Anal. Chem. 1996, 68, 2605-2609
Oxygen Sensors Based on Luminescence Quenching of Metal Complexes: Osmium Complexes Suitable for Laser Diode Excitation Wenying Xu,† Kristi A. Kneas,† J. N. Demas,*,† and B. A. DeGraff*,‡
Departments of Chemistry, University of Virginia, Charlottesville, Virginia 22901, and James Madison University, Harrisonburg, Virginia 22807
Oxygen quenching of a series of Os(II) complexes with r-diimine ligands has been studied in a predominantly poly(dimethylsiloxane) (PDMS) polymer and in Gp-163 (an acrylate modified PDMS). Unlike previous Ru(II) complexes used as oxygen sensors, the Os complexes can be excited by readily available, high-intensity, low-cost, red diode lasers at 635, 650, and 670 nm. Variations in the polymer properties have been made in order to delineate the structural features important for satisfactory use of supports for oxygen sensors. A key factor is matching the hydrophobicity of the sensor and support for optimal compatibility and minimizing the size of low oxygen diffusion domains. Because of their numerous applications,1-14 luminescencebased probes and sensors and their design continue to be of considerable interest. Sensors based on luminescent materials as the sensor element are of particular interest because of their great potential for sensitivity and specificity. In particular, Ru(II) complexes with R-diimine ligands (e.g., 2,2′-bipyridine, 1,10phenanthroline, and their substituted analogues) show great promise as quenchometric oxygen sensors because of their long lifetimes, ease of excitation, and high luminescence quantum yields. However, a glaring deficiency of these systems has been the need for excitation in the blue region of the spectrum. While blue LEDs have been successfully used, they have had limited intensity, although this is changing. We describe here studies †
University of Virginia. James Madison University. (1) Preininger, C.; Klimant, I.; Wolfbeis, Otto S. Anal. Chem. 1994, 66, 1841. (2) Lieberman, R. A., Wlodarcyzk, M. T., Eds. Chemical, Biochemical, and Environmental Fiber Sensors; Proceedings of SPIE; SPIE: Bellingham, WA, 1989; Vol. 1172. (3) Demas, J. N.; DeGraff, B. A. Anal. Chem. 1991, 63, 829A-837A. (4) Berndt, C. W.; Lakowicz, J. R. Anal. Biochem. 1992, 201, 319. (5) Lakowicz, J. R.; Malak, H.; Gryczynski, I.; Castellano, F. N.; Meyer, G. J. Biospectroscopy 1995, 1, 163-168. (6) Terpetschnig, E.; Szmacinski, H.; Lakowicz, J. R. Anal. Biochem. 1995, 227, 140-147. (7) Terpetschnig, E.; Szmacinski, H.; Malak, H.; Lakowicz, J. R. Biophys. J. 1995, 68, 342-350. (8) Li, X. M.; Ruan, F. C.; Wong, K. Y. Analyst 1993, 118, 289. (9) Kaneko, M.; Iwahata, S.; Asakura, T. Photochem. Photobiol. 1992, 55, 505509. (10) Bacon, J. R.; Demas, J. N. Anal. Chem. 1987, 59, 2780-2785. (11) Xu, W.; McDonough, R. C., III; Langsdorf, B.; Demas, J. N.; DeGraff, B. A. Anal. Chem. 1994, 66, 4133-4141. (12) MacCraith, B. D.; McDonagh, C. M.; O’Keefe, G.; Keyes, E. T.; Vos, J. G.; O’Kelly, B.; McGilp, J. F. Analyst 1993, 118, 385. (13) Hartmann, P.; Leiner, M. J. P. Anal. Chem. 1995, 67, 88-93. (14) Li, L.; Walt, D. R. Anal. Chem. 1995, 67, 3746-3752. ‡
