Utilization of Lipophilic Ionic Additives in Liquid Polymer Film Optodes

as components for use in liquid polymer film-based optodes. These ionic additives are known to strongly influence the selectivity behavior of anion-se...
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Articles Anal. Chem. 1997, 69, 990-995

Utilization of Lipophilic Ionic Additives in Liquid Polymer Film Optodes for Selective Anion Activity Measurements Susan L. R. Barker, Michael R. Shortreed, and Raoul Kopelman*

Department of Chemistry, The University of Michigan, Ann Arbor, Michigan 48109-1055

Lipophilic anionic and cationic additives are investigated as components for use in liquid polymer film-based optodes. These ionic additives are known to strongly influence the selectivity behavior of anion-selective electrodes. They also make it possible to construct ion coextraction optodes that can measure anion activities. Theoretical, thermodynamic anion-optode equilibria formalisms were derived to help understand the influence these additives have on the overall optode response and on the selectivity behavior. Neutral and charged anion carrier film configurations are described and tested for two anion ionophores with known modes of action, ruthenium(II) octaethylporphyrin and a vitamin B12 derivative (cyanoaquacobyrinic acid heptakis(2-phenylethyl ester)). These film configurations were further tested using a less well-understood ionophore, indium(III) octaethylporphyrin. Notable is the indium(III) octaethylporphyrin optode, which has a dynamic range appropriate for physiological measurements of chloride at neutral pH values. Also of interest is the optode with the vitamin B12 derivative ionophore, which has a dynamic range appropriate for some physiological measurements of nitrite at neutral pH values. Lipophilic anionic and cationic additives strongly influence the selectivity behavior of anion-selective electrode membranes1-4 and, in some cases, aid in determining the mode of action by which that membrane responds.1 That is to say, additives play an important role in determining whether the ionophore acts as a neutral or charged carrier (defined by the charge of the unbound ionophore). Understanding the mode of action of the ionophore is critical in optimizing the selectivity and response characteristics of both optodes and ion-selective electrodes (ISEs). Ionophores capable of binding multiple anions can sometimes operate in a mixed-mode manner.1,5,6 Here, the ionophore initially acts as a (1) Bakker, E.; Malinowska, E.; Schiller, R. D.; Meyerhoff, M. E. Talanta 1994, 41, 881-890. (2) Huser, M.; Morf, W. E.; Fluri, K.; Seiler, K.; Schulthess, P.; Simon, W. Helv. Chim. Acta 1990, 73, 1481-1495. (3) Rothmaier, M.; Simon, W. Anal. Chim. Acta 1993, 271, 135-141. (4) Schaller, U.; Bakker, E.; Spichiger, U. E.; Pretsch, E. Anal. Chem. 1994, 66, 391-398. (5) Li, X.; Harrison, D. J. Anal. Chem. 1991, 63, 2168-2174.

