J. Phys. Chem. 1994,98, 2367-2376
2367
Photophysical and Electron-Transfer Properties of Pseudoisocyanine in the Hydrophobic Microdomain of an Aqueous Polyelectrolyte Guilford Jones 11’ and Churl Oh Department of Chemistry, Boston University, Boston, Massachusetts 0221 5 Received: October 26. 1993”
The binding of pseudoisocyanine (PIC+) to the polyelectrolyte poly(methacry1ic acid) (PMAA) has profound effects on the photophysical and photochemical properties of this prototypical cyanine dye. The hydrophobic dye was bound in the microdomain of the compact conformation of the polymer in its (uncharged, “hypercoiled”) acid form a t p H < 4.0 in water. Under these conditions, the fluorescence quantum yield for PIC+ was increased 600-fold and its lifetime is extended to 2.7 ns. The dye triplet state observed by flash photolysis provided a very long-lived phototransient (Amx = 640 nm, 50-100-ps decay time). Electron-transfer quenching was investigated using the oxidant tetranitromethane (TNM) which provided thesemioxidized dye radical intermediate (440-nm transient) on cobinding within PMAA hypercoils. The dye was also bound to a covalently modified form of PMAA in which polymer chains were end-labeled with 9-methylanthracene moieties. Electron transfer between anthracene chromophores and PIC+ within the polymer domain was observed.
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Introduction Study of the photophysics of the cyanine dye known as pseudoisocyanine (PIC+) has a relatively long history, as the result of early findings regarding activity of thedye and its relatives in the sensitizationof silver halide emulsions.1 Other more recent applications of PIC+ have involved observation of photoconductivity or electroluminescence of organic thin films of dye aggregates.2 Although the mechanisms of sensitization of photographic films are now somewhat better understood in terms of either hole or electron injection by cyanine dyes in their aggregated forms,Is3-5 it remains curious that, as a class, cyanines have photoproperties that are circumscribed by very rapid nonradiative decay involving torsional motion or *-bond isomerization. For instance, PIC+ is known to have a very short singlet excited-state lifetime and low fluorescencequantum yield in lowviscosity media.6s7 The fluorescencelifetime of monomeric PIC+ in aqueous solution has been reported to be in the range of 11-16 PS.~-’O Diverse results have been reported for the fluorescence lifetime of PIC+ aggregates, which are formed at high concentrations of PIC+ in aqueous media. These lifetimes depend on the concentration, temperature, and excitation intensity but are still very short and rarely exceed a few hundred picoseconds.11 For radiationless deactivation of PIC+, Tredwell and Keary12 proposed a photoisomerization mechanism involving rotation around the methine bridge, which occurs due to the lowering of the bond order of the methine chain in the excited state. By laser flash photolysis, Rentsch et 01.8 observed rapid formation of a 1 ns) which they relatively long-living phototransient ( T identified as the “photoisomer” of PIC+. However, through investigation of the temperature dependence of fluorescence intensityof PIC+ in glycerol solution, Dorn and Muller13 observed two activation energies (320 and 23 15 cm-l), for the radiationless transition, both of which were considered small for an isomerization of this type involving bond rotation in the highly viscous glycerol medium. They favored a more general direct internal conversion path for PIC+ decay via various vibrational modes, including a torsional vibration of the quinoline rings. Although the precise mechanism responsible for the rapid nonradiative deactivation for PIC+ remains in doubt, some rotatory motion around the methine bridge clearly plays an important role in the deactivationprocess. Indeed, l,l’-methylene-2,2’-cyanine, which has a structural similarity to PIC+ but has a rigid methine linkage
-
*Abstract published in Advance ACS Abstracts. February 1, 1994.
H3C COOH
Pseudoisocyanine (PIC+)
PMAA
PlC‘f+
APMAA
Figure 1. Structures of dye (parent and semioxidized forms) and
polyelectrolytes (acid form).
