Polymer-Supported pH Sensors Based on Hydrophobically Bound

These sensor systems have a usable sensitivity over the 2−8 pH range, but are most usable in the 2−6 pH range. Response ... Analytical Chemistry 0...
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Anal. Chem. 1998, 70, 265-270

Polymer-Supported pH Sensors Based on Hydrophobically Bound Luminescent Ruthenium(II) Complexes Jason M. Price,† Wenying Xu,† J. N. Demas,*,† and B. A. DeGraff*,‡

Department of Chemistry, University of Virginia, Charlottesville, Virginia 22901, and Department of Chemistry, James Madison University, Harrisonburg, Virginia 22807

A new luminescence intensity-based sensor-polymer support system is described. The pH-sensitive element is [Ru(phen)2[phen(OH)2]]2+ or [Ru(Ph2phen)2[phen(OH)2]]2+ (phen ) 1,10-phenanthroline, Ph2phen ) 4,7diphenyl-1,10-phenanthroline, and phen(OH)2 ) 4,7dihydroxy-1,10-phenanthroline) immobilized on a polymer built from a four-function cyclic methyl siloxane ring polymer having (SiRCH3O)4 units with poly(ethylene oxide) cross-linkers. Hydrophobic binding of the sensor to the polymer is used to avoid the need for covalent chemistry. These sensor systems have a usable sensitivity over the 2-8 pH range, but are most usable in the 2-6 pH range. Response times for thick films (250 µM) are rapid (several minutes). Recently, there has been much interest in the design and application of remote fiber-optic sensors.1 Luminescence is widely used because of its inherent sensitivity. Such sensors would have great medical and industrial importance in blood gas measurement, in groundwater analysis, in wastewater monitoring, and as pressure sensors in wind tunnels. The measurement of oxygen, pH, and pCO2 is especially important in biological and industrial settings. Highly luminescent Ru(II), Os(II), and Re(I) metal complexes are a promising class of sensor materials. These complexes alter their emissive properties, specifically luminescence intensities and lifetimes, depending on the local concentration of an analyte. For instance, the quenching of their emission intensity or lifetime by molecular oxygen provides a sensitive and reliable oxygen sensor suitable for remote fiber-optic sensing.2-6 Another important area is pH sensing. Many common pH indicators are based on absorption, but for the high sensitivity †

University of Virginia. James Madison University. (1) Topics in Fluorescence Spectroscopy. Probe Design and Chemical Sensing; Lakowicz, J., Ed.; Plenum Press: New York, 1994; Vol. 4. (2) Klimant, I.; Belser, P.; Wolfbeis, O. S. Talanta 1994, 41, 985-91. (3) Carraway, E. R.; Demas, J. N.; DeGraff, B. A.; Bacon, J. R. Anal. Chem. 1991, 63, 337-342. (4) Xu, W.; McDonough, R. C., III; Langsdorf, B.; Demas, J. N.; DeGraff, B. A. Anal. Chem. 1994, 66, 4133-41. (5) Xu, W.; Kneas, K. A.; Demas, J. N.; DeGraff, B. A. Anal. Chem. 1996, 68, 2605-9. (6) Bambot, S. B.; Rao, G.; Romauld M. M.; Carter, G. M.; Sipior, J.; Terpetchnig, E.; Lakowicz, J. R. Biosens. Bioelectron. 1995, 10, 643-52. ‡