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charged, cationic carrier and becomes neutral upon binding the first anion. This is followed by a second binding event whereby the ionophore becomes negatively charged. Thus, the ionophore acts first as a charged carrier and then as a neutral carrier. Metalloporphyrins with two axial ligation sites can sometimes be considered to operate in such a mixed-mode manner. The mixedmode of action may be one contributor to the notoriously nonNernstian behavior of such anion-selective electrodes.1,6,7 Thermodynamic descriptions of the selectivity behavior of cation- and anion-selective optodes have been given which can be reliably used to predict optode behavior.8 These theories are used extensively in the development of cation-selective optodes.9-12 The predominance of cation (rather than anion)-selective optodes is likely due to the availability of highly selective and lipophilic cation ionophores.9 Anion carriers,3,13,14 on the other hand, are still relatively few in number. The response of liquid polymer film anion-selective optodes can be thought of in terms of a twophase liquid-liquid extraction. An anion is extracted from the aqueous phase by an ionophore which is dissolved in an organic liquid polymer, namely plasticized poly(vinyl chloride) (PVC). Electroneutrality requires that either a cation be coextracted with the anion or another anion be expelled. Placing an ideally selective15 proton ionophore (chromoionophore C) in the liquid polymer favors the former mode of action. Upon binding a proton, this chromoionophore undergoes a change in its optical properties (absorbance or fluorescence), which is used to indirectly measure the aqueous analyte ion activity. (6) Park, S. B.; Matuszewski, W.; Meyerhoff, M. E.; Liu, Y. H.; Kadish, K. M. Electronanalysis 1991, 3, 909-916. (7) Badr, I. H. A.; Meyerhoff, M. E.; Hassan, S. S. M. Anal. Chim. Acta 1996, 321, 11-19. (8) Bakker, E.; Simon, W. Anal. Chem. 1992, 64, 1805-1812. (9) Shortreed, M. R.; Bakker, E.; Kopelman, R. Anal. Chem. 1996, 68, 26562662. (10) Bakker, E.; Willer, M.; Pretsch, E. Anal. Chim. Acta 1993, 282, 265-71. (11) Lerchi, M.; Bakker, E.; Rusterholz, B.; Simon, W. Anal. Chem. 1992, 64, 1534-40. (12) Lerchi, M.; Reitter, E.; Simon, W.; Pretsch, E.; Chowdhury, D. A.; Kamata, S. Anal. Chem. 1994, 66, 1713-1717. (13) Stepa´nek, R.; Kra¨utler, B.; Schulthess, P.; Lindemann, B.; Ammann, D.; Simon, W. Anal. Chim. Acta 1986, 182, 83-90. (14) Sugimoto, H.; Ueda, N.; Mori, M. Bull. Chem. Soc. Jpn. 1981, 54, 34253432. (15) Bakker, E.; Lerchi, M.; Rosatzin, T.; Rusterholz, B.; Simon, W. Anal. Chim. Acta 1993, 278, 211-25. S0003-2700(96)00700-7 CCC: $14.00

© 1997 American Chemical Society

using anion carriers with known modes of action and then on a formerly less well understood carrier. In addition, a method for increasing the sensitivity of an anion optode response is demonstrated by means of calculation and experiment. THEORY Anion Carrier Optode Modes of Action. As shown above (Figure 1C,D), lipophilic ionic additives can be included in optode films to maintain the ionic strength within the liquid polymer, leading to a constant mean activity coefficient γ.18 This addition makes it possible to derive thermodynamically more exact expressions for the equilibrium behavior of an optode. The process of anion-proton pair coextraction into such an optode film (Figure 1C,D) can be expressed by simple net ionic equations (eqs 1 and 2). Here, the ionophore L (neutral carrier) or L+ Kcoex

aXaq- + aHaq+ + γL[Lorg] + γC-[Corg-] 798 γLX-[LXorg-] + γCH[CHorg] (1) Kcoex

aXaq- + aHaq+ + γL+[Lorg+] + γC[Corg] 798 Figure 1. Four illustrations of coextraction of an anion-proton pair from aqueous solution into the organic liquid polymer film of the optode. (A) Ion coextraction into the film is followed by binding to a basic chromoionophore and neutral ionophore. The formed ion pair increases the ionic strength in the film. (B) Ion coextraction into the film is followed by binding to an acidic chromoionophore and charged ionophore. The ion pair is dissolved, decreasing the ionic strength in the film. (C) Ion coextraction into the film is followed by binding to an acidic chromoionophore and neutral ionophore in the presence of a lipophilic cationic additive. The ionic strength in the film is a constant. (D) Ion coextraction into the film is followed by binding to a basic chromoionophore and charged ionophore in the presence of a lipophilic anionic additive. The ionic strength in the film is a constant.

