1714
Anal. Chem. 1988, 60,1714-1721
Ion-Selective Electrodes Based on Natural Carboxylic Polyether Antibiotics Koji Suzuki,* Koji Tohda, Hiroshi Aruga, Misako Matsuzoe, Hidenari Inoue, and Tsuneo Shirai* Department of Applied Chemistry, Keio Uniuersity, 3-14-1 Hiyoshi-cho, Kohoku-ku, Yokohama 223, J a p a n The response characteristics of poly(vinyl chloride) (PVC) matrix membrane Ion-selectlve electrodes based on five natural carboxylic polyether antlbiotks, ionomycin, lasalocid, monensln, nlgerlcin, and salinomycin, were examined. These are characterlred by a good response to divalent (alkalineearth-metal) cations such as Ca2+ and Ba2+. However, electrodes based on methyl esters, in which the carboxyl groups of the natural carboxylic polyethers are esterified, responded weakly to divalent cations. This Indicated that the carboxyl group plays an Important role in making a complex with the dlvalent catlon In a ilpophillc polymeric electrode membrane, which is signiflcant for the electrode response to the dlvaient cation. Electrodes based on all the carboxylic polyethers except iasaiockl show “bi-Nernstlan response” for divalent catlons. I n order to explain this unusual response, ion extraction and permeability measurements were carried out with lasalocid and monensln. The general relationship between electrode response and ion extraction or ion-ligand complexatlon is also dlscussed on the basis of the results obtained with lasaiocid and monensin.
Natural carboxylic polyethers are a class of antibiotics produced by various Streptomyces species ( 1 , Z ) . They are mostly open-chain molecules of the polyether type containing heterocyclic rings and a variety of functional oxygen atoms of the carbonyl, carboxyl, ether, hydroxyl, and ketone type. They, therefore, have the ability to form complexes with alkali-metal and alkaline-earth-metal ions. It has been reported ( 1 , 3 , 4 )from crystallographic data that many of these molecules become cyclic by forming hydrogen bonds between a carboxyl group a t one end of the molecule and a hydroxyl group at the other end. These molecules make complexes with such cations and provide ion binding specificity due to the size of the cavity created when the cyclic molecule is formed. Most of these complexes are lipophilic and are classified as “ionophores” ( 5 ) . These characteristics satisfy the requirements of an active ligand for an ion-selective electrode (ISE). Though their biological actions have been actively investigated (1,2,6, 7), only a few applications for ISEs have been reported using A23187 (8), monensin (9), and nigericin (9). Here, we report the detailed response characteristics and response mechanisms of ISEs based on five natural carboxylic polyether antibiotics, ionomycin, lasalocid, monensin, nigericin, and salinomycin, whose chemical structures are shown in Figure 1. In addition, we have also modified the carboxyl group of these carboxylic polyethers by methyl esterification and examined the ion response characteristics of the ISEs by using these metyl esters to better understand the importance of the carboxyl group on natural carboxylic polyethers. Carboxylic polyethers produce negative charges by deprotonation of the carboxyl group, which gives them features different from the so-called “neutral carrier ligand” (10). Therefore, we expected to make ISEs sensitive to divalent cations (alkaline-earth-metal ions) rather than to monovalent cations (alkali-metal ions) by using these carboxylic polyethers. While sensitive electrodes for the divalent cations were actually obtained, unusual response slopes were observed for the electrode based on all the natural carboxylic polyethers except
