Anal. Chem. 1996, 68, 604-610
Development of Chemical Sensing Platforms Based on Sol-Gel-Derived Thin Films: Origin of Film Age vs Performance Trade-Offs Richard A. Dunbar, Jeffrey D. Jordan, and Frank V. Bright*
Department of Chemistry, Natural Sciences and Mathematics Complex, State University of New York at Buffalo, Buffalo, New York 14260-3000
We present a detailed investigation on the evolution and performance of sol-gel-derived thin films as used for chemical sensing platforms. In order to develop an understanding of how the sol-gel matrix affects the entrapped recognition chemistry and determine how and whether the analyte interacts with the sensing element, we have chosen to investigate a simple model probeanalyte system. Specifically, we use static and timeresolved fluorescence spectroscopy to report on the photophysics and O2 quenching of pyrene entrapped within sol-gel-derived thin films as a function of precursor form, processing conditions, and storage time. The results of this year-long study show that the analytical response of the pyrene-doped film/sensor to O2 decreases as a function of storage time. This response decrease results from two separate factors. First, the average bimolecular quenching constant decreases from (1.31.4) × 107 to (0.4-0.6) × 107 M-1 s-1 for fresh and 300day-old films, respectively. Second, the average pyrene excited-state fluorescence lifetime, in the absence of quencher, decreases as a function of storage time. The simultaneous decrease in bimolecular quenching constant and average fluorophore lifetime are directly related to the change in analytical signal (i.e., response). These results demonstrate that single-component sol-gel-derived sensing platforms are unstable over time. However, we find that most of the observed instability occurs during the first month following film preparation. Doping of organic molecules into inorganic glasses is possible due to low-temperature sol-gel processing. To date, sol-gelderived composites have been used for the development of nonlinear optical materials and even bioactive glasses.1-9 Sol-gel(1) (a) Chemical Processing of Advanced Materials; Hench, L. L., West J. K., Eds.; Wiley: New York, 1992. (b) Paul, A. Chemistry of Glasses, 2nd ed.; Chapman and Hall: New York, 1990; pp 51-85. (c) Brinker, C. J.; Scherer, G. W. Sol-Gel Science; Academic Press: New York, 1989. (2) (a) Wung, C. U.; Pang, Y.; Prasad, P. N.; Karasz, F. E. Polymer 1991, 32, 605-608. (b) Zang, Y.; Prasad, P. N.; Burzynski, R. In Chemical Processing of Advanced Materials; Hench, L. L., West, J. K., Eds.; Wiley: New York, 1992; p 825. (3) Jeng, R. J.; Chen, Y. M.; Jain, A. K.; Kumar, J.; Tripathy, S. K. Chem. Mater. 1992, 4, 972-974. (4) (a) Braun, S.; Rappoport, S.; Zusman, R.; Avnir, D.; Ottolenghi, M. Mater. Lett. 1990, 10, 1-5. (b) Shtelzer, S.; Rappoport, S.; Avnir, D.; Ottolenghi, M.; Braun, S. Biotechnol. Appl. Biochem. 1992, 15, 227-235. (c) Zusman, R.; Rottman, C.; Ottolenghi, M.; Avnir D. J. Non-Cryst. Solids 1990, 122, 107-109. (d) Lev, O.; Kuyauskaya, B. I.; Sacharov, Y.; Rotman, C.; Kuselman, A.; Avnir, D.; Ottolenghi, M. Proc. SPIE-Int. Soc. Opt. Eng. 1992, 1716.