Introduction of charged molecules into a hydrophobic, organic film requires careful forethought with regard to counterions. One’s first notion might be to begin with a neutral anion carrier L and a basic chromoionophore C combination (Figure 1A). In this way, as each anion-proton pair is extracted into the organic phase, the counterions (LX- and CH+) are automatically formed. Starting with a charged anion carrier L+ and an acidic chromoionophore C- would be as effective (Figure 1B). A number of examples of such systems exist in the literature.16,17 However, in both systems, a rigorous description of the thermodynamic behavior is hampered by the lack of constant ionic strength in the optode film. The purpose of this work is to explore the role ionic additives play in formulating anion-selective optode films with a predictable optical response. Unreactive lipophilic cations and anions are used here in conjunction with the various chromoionophores and ionophores to prepare anion-selective optode films that maintain a constant ionic strength in the organic phase (Figure 1C,D). New thermodynamic descriptions that permit the measurement of the aqueous anion activity are given for the anion optode response displayed in Figure 1C,D. These formulations were tested first (16) Kuratli, M.; Badertscher, M.; Rusterholz, B.; Simon, W. Anal. Chem. 1993, 65, 3473-3479. (17) Tan, S. S. S.; Hauser, P. C.; Wang, K.; Fluri, K.; Seiler, K.; Rusterholz, B.; Suter, G.; Kru ¨ ttli, M.; Spichiger, U.; Simon, W. Anal. Chim. Acta 1991, 255, 35-44.

γLX[LXorg] + γCH+[CHorg+] (2)

(charged carrier) selectively binds the anion X-, which results in the concomitant coextraction of a proton H+ by an acidic C- or a basic C chromoionophore, respectively. In eqs 1 and 2, concentrations in the film are denoted by brackets, activity coefficients in the film by γ, and activities in the aqueous sample solution by aX- or aH+. The traditional equilibrium constant Kcoex which governs this process is a function of a number of parameters which have each been discussed previously,8 including the acidity constant of the chromoionophore, Ka; the stability constant of the ionophore-anion complex, βLX; the activity coefficients in the organic (film) phase, γ; and the distribution coefficients of the free ions between the organic phase and the aqueous solution phase, k. The two net ionic equations (eqs 1 and 2) can be rewritten with the appropriate mass and charge balance conditions in an analytically more useful form (eqs 3 and 4), where the optical

aX- )

aX- )

( )( )( )( ( )( )( ) ( 1

Kcoex

[Rtot+] - R[Ctot] 1 1-R aH+ R [Ltot] - ([Rtot+] - R[Ctot])

1 1 1-R × Kcoex aH+ R

)

(3)

)

(4)

[Ltot] - ([Rtot-] - (1 - R)[Ctot]) [Rtot-] - (1 - R)[Ctot]

response π ≡ (1-R) (the degree of protonation of the chromoionophore, defined in terms of absorption A or fluorescence9 F) is expressed as a function of the anion (aX-) and proton (aH+) activities. These expressions are useful not only in understanding the selectivity behavior but also in predictably tuning the dynamic (18) Morf, W. E. The Principles of Ion-Selective Electrodes and of Membrane Transport; Elsevier Scientific Publishing Co.: Amsterdam, 1981.

Analytical Chemistry, Vol. 69, No. 6, March 15, 1997

991

Figure 2. An increasing optode sensitivity, displayed as the slope of the theoretical optode response (eqs 3 and 4) for protonation values (π ) 0.5), can be obtained by increasing the concentration of ionophore relative to chromoionophore and ionic additive.

range of the optode response to the desired region of interest.9,11,19 Selectivity. The selectivity coefficient of the optode film is easily determined8 in the case of a singly charged interfering ion Y-. It is simply the ratio of the individual coextraction constants, which reduces to a ratio of the product of the individual distribution coefficients and the ionophore-ion stability constants (eq 5). Mathematical relationships (which govern the selectivity

KX-Y-opt )

KCOEXYKCOEX

X-

≈

kY-βLYkX-βLX-

(5)

coefficients in cases where the interfering ion is multiply charged and where other binding stoichiometries must be considered) can be derived in an analogous fashion by including the appropriate stoichiometric coefficients. Practically, however, extraction of multiply charged anions is not expected.20 Sensitivity Enhancement. Manipulations of the ratios of ionophore to chromoionophore and charge sites can be performed to either enhance the sensitivity of the optode or increase the optode’s dynamic range. In films configured for either charged or neutral anion carriers, increasing the relative amount of ionophore to chromoionophore and ionic additive increases the sensitivity of the optode and decreases its dynamic range. Here, the sensitivity is defined as the slope of the optode response curve (eqs 3 and 4) at π ) 0.5. This dramatic effect is shown (Figure 2) for both optode systems, where the charged site concentration equals the chromoionophore concentration and the ionophore concentration is varied.

