with the use of homogeneous plasticized poly(viny1 chloride) (PVC) membranes, incorporating lipophilized components as the chemically active species.lo In order to reduce the possible influence of surface effects on the response signal, a sensing layer with a thickness of the order of 3-4 pm was proposed. As a consequence, these layers can be treated as bulk phases with uniform properties.Highly lipophilic ionophores are the selective components of the widely used ion-selective elec- trodes and have been incorporated in such optode membranes together with specially designed chromo-ionophores. The free mobility of the complexing agent allows its active participation in the extraction of the substrate. This type of system appears to be well-suited for the detection of many analytes.10-1' In this paper it is shown that not only ionophores, but also lipophilized conventional dye indicators" are useful as cam- + O n leave from the Hunan Univcrsity. Changsha. China. .i_ To whom correspondence should be addressed. plexing agents in such layers.An optode membrane incorpor- ating a lipophilized analogue of the well-known indicator 4-(2-pyridylazo)resorcinol (PAR)'"-" and its response towards Zn'+ ions in a sodium acetate pH buffer are described. Experimental Reagents Aqueous solutions were prepared with water that had been doubly distilled from a quartz still and with salts of the highest purity available. For the preparation of the membrane, PVC (high relative molecular mass), tris(2-ethylhexyl) phosphate (TEHP) and tetrahydrofuran (THF) were used and were obtained from Fluka (Buchs, Switzerland). The buffer solutions used for the characterization of the optode membrane were acetate buffers of pH 4.8 and 5.6 (see Table 10.18 in ref. 16). Synthesis of 1-Octadecyloxy-4-(2-pyridylazo)resorcinol (ETH 2464) A solution of 2.0 g (9.3 mmol) of PAR (Fluka, p.a.), 3.03 g (9.3 mmol) of caesium carbonate (Fluka, puriss., p.a.) and 3.1 g of 1-bromooctadecane (Fluka, purum) in 100 ml of N,N-dimethylformamide (Fluka puriss., p.a.) was stirred for 1 h at 140 "C.The solvent was evaporated and the residue dissolved in 200 ml of CH2C17. This solution was washed with two 200 ml portions of distilled water and the solvent was evaporated again. The residue was purified by flash chroma- tography on silica gel 60 (Fluka) using ethyl acetate as eluent, and on neutral alumina (Woelm; activity grade 1) using hexane, and subsequently ethyl acetate, as eluent. The product was recrystallized twice from hexane to yield 51 mg (0.1 mmol, 1.2%) of the pure product (m.p. 69.5-70.5 "C). The structure of ETH 2464 (see Fig.1) was confirmed by proton nuclear magnetic resonance spectroscopy (300 MHz, CDC13) and fast atom bombardment mass spectrometry. The purity was determined by elemental analysis [Found: C , 74.4; H, 10.0; N, 8.7. Calc. forC2c,H45N307 (467.7): C, 74.5; H, 9.7; N , 9.0%]. Membrane Preparation The optode membranes were prepared from a mixture of 79 mg of PVC, 157 mg of plasticizer (TEHP) and 3.0 mg of ETH58 ANALYST, JANUARY 1992, VOL. 117 ETH 2464 Fig. 1 Structures of the indicator PAR and its lipophilized analogue (ETH 2464) Sample I 1 To detector - 1 To waste Fig. 2 Schematic reprcsentation of the flow-through cell: 1. poly- (propylenc) support with sample inlet and outlet; 2. O-ring seal; 3, quartz glass support; 4, ion-sensing optodc membrane; 5 , Plexiglas cell wall; and 6 , screw for fixation 2464.The membrane components were dissolved in 2 ml of freshly distilled THF. By means of a spin-on device, two identical membranes of approximately 4 vm thickness were cast onto two quartz glass plates, which were subsequently mounted in the specially designed flow-through measuring cell (see Fig. 2).".17 Apparatus Two quartz glass plates without membranes were mounted in the reference cell. Both measuring and reference cells were then placed in a conventional double-beam ultraviolet-visible spectrophotometer (Perkin-Elmer, Model Lambda 2, Kiis- nacht, Switzerland) to measure the absorbances. A specially designed base for the cells was flushed with thermostated water to keep the temperature constant at 25 "C.Procedure Except for the response behaviour, which was studied under batch conditions, all other measurements were carried out in the flow-through mode. The sample solutions were pumped (SJ-1211 Perista Mini-Pump, ATTO, Japan) through the measuring and reference cells at a constant flow rate of 1 ml min-1. The absorbance values A l and A. were determined with the optode membrane in contact with 0.1 rnol dm-3 HCI and 0.1 rnol dm-3 ZnS04, respectively. Table 1 Effect of diffcrcnt metal ions on the absorbance values of the optodc mcmbranc. in contact with a fixed concentration of 3 X rnol dm-3 ZnSO, in an acctatc buffer at pH 4.8 Metal ion Zn2+ K+ Na + Caz+ Mg2+ ~ 1 3 + Fe3+ Ni*+ Mn2+ Pb2+ Cd? + co2+ Cu2+ Concentration/ rnol dm-3 0.1 0.1 3 x 10-2 3 x 10-2 3 x 10-5 3 x 10-3 3 x 10-3 3 x 10-3 3 x 10-3 3 x 10-3 3 x 10-3 3 x 10-3 3 x 10-4 3 x 10-4 