lasalocid. The ISEs based on these carboxylic polyethers show a linear response slope of approximately 60 mV/activity decade (a.d.) a t 25 “C for divalent cations such as Ca2+and Ba2+,which corresponds to twice the usual Nernstian theoretical response slope (theoretical slope for divalent cations: 29.6 mV/a.d. at 25 “C). In order to explain the reason for this peculiar response, ion extraction and ion permeability measurements were carried out in addition to electromotive force (emf) measurements with two typical carboxylic polyethers, lasalocid and monensin, which show different response slopes for Ba2+ as a divalent cation. Furthermore, the general relationship between ISE response and ion extraction or ion-ligand complexation, which contributes to the generation of an emf at the ISE membranesample solution interface, is also discussed on the basis of results obtained with the two typical carboxylic polyethers. EXPERIMENTAL SECTION Reagents. All chemicals used were reagents of the highest grade commercially available. Distilled and deionized water had resistivities more than 1.5 X lo7Q cm at 25 “C. The active ligands (ionophore) used for ISEs were ionomycin (Calbiochem,American Hoechst Corp., La Jolla, CA), lasalocid (Aldrich Chemical Co., Milwaukee, WI), monensin (Sigma Chemical Co., St. Louis, MO), nigericin (Sigma), salinomycin (Pfizer Taito Co., Ltd., Tokyo, Japan), and valinomycin (Sigma). All ligands were purified by reversed-phased high-performanceliquid chromatography (HPLC) (column: D-ODs-5, 20 (0.d.) X 250 mm (YMC Co., Ltd., Kyoto, Japan); eluent: acetone, 5 mL/min) with refractive index (RI) detection (RID: SE-51 (Showa Denko Co., Ltd., Tokyo)). Ion-free ligand crystals of the natural carboxylic polyethers required for esterification, extraction, and permeability experiments were prepared by evaporation of chloroform after twice extracting with 1 N HCl/chloroform (1/1)and water/chloroform (l/l).The free forms were confirmed by checking their melting points ( I l ) , IR spectra [KBr; u CO, 1700-1710 cm-* (acid form, dimer)), and ‘H NMR data (12-16). Methyl Esterification of Carboxylic Polyethers. Carboxyl groups of all the studied natural carboxylic polyethers were methyl esterified according t o the method reported by Ono et al. (17). The products were finally purified by silica gel column chromatography with benzene/ethyl acetate (2/1) followed by reversed-phase HPLC (eluent: acetone) with RI detection. Satisfactory elemental analyses and ‘HNMR and IR spectroscopic data were obtained for all methyl esters (all oily liquids) derived from the five natural carboxylic polyethers investigated. Preparation of ISE. Ion-sensitive membranes of the PVC matrix type prepared according to the procedure of Moody and Thomas (18)were used. The membrane compositions were 3 wt % active ligand (ionophore), 70 wt % membrane solvent (dibutyl sebacate (DBS), Tokyo Chemical Industry Co., Ltd., Tokyo), and 27 w t 7” PVC (high molecular weight type, Sigma Chemical Co.). Membrane thicknesses were ca. 100 pm. A 7-mm-diameter circle was cut out of the prepared membrane and mounted on a Philips IS-561 electrode body for ISE preparation. the reference electrode was a double-junction type based on an Ag-AgC1 electrode (HS-305DS, TOA Electronics,Ltd., Tokyo). Thus, the electrode cells for the emf measurements were as follows: Ag; AgC1, 3 M KCl10.3 M NH4N031sampleJmembrane10.1 M MCl or MCl,, AgCl; Ag M: strongest sensitive metal cation for each electrode Measurement of Electrode Potential. The ISE response potential (emf)was measured at 25 f 0.5 “C with a digital mV/pH
0003-2700/88/0360-1714$01,50/00 1988 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
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Sal inomycin Flgure 1. Chemical structures of the natural carboxyic poiyether antibiotics discussed.