EXPERIMENTAL SECTION The following were used as received from Fluka (Ronkonkoma, NY): high molecular weight poly(vinyl chloride) (PVC), 2-nitrophenyl octyl ether (o-NPOE), bis(2-ethylhexyl) sebacate (DOS), chromoionophore II or 9-(dimethylamino)-5-[4-(16-butyl-2,14-dioxo3,15-dioxaeicosyl)phenylimino]-5H-benzo[a]phenoxazine (ETH (19) Barker, S. L. R.; Shortreed, M. R.; Kopelman, R. Proc. SPIE-Int. Soc. Opt. Eng. 1996, 2836, 304-310. (20) Ammann, D.; Huser, M.; Kra¨utler, B.; Rusterholz, B.; Schulthess, P.; Lindemann, B.; Halder, E.; Simon, W. Helv. Chim. Acta 1986, 69, 849854.

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2439), chromoionophore IV or 3-hydroxy-4-[(4-nitrophenyl)azo]phenyl octadecanoate (ETH 2412), nitrite ionophore I (NI I) or cyanoaquacobyrinic acid heptakis(2-phenylethyl ester), tridodecylmethylammonium chloride (TDDMACl), and potassium tetrakis[3,5-bis(trifluoromethyl)phenyl]borate (KTFPB). Indium(III) octaethylporphyrin chloride (In(OEP)Cl) was obtained from Midcentury (Posen, IL). Ruthenium(II) octaethylporphyrin carbonyl (Ru(OEP)CO) was used as received from Aldrich. Standard Solutions. Measurements were performed in 10 mM 2-amino-2-(hydroxymethyl)propane-1,3-diol (TRIS), 50 mM 4-morpholineethane sulfonic acid (MES), or KH2PO4 buffer solutions (Aldrich). Solutions of 1.019 N HCl and 0.991 N NaOH were used as received from Aldrich to adjust the buffer pH. Standard solutions were prepared with NaCl (Sigma), NaNO2, NaSCN, NaBr, KClO4, and KNO2 (Fluka). All solutions were prepared with 18 MΩ water (Barnstead I Thermolyne Nanopure II system, Dubuque, IA). Silanization. Prior to application of the optode film cocktail, the slides were immersed in 1% (v/v) 3-(trimethoxysilyl)propyl methacrylate in 1 mM acetic acid in toluene at 70 °C for 1 h in a well-ventilated fume hood. The glass was rinsed with dry toluene and baked at 140 °C for 10 min. Film Preparation. All optode films were prepared with 30 mmol/kg ionophore, 15 mmol/kg chromoionophore, and 15 mmol/kg lipophilic cationic (TDDMACl) or anionic (KTFPB) sites unless otherwise noted. All films were prepared with 66 wt % o-NPOE or DOS and 33 wt % PVC. The components were dissolved in freshly distilled tetrahydrofuran (THF, Aldrich). The optode samples were prepared by dipping the silanized glass slides repeatedly (∼5 times) into the film cocktail solution. Each consecutive pass consisted of only 0.1 s contact with the solution,followed by a 10 s evaporation period. The THF was allowed to evaporate completely before the films were tested. Optodes were viewed with an optical microscope, and the film thickness was found to be less than 5 µm. The exact porphyrin film compositions are as follows: for the response and selectivity experiments, the NI I film contained 3.1 mg of NI I, 1.8 mg of KTFPB, 1.4 mg of ETH 2439, 81.9 mg of DOS, and 41.2 mg of PVC; the Ru(OEP)CO film contained 2.9 mg of Ru(OEP)CO, 1.2 mg of TDDMACl, 1.1 mg of ETH 2412, 91.2 mg of o-NPOE, and 46.4 mg of PVC; the first In(OEP)Cl film contained 2.5 mg of In(OEP)Cl, 1.6 mg of KTFPB, 1.3 mg of ETH 2439, 79.5 mg of o-NPOE, and 34.9 mg of PVC; the second In(OEP)Cl film contained 4.3 mg of In(OEP)Cl, 1.7 mg of TDDMACl, 1.6 mg of ETH 2412, 129.7 mg of o-NPOE, and 65.0 mg of PVC. Films were also prepared which contained no ionophore. The first contained 1.29 mg of TDDMACl, 1.18 mg of ETH 2412, 121.4 mg of o-NPOE, and 61.0 mg of PVC. The second contained 14.7 mg of ETH 2439, 110.4 mg of o-NPOE, and 54.5 mg of PVC. For the sensitivity experiments, the optode film compositions are as follows: the low-sensitivity film contained 6.11 mg of NI I, 1.61 mg of KTFPB, 0.98 mg of ETH 5350, 221.93 mg of DOS, and 110.62 mg of PVC; the high-sensitivity film contained 15.10 mg of NI I, 1.89 mg of KTFPB, 1.13 mg of ETH 5350, 252.86 mg of DOS, and 126.44 mg of PVC. Absorbance Measurements. Absorbance measurements were made using a Shimadzu UV-160U UV/visible spectrophotometer controlled by a DTK computer using UV-160 Plus software. Optode film-coated glass slides were placed in 1 cm cuvettes. These optodes were rinsed three times in each new

