Long-Lived Potassium Ion Selective Polymer Membrane Electrode Oliver H. LeBlanc, Jr." and W. 1.Grubb General Electric Research and Development Center, Schenectady, N.Y. 1230 1
Long-lived potasslum-selectlve electrodes were constructed from membranes incorporatlngpotassium vallnomycin tetraphenylborate salt In a polymeric matrix. The polymer was a block copolymer of poly( bisphenol-A carbonate) and poly(dimethylsiloxane) with sufflclent cyanoethyl substitution in the latter to provlde a dlelectrlc constant of 5.2. Nearly Ideal Nernstian response to K+ from lov5 to lo-' N was observed over more than 3 years of exposure to neutral electrolytes.
Following the discovery by Mueller and Rudin (1)that the macrocyclic antibiotic Valinomycin induces specific K+ ion conductance through lipid bilayer membranes, Stefanac and Simon (2),and many others since, employed Valinomycin as a neutral carrier to construct practical K+ -ion-selective electrodes; see the review by Buck ( 3 ) . In 1970 ( 4 ) , electrodes incorporating Valinomycin as the K + ion carrier were prepared in a special elastomeric polysiloxane poly(bispheno1-A carbonate) block copolymer matrix ( 5 )prepared by the late Johannes F. Klebe of our laboratories. The use of this copolymer membrane in pH-sensing electrodes for in vivo biomedical applications has been described (6). Such block copolymers containing about 50% of each constituent polymer are actually heterogeneous, two-phase systems, the poly(si1oxane) blocks comprising a continuous, amorphous phase through which molecular transport occurs rapidly, as in all silicones, while the poly(carbonate) blocks form a discontinuous, crystalline phase that cross-links the structure (7,8). Because such cross-linking is destroyed by poly(carbonate) solvents, such as methylene chloride, these elastomers are solvent castable, a useful property. If such copolymers contain only dimethylsiloxane moieties in the poly(si1oxane) blocks, they do not perform well in ionselective electrodes as matrices for ion-specific carriers. Presumably, their dielectric constants (2.5-3.0) are too low to permit significant ion unpairing or electrical charge injection at the aqueous/membrane interfaces (9).Adding ionic carriers of various sorts to them yielded membranes of very high electrical resistances and erratic transmembrane potentials between aqueous solutions containing the transportable ions. Accordingly, Klebe synthesized modified copolymer materials in which the poly(si1oxane) blocks were a random sequence of dimethylsiloxane and cyanoethylmethylsiloxanes.Sufficient cyanoethyl groups to yield a dielectric constant between 4 and 13 gave copolymers which performed well in ion-selective electrodes. Klebe also achieved improved hydrolytic stability of these copolymers by employing a carbamate linkage, rather than the more customary aryl-oxy-silicon linkage between the poly(si1oxane) and poly(carbonate) blocks. Synthetic procedures are described in detail in Ref. 4.
K+ ion selective electrodes formed by simply incorporating neutral Valinomycin (Calbiochem, LaJolla, Calif.) into films of the above polymers showed some K+ response, but resistances were so high (?lo9 Q ) that measurements of transmembrane potentials were exceedingly difficult. Using the salt formed between the K+ : Valinomycin complex cation and the 1658
*
tetraphenylborate anion yielded much lower resistance membranes with good K+-response characteristics.