meter (HM-ME, TOA) and recorded by a personal computer (PC9801-VM2, NEC Corp., Tokyo) with an interface (KADEC 1, KONA System Co., Ltd., Sapporo, Japan) and associated software (PHPRO, Ver. 1.01, KONA). The emf that remained constant within f0.5 mV for at least 3 min (in this condition at least 95% response was attained) was used as the electrode potential. The sample was a cation chloride solution unless otherwise stated. The pH-adjusting buffer was 0.1 M Tris/HCl (pH 7.0-10.0). The pH of the other solutions, below 7.0, was adjusted only with HC1. Activity coefficients were calculated from the equation proposed by Kielland et al. (19). When a pH buffer was used, the activity was obtained from the Debye-Huckel equation by using the ion strength of the buffer and ions for calculations (20). The selectivity coefficients of ISE (KijPot:i, primary ion; j, interfering ion) were calculated from response potentials in a lo-' M cation chloride solution (pH 7.0, Tris buffer) by the separate solution method (SSM; 21, 22). E x t r a c t i o n Measurement. The following experiment was carried out to determine the stoichiometry of the ion-ligand complex in an electrode membrane and the ion extraction ability of the ligand into the membrane. A DBS solution containing a certain amount of ion-free carboxylic polyether (1 X lo-* M, 2 mL (2 X mol)) and a cation chloride solution ((0-2) X lo-' M, 2 mL ((0-4) X 10" mol); pH 7 or 9.5,O.l M Tris/HCl), which corresponds to 0-2 times the weight of the ligand, were mixed in a 10-mL test tube for centrifugation. After the tube was shaken vigorously for 1 min, followed by centrifugation, the remaining ion in the aqueous phase was determined with an atomic absorption spectrophotometer (Type 170-30, Hitachi Co., Ltd., Tokyo) or ion chromatograph (IC-100,Yokogawa Electric Works, Tokyo). Ion P e r m e a b i l i t y (Electrodialysis) Measurement. Two 30-mm-cubic polystyrene compartments separated by a PVC matrix membrane (10-mm circle) especially prepared for the ISEs were used. A 15-mL sample was introduced into each compartment. A constant voltage (-40 V) was applied for -24 h by a potentiostat (Model 2001, Toho Technical Research Co., La., Tokyo) through two Ag-AgC1 electrodes dipped in the solutions. Constant stirring was maintained. The resulting current or applied coulombs were measured. This measurement had been performed under a N2 atmosphere, with dissolved oxygen in the sample solution being removed before use. The concentration of ions in
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988 I I Valinomycin
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Table I. Response Characteristics of ISEs Based on Natural Carboxylic Polyether Antibiotics and Their Methyl Esters
Nigericin methyl ester
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most strongly sensitive cation speciesn
linear response region, M
response slope, mV1a.d.
Ca2+ Ba2+ Ba2+
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58
Ion selectivity factors (Kijwt)are shown in Figure 5 and Figure 3b,c. Test solutions contain 0.1 M Tris/HCl (pH 7).
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-6
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to the strongest sensitive cation species from 1 X to 1 X 10-1 M was within 30 s. The ion selectivity factors of these electrodes are shown in Figure 5 and Figure 3b,c and are indicated as a function of the crystal ionic radii (24) of cations. (The selectivity coefficients (SSM; 0.1 M cation chloride) determined from the response of the ionomycin electrode to Ca2+and the responses of monensin, nigericin, and salinomycin electrodes to Ba2+were calculated as CaL+ and BaL+ (monovalent ion species; L represents ligand). The reason is explained in detail in the latter part of this report. These ionic radii were plotted a t the values of Ca2+and Ba2+.) Ionomycin Electrode. The carboxylic polyethers investigated are all monobasic acids except for the ionomycin molecule. This ligand is dibasic, having an enolized P-diketone in addition to the normal carboxyl group (25). Ionomycin is known to form a stable complex with Ca2+,which has been reported as a 1:l ligand-cation complex (strictly, a 2:2 complex form (12)). Actually, this ligand-based electrode shows high sensitivity to Ca2+. For monovalent cation selectivity, the electrode responds strongly to Li+. The Li+ selectivity on the response of divalent cations can be improved by using the
-1
l o g aga2+ Figure 4. Response curves for Ea2+ obtained by ISEs based on valinomycin, nigericin, and nigericin methyl ester.