of the heterocyclic aromatic rings, has intense emission even at room temperature in fluid media.14 Moreover, with the use of glasses at low temperature, fluorescence, and even phosphorescence emission for PIC+ are readily observed.13J5 A recent study of PIC+ in ethylene glycol-water at 4.2 K revealed triplet properties of both monomeric and aggregated forms of the dye, including phosphorescence and delayed fluorescencedetection of magnetic resonance.16 The aqueous medium of the polyelectrolyte poly(methacry1ic acid) (PMAA) has been the object of considerable scrutiny in recent years. The “hypercoiled” c o n f ~ r m a t i o n ~of ~ -the l ~ polymer in its acid form displays properties reminiscent of a globular protein and is capable of very effective solubilization and compartmentalized reaction of nonpolar compound~.~O-25 The conformational transition of PMAA from the hypercoil to a more extended, rodlike, form has also been of particular A variety of studies have used fluorescence probes for observation of the change in conformation, or for hydrophobic or electrostatic assembly of photochemicalreactants to assist electron- or energytransfer processes in the polymer microdomain.17-23929-36 In the present paper, the binding of PIC+ to PMAA in aqueous solution is described in terms of dramatic changes in photophysical properties observed under certain conditions. Upon binding to PMAA in slightly acidic solutions (pH < 5.0), a much longer excited-state lifetime is conferred on the dye, allowing the observation of relatively robust fluorescence and triplet formation
0022-3654/94/2098-2361$04.50/0 0 1994 American Chemical Society
Jones and Oh
2368 The Journal of Physical Chemistry, Vol. 98, No. 9, 1994 in a fluid medium at room temperature. By laser flash photolysis, photochemical electron transfer for PIC+ in the aqueous PMAA medium has been investigated, and the transient spectra for the important triplet and radical-ion intermediates for PIC+ (microsecond time domain) have been identified. Additionally, the effects of polymer compartmentalization on the electron transfer between PIC+ and electroactive reagents that are hydrophobically cobound with dye, or covalently attached to PMAA, have been studied. Experimental Section Materials. Tetranitromethane (TNM) was purchased from Aldrich Chemical Co. and used as received. Pseudoisocyanine (PIC+) iodide salt was purchased from Kodak and transformed into the chloride salt by adapting an ion-exchange method previously de~cribed.3~ It was then recrystallized from methanolethyl ether three times. Poly(methacry1ic acid) (PMAA) was prepared by AIBN-initiated polymerization38 of freshly distilled methacrylic acid (MAA) in D M F with continuous nitrogen bubbling at 60 OC for 12 h and purified by multiple precipitation from methanol on addition of ethyl ether. The polymer was fractionated according to the procedure described by Fl0ry3~ except that each polymer solution was homogenized by cooling. The second fraction with the number-average molecular weight of 25 000 (determined by glass v i s ~ o m e t r y ~was ~ ) used in subsequent experiments. The 9-methylanthracene end-labeled PMAA (APMAA) was synthesized by AIBN-initiated radical polymerization of MAA in the presence of 9-(bromomethy1)anthracene (9-BMA) in dimethylformamide as described by Holden and G ~ i l l e t . ~The ’ mole percentage for anthracene-group incorporation in the polymer product varied dramatically depending on the composition of the reagents, the monomer (MAA), the radical transfer agent (9-BMA), and the initiator (AIBN). The highest labeling percentage was achieved starting with a 1OO:lO:l weight ratio of MAA:9-BMA:AIBN. Except for the composition of reagents, theother experimental conditions wereidentical to those employed for the synthesis of PMAA. In a typical experiment, 1.0 g of crude APMAA (after multiple precipitation) was obtained from 2.5 g of MAA and 0.25 g of 9-BMA on reaction at 60 OC for 12 h in DMF.41 The crude APMAA was then subjected to gel filtration using a polyacrylamide gel with fractionation range 1 500-20 000 (Bio-Gel P-10 from Bio-Rad) to ensure that the polymer is free of any unreacted 9-BMA. The eluent was then freeze-dried. The number-average molecular weight of the final APMAA sample determined by glass viscometry was 4500 (or 52 residues per polymer on average); the mole fraction of the anthracene labeling group was determined from the absorption spectrum of the sample to be 1.25%. Therefore, on average, about half of the polymer chains are terminally labeled with a methylanthryl group. In this analysis, the extinction coefficient of 9-methylanthracene (9.7 X 103M-1 cm-1 at 386 nm in heptane) was used as a reference. Instrumentation. Measurements of pH were made on a Fisher Accumet pH meter. Absorption spectra were measured using a Beckman Model DU-7 spectrophotometer. Both steady-state emission spectra and emission lifetimes were obtained using a Model 48000 phase-shift fluorometer from SLM Instruments. Although the basic layout of the laser flash photolysis system in this laboratory has been described previ0usly,~2several important changes recently made for the system require a more detailed description. The transient absorption is now monitored by a pulsed 150-W Xenon lamp that improves the signal to noise ratio. Data acquisition is carried out by using an Apple Macintosh IIci connected to the CAMAC electronic modules that are housed in a Kinetic Systems Model 1500 CAMAC crate. The PMT (Hamamatsu R928) signal is digitized by using either a LeCroy TR8818 100 MHz 8-bit transient recorder (8818) or a LeCroy 6880A 1.3 GHz 8-bit waveform digitizer (6880). The 8818 has