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processed glasses are ideal support matrices for organic and inorganic molecules because they combine the manufacturing versatility of organic polymers with the favorable chemical and optical characteristics of silica gel. These properties are especially important in photometric sensing, where optical transparency is of paramount importance. Low-temperature sol-gel processing involves the hydrolysis and condensation of metal or semi-metal alkoxide monomers, commonly tetramethoxysilane [TMOS, (CH3O)4Si] or tetraethoxysilane [TEOS, (CH3CH2O)4Si], to produce a colloidal suspension (the “sol”); gelation (polycondensation of the “sol” to form a wet siloxane network); and drying to form the xerogel (i.e., the dry gel). The sol-gel process is traditionally used to prepare large monolithic glasses (i.e., optical lenses and mirrors), fibers, and thin films. However, it is possible now to dope these sol-gelderived materials with a plethora of organic reagents applicable to chemical sensing.4-7 Comprehensive reviews on organically doped sol-gel composites and their potential for the development of chemical sensors are available.7 Recently, our group has focused on developing an understanding of the behavior of model dopants sequestered within solgel-derived composites6e-i and have actually used these composites as platforms for removing ribonuclease from aqueous solution,6d antibody-based biogels,6a and biosensors for rapid quantification of glucose6b and urea.6c In the course of this work, two key issues have become readily apparent. First, most work to date has focused on the entrapment of organic molecules or recognition (5) (a) Ellerby, L.; Nishida, C.; Nishida, F.; Yamanaka, S. A.; Dunn, B.; Valentine, J. S.; Zink, J. I. Science 1992, 25, 1113-1115. (b) Yamanaka, S.; Ellerby, L.; Nishida, C. R.; Dunn, B.; Valentine, J. S.; Zink, J. I. Chem. Mater. 1992, 4, 495-497. (c) Wu, S.; Ellerby, L. M.; Cohan, J. S.; Dunn, B.; El-Sayed, M.; Valentine, J. S.; Zink, J. I. Chem. Mater. 1993, 5, 115-118. (6) (a) Narang, U.; Wang, R.; Prasad, P. N.; Bright, F. V. Anal. Chem. 1993, 65, 2671-2675. (b) Narang, U.; Prasad, P. N.; Bright, F. V.; Kumar, A.; N. D.; Malhotra, B. D.; Kamalasanan, M. N.; Chandra, S. Anal. Chem. 1994, 66, 3139-3144. (c) Narang, U.; Prasad, P. N.; Bright, F. V.; Kumar, A.; Kumar, N. D.; Malhotra, B. D.; Kamalasanan, M. N.; Chandra, S. Chem. Mater. 1994, 6, 1596-1598. (d) Narang, U.; Rahman, M. H.; Wang, J. H.; Prasad, P. N.; Bright, F. V. Anal. Chem. 1995, 67, 1935-1939. (e) Jordan, J. D.; Dunbar, R. A.; Bright, F. V. Anal. Chem. 1995, 67, 2436-2443. (f) Narang, U.; Dunbar, R. A.; Bright, F. V.; Prasad, P. N. Appl. Spectrosc. 1993, 47, 1700-1703. (g) Narang, U.; Bright, F. V.; Prasad, P. N. Appl. Spectrosc. 1993, 47, 229-234. (h) Narang, U.; Wang, R.; Prasad, P. N.; Bright, F. V. J. Phys. Chem. 1994, 98, 17-22. (i) Narang, U.; Jordan, J. D.; Bright, F. V.; Prasad, P. N. J. Phys. Chem. 1994, 98, 8101-8107. (7) (a) Lev, O. Analusis 1992, 20, 543-553. (b) Dave, B. C.; Dunn, B.; Valentine, J. S.; Zink, J. I. Anal. Chem. 1994, 66, 1120A-1127A. (c) Lev, O.; Tsionsky, M.; Rabinovich, L.; Glezer, V.; Sampath, S.; Pankratov, I.; Gun, J. Anal. Chem. 1995, 67, 22A-30A. (8) Wolfbeis, O. S.; Rodriquez, N. V.; Werner, T. Mikrochim. Acta 1992, 108, 133-135. (9) Li, X. M.; Ruan, F. C.; Wong, K. Y. Analyst 1993, 118, 289-291. 0003-2700/96/0368-0604$12.00/0
© 1996 American Chemical Society