a complex with a divalent cation in a lipophilic electrode membrane phase. The cation response characteristics of ISEs based on all the natural carboxylic polyethers investigated are summarized in Table I. The stabilities (noise) of all the ISEs shown in Table I were less than 0.1 mV, and the response time (95% response) I
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
methyl ester of this ligand (see Figure 5a,b). Lasalocid Electrode. This electrode is highly selective for Ba2+,and its reponse slope for Ba2+is 29 mV1a.d. Among all the investigated ISEs based on natural carboxylic polyethers, this is the only electrode that responds to divalent cations with a Nernstian theoretical response (theoretical reponse slope: 29.6 mV1a.d. at 25 "C). Lasalocid is one of the few carboxylic polyethers that form dimeric complexes with monovalent cations as well as with divalent cations (3, 13, 26, 27). The Ba2+ complex of lasalocid coordinates two ligand molecules with the cation unsymmetrically (26). The more interesting coordination behavior is that there are no hydrogen bonds from one ligand to the other, whereas other dimeric complex-forming ligands such as monensin and nigericin have intermolecular hydrogen bonds (3). Another unique feature observed for the lasalocid electrode is that its ion selectivity and sensitivity are independent of solvent polarity. The selectivity was almost the same whether a low-polarity solvent, DBS (e8 = 4; eB is specific inductive capacity), or a relatively high polarity solvent, 2-nitrophenyl octyl ether (NPOE; tg = 24), was used as the membrane solvent. The electrodes using the other four natural carboxylic polyethers with NPOE gave neither sufficient linear reponse against any cations nor apparent selectivity. It has also been reported by NMR studies that the backbone configuration of the lasalocid-Ba2+ complex is unchanged with solvent polarity (13). Monensin and Nigericin Electrodes. Monensin and nigericin have similar skeletons and very similar configurations in their complexed form. For their backbone lengths, monensin and nigericin prefer Na+ and K+, respectively. Though emf measurements have already been performed (9) on these ligands with the other membrane solvents we investigated, our results include some other interesting facts. ISEs based on monensin and nigericin are sensitive to Ba2+as well as to the monovalent cations, such as Na+ and K+. Furthermore, these electrodes show unusual responses to Ba2+in that the response slopes of monensin and nigericin electrodes are 58 and 57 mV/a.d., respectively. These slopes are twice the theoretical Nernstian response (the reason for this peculiar response will be fully discussed later in this report). Other facts are related to the improvement in selectivity of these electrodes. We modified the carboxyl groups of the carboxylic polyethers, including monensin and nigericin, by esterification and compared the ion selectivities of the electrodes based on carboxylic polyethers and those based on their methyl esters. As shown in Figure 5e,f and Figure 3b,c, the selectivities for monovalent cations of electrodes based on monensin and nigericin methyl esters are similar to those of the electrodes using the acid forms of monensin and nigericin. On the other hand, ion selectivities for divalent cations are significantly decreased by esterification. Thus, the Na+- and K+-ISEs, having high selectivity against divalent cations, could be obtained by using the methyl esters of monensin and nigericin, respectively. In this case, the improvement in the pH effect on the nigericin electrode, shown in Figure 2b,c as an example, is an additional advantage for these electrodes. Salinomycin Electrode. The crystallographic structure of the ion-ligand complex of salinomycin has not yet been reported except for the p-indophenacyl derivative (28). As shown in Figure 5g, a salinomycin electrode is highly selective for Ba2+versus other monovalent and divalent cations. The response slope for the divalent cation is approximately 60 mV1a.d. (58 mV/a.d.), which is the same for the monensin and nigericin electrodes for Ba2+. Naphthenic Acid Electrode. Naphthenic acid has a carboxyl group at the end of the molecule but no ether oxygens along its backbone. This molecule was used to compare re-