3 x 10-3 3 x 10-4 Absorbance 0.2178 0.2178 0.2178 0.1809 0.2125 0.1812 0.2129 0.1743 0.2061 0.2188 0.2000 0.2179 0.2154 0.2238 0.2603 0.2809 Absorbance change (%) 0 0 -16.9 -2.4 -16.8 -2.2 -20.0 -5.4 +0.5 -8.2 +0.05 -1.1 +2.7 +19.5 +29.0 - For the selectivity measurements, the membrane was equilibrated with a 3 X 10-5 rnol dm-3 Zn'+ sample solution (pH 4.8), before the solutions with the respective metal ion as background were injected (see Table 1).The absorbance readings for the selectivity studies were taken 15 min after the sample had been changed. Owing to the strong binding, Cu and Co were removed from the membrane by pumping 1 x 10-2 rnol dm-3 HCI through the measuring cell.In this way, Cu could be removed within 30 min, whereas more than 6 h were required to remove Co. Results and Discussion Principle of Operation The optode membrane described here belongs to the class of ion-exchange systems described previously. 1 0 The extraction of Zn'+ from the aqueous sample solution into the membrane phase and its complexation by the lipophilized metallochromic indicator ETH 2464 (HL) proceed with the loss of two protons from the hydroxy groups of two indicator molecules. This ion-exchange process is determined by the electroneutrality in the organic membrane phase. Fig. 3 shows the absorption spectra of the optode membrane incorporating ETH 2464, as obtained after equilibration with pH-buffered solutions (acet- ate buffer, pH 4.8) containing different ZnS04 concentra- tions.Obviously, the formation of the Zn complex induces a bathochromic shift of the absorption band from 380 nm (protonated form) to 523 nm. The over-all equilibrium between the organic membrane phase (org) and the aqueous sample solution (aq) is described as follows: K,,CI, ZnL?(org) + 2H+(aq) e 2 H L ( o r g ) + Zn2+(aq) (1)ANALYST, JANUARY 1992. VOL. 117 59 0.4 0.3 0) C m + 0.2 s a I] 0.1 0 I 1 I I 1 1 260 340 420 500 580 660 Unm Fig. 3 Absorption spectra of two 4 pm thick optode membranes after equilibration with different ZnSO, concentrations in an acetate buffer at pH 4.8. The ligand shows an absorbance maximum in its protonated form at 380 nm and the Zn complex an absorbance maximum at 523 nm. A, Buffer [ZnSO,]; B, 1 X 10-6; C, 3 x D, 1 X 10-5; E, 3 x 10-5; F, 1 x 10-4; G, 3 x lo-,; H, 1 x 10-3; and I, 3 x 10-3 mol dm-3 The corresponding equilibrium constant Kexch depends on the complex formation constant and on the distribution coef- ficients of the H+ and Zn?+ ions between the aqueous sample solution and the organic membrane phase.It can be assumed that a 1 : 2 Zn2+-ligand complex is formed in the optode membrane, as this stoichiometry was found in aqueous solutions for both indicator dyes, 1-(2-pyridylaz0)-2-naphthol (PAN) and PAR,13 and also in chloroform for PAN.18 Substituting the activities of the species in the membrane phase by their concentrations,I0.l I and with the introduction of a [the ratio of the concentration of the protonated ligand relative to the total amount of ligand (LT) present in the membrane phase] the response of the optode membrane can be given by: ffL” = (UH)’Kcycl,( 1 - CY)/(2CY’Lr) (2) where aL,, and aH denote the activities of the Zn’+ and the H+ ion, respectively.The high concentration of the acetate buffer in the sample solution provides a nearly constant ionic strength and hence the activity coefficient of Zn?-+ can be assumed to be constant. About 60% of the total Zn in the sample solution is complexed by the acetate anion.19 However, in the concentration range considered the relative amount of free Zn’+ does not change significantly, and hence it is approximately proportional to its total concentration c::. If LT and Kexcl, are assumed to be constant over the whole dynamic range, and if all the constant values are summarized in K ’ , then eqn.(3) can be derived from eqn. (2): (3) t o t CZ” = K’(1 - a)/& The measured absorbance A is directly related to a, if the optode membrane obeys Beer’s law: CY = (A0 - A)/(Ao - A , ) (4) where A I and A,) are the limiting absorbance values for CY = 1 (fully protonated ligand L) and CY = 0 (fully complexed ligand L), respectively. Response Behaviour In Fig. 4 the relative absorbance values, a, are given as a function of log ~2: for two different pH values. The curve fittings for the experimental points were calculated from eqn. (3) with log K’ = -4.92 (pH 4.8) and -6.40 (pH 5.6), respectively. The good correlation of the measured data with the theory confirms the validity of the assumptions made in 1 .o 0.8 0.6 0.4 0.2 0 8 I I I -8 -6 -4 -2 Log c g Fig.4 Relative absorbance values at 523 nm as a function of log c:’,: at A, pH 4.8; and B , pH 5.6. The curves fitting the experimental points were calculated from eqn. (3). Membrane: ETH 2464 + TEHP + PVC t 0 B 4 rnin - 10.02 I 0.3038 t 0.2686 4 rnin - Io.02 1 x 10-3 rnol drn-3 ZnS04 t t t 0.2687 0.2681 0.2677 3 x 10-4 rnol dm-3 ZnS04 Time - Fig. 5 membranes at 523 nm after several concentration step changes Absorbance