chemistries within sol-gel-derived monoliths. Unfortunately, monolith-based sensors are not practical for real-world detection because of the inherently long response times associated with the long, tortuous diffusion path length of the analyte and the limited accessibility of an entrapped chemical sensing element to the analyte. Therefore, in order to shorten the response times and improve sensor performance, diffusion path lengths must be decreased and the accessibility of the sol-gel-entrapped sensing element increased. This can, in principle, be accomplished by preparing porous sol-gel thin films.6b,c A second problem we have identified is associated with the fact that these sol-gel-derived materials are dynamical and evolution/aging of the sol-gel composite actually modulates the performance of any dopant entrapped within a sol-gel-derived material. In an effort to determine how the sol-gel matrix affects the sensing element (SE), determine the physicochemical properties of the local environment surrounding the SE, and quantify the ability of the analyte to diffuse to the SE, we have carried out a long-term series of experiments on a model SE/analyte system formed within a sol-gel-derived thin-film architecture. Specifically, we have investigated sol-gel-derived thin films doped with the fluorescent probe pyrene as they age and are subjected to O2 (analyte). Pyrene was chosen for several reasons. First, its photophysics have been extensively studied and are well understood.10 Second, the fluorescence intensity ratio of the I1 to I3 peaks (I1/I3) is sensitive to the physicochemical properties of the immediate environment surrounding the pyrene.10 For example, the I1/I3 value ranges from 0.41 in the gas phase to 1.95 for pyrene in dimethyl sulfoxide.10d Therefore, I1/I3 is a convenient “scale” of the dipolarity of the local environment surrounding the pyrene molecule. Hence, one can use I1/I3 to report on changes taking place in the sol-gel-derived film as a function of storage time and processing conditions. Finally, pyrene exhibits a relatively long excited-state fluorescence lifetime and is effectively quenched by O2.10 As a result, one can use the static and dynamic pyrene (Py) fluorescence to obtain key information on the efficiency of O2 quenching (Py* + O2 f Py + O2) and the accessibility of the pyrene residue to quencher/analyte.11 It is well-known that the physical characteristics (e.g., porosity) of the final xerogel depend on the size of the colloidal particles prior to gelation.12 For example, large particles eventually aggregate and give rise to highly porous silica xerogels and large particles can be produced preferentially by using longer hydrolysis times.13 As a result, we have investigated hydrolysis time as the key processing condition. Of course, there are many other process variables (e.g., temperature, pH, precursor molar ratios) that could be used to control average pore size and distribution. However, manipulation of more than one variable at a time is by (10) (a) Birks, J. B. Photophysics of Aromatic Molecules; Wiley-Interscience: New York, 1970. (b) Forster, T. H.; Dasper, K. Z. Phys. Chem. 1954, 1, 275279. (c) Birks, J. B.; Dyson, D. J.; Munro, I. H. Proc. R. Soc. A 1963, 275, 575-588. (d) Dong, D. C.; Winnik, M. A. Can. J. Chem. 1984, 62, 25602565. (e) Dong, D. C.; Winnik, M. A. Photochem. Photobiol. 1982, 35, 1719. (f) Ham, J. S. J. Chem. Phys. 1953, 21, 756-759. (g) Lochmu ¨ ller, C. H.; Wenzel, T. J. J. Phys. Chem. 1990, 94, 4230-4235. (h) Yamanaka, T.; Takahashi, T.; Kitamura, T.; Uchida, K. Chem. Phys. Lett. 1990, 172, 2932. (i) Yorozu, T.; Hushino, M.; Imamura, M. J. Phys. Chem. 1982, 86, 44264429. (j) Winnik, F. Chem. Rev. 1993, 93, 587-614. (k) Li, M.; Pacholski, M. L.; Bright, F. V. Appl. Spectrosc. 1994, 48, 630-637. (11) Lakowicz, J. R. Principles of Fluorescence Spectroscopy; Plenum Press: New York, 1983. (12) Hench, L. L.; West, J. K. Chem. Rev. 1990, 90, 33-72. (13) Greenblatt, M.; Buckley, A. M. J. Non-Cryst. Solids 1992, 143, 1-13.