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Figure 6. Response curves for Ba2+ obtained by ISEs based on
lasalocid and monensin. sponse characteristics with those of other carboxylic acid based electrodes. As shown in Figure 5i, the electrode using naphthenic acid shows no specific selectivity. The response slope of this electrode is Nernstian for both monovalent and divalent cations such as K+ and Ba2+,ranging from 5 X to 1 X 10-1 M and 2 X to 1 X lo-' M, respectively. These responses indicate that esters in the backbone structures of natural carboxylic polyethers apparently play an important role in the formation of the "selective" ion complex. Figure 5 and Figure 3b,c show a comparison of ion selectivities of ISEs based on natural carboxylic polyethers and those based on their methyl esters. The response characteristics of electrodes using methyl esters are summarized in Table I. As mentioned above, divalent cation sensitivity is markedly decreased by esterification of the carboxyl group of the natural carboxyl polyethers. A more interesting fact is that the monovalent cation selectivity of these electrodes is almost unchanged by esterification. This indicated that the ion complex responsible for electrode response is the same for both types of electrodes; that is, the ion complex is a species formed by an electrically neutral ligand and an appropriate monovalent cation for the ligand. The response curve of K+ obtained by the nigericin electrode shown in Figure 2b also validated this assumption, because the electrode apparently to 1 X lo-' M, even with responds to K+ ranging from 7 X a sample pH of 2.5. At this pH, deprotonation occurs only weakly a t the carboxyl group of nigericin. As shown in Table I, all electrodes based on natural carboxylic polyethers except the lasalocide electrode gave unusual "bi-Nernstian" response slopes for some divalent cations such as Ca2+and Ba2+. To understand this peculiar response, ion extraction and ion permeability measurements were carried out with two typical carboxylic polyethers, lasalocid and monensin. These two polyether-based electrodes show different response slopes for divalent cations. Typical response curves for Ba2+,which is a divalent cation, are shown in Figure 6. A lasalocid electrode shows normal Nernstian response for Ba2+. On the other hand, a monensin electrode gave an unusual response slope of approximately 60 mV1a.d. (58 mV1a.d. at 25 "C) for the divalent cation, which corresponds to twice the theoretical Nernstian response slope (theoretical slope: 29.6 mV1a.d. at 25 "C). The response mechanism of ISEs based on carboxylic polyethers was therefore examined in association with these two typical ligands. Monensin is a well-known carrier that transports Na+ actively, using a pH gradient as the driving force (29). Therefore, the effect of active ion transport on the electrode response has been examined with this ligand. The active Na+ transport occurs across a lipophilic membrane containing monensin placed between Na+ solutions with the pH adjusted to 7 and 1.7 (29). However, when one uses an electrode having a
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988 14,
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1
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v
( v o l t . V)
Flgure 8. Applied voltage and resulting current curves ( V - I curves) of ISE membranes containing lasalocid and monensin.
1 2 L(x!fj Amount of ca:ion cnioiide in the aqueous p h s e beio-e ex!ract;on (TO!:
Figure 7. Degree of extraction of barium ion from the aqueous phase (pH 7, 0.1 M Tris/HCI) into the ISE membrane phase (DBS) with lasalocid and monensin.