S0003-2700(97)00784-1 CCC: $15.00 Published on Web 01/15/1998

© 1998 American Chemical Society

required on small fiber-optic probes or evanescent wave systems, much more sensitivity is preferred. Currently, much pH sensor research has focused on organic fluorophores utilizing fluorescence decay time7-9 or emission intensities.10 Potentially, metal complexes have attractive characteristics for pH sensors. While oxygen sensors built on platinum metal complexes are now becoming a mature technology, pH sensors based on such systems are still in their infancy. For direct sensing of pH by the complex, the simplest strategy is to include a pH-sensitive functionality on the metal complex, which alters the emissive properties with pH changes. There are a number of luminescent inorganic complexes that show pH-sensitive luminescence characteristics. These include Ru(bpy)2L2 complexes where L is an R-diimine ligand with pyridine, carboxylic acid, amine, or phenol substituents. This may be coupled to,11-13 or electronically separated from, the ligand by a methylene group.14,15 For the amines and phenols, the degree of protonation alters the luminescence quenching due to a photoinduced electron transfer. However, there have been no reports to date of attempts to incorporate such systems into practical polymeric supports. We report on a system consisting of [Ru(phen)2[(phen)(OH)2]]2+ or [Ru(Ph2phen)2[(phen)(OH)2]]2+ (phen ) 1,10phenanthroline, Ph2phen ) 4,7-diphenyl-1,10-phenanthroline, and phen(OH)2 ) 4,7-dihydroxy-1,10-phenanthroline) (Figure 1A) immobilized in a D4TMI-PEG-Jeffamine polymer network (Figure 1B). In solution, this class of Ru(II) complexes shows pronounced emission pH sensitivity over the 0-6 pH range.16 We will show that these complexes are a new class of pH sensors suitable for (7) Draxler, S.; Lippitsch, M. E. Anal. Chem. 1996, 68, 753-7. (8) Draxler, S.; Lippitsch, M. E.; Leiner, M. J.-P. Sens. Actuators B 1993, 11, 421-4. (9) Bambot, S. B.; Sipior, J.; Lakowicz, J. R.; Rao, G. Sens. Actuators B 1994, 22, 181-8. (10) Parker, J. W.; Laksin, O.; Yu, C.; Lau, M.-L.; Klima, S.; Fisher, R.; Scott, I.; Atwater, B. W. Anal. Chem. 1993, 65, 2329-34. (11) Kalyanasundaram, K.; Nazeeruddin, M. K.; Graetzel, M.; Viscardi, G.; Savarino, P.; Barni, E. Inorg. Chim. Acta 1992, 198-200, 831-9. (12) Zheng, G. Y.; Wang, Y.; Rillema, D. P. Inorg. Chem. 1996, 35, 7118-23. (13) Thompson, A. M. W. C.; Smailes, M. C. C.; Jeffery, J. C.; Ward, M. D. J. Chem. Soc., Dalton Trans. 1997, 5, 737-43. (14) Grigg, R.; Norbert, W. D. J. A. J. Chem. Soc., Chem. Commun. 1992, 18, 1300-2. (15) Grigg, R.; Holmes, J. M.; Jones, S. K.; Norbert, W. D. J. A. J. Chem. Soc., Chem. Commun. 1994, 2, 185-7. (16) Giordano, P. J.; Bock R. C.; Wrighton, M. S. J. Am. Chem. Soc. 1978, 100, 6960-5.

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the changing pH in a surrounding matrix. As cross-linked polymers, they are insoluble. Finally, the siloxane ring structure, coupled with the phenyl rings, potentially provides a very hydrophobic pocket for anchoring the hydrophobic region on our metal complexes while providing a water-swellable domain for the pH-sensitive functionality. Our results show that this prediction works and provides functional pH sensors.

Figure 1. (A) Structures of [Ru(phen)2[phen)(OH)2]]2+ and [Ru(Ph2phen)2[phen)(OH)2]]2+ pH-sensitive metal complexes. (B) Structure of components used to build network polymers.