no means straightforward. In this work, the long-term accessibility of the fluorescent sensing element/probe (pyrene) entrapped within sol-gel-derived thin films to O2 is investigated as a function of precursor form and hydrolysis time. The ease with which O2 can diffuse into the sol-gel-derived film and quench pyrene and the accessibility of the pyrene and the ability of the O2 to reach the pyrene SE are investigated as a function of film age. In this way we provide information on how the actual sol-gel matrix affects the entrapped SE, we determine how or whether the analyte interacts with the dopant, and we explain the exact origin of changes in the analytical response associated with this simple O2 sensor over 1 year of storage. THEORY Time-Correlated Single-Photon Counting. The theory and technique of time-correlated single-photon counting has been described in detail in the literature.14 A brief discussion of the method of lifetime recovery follows. The time-resolved fluorescence intensity [F(t)] can generally be described by a series of discrete exponential decays of the form n
F(t) )
∑R e
-(t/τi)
(1)
i
i
where Ri is a preexponential factor for component i having excitedstate lifetime τi. This is the decay that one would observe if one used an infinitely sharp excitation pulse (i.e., a Dirac delta), fast detection, and/or a pulse whose width is significantly shorter than the sample decay time. In more realistic situations, the observed decay of fluorescence [R(t)] is convolved with the excitation lamp pulse [L(t)]:
R(t) )
∫ L(t′)F(t - t′) dt t
0
(2)
The sought after impulse response function [F(t)] is generally determined from the convolved data by the method of nonlinear least-squares analysis.14 In this approach, R(t) is calculated using initial values of Ri, τi, and the actual, experimentally measured lamp profile. The calculated values of Rc(t) are then compared to the experimental R(t) values and the goodness-of-fit is determined by minimization of the χ2 function: n
χ2 )
∑w [R(t) - R (t)]
2
i
c
(3)
i)1
where wi is a statistical weighting factor that accounts for the uncertainty in each R(t) value. If wi accurately describes the uncertainty in R(t), then the model is judged by the closeness of χ2 to unity and random distribution of residuals [Rc(t) - R(t)] about zero. Average Lifetime. In many situations the excited-state intensity vs time profile is not described by a single-exponential decay law. Under these circumstances, when one does not have any a priori information on the origin of the individual decay terms (e.g., τ1, τ2), it is often best to refer to an average excited-state (14) Birch, D. J. S.; Imhof, R. E. In Topics in Fluorescence Spectroscopy; Lakowicz, J. R., Ed.; Plenum: New York, 1991; Vol. 1, Chapter 1.
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lifetime (〈τ〉):
〈τ〉 )
∑R τ /∑R τ 2
i i
i i
(4)
Collisional Quenching. In collisional quenching, a quencher and fluorophore must meet during the fluorophore’s excited-state lifetime. The fluorophore returns to the ground state, upon contact with the quencher, without the emission of a photon. Collision quenching of fluorescence is most simply described with the Stern-Volmer expression:11,15
F0/F ) 1 + kqτ0[Q] ) 1 + KD[Q]
(5) Figure 1. Pyrene-doped sol-gel-derived thin-film apparatus.
where F0 and F are the fluorescence intensities in the absence and presence of quencher, respectively, [Q] is the analytical concentration of quencher, kq is the bimolecular quenching constant for the interaction of the fluorophore and quencher, τ0 is the excited-state fluorescence lifetime of the fluorophore in the absence of quencher, and KD is the Stern-Volmer quenching constant. If the system is well-behaved, a plot of F0/F (or τ0/τ) will be linearly dependent on the concentration of quencher [Q]. However, this simple law is generally only followed if there is a single emissive species that is equally accessible to quencher.16 EXPERIMENTAL SECTION Materials and Reagents. The following chemicals were used: tetramethoxysilane (TMOS), tetraethoxysilane (TEOS), and pyrene (99%) (Aldrich); methanol and KOH (J. T. Baker), ethanol (200 proof) (Quantum); and HF, HCl, K2HPO4, and KH2PO4‚2H2O (Fisher). Nitrogen/oxygen gas mixtures were purchased from Matheson. Fused-silica, multimode optical fibers (1000-µm core diameter) were purchased from