several sampling frequencies available from an internal clock; extra sampling frequencies are provided by an external LeCroy 8501 programmable 3-speedclockgenerator. A Kineticsystems 3082 12-bitoutput register module controls the shutter and pulser of the monitoring lamp. The interfacing program, LASER, has been developed using THINK C and THINK Class Library (TCL) from Symantec Co. The LASER program implements a full Macintosh graphical user interface and provides simple ‘point-and-click” operation of the system, involving data acquisition, manipulation, and analysis. The data-fitting algorithm in LASER employs the Marquart method, an iterative nonlinear least-squares fitting procedure. Initial parameter values, the pre-exponential factors and decay constants, are provided for exponential fittings by the method of successiveintegration.43 Currently, most types of kinetic processes commonly encountered, such as first-order (or single-exponential), second-order, mixed first- and second-order, and multexponential (with up to three exponential terms), including rise-and-fall kinetic data are incorporated into the data analysis options. General Methodology. An aqueous stock PIC+ solution with an approximate concentration of 2 mM was prepared and stored in a refrigerator. A 10pM PIC+ solution was prepared by diluting the stock PIC+ solution, and the exact concentration of dye was determined by using the known extinction coefficient of PIC+ in waterat 520nm(6.4X 104M-lcm-l). The0.5MPMAAsolution was prepared by dissolving the required amount of PMAA in deionized water; the resulting gellike mixture was kept in a refrigerator to homogenize a t least a day before use. For the mixture of PIC+ and PMAA with varying RID (the molar ratio of polymer residues to dye), a series of PMAA solutions with varying concentrations (1, 2, 5 , 10, 50, and 100 mM residues) was prepared from the stock of PMAA solution and mixed with an equal amount of 10pM PIC+solution. All spectrophotometric measurements were carried out while the sample holder was thermostat-controlled to 20 OC. For emission studies, a 1 X 1 cm all-face-polished quartz cell was used for right angle detection. The samples were excited a t 460 nm, and emission was monitored for a spectral range from 500 to 750 nm; the monochromator slit widths were set at 4 and 16 nm,for the inlet and outlet paths, respectively; the optical density at the excitation wavelength was no greater than 0.1 under all fluorescence measurement conditions. Fluorescence Quantum Yield Measurements. Fluorescence quantum yields (apf)weredetermined by comparing the spectrally corrected emission intensity of the sample to that of a fluorescence standard by using the following equation (with no refractiveindex correction):44
where I(;) and A stand for the emission intensity as a function of wavenumber and the number of absorbed photons, respectively; the subscript (s) denotes the standard. In the equation above, A can be substituted by the optical density, when the OD is reasonably small ( 6.0 for which the polymer adopts the extended rod conformation.17-19 The electrostatic interaction of PIC+ with charged polyanions, including poly(sodium methacrylate) at basic pH, has been studied p r e v i o u ~ l y , including 5 ~ ~ ~ ~ observation of PIC+ J-aggregates,56,57which can be identified by a sharp absorption band (J-band) about 50-nm red-shifted from the monomer absorption maximum (520 nm). Theaggregation is favored under high dye loading conditions (low values for the ratio of polymer residue to dye concentration, RID) and gradually disappears as RID increases; with PMAA, for instance, PIC+ J-aggregates can be optimized a t RID 6 and pH 8. Under all conditions employed for this study, however, J-aggregation was not favored due to relatively low dye loading conditions (high R I D S ) . The effect of dye binding to polymer on the emission and absorption of the dye are shown in Figures 2 and 3. Upon binding to hypercoiled PMAA, the absorption maximum shifts to the red by 6-8 nm, and the peak narrows; the extinction coefficient at its maximum is more than 20% greater than that of free PIC+ (Figure 2). The vibronic features in the absorption spectra are also somewhat better resolved upon binding. Figure 3 shows the effects of pH on the absorption maximum of PIC+ in the presence of PMAA at RID = 2000. The sharp decrease of the absorption
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maximum from pH = 5.5 to pH = 6.0 should be due to the PMAA conformational transition,I7-l9and further subtle changes of absorption around pH = 7 can be attributed to a low level of aggregation of PIC+ and partially charged PMAA. The degree of ionization for PMAA in water in the absence of dye (and at low ionic strength) is 0, 0.08, and 1 for pH 4.0, 6.0, and 8.0, respe~tively.~~ At high p H (18),the absorption spectrum closely resembles that of free PIC+. While the effect of PMAA binding on absorption is moderate, the effects on the emission properties of PIC+, such as the fluorescence lifetime and quantum yield, are much more pronounced. Emission data and other photophysical properties of bound and free PIC+ are compiled in Table 1. Figure 2 shows the corrected emission spectra of PIC+ along with the absorption spectra in the absence and presence of PMAA. The effect of pH (and the conformation transition) on dye fluorescence intensity along with dye absorptivity is also shown in Figure 3. Note that the emission quantum yield of PIC+ increases several hundred fold upon binding to hypercoiled PMAA. Binding Constant for PIC+-PMAA Complexation. Several methods for characterizing the binding constants for well-defined binding equilibria, using steady- state fluorescence measurements, have been described p r e v i o u ~ l y .Since ~ ~ it is quite complicated to describe generally the binding of a dye to a polydisperse polymer
-
Pseudoisocyanine in an Aqueous Polyelectrolyte
The Journal of Physical Chemistry, Vol. 98, No. 9, 1994 2371
TABLE 1: Some Photophysical Properties of PIC+ with and without PMAA parameter without PMAA with PMAA“ h,,(abs) 520 nm 526 nm 6.4X lo4 M-1 cm-I 7.7 X lo4 M-1 cm-1 h X X,(em) 542 nm 550 nm @f 6.8X 1t5 4.1 X 1p2 ‘f 11-16 psb 1.6nsc 0.03& 0.02 0.35f 0.01 anisotropy ( 7 ) a Aqueous polymer solutions with R/D 2000 and pH 4.0. From refs 8-10. e Average lifetime based on double-exponential analysis.