Figure 2. Acid/base reactions relevant to a pH-sensitive molecule with two acidic protons.

remote sensing. Figure 2 shows the relevant acid dissociation chemistry. Luminescence sensors report pH changes in lifetime or spectral changes including brightness and/or color. The phen(HO)2 ligand has two acidic protons. Our system is a diprotic acid, so there are two excited-state pK*’s. If the exchange rates between different excited species is rapid compared to the lifetimes, then only a single-exponential decay with a lifetime weighted by the relative amounts and lifetimes of the different components will be observed. For a slow exchange, there will be multiple lifetimes. A practical system would require a polymer support that allowed free access of the analyte to the sensor molecules. The D4TMI-PEG-Jeffamine polymer networks were selected for several reasons. First, they are very hydrophilic and swell substantially in water, which allows equilibration of embedded sensors with 266

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EXPERIMENTAL SECTION Materials. The 4,7-dihydroxy-1,10-phenanthroline was obtained from G. Frederick Smith. This was an old sample that is much purer than that currently commercially available. The Ru(phen)2Cl2 and Ru(Ph2phen)2Cl2 were either available from previous work or were synthesized by standard methods.17 The D4TMI-PEG-Jeffamine network polymers were previously synthesized.4 Synthesis of [Ru(phen)2[(phen)(OH)2]](ClO4)2. Both complexes were synthesized similarly. We describe only the synthesis of [Ru(phen)2[(phen)(OH)2]](ClO4)2. A three-necked flask was evacuated and then flushed with nitrogen to remove any oxygen. Column-purified, TLC-pure Ru(phen)2Cl2 (113 mg) and (phen)(OH)2 (91 mg) were added to the flask along with ∼44 mL of a 75% (v/v) ethanol/water. The reaction mixture was bubbled with nitrogen for 10 min and then sealed and kept under a nitrogen atmosphere for the duration of the reaction. The mixture was refluxed in an oil bath at 98 °C for ∼24 h. The volume was reduced to one-fourth its original value, and the product was precipitated by adding an excess of saturated aqueous NaClO4. The red-brown crystals were collected by vacuum filtration. Inorganic salts were removed by dissolving the crystals in acetonitrile and filtering off the insoluble salts by vacuum filtration. The acetonitrile was removed by Rotovac, and the crystals were vacuum-dried. The product was recovered in 80% yield. Absorption and emission spectra were consistent with published data.16 Only a single spot was seen on TLC. Elemental analysis for a trihydrate. Anal. Calcd for C36H30N6O13Cl2 Ru: C, 46.69; H, 3.24; N, 9.07. Found: C, 46.69; H, 3.09; N, 9.66. For the [Ru(Ph2phen)2(phen)(OH)2]Cl2, precipitation was with NaCl, careful drying of the complex, rinsing with ether, and washing out excess salts and ligand with dry acetonitrile. Elemental analysis for a trihydrate. Anal. Calcd for C60H46N6O5Cl2 Ru: C, 65.34; H, 4.20; N, 7.62. Found: C, 65.59; H, 4.32; N, 7.63. Instrumentation. Absorption spectra were recorded on an HP 8452A. Fluorescence spectra were measured on a Spex Fluorolog 2+2 spectrofluorometer. The complexes were excited at isosbestic points in the metal-to-ligand charge transfer (MLCT) transitions as a function of pH. Solution and polymer emission data were collected with either front surface or right angle detection. To measure the pH dependence, films were first equilibrated at different pH’s and then mounted between quartz slides or in a quartz cell and placed in the spectrofluorometer. A pulsed N2 laser (337 nm) decay system was used to measure room-temperature τ values, and a Tektronix TDS-540 digital oscilloscope was used for recording transients. The emissions were monitored at 640 nm. The emission was monitored at the (17) Sprintschnik, G.; Sprintschnik, H. W.; Kirsch, P. P.; Whitten, D. G. J. Am. Chem. Soc. 1977, 99, 4947-53.