sample solution prior to a 2 min equilibration period before measurement. The pH of the test sample solutions was measured with a calibrated glass electrode (Ingold). The films were allowed to equilibrate for 5-15 min prior to initial use in order to obtain a stable starting absorbance value. The absorbance maximum of the protonated ETH 2439 was measured at 665 nm, and the absorbance maximum of the basic form of the ETH 2412 was measured at 550 nm. Calculations. Activity coefficients were calculated according to the two-parameter Debye-Hu¨ckel formalism of Meier.21 Kcoex values were calculated from experimental data according to eq 3 or eq 4 for intermediate π values. The absorbance spectra were resolved using principal component analysis with the aid of MATLAB version 4.2c.1 software (The Mathworks, Inc., Natick, MA). RESULTS AND DISCUSSION To test the viability of the aforementioned theoretical predictions, it is important to select ionophores whose carrier modes of action are well established. A number of cobyrinates13,22 and metalloporphyrins1,2,5-7,20,23-26 have been prepared which, when incorporated into ion-selective electrode and optode films, yield selectivity patterns that deviate from the classical Hofmeister series. Selectivity patterns and UV/visible spectroscopic data confirm direct associative interaction between the analyte anion and the metal center of many of the complexes. The charge of the metal center and the availability of two axial ligand sites help determine the carrier mode. Metal(IV) (e.g., Sn(IV)) porphyrins can act only as charged carriers because of the charge on the metal.7 The Sn(IV) porphyrins bind two anionic ligands (which complicates the stoichiometry) and are, therefore, not good candidates for model charged carriers. The vitamin B12 derivative ionophore1,4,13,20 is known to act solely as a monovalent charged carrier.4,27 It has excellent film characteristics1,4,13,20 (near theoretical Nernstian slopes and good membrane lifetime) and is used here as a model compound. Porphyrins with metal(II) centers can act only as neutral carriers,1 of which Ru(OEP)CO is an excellent example.2 The greatest difficulty in assigning a mode of action comes with the metal(III)-centered porphyrins.1,6,26 The charge of the porphine ring is -2, and the coordination number of the metal is typically 6. The possibility of binding two axial ligands, which may be either neutral (e.g., water or pyridine) or anionic, opens the possibility for both charged and neutral carrier binding. Cobalt(III) porphyrins were once thought to act as charged carriers.2 Recently, a detailed study of the role of axial ligation on the potentiometric response of Co(TPP)+ was performed,26 and this ionophore is now known to act as a neutral carrier.1,26 Until recently, In(OEP)Cl was thought to act either as a neutral carrier or in a mixed-mode fashion6 due to its nonNernstian electrode response. Recent studies,1 including this work, confirm, however, that this ionophore acts solely as a (21) Meier, P. C. Anal. Chim. Acta 1982, 136, 363-368. (22) Schulthess, P.; Ammann, D.; Simon, W.; Caderas, C.; Stepa´nek, R.; Kra¨utler, B. Helv. Chim. Acta 1984, 67, 1026-1032. (23) Wang, E. J.; Meyerhoff, M. E. Anal. Chim. Acta 1993, 283, 673-682. (24) Chaniotakis, N. A.; Park, S. B.; Meyerhoff, M. E. Anal. Chem. 1989, 61, 566-570. (25) Chaniotakis, N. A.; Chasser, A. M.; Meyerhoff, M. E.; Groves, J. T. Anal. Chem. 1988, 60, 185-188. (26) Malinowska, E.; Meyerhoff, M. E. Anal. Chim. Acta 1995, 300, 33-43. (27) Schaller, U.; Bakker, E.; Spichiger, U. E.; Pretsch, E. Talanta 1994, 41, 1001-1005.