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where A@ and A@, are the change of apparent fluorescence quantum yield and its limiting (maximum) value, respectively. Therefore, by fitting the data of fluorescence quantum-yield change as a function of total monomeric polymer concentration to eq A9, one may obtain the extrapolated maximum value of quantum-yield change (A@,) and the effective binding constant (K). Figure 4 shows the plot of the quantum-yield change versus monomeric polymer concentration. The best fit was obtained by a nonlinear least-squares procedure with the monomeric polymer concentration being the weighing factor to take into account the approximation in eq A6. The values of the phenomenological binding constant and maximum emission quantum yield, K and A$,, from the fit are 1700 M-1 and 0.043, respectively.
TABLE 2 Emission Lifetimes for PMAA-Bound PIC+, Determined Using a Two-Component (Double-Exponential) Analysis of the Data Obtained from Phase-Shift Fluorometry* T(short) Ashort) ‘(long) Along) reduced av RID (ps) (%) (ns) (%) x2 lifetime (ns) 100 175 61 1.51 33 8.9 0.64 200 394 64 2.00 36 1.03 0.91 500 614 56 2.31 44 0.50 1.39 1000 707 51 2.63 43 0.52 1.53 2000 736 55 2.71 45 0.82 1.62 5000 665 49 2.65 51 0.79 1.68 10000 810 55 2.15 45 0.74 1.68 [PIC+]= 5 pM in aqueousPMAA atvariousR/Dratios (thenatural pH of the samples varied from 4.8 at RID = 100 to 3.6at RID = 1000); LX= 460 nm; X,, 1 550 nm. Fluorescence-Lifetime Measurements. The phase-shift and demodulation data for emission obtained for PIC+-PMAA samples can be fitted to a double-exponential decay quite well (Table 2). This result may indicate that there are two distinct polymer regions for dye binding, one in the core and the other near the surface of the PMAA hypercoil. However, given the complexity of the PIC+-PMAA system and the likely heterogeneity of binding sites, other methods of evaluation of fluorescence decay data were employed, especially since it is known that the phase-shift and demodulation data from a sample made up of components with a broad distribution of lifetimes can be generally fitted to a double exponential.61 A simulation study61 has also shown that the results of a double-exponential fitting to data from a distributed-lifetime sample were quite dependent on the set of modulation frequencies used to obtain the data. In general, a double-exponential fit to simulated data from a symmetric distribution is expected to give two components of equal amplitudeswith lifetimes symmetrically located with respect to the original distribution. However, asymmetric results are often obtained due to the set of frequencies used that weighted the components of the distribution nonuniformly. For a given distribution function the sets of frequencies that weight all the components of the distribution uniformly must be chosen depending upon the shape of the particular distribution. However, there is no standard set of frequencies that uniformly weights all the components of the distribution regardless of its shape. The analysis of the data in terms of probability density and lifetime distribution functions (vide infra) was found to be much less sensitive to such systematic errors due to the weighting of the data.6l In order to test if that is indeed the case, the lifetime data have been also analyzed by distributed-lifetime analysis (DLA) .61 In DLA, not just a few discrete lifetimes, but a range of lifetimes, whose fractional contribution to the total emission is defined by a function (a lifetime-distribution function), are parametrized in data fitting. Among several lifetime-distribution functions tested-including Lorentzian and Gaussian functions-the Lorentzian function gave the best results. Although the goodness of fit (x2)was similar to the results of double-exponential analysis, the lifetime distribution was unacceptably broad (typically greater than 1 ns), suggesting that the proper lifetime-distribution function should be more complicated. Since use of the more complicated lifetime-distribution function may result in errors due to overparametrization, further quantitative analysis of PIC+-PMAA lifetimedata was carried out using the double-exponential analysis method. The results of double-exponential analysis were far better than those of single-exponential analysis. Triplet-exponential analysis did not improve the goodness of fit much at all and often resulted in an insignificant contribution from the third component. One should bear in mind that the DLA analysis suggests that the appearance of two lifetimes in the PIC+-PMAA system is not the result of the systematic error associated with the selection of a particular set of modulation frequencies as discussed above. Of special interest was the apparent smooth shift of the component and average lifetimes (Table 2) with the polymer to dye
2372 The Journal of Physical Chemistry, Vol. 98, No. 9, 1994
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A@ Figure 5. Plots of the fluorescence anisotropy versus the fluorescence quantum yield of PIC+ upon binding to PMAA ([PIC+] = 5.0 p M ) at various R / D values (0, 100, 200, 500, 1000, 2000, 5000, and 10000 in the order of increasing emission quantum yield).