S0003-2700(96)00083-2 CCC: $12.00
© 1996 American Chemical Society
on analogous Os(II) complexes that have intense red absorptions15-18 and that can be excited with low-cost, highintensity red diode lasers, which are currently widely used as laser pointers and have powers in the multi-milliwatt range in off-theshelf items. In solution, these osmium complexes have demonstrated relatively long excited state lifetimes and a reasonable level of oxygen quenching.19 A recent study has shown that [Os(terp)2]2+ (terp ) 2,2′,2′′-terpyridine) can function as a red excitable oxygen sensor, albeit with much poorer sensitivity than Ru(II) sensors.20 As an added benefit, the osmium complexes are probably more photochemically robust than their Ru(II) analogues because of the larger energy gap between the emitting state and the photochemically destructive upper dd states.21-23 These new sensor complexes, while not as sensitive to oxygen quenching as the ruthenium complexes, possess large enough quenching responses and long enough lifetimes to be useful as sensors. Consistent with our earlier results, sensing properties have complex dependencies on polymer structure, but the quenching properties do satisfy some of our earlier guidelines.24,25 Our results expand substantially on the number of oxygen-sensitive Os(II) complexes and our understanding of the sensor-probe interactions. Further, one of our systems has over an order of magnitude larger response than the system reported earlier. As has been shown20 and discussed,26,27 red-absorbing oxygen sensors have another application. They can be used in vivo as (15) Dwyer, F. P.; Gibson, N. A.; Gyarfas, E. C. J. Proc. R. Soc. New South Wales 1950, 82, 68-70. (16) Buckingham, D. A.; Dwyer, F. P.; Goodwin, H. A.; Sargeson, A. M. Aust. J. Chem. 1964, 17, 325-336. (17) Bryant, G. M.; Fergusson, J. E.; Powell, H. K. J. Aust. J. Chem. 1971, 24, 257-273. (18) Sauvage, J.-P.; Collin, J.-P.; Chambron, J.-C.; Guillerez, S.; Coudret, C.; Balzani, V.; Barigelletti, F.; De Cola, L.; Flamigni, L. Chem. Rev. 1994, 94, 993-1019. (19) Demas, J. N.; Harris, E. W.; Flynn, C. M., Jr.; Diemente, D. J. Am. Chem. Soc. 1975, 97, 3838-3839. (20) Bambot, S. B.; Rao, G.; Romauld, M.; Carter, G. M.; Sipior, J.; Terpetchnig, E.; Lakowicz, J. R. Biosens. Bioelectron. 1995, 10, 643-652. (21) Kober, E. M.; Sullivan, B. P.; Dressick, W. J.; Caspar, J. V.; Meyer, T. J J. Am. Chem. Soc. 1980, 102, 7385-7387. (22) Kober, E. M.; Caspar, J. V.; Lumpkin, R. S.; Meyer, T. J. J. Phys. Chem. 1986, 90, 3722-3734. (23) Kober, E. M.; Marshall, J. M.; Dressick, W. J.; Sullivan, B. P.; Caspar, J. V.; Meyer, T. J. Inorg. Chem. 1985, 24, 2755-2763. (24) Xu, W.; Schmidt, R.; Whaley, M.; Demas, J. N.; DeGraff, B. A.; Karikari, E. K.; and Farmer, B. L. Anal. Chem. 1995, 67, 3172-3180. (25) Demas, J. N.; DeGraff, B. A.; Xu, W. Anal. Chem. 1995, 67, 1377-1380. (26) Papkovsky, D. B.; Ponomarev, G. V.; Trettnak, W.; O’Leary, P. Anal. Chem. 1995, 67, 4112-4117. (27) Vinogradov, S. A.; Wilson, D. F. J. Chem. Soc., Perkins Trans. 2 1995, 103111.