EXPERIMENTAL The electrodes were constructed using the poly(si1oxane)-poly(bisphenol-A carbonate) block copolymer preparation specifically designated as example number 2 in Ref. 4. This contained 52% siloxane, with 2.2 dimethylsiloxanes per cyanoethylmethylsiloxane with a static dielectric constant of 5.2, and an intrinsic viscosity of 0.3 dl/g in chloroform at 25 O C . The carrier salt was prepared from potassium tetraphenylborate precipitated from aqueous solutions of KC1 and sodium tetraphenylborate. After careful washing and drying 3.3 mg (9.2 mmol) of this salt plus 12.2 mg (10.9 mmol) of Valinomycin were dissolved in 8 ml of methylene chloride in about 3 h. (Potassium tetraphenylborate is insoluble in methylene chloride, but the complex salt is soluble.) The complex salt formed more rapidly in the mutual solvent acetone, which was removed in vacuo before dissolving the salt in methylene chloride. The complex salt in methylene chloride was added to methylene chloride solution of the block copolymer to obtain about 3%solids in solution consisting of 3.0 parts salt to 100 parts copolymer by weight. After filtering through a glass frit to remove any particles, the solvent was partially evaporated until there was about 7% solids solution. This solution was poured onto a glass plate, allowing it to spread freely, and allowed to dry overnight with a Petri dish cover to slow solvent evaporation. The resulting film, 60-90 p m in thickness, was peeled from the plate. It was tough, rubbery, colorless, and slightly cloudy, probably indicating that the complex salt was present slightly in excess of its solubility in the copolymer. Small circular disks 6 mm in diameter, punched from this film with a stainless steel punch, were incorporated in two electrodes whose structure is shown in Figure 1.The electrode body was a 10-cm length of Pyrex tubing, 5.0-mm 0.d. and 3.3-mm id., with ends ground flat. One end of the tube was coated with a small quantity of silicone rubber cement (RTV 108, General Electric, Waterford, N.Y.), the tube mounted in a micromanipulator, and the cement-coated end lowered into concentric contact with the slightly larger 6.0-mm diameter disk of the K+ -sensitive film. The silicone rubber cement was allowed to cure overnight, forming the first of two seals between glass and film. Making of the second seal was simplified by use of a shaped sleeve. This had been preformed from heat-shrinkable polyolefin tubing (Flexite PO-135, expanded i.d. 6 . 3 mm, recovered 3.2 mm, L. Frank Markel & Sons, Norristown, Pa.) by shrinking it onto a Teflon polymer rod machined to two diameters of 5.0 and 6.0 mm with a shoulder between. The Pyrex glass tube was held vertically with the membrane at the top, and the shaped sleeve positioned about half-way up the tube with its larger diameter facing upward. The annular space between the sleeve and the tube was filled by injecting more of the RTV-108 silicone from a syringe. The sleeve was then slid upwards until it touched the membrane. The outer edges of the membrane were gently pressed down to make good contact with the silicone rubber cement, which then was allowed to cure overnight. Thus, two independent seals were formed sequentially, providing better assurance against pinhole leaks. The silicone rubber cement used to make these seals adheres extremely well to poly(si1oxane) materials, and also to clean glass surfaces. An internal reference element consisting of a chloride silver wire was sealed into a sized piece of the heat-shrinkable polyolefin tubing. A vent hole was provided in this to facilitate final assembly of the electrode, after it was filled with a reference electrolyte solution, by simply sliding the snugly-fitting polyolefin tubing onto the open end of the Pyrex glass tube. The two electrodes constructed on November 11,1970, have had slightly different histories. Electrode A was stored wet a t room temperature, filled with and immersed in a solution of either 4 mM KC1
ANALYTICAL CHEMISTRY, VOL. 48, NO. 12, OCTOBER 1976
Table I. Resistance as Function of Time Elapsed Since Beginning of Continuous Storage in 0.15 M NaCl, 0.01 M KCI Electrode A
iNTERNAL ELECTROLYTE
Kf‘
Figure 1. Construction of the
10-6
Flgure 2.
PERMEABLE MEMBRANE
Electrode B
Date tested
Time, days
Resistance,
Time,
Resistance,
MO
days
MO
11/25/70 12/10/70 1/20/71 4/22/71 5/11/72 11/13/72 5/17/72 11/19/73 2110176
14 29 70 103 547 733 917 1104 1917
22 20 27 384 570 754 941 1754
22 17
21 35 35 30
35 29
18 18
6
electrodes.