Fiberguide. Sol-Gel Film Preparation. TEOS sol-gel stock solutions were prepared by mixing TEOS (20.83 g), deionized water (3.60 g), ethanol (9.21 g), and HCl (16 µL of 1.00 M HCl). TMOS solgel stock solutions were prepared similarly by mixing TMOS (15.20 g), deionized water (3.60 g), methanol (6.40 g), and HCl (16 µL of 1.00 M HCl). Once the stock solutions were prepared they were individually transferred to Erlenmeyer flasks and mechanically stirred at room temperature for 1 or 5 days (hydrolysis times). Films were prepared from these solutions. To generate a substrate suitable for film casting, we prepared 6-cm segments of optical fiber by mechanically removing a 2-cm section of the protective sheath from the distal end of the optical fiber. In a second step, we removed the actual cladding by soaking the 2-cm section for 20 min in HF. ESCA and simple wetability studies were used to determine the point when all the cladding was removed and only clean, bare silica remained. If residual cladding were not removed, water would not wet the “bare” silica well. Prior to actual film casting, a 2.50-mL aliquot of hydrolyzed TEOS or TMOS was added to 1.25 mL of 0.010 M pH 6.0 phosphate buffer, and the mixture was stirred well. Immediately prior to film casting, we added a 50-µL aliquot of 14.9 mM pyrene in ethanol to the above mixture with stirring. Thin sol-gel films were subsequently prepared by immersing the clean fiber-optic (15) Eftink, M. R. In Topics in Fluorescence Spectroscopy; Lakowicz, J. R., Ed.; Plenum: New York, 1991; Vol. 2, Chapter 2. (16) Xu, W.; McDonough, R. C., III; Langsdorf, B.; Demas, J. N.; DeGraff, B. A. Anal. Chem. 1994, 66, 4133-4141.
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surface into the buffered TEOS or TMOS sol-gel solutions and withdrawing the fiber optic at a rate of 1 mm/s. Films were investigated using SEM, and the average film thickness was always 0.5 ( 0.1 µm. The morphology of these films, as sensed by SEM (not shown), did not change detectably over the course of these experiments; however, the film coverage is generally uniform and there were no detectable regions of composite inhomogeneity. Moreover, when different regions of the same film were probed, the experimental data were statistically equivalent. After dip casting the films were stored in a desiccator at room temperature in the dark. Under these preparation conditions, there were no complications from excimer-like emission or photoinstability. Triplicate samples were prepared and tested. The results shown are the average of these three discrete samples. Instrumentation. All steady-state fluorescence measurements were made with an SLM 48000 MHF spectrofluorometer. A xenon arc lamp was used as the excitation source (332 nm). Emission spectra and slow-time acquisitions were obtained using a monochromator for wavelength selection with photomultiplier tube detection. A 345-nm long-pass filter was used in concert with the emission monochromator to reduce the amount of scattered light reaching the detector. In order to resolve the vibronic bands within the pyrene emission envelop, the excitation and emission bandpasses were set at 8 and 2 nm, respectively. The slow-time acquisition traces were acquired with excitation and emission bandpasses at 2 and 16 nm, respectively. The emission monochromator was set at 393 nm for all slow-time data acquisitions. All time-resolved measurements were made using an IBH 5000W SAFE multiplex time-correlated single-photon-counting fluorescence lifetime spectrometer. A nanosecond flashlamp operating with N2 at 40 kHz was used as the excitation source. The excitation wavelength (337 nm) was selected using an monochromator. The emission was monitored using a 345-nm long-pass filter and the emission monochromator set at 390 nm. The fluorescence decay traces were analyzed using commercial software from Globals Unlimited.17 Measurement Scheme. The pyrene-doped sol-gel films were mounted within a quartz cuvette as shown in Figure 1. The film-coated fiber optic was aligned with respect to excitation and emission beam axis within the spectrofluorometers and held in place within a drilled Teflon cuvette cap. Two other holes in the (17) Beechem, J. M.; Gratton, E.; Mantulin, W. W. Globals Unlimited; Laboratory of Fluorescence Dynamics, University of Illinois at UrbanasChampaign, 1992.