Figure 3. Degree of protonation π ) (1 - R) vs log(aH+aX-) ) -pH - pX. Vitamin B12 derivative ionophore optode response in a charged carrier optode film configuration. The curves are plots of eq 3. 4, Nitrite; O, thiocyanate; and 0, perchlorate. Due to overlap in the absorbance spectra of the ionophore and the chromoionophore which does not occur in the fluorescence spectra, nitrite and thiocyanate data points were measured via fluorescence.19 Table 1. Anion Selectivity Values (versus Perchlorate) (Logarithmic Units) for Three Different Optode Films and Their Corresponding ISEsa vitamin B12 derivative ionophore (NI I) Ru(OEP)CO ionophore SalClO4SCNNO3NO2IBrCl-

optode

ISE4,b

0 2.9

0 2.1 -1.6 1.8 -0.5 -1.5 -1.9

3.3

In(OEP)Cl TDDMACl

optode ISE2 optode ISE1 0 0.9 -2.5

-2.5

0 0.3 -3.0 -2.1 -1.6 -3.9 -5.1

2.9 0 3.0

ISE1

3.3

0 3.8 -1.6 3.3 3.2

-1.4 0 -1.4 -3.0 -3.8 -1.4

2.3

2.5

-4.7

a The so-called Hofmeister selectivity pattern is given on the basis of the ISE response with a film containing a dissociated cationic ionexchanger (TDDMACl). b o-NPOE was used as plasticizer.

charged carrier. The mode of action of the indium(III) porphyrin ion carrier is confirmed by the optode response in accordance with the theoretical predictions. These studies are discussed in greater detail below. Charged Carrier Mode of Action (Figure 1D). In the charged carrier optode system, negatively charged lipophilic additives (tetraphenylborate salts) are used as counterions for both the ionophore and the chromoionophore. The ionophore is positively charged in its unbound form, while the basic chromoionophore is neutral in its unbound form. Here, as an anionproton pair is extracted, an ionophore molecule becomes neutral upon anion binding, and a chromoionophore binds a proton to become cationic. In this way, the net number of charges in the system is constant. The net ionic reaction is shown in eq 2, and the expression defining the expected analytical response is shown in eq 4. This theoretical curve is plotted with the experimental data in Figure 3. The selectivity pattern (Table 1) is in good agreement with that of ISEs employing the same ionophore,4,27 noting the use of a different plasticizer. To further prove that the vitamin B12 derivative ionophore is, indeed, a charged carrier, this ionophore was placed in an optode Analytical Chemistry, Vol. 69, No. 6, March 15, 1997

993

Figure 4. Degree of protonation π ) (1 - R) vs log(aH+aX-) ) -pH - pX. Ru(OEP)CO optode response in a neutral carrier optode film configuration. The curves are plots of eq 4. O, Perchlorate; 0, bromide; ], thiocyanate; and 3, nitrate.