Figure 6. Transient spectra obtained on laser photolysis of BP in 50/50 (v/v) acetonitrile-water (& = 355 nm; [BPI = 5.0 mM). concentration ratio, RID. The effect is most likely due to the binding of dye to a polymer coil that takes on a continuously variable shape as more polymer is added and pH falls to a limiting value (for this concentration range) of pH about 3.5 where the (uncharged) polyelectrolyte is optimally hypercoiled. Not observed in the case of PIC+ binding is a discontinuityin the spectral signature associated with dye complexation with partially coiled, discrete PMAA structure (a cooperative binding and folding) that has been noted with several ~hromophores.~~-~~~~~~62~63 Fluorescence anisotropy ( r ) was monitored as a function of PMAA binding (reflected in parallel with emission quantum yield, 9,Figure 5 ) . One can notice that when PMAA is present, r for PIC+ is almost independent of fbund, the fraction of dye bound to polymer, if A 9 can be considered as a good representation of fbund (vide supra). This result is obtained because the anisotropy values from bound species are associated with the higher fluorescence intensities, and polymer complexes are much larger in effective size (larger rotational correlation timess1). Indeed, regardless of the fraction of bound species, the anisotropy data closely approach the fully bound state property. Pseudoisocyanine Triplet via Sensitization of Dye in Acetonitrile-Water. Because of the highly efficient nonradiativedecay of its singlet excited state, the formation of PIC+ triplet via direct excitation is very unlikely especially in nonviscous solvents. In Figure 7. Transient spectra obtained on laser photolysis of BP in the fact, free PIC+ in aqueous solution did not show any detectable presenceof PIC+ in 50/50 (v/v) acetonitrile-water (hx = 355 nm; [BPI transient in the microsecond time regime on laser flash photolysis. = 5.0 mM; [PIC+] = 5.0 pM). However, formation of the PIC+ triplet can be sensitized by benzophenone (BP) in AN-water. Figures 6 shows the transient photolysis, indeed, revealed sizable transients for the microsecond spectra resulting from 355-nm excitation of a sample containing time regime. Figure 8 shows the transient spectra from PIC+ 5.0 mM BP in AN-water (50/50 (v/v)) with 5 mM PIC+. In the bound to PMAA at RID = 2000 and hx= 532 nm. The negative presence of PIC+, BP triplet decay (520-nm transient52) is AOD corresponds to the PIC+ ground-state bleaching and accompanied by development of a transient absorption band in recovery. The positive absorption around 640 nm, with similar the 600-nm spectral region, along with negative optical density spectral characteristics to the BP-sensitized transient in ANdue to PIC+ bleaching at 500 nm (Figure 7). Since most 355-nm water, is assigned to the PIC+ triplet. The triplet should be also excitation is absorbed by BP in this experiment, and since BP is responsible for the relatively weaker absorption in the 400-nm transparent in the visible region, any negative optical density in region, as its formation and decay profile matches that of the the visible region should be unambiguously due to the loss of main triplet absorption band at 640 nm. ground-state PIC+ via photosensitization. This finding is conFor this air-saturated sample, the triplet decay is mostly induced sistent with a mechanism of triplet energy transfer from BP (ET by triplet oxygen, and the decay of triplet is accounted for almost = 68 k ~ a l / m o l toPIC+ ) ~ ~ (ET= ca. 43 kcal/mol).16 The transient entirely by the recovery of the ground state. By purging the observed in the 600-nm region is assigned to the triplet state of sampleof oxygen, the lifetimeof the transient at 640 nm increased PIC+. significantly (Figure 9). The decay of triplet in the presence of PseudoisocyanineTriplet in the PMAA Polymer Domain. Since oxygen can be best fitted to a double-exponentialdecay with the PIC+ in the hydrophobic polymer domain has a singlet lifetime following parameters: 21 ps (45%) and 89 ps (55%). This result extended several hundred times longer than that of free PIC+ in is consistent with the earlier finding in the PIC+ emission lifetime water, it was of interest to determine whether the dye triplet yield analysis indicating that the dye experiences two binding envialso increases significantly in such an environment. Laser flash
Pseudoisocyanine in an Aqueous Polyelectrolyte
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Time (p) Figure 9. Transient decay for PIC+-PMAA monitored at 640 nm with and without oxygen (hx= 532 nm; [PIC+] = 5.0 pM; aqueous PMAA solutions with RID = 2000 and pH = 4.0). The best fit was obtained with a double-exponentialdecay and a second-order decay for the decay with and without oxygen, respectively (see text). ronments. In the absence of oxygen, the half-life of the triplet decay increased to 670 ps. From the ratio of transient optical densities associated with the ground state (bleaching a t 530 nm; assumed t = 7.3 X lo4 M-' cm-I) and triplet state (640nm transient) of PIC+, measured at 3-10 ps following a 532-nm laser pulse and extrapolated to zero time, the extinction coefficient of triplet PIC+ at 640 nm was estimated to be 2.8 X lo4M-1 cm-l. The intersystemcrossing quantum yield for PIC+ in aqueous PMAA was determined to be 0.16 by comparative flash irradiation of