maximum of the acidic form. Each decay curve was the result of 400 averages. All spectral and lifetime data were acquired at room temperature (∼22 °C) in air-saturated solutions. Measurements of pH were made on a temperature-compensated Corning Model 340 pH meter, which was calibrated frequently with commercial buffers. Sensor Film Preparation. Each polymer was loaded by swelling a film (∼200-250 µm thick) of the polymer in ∼10-15 mL of a 12 µM aqueous solution (phen complex) or 15% aqueous ethanol solution (Ph2phen complex) of the complex for a period of 24-36 h. The complexes bound very tightly to the polymer; at the end of the doping, the solution was colorless while the polymer was highly colored. Initially the polymer exhibited a peach-orange color that changed to yellow as the pH was decreased. Visually the complex was homogeneously distributed. Samples were stored wet at near-neutral pH’s and in the dark. Titrations and Equilibration Times. Phosphate buffers were used for pH 6-8 and acetate buffers were used for pH 3-5. HCl and NaOH were used for more acidic and basic regions. Solution titrations were done at 14 µM complex in 50 mM sodium chloride to control the ionic strength. Titrations were performed by monitoring the emission intensity at the maximum of the acidic form of the complex. Response times of the polymer film phen sensors to step changes in pH were obtained by suspending the J-2000 polymer in a cuvette. The emission intensity was monitored at the acidic emission maxima of 620 nm as a function of time following insertion into solutions of different pH’s. RESULTS AND DISCUSSION Solution Data. Absorption spectra for [Ru(phen)2[(phen)(OH)2]](ClO4)2 have been shown earlier.16 The peaks in the 400500 nm region are MLCT transitions arising from an electron promotion from the Ru metal-centered t2 orbital to a phenanthroline π* antibonding orbital. The more intense peaks around 250260 nm are due to ligand π-π* transitions. Isosbestic points are maintained over the pH range 0-12. Similar results were obtained for [Ru(Ph2phen)2[(phen)(OH)2]](ClO4)2, but because of the low solubility in water, measurements were made in 15% (v/v) ethanol/water. Even in this solvent mixture, the complex precipitated at pH’s less than 2, presumably because of the high chloride concentration. Figure 3 shows the emission spectra of [Ru(phen)2[(phen)(OH)2]](ClO4)2 at various pH values between 2.4 and 10.5. Not shown are the spectra below pH 2, because the intensities were so large that it would have made the high-pH data hard to visualize. These lower pH data were over a factor of 2 more intense than the highest curve shown. Similar emission results were obtained for the[Ru(Ph2phen)2[(phen)(OH)2]](ClO4)2 in 15% ethanol except that the pH range was more limited at low pH due to solubility. It is clear that as each of the complexes is deprotonated the emission intensity decreases and the wavelength red shifts. The much lower luminescence efficiency of the deprotonated form can probably be attributed to the lower excitedstate energy. The energy gap law predicts that radiationless deactivation will be much more efficient the closer the emitting state is to the ground state, and that was observed. The MLCT transition has been assigned to an electron promotion to the phen

Figure 3. Room-temperature uncorrected emission spectra of an aqueous 14.2 µM solution of [Ru(phen)2[(phen)(OH)2](ClO4)2. From top to bottom, the pH’s are 2.43, 2.80, 3.25, 3.62, 4.00, 4.34, 4.75, 5.14, 5.55, 6.51, 6.77, 8.04, and 10.5. Low-pH curves are not shown for clarity. For pH 1.55, the intensity was 123 000.

Figure 4. Relative emission intensity vs pH of [Ru(phen)2[(phen)(OH)2]](ClO4)2 in an aqueous solution (+) and J-2000 polymer supported (]) in water. All points are relative to the highest intensity point at pH -0.15. The lines have no physical significance and are provided to aid visualization.