IO-^ I,+ 163 10-2 EXTERNAL KCI CONCENTRATION (MOLES/f)
16’
Electrode potential as a function of K+ ion concentration
Internal electrolyte: 140 mM NaCl plus 4 mM KCI. Reference electrode: saturated KCI plus 140 mM NaCI. The K+ calomel. External solution: (0)KCI alone; (0) ion concentration in Hyland serum was calculated by Interpolation from the potentials measured in it, in 1 mM KCI plus 140 mM NaCI, and in 10 mM KCI plus 140 mM NaCi. Value found: 4.1 mN; Hyland assay: 4.3 mN and 140 mM NaCl or 10 mM KCl and 150 mM NaCI. Electrode B was stored dry for approximately five months and in the same manner as Electrode A since then. From time to time, the electrodes were briefly removed from storage for testing. The internal electrolyte solutions were renewed if necessary. Electrode potentials were conventionally measured against a reference electrode using high impedance electrometers or pH meters. Electrode resistances were measured by a variable frequency audio bridge with extrapolation to zero frequency or by measuring the electrode potentials as a function of imposed shunt load resistances. All data were obtained a t room temperatures, 23 k 2 “C.
RESULTS AND DISCUSSION Static potential response of electrode A to variations in external K+ ion concentrations is shown in Figure 2; the characteristics of electrode B were indistinguishable. The same response, repeatedly observed for both electrodes for a period of three years was nearly linear from to lom5N [K+] and not exactly linear at higher concentrations because of the variation of activity coefficients. For example, the response was 56 f 2 mV when the external solution was changed from 1 mM KC1 to 10 mM KC1 a t 23 OC. This is Nernstian behavior considering the activity coefficient differences between these two solutions. T o determine if the electrodes are useful in the determination of [K+] in whole blood plasma, separated plasma, or serum, in which Na+ is present (a140 mN Na+ vs. =4 mN K+), potentials were determined in the presence and absence of 140-150 mM NaCl. As Figure 2 shows, the addition of NaCl generally decreased the electrode potential, as expected for activity coefficient effects, rather than increasing them, as would be expected for Na+ ion interference. A test of reconstituted human blood serum (Hyland Clinical Control Serum, Hyland Division, of Travenol Laboratories, Los Angeles,
Calif.) is shown in Figure 2. The agreement with the Hyland assay shows that these electrodes can be used to determine K+ ions in plasma or serum. For three years after they were constructed, the response time of both electrodes was such that steady potentials were obtained in less than 1min after changing the external solutions. This was probably limited by the time of mixing. Electrical resistance was monitored as a sensitive measure of the stability of ion selective electrodes (probably more sensitive than the electrode potential itself). The resistance will undergo changes due to deteriorations in either the “liquid membrane” matrices or the carrier systems, or due to the development of pinhole leaks, long before any of these phenomena lead to detectable changes in electrode potential characteristics. The resistance data obtained on the present electrodes are listed in Table I. They demonstrate that the electrodes were surprisingly stable for 3 years, with detectable changes occurring a t 5 years. At 5 years of age, both electrodes had deteriorated, as judged by all criteria for evaluating their performance. The resistances had noticeably decreased. The potential response to changes in external [K+]was now sluggish, steady potentials being reached only after as long as 5-10 min. The static response was now low; the potential now changed only +50 mV for electrode A and +54 mV for electrode B when the external solution was changed from 1mM KCl to 10 mM KC1. The reason for this ultimate deterioration in performance is not certain, but all the observations would be consistent either with the development of Na+ (or C1-) interference in the membrane proper or with the development of pinhole leaks around or in the membrane. The latter is considered more probable. CONCLUSION The long life of these electrodes is remarkable. Clearly, the poly(si1oxane)-poly(bispheno1-A carbonate) block copolymer synthesized by Klebe is quite stable in contact with neutral aqueous solutions. The potassium Valinomycin tetraphenylborate complex salt remained active in the membrane during a 3-year period without dissolving into the aqueous solutions or hydrolytically decomposing. Similar long life behavior has also now been observed with ion-selective electrodes constructed using plasticized poly(viny1 chloride) as the “liquid membrane” material (10-1 7), which suggests that the phenomena may be general to electrodes constructed with noncrystalline polymer matrices. The electrodes of the present paper have utility in biomedical applications such as blood serum K+ determinations. The deterioration in transmembrane potential and response time properties and the accompanying decrease in electrical resistances at 5 years’ life may indicate the onset of electrolytic shorting paths through or around the membranes. The use of
ANALYTICAL CHEMISTRY, VOL. 48, NO. 12, OCTOBER 1976
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thicker membranes or even better adhesive or sealing procedure might yield electrodes with longer life. Finally, the elastomeric copolymer applied in this work may be useful as a host matrix for other ion-selective carriers, providing improved stability and lifetime.