methylene blue in water (triplet transient a t 420 nm with @ISC assumed to be 0.5253). Quenching of PIC+ Triplets with Electron Acceptors Internal or External to the Polymer Domain. The extended singlet lifetime and the significant yield of triplet make polymer-bound PIC+ potentially more photoreactive. In the presence of a cobound electroactive reagent, photochemistry may be observed due to an assumed proximity appropriate for the electron-transfer step within the polymer domain. Transient spectra are shown in Figure 10, obtained under experimental conditions identical to those employed to observe the triplet transient spectra, except for the presence of the electron acceptor reagent, tetranitromethane (TNM).64 The new transient absorption around 440 nm, which grows during the observation time, is assigned to the semioxidized
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Time (p) Figure 11. Transient decay for PIC+-PMAA in the presence of TNM monitored at 440 nm (X,= 532 nm; [PIC+] = 5.0 pM, [TNM] = 0.42 mM; aqueous PMAA solutions with RID = 2000 and pH = 4.0). PIC+ radical; one-electron reduction of TNM by PIC+ is consistent with the formation ofthe T N M reduction product, trinitromethide. The latter has an absorption maximum near 350 nmU and is responsible for the transient absorption below 400 nm. Other changes immediately obvious are the faster PIC+ triplet decay and the lack of recovery of the ground state. The very modest ground-state recovery (up to 40 p ) probably results from the triplet quenching by oxygen molecules, which competes with T N M oxidation. Purging the sampleof oxygen leads toa stable transient bleaching at 530 nm for the time regime up to 50 ps. The transient growth at 440 nm is shown in Figure 11. The fit obtained from nonlinear least-squares analysis of the last 990 points of decay (growth) to a single-exponential function with nonzero base line yielded the following results: baseline = 0.012; pneexponential factor = 8.0 X 10-3; growth rate constant = 1.1 X lo5 s-1. The fact that the transient decay a t 440 nm extrapolates to nonzero optical density for a time immediately following the laser pulse indicates that not only the triplet state but also the singlet state of PIC+ has been quenched (oxidized) by TNM. In order to verify the oxidation of PIC+ excited singlet by TNM, the changes of PIC+ fluorescence intensity as a function of T N M concentration were studied, and a Stem-Volmer plot of the results
Jones and Oh
2314 The Journal of Physical Chemistry, Vol. 98, No. 9, 1994 1.25
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SCHEME 1: Photochemistry of PIC+ in the Presence of TNM PIC+ 'PIC" + C(N02)4 'PIC+*
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+ Products
PIC+
is shown in Figure 12. In the presence of the electron acceptor, the fluorescence intensity of PIC+ was indeed reduced with increased TNM concentration up to 0.3 mM, before reaching a plateau; the reduction in fluorescence a t [TNM] = 0.42 mM was about 25%. The nonlinear behavior of the fluorescence quenching at higher T N M concentrations seems to be due to the saturation of the solution with T N M as indicated by the increased light scattering noted on visual inspection of the mixture. Obviously, PMAA can accommodate water-insoluble T N M in its hydrophobic polymer matrix only to a limited extent (-0.3 mM). On the other hand, the fluorescence lifetimes in the absence and presence of 0.42 mM T N M were virtually identical: 2.8 ns/720 ps (40%/60%) and 2.8 ns/700 ps (41%/59%), respectively. Therefore, the mechanism of the fluorescence quenching is mostly static (no diffusional component); Le., PIC+ is quenched by neighboring (cobound) T N M molecules or remains fluorescent a t more remote sites. A reaction mechanism for the oxidation of PIC+ by T N M is summarized in Scheme 1. The products of photooxidation by TNM are often complicated mixtures and include ring substitution (nitration) and addition;64 one electron oxidation of cyanines has also been shown to result in stable dimers of the radical dications.65 The sample of PIC+ in PMAA (at RID = 2000) was also mixed with 1-10 mM methylviologen (MV2+) and wassubjected to laser flash photolysis. In contrast to the results with TNM, PIC+ triplet decay monitored a t 640 nm (up to 500 ps) was unaffected in the presence of the viologen acceptor and none of the reduced species (MV+, A,, = 396 nm) was observed. The emission lifetime of PIC+ was also measured to be identical with and without MV2+, indicating that MV2+ did not quench the PIC+ excited singlet. These results can be attributed to the fact that the less hydrophobic MV2+ is not readily incorporated into the PMAA hypercoil (pH I4.0). The results provide a contrast with findings for zwitterionic viologen and PMAA-bound chromophores. The latter more hydrophobic electron acceptor can
380
4W
420
440
460
480
5W