Analytical Chemistry, Vol. 68, No. 15, August 1, 1996 2605
oxygen sensors without the need for a fiber coupler. Tissue is reasonably transparent at longer wavelengths, and embedded sensors can be excited directly through the skin. EXPERIMENTAL SECTION Emission Spectra. Most luminescence spectra and quenching measurements were carried out on a Spex Fluorolog 2 + 2 spectrofluorometer using excitation wavelengths of 480-502, 630, 650, and 670 nm.11 Corrected luminescence spectra were measured on a SPEX Fluoromax spectrofluorometer. Lifetime measurements were made with the nitrogen laser-based decay time instrument described earlier.11 Synthesis of Osmium Complexes. Four representative osmium complexes were studied: [Os(Ph2phen)3]Cl2, [Os(Ph2phen)(phen)2](PF6)2, [Os(Me2phen)(phen)2](PF6)2, and [Os(phen)3](PF6)2. The abbreviations used are as follows: phen, 1,10phenanthroline; Ph2phen, 4,7-diphenyl-1,10-phenanthroline; and Me2phen, 4,7-dimethyl-1,10-phenanthroline. All complexes were prepared by reaction of Os(L)2Cl2 with the appropriate R-diimine ligand. Synthesis and purification were patterned after literature methods.28,29 Os(phen)2Cl2 and Os(Ph2phen)2Cl2 were prepared by the reaction of (NH4)2OsCl6 and 2 equiv of the organic ligand in refluxing ethylene glycol. A sample preparation is as follows: 139 mg (0.316 mmol) of (NH4)2OsCl6 (99.99% Alfa) and 210.6 mg (0.633 mmol) of Ph2phen (G. Frederick Smith Chemical Co.) were added to the reaction flask, which was evacuated and flushed with nitrogen two times. Next, 10 mL of ethylene glycol (Mallinckrodt) was added to the flask. The reaction mixture was purged with nitrogen for 5-10 min. After the reaction mixture was refluxed for about 40 min under nitrogen, 10 mL of saturated aqueous sodium dithionite (sodium hydrosulfite) solution was added to the cooled reaction mixture, and the mixture was stirred for 5-10 min in order to reduce any Os(III) to Os(II). A dark black precipitate was isolated by filtration and extensively washed with water and ethyl ether. The yield was about 90%. The products of this reaction depend on the reaction conditions, reaction medium, temperature, and reaction time. If the reaction time was 24 h or longer, then the reaction would yield a number of products. For example, using Ph2phen as the ligand, it is possible to obtain Os(Ph2phen)2Cl2, Os(Ph2phen)3Cl2, and other, unknown species. Column chromatography was used to purify Os(phen)2Cl2 and Os(Ph2phen)2Cl2. Neutral alumina or basic alumina activity 1 (Fisher) was used as the column support, and acetonitrile/ethanol mixtures were used as the eluant. The separation was monitored by TLC. Solvents were removed by rotary evaporation. [Os(Me2phen)(phen)2](PF6)2, [Os(Ph2phen)(phen)2](PF6)2, [Os(phen)3](PF6)2, and [Os(Ph2phen)3]Cl2 were prepared under oxygen-free conditions as follows: For the first three complexes, Os(phen)2Cl2 was reacted with Me2phen, Ph2phen, or phen (1: 1.1 metal ligand ratio) in a 1:1 ethanol/water solution. For [Os(Ph2phen)3]Cl2, [Os(Ph2phen)2]Cl2 was reacted with a large excess of Ph2phen (1:2). Sufficient solvent was used to dissolve all the reactants during refluxing. Usually the reaction required 4-8 h or longer to complete the conversion, which was monitored by TLC. For [Os(Ph2phen)3]Cl2, it was necessary to monitor the reaction closely, as too long a reaction period gave increasing (28) Johnson, S. R.; Westmoreland, T. D.; Caspar, J. V.; Barqawi, K. R.; Meyer, T. J. Inorg. Chem. 1988, 27, 3195-3200. (29) Giuffrida, G.; Calogero, G.; Guglielmo, G.; Ricevuto, V.; Ciano, M.; Campagna, S. Inorg. Chem. 1993, 32, 1179-1183.
2606 Analytical Chemistry, Vol. 68, No. 15, August 1, 1996
Figure 1. Prepolymer used.
amounts of a red-emissive species. Except for the [Os(Ph2phen)3]Cl2, which precipitated directly from the reaction mixture, the complexes were precipitated as hexafluorphosphate salts upon addition of an excess of saturated aqueous NH4PF6 (Strem Chemicals, 99.5%). After evaporation, the solid was washed with water and ethyl ether and then dried. Purification was done by column chromatography. Basic alumina activity 1 (Fisher) was used as the column support. For the first three complexes, CH2Cl2/ethyl alcohol/ethyl ether in a 1:2:1 ratio was used as the eluant. The results were monitored by TLC, luminescence intensity measurements, and UV-visible spectra. [Os(Ph2phen)3]Cl2 was purified on basic alumina activity 1 using acetonitrile/ethanol mixtures; initially, a low ethanol concentration was used and gradually increased. A dark brown portion (80%), a golden yellow, more emissive portion (