Figure 2. Fluorescence of a typical pyrene-doped sol-gel-derived thin film cycled between (s) N2, (- - -) O2, and (‚‚‚) N2 purges (A). (B) Fluorescence intensity vs time profiles for a typical pyrene-doped solgel-derived thin film cycled between N2 and O2 purges.
cap allowed the system to be effectively purged with N2 or O2 and vented to atmosphere without disturbing the sample alignment. RESULTS AND DISCUSSION Steady-State Fluorescence. All measurements were made in the quartz cuvette (Figure 1) described in the Experimental Section. Figure 2A shows the fluorescence spectra of a typical pyrene-doped sol-gel-derived film stored for 1 month. Three spectra are presented, illustrating the fluorescence of the film cycled between N2 and O2 purges. These results show several interesting features. First, O2 can diffuse into the sol-gel film matrix and quench a portion of the pyrene fluorescence. In fact, the analytical response (R) is approximately 50%. Thus, a significant population of the pyrene molecules entrapped within the thin, sol-gel-derived composite are accessible to and can be quenched by O2. Second, the film functions as a true chemical sensor and reversibly responds to cycling between N2 and O2. Third, the key vibronic features inherent to the pyrene emission are evident and there is no detectable excimer-like emission. In addition, Figure 2B shows a typical fluorescence intensity (393 nm) vs time profile as the film is cycled between N2 and O2 purges. The response and the response time of the films are estimated from these data. This provides us with a means to measure or estimate the population of pyrene molecules accessible to O2 and the ease with which O2 can diffuse into the sol-gel matrix and quench pyrene. (Note: A significant fraction of the observed response time arises from the time it takes to sweep out residual gases from the tubing lines and the cuvette.) Additional film characterization was performed by determining the I1/I3 ratio, response, and the response times as a function of film storage time. Figure 3A presents the I1/I3 ratio for pyrene in TMOS- and TEOS-derived thin films (1- and 5-day hydrolysis) as a function of storage time. In an effort to calibrate the local
Figure 3. Pyrene I1/I3 ratio (A) and percent change in response (B) for a pyrene-doped TMOS- and TEOS-derived films (processed under 1- and 5-day hydrolysis times) as a function of storage time (TMOS: 1 day, 3; 5 days, O. TEOS: 1 day, 1; 5 days, b).
polarity scale, we also present benchmark I1/I3 values for dilute solutions of pyrene dissolved in liquid water, methanol, or ethanol. These values are shown and represented by horizontal arrows in the figure. Inspection of these results shows also that the I1/I3 ratio reproducibly decreases as a function of storage time. This is consistent with a slow, time-dependent decrease in the dipolarity of the local microdomain surrounding the average pyrene moiety. The I1/I3 ratio for the TMOS-derived films decreases from an initial value of ∼1.6 to 1.3, over a time period of 1 month. Therefore, the I1/I3 ratio senses an evolution of the composite that affects the average pyrene molecular environment. Interestingly, this evolution occurs over a time period of 1 month even though these films appear dry within a few minutes after being cast. After 1 month of aging, the time-dependent changes slow significantly. It is important to note that the aging/drying process for a monolith, under the same experimental conditions, is on the order of several months.18 The initial I1/I3 ratio is ∼1.2 for the TEOS-derived films, and it decreases to a value of ∼1.0 over the duration of these experiments (1 year). In all cases, the changes in the I1/I3 values are consistent with the expulsion of polar solvent from the matrix and a redistribution of the average pyrene molecules to a less dipolar (compared to liquid water) microdomain. However, the fact that the final domain is apparently alcohol-like may arise from the fact that there are significant numbers of residual silanol groups within dry xerogel composites.1 In order to determine the long-term stability and the performance of these films, the response and the response time were measured as a function of storage time. Figure 3B illustrates the film response to pure O2 as a function of storage time. Close inspection of these results shows that most of the decrease in response again occurs during the first month following film (18) Kaufman, V. R.; Avnir, D. Langmuir 1986, 2, 717-732.