Wavelength (nm) Figure 13. Transient spectra observed on photolysis of APMAA (a) at pH 4.8 and (b) at pH 8.3 (hx = 355 nm; [APMAA] = 1.0mM-polymer residue concentration): recorded 1,2,3,4,5,7, and 9 ps after the laser flash in the order of increasing line thickness. be cobound under certain (lower pH) conditions.21.22.66 The important differences may also have to do with the lower driving force for singlet electron-transfer quenching for visible absorbing PIC+ that could potentially thwart a "long-range" electron transfer involving acceptor (e.q., MV2+)bound at a more remote site near the polymer surface. Photochemistry of Anthracene End-Labeled PMAA. In order to investigate the effects of the conformational change on the dynamics of the electron or energy transfer within the polymer microdomain, a second species that would react with PIC+ upon independent excitation was incorporated via covalent attachment to polymer. A modified PMAA containing a 9-methylanthracene group at the end of the polymer chain (APMAA) was synthesized and used to study its photoreaction with cobound PIC+. At pH 4.8, the transient observed for APMAA on 355-nm laser photolysis is the triplet state of the pendant methylanthryl group, which displays the well-known anthracene T-T absorption at about 430 nm.67,68 At pH = 8.3, the features of the transient spectra from APMAA are almost identical to those obtained at pH = 4.8 (Figure 13) except that the peak maximum is blue-shifted by a few nanometers and the decay rate is faster. The spectral shift can be attributed to the polarity change around the methylanthryl group upon unfolding of the polymer; the T-T absorption maximum for methylanthracene appears at 430 and 424 nm for the samples in benzene67and respectively. The faster triplet decay rate is also due to the exposure of the methylanthryl group to the water medium, where molecular oxygen can more freely encounter the pendant group. Some interesting changes in phototransient behavior were observed on "loading" APMAA with PIC+, although transient signals were weak and not well resolved. The experimental difficulties had to do with the relatively low absorptivity of the anthracene chromophore at the excitation wavelength (a new level of incorporation) and the moderate yield of triplets (10.5) expected for the methylanthryl g r o ~ p . ~In ~the , ~presence ~ of PIC+ (Figure 14), the transient spectra show elevated transient absorption near and below 410 nm, consistent with the formation of the reduced methylanthryl group (the anthracene radical anion absorbs at 410 nm69). A peak in the 440-450-nm region that we assign to the semioxidized PIC+ (uidesupru)along with a negative AOD due toPIC+ bleachingcanalso beobserved. What is perhaps most important is that the PIC+ transient bleaching that is unmistakable occurs in the submicrosecond time regime for APMAA at p H = 4.8, where the hypercoiled conformation dominates, and, by contrast, this bleaching signature is somewhat more intense and slower to develop for photolysis of PIC+-
The Journal of Physical Chemistry, Vol. 98, No. 9, 1994 2375
Pseudoisocyanine in an Aqueous Polyelectrolyte
0.wz
360
380
400
420
440
460
480
500
Wavelength (nm) Figure 14. Transient spectra observed on photolysis of APMAA in the presence of PIC+ (a) at pH 4.8 and (b) at pH 8.3 (Lx= 355 nm; [PIC+] = 10 pM, [APMAA] = 1.0 mM-polymer residue concentration): recorded 1, 2, 3, 4, 5, 7, and 9 ps after the laser flash in the order of increasing line thickness. APMAA at higher pH (Figure 14). Thelatter result isconsistent with the imposition of a diffusional component (some movement of bound counterions) for the quenching of anthracene triplet at polymer ends. On the other hand, ion-radicals could be formed as the result of (static) singlet quenching of the methylanthryl group when polymer has cobound PIC+ incorporated in the hypercoil. The spectral shift observed for the anthracene triplet is consistent with the positioning of this end group for polymer chains at a somewhat buried, less exposed site (along with PIC+). The fluorescence intensities measured a t the two extremes of pH indicate that the low-pH condition indeed yields a more efficient quenching: reduced by 57% and by 44% a t pH = 4.8 and 8.3, respectively. The apparently lower yield of the radicals at the lower pH, however, suggests that some fraction of quenching by an initial forward electron transfer is negated by charge recombination. Mixed results have been obtained in several studies regarding the efficiency of charge separation for cobound species in polyelectrolyte hypercoils. Delaire et al.21J2,66970in studies with diphenylanthracene (DPA) covalently bound to PMAA, found that when the DPA moiety is quenched by either methyl viologen or by a neutral (zwitterionic) viologen, the quenching efficiency is reduced a t lower pH, but the net charge separation efficiency is elevated significantly. They concluded that the "hydrophobic protection" afforded in PMAA at low pH was more effective in enhancing charge separation than simple electrostatic repulsion of the reduced quencher. Stramel et ~ l . , ~however, l found no enhancement of chiirge separation a t low pH, in their studies of the viologen quenching of pyrene, covalently bound to PMAA. More recently, Chatterjee et ~1.26also showed, in their studies of electron transfer involving a zwitterionic viologen and phenanthrene or naphthalene chromophores covalently bound to PMAA, that the yield of charge-separated products tends to increase with polymer deprotonation (charging or uncoiling). The results here, anthracene quenching by PIC+ and PIC+ quenching by TNM, emphasize the importance of static quenching of singlet excited states in the polymer domain. The dynamic (diffusional) quenching involving triplet states is more efficient for the unfolded polymer conformation.