LITERATURE CITED (1) P. Mueller and D.0. Rudln, Biochem. Biophys. Res. Commun., 26, 398 (1967). (2) J. Stefanac and W. Simon, Microcbem. J., 12, 125 (1967). (3) R. P. Buck, Anal. Chem., 46, 28R (1974). (4) J. F. Brown, 0. H. LeBlanc, and W. T. Giubb, US. Patent 3,767,533(October 23, 1973). (5) H. A. Vaughan, J. Polymer Sci., Part 6, 7, 569 (1969). (6) 0. H. LeBlanc, J. F. Brown, J. F. Klebe, L. W. Niedrach, G. M. J. Slusarczuk, and W. H. Stoddard, J. Appl. Physiol., 40,644 (1976).
(7) R. P. Kambour, J. PolymerSci., Part6, 7, 573(1969). (8) D. G. LeGrande, J. Polymer Sci., Part 6, 7, 579 (1969). (9) R. M. Fuoss and C. A. Kraus, J. Am. Chem. Soc., 57, 1 (1935). (IO) G. J. Moody, R. B. Oke, and J. D. R. Thomas, Ana/yst(London),95, 910 (1970). (1 1) R. W. Cattrall and H. Freiser, Anal. Chem., 43, 1905 (1971). (12) J. E. W. Davies, G. J. Moody, and J. D. R. Thomas, Ana/y.st(London),97, 87 (1972). (13) H. James, G. D. Carmack, and H. Freiser, Anal. Chem.. 44, 856 (1972). (14) G. H. Griffiths, G. J. Moody, and J. D. R. Thomas, Ana/ysf (London),97,420 (1972). (15) J. Ruzicka, E. H. Hansen, and J. C. Tjell, Anal. Chim. Acta, 67, 155 (1973). (16) U. Fiedler and J. Ruzicka, Anal. Chim. Acta, 67, 179 (1973). (17) G. D.Carmack and H. Freiser, Anal. Chem., 47, 2249 (1975).
RECEIVEDfor review April 19, 1976. Accepted June 24, 1976.
Determination of Subnanogram Amounts of Fluoride with the Fluoride Electrode Alan S. Hallsworth,* John A. Weatherell, and Dan Deutsch Department of Oral Biology, Dental School, University of Leeds, Leeds LS 1 3EU. England.
An extremely sensltive method of determlnlng fluoride wlth the Orion electrode is descrlbed havlng an absolute detectlon limit of 10 pg (IO-" g) of F-. One-mlcrollter volumes of sample solution are confined as thin layers between the fluoride-Ion sensing element of a standard Orion electrode and the flat sleeve-junctlon of a calomel reference electrode mounted g) of Fimmediately below it. One hundred picograms ( have thus been determined wlth a precislon (relative standard deviatlon) of 5%: the precision improvlng to 2.8% at the 1-ng level. Sample preparation procedures and a fluoride mlcrodiffusion technique of great sensltlvlty and range (