Figure 5. Degree of protonation π ) (1 - R) vs log(aH+aX-) ) -pH - pX. In(OEP)Cl optode response in a charged carrier optode film configuration. The curves are plots of eq 3. 0, Salycilate; O, iodide; 4, thiocyante; and ], chloride.

film configured for a neutral carrier. This film contained cationic lipophilic additives (quaternary ammonium salts) and an acidic chromoionophore. Here, the selectivity pattern is expected to be that of a dissociated ion exchanger (Hofmeister selectivity). UV/ visible spectroscopic data (not shown) were not useful in determining whether or not specific binding between the vitamin B12 derivative ionophore and the anions occurred. This was due to severe overlap of the ionophore absorbance spectra with the absorbance spectra of both forms of the chromoionophore. A precursor of the vitamin B12 derivative ionophore was stated22 to undergo a shift in its UV/visible spectrum associated with a structural change upon binding. This spectroscopy was, however, not discussed in detail. Neutral Carrier Mode of Action (Figure 1C).8 In the neutral carrier optode system, positively charged lipophilic additives (quaternary ammonium salts) are used as counterions for both the ionophore and the chromoionophore. Here, the resulting anion-ionophore complex is negatively charged, as is the chromoionophore in its basic form. As a proton-anion pair is extracted from the aqueous sample solution into the liquid polymer film, the chromoionophore turns into its neutral, protonated form, and the ionophore binds an anion to become negatively charged. Again, the net number of charges in the film is unchanged. The net ionic reaction is shown in eq 1, and the expression defining the expected analytical response is shown in eq 3. The neutral carrier optode (Ru(OEP)CO, TDDMACl, ETH 2412) responses to changes in the anion activity correspond well with the theoretical predictions (Figure 4), and the selectivity values (Table 1) are in good agreement with those of a previously prepared ISE2 (Ru(OEP)CO, 10 mol % TDDMACl, o-NPOE, PVC). Attempts were made to test the response of the neutral carrier, Ru(OEP)CO, in optode films configured for charged ion carriers. As expected, the films yielded a Hofmeister selectivity pattern, indicative of nonspecific ion exchange without ionophore-induced selectivity. In addition, the response of such a film (Ru(OEP)CO, ETH 2439, o-NPOE, and PVC) was found to be exactly the same as the response of a film prepared without ionophore (ETH 2439, o-NPOE, and PVC), indicating a complete lack of stabilization of the anion by the ionophore.

It has been reported2 that the absorption spectrum of Ru(OEP)CO undergoes a shift in the absorbance maximum (550 nm) upon binding SCN- but not with ClO4- or Cl-. The results (not shown) for the optode film described here are at odds with these findings, however. The absorbance maximum of the Ru(OEP)CO increased and shifted to shorter wavelengths upon binding with SCN-, ClO4-, Br-, and NO3-. Unknown or Mixed-Mode Carrier Optode Films. Prior to the discovery of the impact of lipophilic ionic additives on the selectivity behavior of anion-selective electrodes,1 the modes of action by which specific anion carriers acted were speculative. The mysterious non-Nernstian slopes have led some researchers to hypothesize a mixed mode of action whereby both neutral and charged forms of a carrier were capable of selective anion extraction from the sample solution into the organic phase. Lipophilic cationic and anionic additives have only recently shed light on the true carrier modes of action. The porphyrin In(OEP)Cl, once thought to be a neutral carrier,6 has now been shown to behave as a charged carrier1 in ISE membranes. To experimentally determine the carrier mode of action of an ionophore, the ionophore in question can simply be placed in two different optode films, one configured for a charged carrier and the other configured for a neutral carrier. The mode of action is determined according to when one of the two configurations has a selectivity pattern that deviates from the classical Hofmeister series selectivity. In the case of In(OEP)Cl, an optode film configured for a charged carrier (Figure 5) showed enhanced selectivity toward Cl-. The optode selectivity pattern found for In(OEP)Cl is quite comparable (see Table 1) to that found using ISEs with similar compositions.1 It is interesting to note that spectroscopic shifts of peaks in the visible absorbance spectrum of the indium porphyrin in optode films configured for a neutral anion carrier did occur. ISE studies1 indicate that the indium porphyrin acts as a charged carrier with ISE membranes containing the neutral form of the indium porphyrin, yielding a Hofmeister selectivity pattern. Optode films with the indium porphyrin configured for a neutral carrier have a Hofmeister selectivity pattern as well; however, shifts in the indium porphyrin peaks upon addition of SCN- to the aqueous sample solution indicate that there is ion exchange occurring at the fifth