hydrophobic to be easily incorporated into the hypercoil. For various polymer-residue-to-dye concentration ratios (RID), several photophysical properties of PIC+ have been recorded, e.q. fluorescence yield, lifetime, and polarization. The dye fluorescence is a sensitive probe of polymer microdomain and reveals a high local viscosity for the compact form of PMAA. The profile of fluorescence decay (double exponential) is consistent with the participation of at least two microenvironments for dye encapsulation which are more or less exposed to an aqueous or interfacial phase (about 0.7- and 2.7-11s lifetimes, respectively). The singlet excited state of PIC+ bound to the PMAA hypercoil lives long enough to yield the dye triplet state in relatively high yield. The resulting triplet from laser photolysis (532-nm excitation) is compared with the PIC+ triplet formed by photosensitization and further characterized by various kinetics and quenching experiments. The extinction coefficient for the PIC+ triplet is determined to be 2.7 X lo4 M-' cm-l at 640 nm, and the decay lifetime (in the compact PMAA coil) in the presence of oxygen is ca. 50 ps. A sacrificial electron acceptor, TNM, cobound to the PMAA hypercoil oxidizes both singlet and triplet excited states of PIC+ by a static quenching mechanism. Studies of methyl-anthracene, end-labeled on PMAA, with PIC+ at two different pH's (4.8 and 8.3) show that the singlet quenching of the methylanthryl group by PIC+ oxidation at the lower pH (hypercoiled PMAA) is followed predominantly by the backelectron transfer and return to the ground state and produces only limited yields of radicals. The quenching of the triplet methylanthryl group by PIC+ is observed only at higher pH, where cobound groups are presumed to have higher mobility.
Acknowledgment. Support of this research by the Department of Energy, Office of Basic Energy Sciences, is gratefully acknowledged. C.O. also wishes to thank the National Institutes of Health for support through a biophysics training grant. Appendix Analysis of Dye-Polymer Binding Equilibria. Consider the equilibrium for dye complexation with polymer binding site i. The microscopic binding constant, k, can be expressed in terms of the concentration of bound and free site i: k = [bound]/[free][D]
(All
where [D] is the free dye concentration. The fractional saturation of site i can then be expressed in terms of k: si =
[bound] [free] [bound]
+
--
[bound]/[free] 1 + [bound]/[free]
-- k[DI 1 + k[D]
A similar expression may be written for each of the n identical sites. Then adding these n expressions together yields n
s=
nk[DI
Csi= ns, = 1 + k[D]
(A3)
i
where 3 is the average number of dye molecules per polymer. Rearranging the above equation yields a linear relationship between Sf [D]and 3: S/[D] = n / k - s / k
(A41
Summary
If [PI0 is the total polymer concentration, by noting that S[P]o is equivalent to the total bound dye concentration, [BD], the fraction of dye bound to polymer,fbund, can be expressed in terms of 3:
The aqueous PMAA that exists as a hypercoil below pH = 5 provides a rigid matrix for solubilization of hydrophobic compounds. Even the charged cyanine dye, PIC+, is sufficiently
fb,,nd
is readily accessible from experimentally observable quan-
2316 The Journal of Physical Chemistry, Vol. 98, No. 9, 1994 tities (uide infra), as is 3. Once 3 values are determined, one can extract values for n and k from the plot of S/[D] versus S for varying dye concentrations (Scatchard plot)."' However, in this study, the dye concentration is held constant, and RID is varied instead. Therefore, needs to be expressed in terms of [P]o.This can be done by using the relationship [D] = [D]o - [BD] = [D]o - J[P]o. Applying this to eq A3 results in a quite complicated relationship between 3 and [Plo. However, under the typical conditions employed in this study, the relationship can be much simplified; that is, the total available binding sites, n[PIo, are much greater than [D] under low dye loading conditions (high RID), and hence, the chance for a particular binding site to be occupied by a dye is much less than 1. In other words, k[D] = [bound]/[free]