response versus time for two 4 pm thick optode eqn. (3). In order to avoid depletion of the analyte close to the optode membrane, which would result in a large concentra- tion gradient in the sample solution, a sufficient amount of the analyte must be ensured. For this reason, the steady state was reached significantly faster for a total Zn concentration of < I X 10-5 rnol dm-3, when the sample solution was pumped through the measuring cell.At low Zn concentrations a reduction of the total amount of the indicator dye would of course also lead to shorter response times, but unfortunately at the expense of sensitivity. Short-term Reproducibility The reproducibility of the optical signals was evaluated by repetitively changing between two samples of 1 x 10-’ and 3 x 10-4 rnol dm-3 ZnSOj in an acetate buffer at pH 4.8. Thc absorbance response at 523 nm versus time is shown in Fig. 5 . The mean absorbance values and their standard deviations were determined after 5 min and were found to be repro- ducible to 0.3037 * 0.0005 (1 x 10-3 rnol dm-3 ZnS04) and to 0.2683 k 0.0005 (3 x rnol dm-3 ZnS04). Response Time The tgS values are reached within 5 min after concentration step changes from 1 x 10-3 to 3 x 10-4 rnol dm-3 ZnS04 and vice versa, as illustrated in Fig.5. The optode membranes described here exhibit a longer response time than the membranes that incorporate electrically neutral ionophores60 ANALYST, JANUARY 1992, VOL. 117 and which reach the final steady state within 3-4 s.IO-I1 The response time of such optodes is mainly governed by diffusion processes in the bulk of the membrane. For the optode membranes with ETH 2464, the rate of complex formation and particularly the rate of complex dissociation (see Fig. 5 ) might play an important role. Lifetime The absorbance signal at 523 nm for the optode membrane in contact with a 3 x 10-4 mol dm--? ZnS04-acetate buffer solution (pH 4.8; flow rate, 1 ml min-*) was recorded over a period of 10 h.From the absorbance values taken every 30 min ( n = 21) a mean absorbance of 0.2682 and a standard deviation of k0.0004 were obtained. No significant loss of the ligand was observed during this period of time. Selectivity The selectivity of optode membranes incorporating lipophilic dye indicators is expected not to deviate significantly from the selectivity behaviour of the less lipophilic analogues used in conventional extraction systems. The influence of other metal ions on the absorbance value at 523 nm, at a level of 3 x 10-5 mol dm-3 Zn2+, was investigated (see Table 1). As the rate of complex dissociation was slow for Cu, and complex dissocia- tion almost irreversible for Co, the selectivity towards these metal ions was determined at the end of the selectivity measurements.An exact description of the selectivity of the optode membrane described here can only be given for ions of the same charge and for chelates with the same stoichiometry and the same spectral properties. For most of the interfering ions considered, these prerequisites are unfortunately not fulfilled.ls Further, the activities of the interfering ions in the sample solution should be known, which requires certain assumptions and extensive calculations. The indicator acts as a non-selective chelating agent towards many metal ions. However, by analogy with extraction systems, the inter- ference from metal ions can be reduced by prior extraction, by masking with certain ligands or by an appropriate choice of the pH.'" Conclusions It has been shown that a lipophilized analogue of the classical indicator dye PAR is well-suited for the construction of an optode membrane based on plasticized PVC.The system described here offers significant advantages over other optodes reported to date, which make use of indicator dyes. These membranes might not only be used as reversibly functioning sensing layers, but also as simple extraction devices for the preconcentration of certain metal ions. Preconcentration by conventional solvent extraction provides a useful, but often sophisticated, means of improving the sensitivity of a determination, e.g., in atomic absorption spectrometry." The extraction of metal ions from a largc volume of the aqueous phase into a small volume of the organic liquid can be a necessary step in trace determinations or can enable the separation of the metal ions from an interfering matrix.An efficient phase separation, e.g., no solvent entrainment phenomena after the extraction, is an important criterion in the design of an extraction system, which is fulfilled in the system described here. There are many possibilities for introducing such optode membranes into other fields of application, e.g., in the titration of trace amounts of metals. This work was partly supported by the Swiss National Science Foundation. The authors thank S. Tan for careful reading of the manuscript. 1 2 3 4 5 6 7 8 9 10 I 1 12 13 14 15 16 17 18 19 20 21 References Seitz.W. 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