structure satisfying this condition (inner filling solution, pH 1.7 (0.1 M NaCl); see Experimental Section), the electrode response for Na+ is almost the same as when the ISE has an inner filling solution of pH 7 . Thus, the phenomenon is not a direct influence on the unusual response of the monensin electrode. The unusual response cannot be explained by assuming a peculiar absorption of monensin at the lipophilic ISE membrane-sample solution interface, because the monensin electrode gave just twice the Nernstian linear response slope for Ba2+. Very similar behavior was also observed for ISEs based on three other natural carboxylic polyethers: ionomycin (for Ca2+),nigericin (for Ba2+),and salinomycin (for Ba2+). The degree of extraction of Ba2+from the aqueous phase of pH 7 into the lipophilic ISE membrane phase has been examined with the two ligands. As shown in Figure 7a, lasalocid apparently forms a 2:l ligand-Ba2+ complex and extracts over 95% of the Ba2+. However, as shown in Figure 7b, monensin does not show apparent extraction ability for Ba2+at the aqueous sample of pH 7 . However, an electrode using monensin actually responded to Ba2+in a sample that was pH 7 (see Figure 6). As shown by the dashed lines in Figure 7b, when the sample pH is increased to 9.5, monensin slightly extracts Ba2+. In addition, the order of cation extraction ability by monensin is Mg2+over Ba2+(Mg2+> Ca2+ > Sr2+> Ba2+),which is the reverse order of the ion selectivity of the ISE using this ligand (Ba2+> Sr2+> Ca2+> Mg2+;see Figure 5 ) . Cation complex formation of monensin was also examined by measuring the melting points of monensin free acid and magnesium and barium chloride salts, which were 102-105 (ref 11: 103-105 "C), 164-176, and 99-103 OC, respectively. These results were also confirmed with IR spectrophotometry. The IR spectrum (KBr) for the barium salt was almost the same as that for the free acid.
The applied voltage and resulting current curves (V-I curves) have been measured with two electrode membranes containing lasalocid or monensin. In this case, the same or 1 X M) was set in each concentration of Ba2+(1X compartment for the ion permeability measurement (see Experimental Section). As shown in Figure 8, a linear relation was obtained for the membrane containing lasalocid. This indicated that lasalocid easily extracted Ba2+at one side of the ISE membrane and released the cation smoothly at the other side of the membrane in addition to the smooth diffusion of the extracted ion-ligand complex. On the other hand, the slope of the V-I curve for Ba2+with a membrane containing monensin is very small compared witli that of lasalocid, which suggests that monensin does not easily form a complex with Ba2+. Ion permeability was measured with two electrode membranes containing lasalocid and monensin. Though this method dealt with the nonequilibrium state of ions a t the electrode membrane-sample solution interface, which is far from equilibrium at zero current, it gave helpful information on the charged complex species in the electrode membrane. Figure 9 shows the relationship between ion permeability of divalent cations (transport selectivity ratio, K~,jt')by an electronic potential gradient (30 V, 24 h) and ion response selectivity of the electrodes (potentiometric selectivity coefficient, Pat) using lasalocid and monensin. In these meaM Mg2+,Ca2+, surements, four mixed divalent cations (1 X Sr2+,and Ba2+)were introduced into one compartment, while ion permeation into the other compartment was monitored (see Experimental Section). The results indicated that the order of ion-permeating selectivities (Ba2+> Sr2+> Ca2+> Mg2+)is compatible for both ligands. The order is the same as that for the potentiometric selectivity of the ISE using these ligands (see Figure 9). (In the ion permeability measurement (30 V, 24 h) using a blank membrane not containing a ligand, the permeating amount of divalent cations (Mg2+,Ca2+,Sr2+, Ba2+)was not detected (510-7 M)). In addition, the transport of Ba2+through the ISE membrane incorporating monensin was examined. In this case, 1 X M Ba(N03)2 and deionized water were placed in each compartment for the ion permeability measurement. Monensin allowed the transfer of 1.59 (f0.03) X mol of Ba2+with the application of 1.53 (f0.005) X faraday (means of three runs). However, the amount of NO3- transferred was only 1.20 (A0.05) X mol, which was about one-tenth in comparison with the cation transferred simultaneously. In order to examine the importance of the counterion (anion), the electrode response curves for Ba2+ (ranging from 1 X to 1 X M) with four different anions (CH,COO-, C1-, NO3-, and SCN-) were also
ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
I
I
Lasalocid
Monensin
ISE membrane
Sample solution (Aqueous phase)
OBS PVC
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1719
( Lipophilic phase)
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Ca2*
1
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Figure 10. Illustration of ion-ligand complexation at the ISE membrane-sample solution interface.