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Table 1. Recovered Stern-Volmer Parameters for Pyrene in TMOS- and TEOS-Derived Thin Filmsa TMOS
TEOS
1 day
a
KD (M-1) 〈kq〉 (107 M-1 s-1)
4.73((0.20) 1.33((0.14)
KD (M-1) 〈kq〉 (106 M-1 s-1)
0.827((0.050) 4.22((0.54)
5 days
1 day
5 days
1-Day Storage 3.77((0.07) 1.37((0.04)
4.47((0.13) 1.35((0.10)
3.77((0.25) 1.38((0.10)
300-Day Storage 0.662((0.027) 4.19((0.33)
1.14((0.039) 5.23((0.54)
1.00((0.025) 5.56((0.18)
Following hydrolysis times of 1 and 5 days.
preparation. Interestingly, the actual response time remains essentially constant (∼10 s) over this same time period (results not shown). These particular results correlate well with previous work from our laboratory on sol-gel-based sensors, biosensors, and dynamical measurements within sol-gel-derived monoliths.6b,c,f,h,i That is, the greatest change in sensor response and/ or dynamics occurs within the first month following initial film preparation. Stern-Volmer Analysis. In the current work, steady-state quenching measurements provide information on the sensitivity of these pyrene-doped films to O2 quenching and yield insight into how the evolving sol-gel film matrix affects the sensitivity. All Stern-Volmer plots for the O2 quenching of pyrene in TMOSand TEOS-derived thin films (not shown) were linear. These results suggest a single fluorophore class accessible to molecular O2.11,15 Nonlinear Stern-Volmer plots usually are suggestive of two or more fluorophore populations with different accessabilities and/or excited-state lifetimes. The KD value associated with each film to O2 quenching are collected in Table 1. Inspection of Table 1 shows that the sensitivity, KD, decreases 4-6-fold after 300 days of storage for all films. Thus, the fact that there is a decrease in response (Figure 3B) is a direct result of the film sensitivity changing with storage time. Of course, one must now question the reason(s) for the observed loss in sensitivity: (a) a change in pyrene accessibility by O2 and/or (b) a lower rate of O2 diffusion within the sol-gel-derived composite (see eq 5). These particular questions can be addressed with information on the excited-state intensity decay kinetics of pyrene entrapped within the sol-gel-derived films. Time-Resolved Fluorescence. Steady-state fluorescence provides information on the average emission and cannot generally be used to extract information on the number and/or type of individual fluorescent centers within a system. In addition, one must know the excited-state fluorescence lifetime in the absence of quencher (τ0) to estimate the bimolecular quenching constant (cf. eq 5). Given this, we carried out a series of time-resolved fluorescence experiments on the pyrene entrapped within the TMOS- and TEOS-derived films. Panels A and B of Figure 4 present typical time-correlated single-photon-counting data sets for 1-day-old TMOS- and TEOS-derived films (1-day hydrolysis), respectively, in the absence of quencher. All data sets were acquired until there were at least 10 000 counts in the peak channel. The solid points are the experimental data, and the traces represent the best fit (vide infra) to multiexponential decay laws. Tables 2 and 3 collect the recovered fits associated with the time-resolved data for TMOS- and TEOS-derived films, hydrolyzed for 1 or 5 days, after 1 or 300 days of storage. In Tables 2 and 3 608 Analytical Chemistry, Vol. 68, No. 4, February 15, 1996
Figure 4. Typical excited-state intensity traces for TMOS- (A) and TEOS-derived (B) sol-gel films after 1 day of storage. The best fits are denoted by the solid-line trace through the experimental data (points). The residuals are plotted below the decay profiles. The recovered lifetime parameters and the average lifetimes are given in each decay curve. Table 2. Recovered Excited-State Decay Parameters for Pyrene in TMOS- and TEOS-Derived Thin Films (1-Day Storage)a hydrolysis precursor time (day) TMOS TEOS TMOS TEOS a
1 1 5 5
single τ1 χ2 349 252 227 228
τ1
double τ2 χ2
τ1
1.24 161 381 1.01 1 2.65 173 406 1.05 96 3.91 118 311 0.98 24 4.77 99 302 1.10 25
triple τ2 τ3 140 218 160 159
376 623 387 398
χ2 1.02 1.11 1.01 1.12
Lifetimes are in nanoseconds.
we have purposely omitted (for clarity) the fits for the replicate films and the recovered preexponential factors. In general, the agreement between samples is very good (