994 Analytical Chemistry, Vol. 69, No. 6, March 15, 1997

axial ligand site. This would be expected on the basis of the enhanced sensitivity of the charged indium porphyrin for SCNversus Cl-. No associated chromoionophore response is expected, as the ion exchange need not be accompanied by concomitant extraction or expulsion of a proton. The enhanced selectivity of films with In(OEP)Cl (compared to dissociated ion exchangers) has been shown to be useful in the determination of chloride in serum samples.6 The physiological range of chloride is 90-120 mM.6 Using a typical pH of 7.4, the required activity product for determination of chloride with an optode using a coextraction mode of action is log((aH+)(aCl-)) ) -8.4. The particular optode composition used in this article would be appropriate for measurements in this range. Measurements of intracellular chloride would also be possible. The typical intracellular chloride concentrations28 are 5-15 mM, with intracellular pHs29 being approximately 7.2-7.3. The required activity product is, therefore, -9.2 which is also within the range of this sensor. Sensitivity Enhancement of Optode Films. Increasing the relative concentration of ionophore to chromoionophore and ionic additives is expected to enhance the sensitivity (or slope) of the optode response curve (see Figure 2). To test this hypothesis, two different optode films were prepared, having different relative concentrations of ionophore to chromoionophore and ionic additives (eq 6). The film compositions (n ) 2.0 and 4.2) do, indeed,

n)

[Ltot] [Ctot]

[Ltot] )

[Rtot]

(6)

have different slopes (Figure 6) as expected. There is, however, an apparent shift in the equilibrium constant (log Kexch) from -11.4 to -11.0, coupled with a slight increase in the absolute detection (28) Alberts, B.; Bray, D.; Lewis, J.; Raff, M.; Roberts, K.; Watson, J. D. Molecular Biology of the Cell, Vol. 2; Garland Publishing: New York, 1989. (29) Ammann, D. Ion-Selective Microelectrodes; Springer-Verlag: Berlin-Heidelberg, 1986; p 346.

Figure 6. Nitrite-selective optode response for two films displaying different sensitivity: O, higher (n ) 4.2) and 0, lower (n ) 2.0). The sensitivity of an anion-selective optode film can be increased by increasing the relative concentration n of ionophore to chromoionophore and ionic additive (eq 6). (see Experimental Section for exact film compositions.)

limit with increasing n. An increase in the detection limit is, of course, expected but would be smaller if the equilibrium constants for both optode films were identical. ACKNOWLEDGMENT We gratefully acknowledge the useful discussions we had with Prof. Eric Bakker, Dr. Ulrich Schaller, and Prof. Mark Meyerhoff and members of his group. We acknowledge NIH Grant 1 RO 1 GM50300 01 for financial support. Received for review July 18, 1996. Accepted January 9, 1997.X AC960700F X

Abstract published in Advance ACS Abstracts, February 15, 1997.

Analytical Chemistry, Vol. 69, No. 6, March 15, 1997

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