Flgure 9. Comparison of divalent cation transport selectivity ( K b { ) of ISE membranes containing lasalocid and monensin by an electronic of ISEs based gradient (30 V, 24 h) and response selectivity (Kh,.ip”t) on lasalocid and monensin.
examined. However, differences were not observed in their responses even though SCN-, as a relatively high lipophilic anion, was used as a conterion for Ba2+. These facts indicated that the barium ion transfers across the membrane containing monensin as a monovalent ion species. However, the counteranion is not thought to contribute to the charge conduction (ion conduction by the ion itself or the charged ion-ligand complex) through the membrane. To summarize the results from a series of measurements on two typical carboxylic polyethers, lasalocid and monensin, the steps of ion complex formation for these ionophores at the ISE membrane-sample solution interface are shown in Figure 10. In the case of the lasalocid electrode, the response mechanism is basically similar to the cation exchange by a lipophilic negatively charged ion, though the ion selectivity is determined by the steric structure of the ligand. The complexation and extraction of Ba2+by lasalocid is fast and smooth. In this case, the generation of emf is based on the divalent cation itself as a charge conductor in the ISE membrane. On the other hand, monensin forms a weak complex with Ba2+ as a 1:l ligand-cation complex form. This Ba2+ complex has a monovalent positive charge ((Ba2+L-)+;L is ligand) and contributes as a charge conductor through the ISE membrane. (The assay of a 1:l Ba2+-monensin complex was not possible from the Ba2+extraction measurement of monensin. However, the ion permeability measurement on the membrane containing monensin proved that the barium ion transfers across the membrane as a monovalent ion, and in this case existence of a 1:l Ba2+-monensin complex in the membrane is presumed from the fact that the transfer of anion across the membrane is small (it was approximately one-tenth of the amount of Ba2+transfer when the anion was NO,-).) Monensin also forms a 2:l ligand-divalent cation complex with Mg2+(see Figure 7 ) . (The formation of a 2:l monensin-Mg2+
complex is presumed from the Mg2+extraction measurement (pH 9.5) with monensin. The presence of this complex formation is also supported by the IR spectrum and melting point data.) However, the Mg2+ complex is electrically neutral (Ba2+2L-),and its dissociation is small in the lipophilic electrode membrane, so that the complex does not contribute as a charge conductor in the membrane. Furthermore, the monensin electrode also responds to a monovalent cation (e.g., Na+). In this case, the emf is produced by a positively charged complex ((Na+L)+)formed with neutral monensin and the monovalent cation, but not with a neutral complex ((Na+L-)) formed by the negatively charged monensin and the cation illustrated in Figure 10. More generally speaking, there are two cases, one in which the ion complex formation or ion extractability agrees and one in which it does not agree with the potentiometric ion selectivity. The former case applies as long as the ion extracted by the simple ion-exchange reaction or extracted ion-neutral ligand complex is the major species for charge conduction in the electrode membrane. The latter case, as shown in Figure 10, is for a monensin electrode; divalent cations (e.g., Ba2+)form weak complexes with the negatively charged polyethers in such a way that a neutral polyether (e.g., valinomycin (30)) form complexes with the monovalent cations. In this case, neutral complexes (e.g., Mg2+-monensin complex) do not contribute to the charge conduction across the membrane. These summaries for lasalocid and monensin electrodes can lead to the general conclusions listed in Figure 11. Though the ISE response is, of course, based on the Donnan equilibrium for ions at the ISE membrane-sample solution interface, the important point for generation of an emf that determines the potentiometric response is how the ion is extracted into the membrane or forms the charged complex at the interface. Each case in Figure 11,of course, requires fast diffusion. of ions into the ISE membrane as well as a fast release to the aqueous phase on the other side of the membrane for “smooth charge conduction. Measurements of the electric circuit revealed very small currents (ca., 10-16-10-13 A). If the electrode responds to a monovalent cation, there
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
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