ligand. The red shift on deprotonation is attributed to destabilization of the t2g level.16 For the polymer emission, the spectra were very similar to the solution data. Sample placement reproducibility was poorer, and the trends are more clearly seen in the solution data. Normalized titration curves are shown in Figures 4 and 5. The inflection points are near pH 2-2.5. Lifetime decay curves for air-saturated acidic and basic solutions of [Ru(phen)2[(phen)(OH)2]]2+ were also obtained. The decay curves were fit well with a single exponential that yielded lifetimes of 720 ns for the protonated form (pH 12). This establishes that the complex is free of any highly emissive [Ru(phen)3]2+ that may have contaminated the Ru(phen)2Cl2. The single-exponential decay coupled with the shifting emission spectra implies a rapid excited-state equilibrium with respect to the individual decay times of the three species. If this equilibrium were slow, one would expect a multiexponential decay with preexponential factors corresponding to the relative concentrations of the different forms. Aqueous solutions of [Ru(phen)2[(phen)(OH)2]](ClO4)2 near neutral pH are quite stable, at least in the dark. Based on the absorbance of the MLCT band at 430 nm, the complex decomposed no more than 3.5% in 24 days. [Ru(Ph2phen)2(phen)(OH)2]Cl2 was stable in solution for months and has shown a consistent response to changes in pH. Polymer Characteristics. The polymers were chosen because they have both a water-swellable hydrophilic region and a hydrophobic region that we hoped would bind the sensor molecules through hydrophobic interactions. In both polymers, J-900 and J-2000, the poly(ethylene glycol) (PEG) cross-linker provided for water swelling and transport of protons throughout the polymer. The J-2000 polymer appeared to swell to a larger volume probably because of the larger PEG chain. It seems likely that the very hydrophobic siloxane ring and substituents (the isopropyl group attached to the phenyl group) provide a hydrophobic binding pocket for the hydrophobic phen ligands. The PEG chain and urea functionalities provide a more hydrophilic environment for the sensing phenol portion of the complex. Since the phenanthroline group is far too large to fit into the siloxane ring, the binding must be due to a hydrophobic interaction between the siloxane ring/organic core and the phenanthrolines coupled with potential hydrogen-bonding interactions between the hydroxyl groups of the complex and the amine/PEG groups of the polymer. The [Ru(Ph2phen)2[(phen)(OH)2]]Cl2 results (vide infra) provide further evidence for this binding model. 268 Analytical Chemistry, Vol. 70, No. 2, January 15, 1998

Despite the ease of loading, there were slow but bothersome leaching problems for the [Ru(phen)2[(phen)(OH)2]](ClO4)2 if the polymer was kept in bulk aqueous solutions for long periods of time. A piece of J-2000 polymer stored in distilled water lost nearly all of its yellow color in less than two weeks. Over the period of several hours necessary to make a titration, sensor stability was tested by returning the pH to a previous value. No changes beyond those associated with positioning reproducibility (∼(10%) were observed. A J-900 polymer film stored in a large volume of distilled water showed noticeable leaching (∼20-30%) in several hours as judged by a decrease in the MLCT absorbance. Because of the leaching with the [Ru(phen)2[(phen)(OH)2]](ClO4)2, we reasoned that the much more hydrophobic [Ru(Ph2phen)2[(phen)(OH)2]]Cl2 complex would bind more tightly and resist leaching. The [Ru(Ph2phen)2[(phen)(OH)2]]Cl2 was absolutely stable in the J-2000 polymer, which supports our hydrophobic binding model. No leaching was observed over a period of two months in 5% ethanol. In water, of course, no leaching would be observed. This result is not surprising given the insolubility of the complex in pure water coupled with the extreme hydrophobicity of the binding site. This shows that reliable sensors are amenable to rational design. Precautions were taken to store the loaded polymers away from light, although there does not seem to be any short-term photobleaching problems. Using 415 nm irradiation in the fluorometer with wide excitation slits, the emission intensity of the protonated [Ru(phen)2[(phen)(OH)2]](ClO4)2 (pH 100 ns) in the easily excitable blue region.14,15 These results encourage continued research in this area with related systems. Future work will be directed toward other pH(18) Demas, J. N.; Degraff, B. A. Sens. Actuators B 1993, 11, 35-41.

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sensitive substituents, with an operating range at physiological pH’s, and metal-sensitive systems. Further work is in progress.

22225). Finally, J.M.P. thanks Kristi Kneas and Yvette Clark for technical assistance.

ACKNOWLEDGMENT

Received for review July 22, 1997. Accepted October 22, 1997.X

We thank the National Science Foundation (Grants CHE 9118034 and CHE-94-19074) for partial support. J.M.P. acknowledges summer support under an NSF REU program (Grant CHE-93-

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AC9707848 X

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