8308
J. Phys. Chem. B 2000, 104, 8308-8320
Changes in Carbon-Carbon and Carbon-Nitrogen Stretching Force Constants in the Macrocycles of Bacteriochlorophyll a and Bacteriopheophytin a upon Triplet and Singlet Excitation: Resonance-Raman Spectroscopy and Normal-Coordinate Analysis of the Unlabeled and Totally 15N-, 13C-, and 2H-Labeled Species Tokutake Sashima,† Leenawaty Limantara,‡ and Yasushi Koyama*,† Faculty of Science, Kwansei Gakuin UniVersity, Uegahara, Nishinomiya 662-8501, Japan, and Faculty of Science and Mathematics, Satya Wacana Christian UniVersity, Jalan Diponegoro 52-60, Salatiga 50711, Indonesia ReceiVed: February 18, 2000; In Final Form: May 8, 2000
The Raman spectra of bacteriochlorophyll a and bacteriopheophytin a in the S0, T1, and S1 states were recorded for the unlabeled species and the totally-labeled species with the 15N, 13C, and 2H isotopes, and an empirical normal-coordinate analysis of the Raman data was performed to establish the assignments of Raman lines in the 1650-1200 cm-1 region and to determine the stretching force constants of the carbon-carbon and the carbon-nitrogen bonds within the macrocycle. By the use of the stretching force constants as a scale, changes in bond order upon triplet and singlet excitation were characterized in both BChl a and BPhe a as follows. (a) For BChl a, upon triplet excitation, the carbon-carbon bonds in the methine bridge (Ca-Cm) and those in the pyrrole rings I and III (Ca-Cb and Cb-Cb) decrease in bond order, whereas the carbon-nitrogen bonds in all the pyrrole rings (Ca-N) and the carbon-carbon bonds in rings II and IV (Ca′-Cb′) increase in bond order. Upon singlet excitation, the Cb-Cb bonds substantially increase in bond order at the expense of decrease in the bond orders of the Ca-Cm and Ca-Cb bonds; the Ca′-Cb′ and the Ca-N bonds also increase in bond order as in the case of triplet excitation. (b) For BPhe a, upon triplet excitation, changes in bond order similar to those in the case of BChl a take place. Upon singlet excitation, the Cb-Cb bonds drastically increase in bond order at the expense of decrease in the bond orders of the Ca-Cm, Ca-Cb, and Ca-N bonds. Change in the stretching force constant of each bond, upon singlet or triplet excitation, varies in magnitude depending on its location in the macrocycle, a fact which suggests that the HOMO and the LUMO are not completely delocalized. Possible relevance of this result to the pigment arrangements in the bacterial photoreaction center is discussed.
Introduction Changes in the electronic structures of photosynthetic pigments upon electronic excitation are expected to play a most important role in the primary processes in photosynthesis. In the bacterial photoreaction center, charge separation takes place upon photoexcitation of the special-pair bacteriochlorophylls (BChls).1 Here, the pair of BChl molecules must be arranged spatially to maximize the overlap and mixing of the LUMOs of the electron-donor and the electron-acceptor BChls to facilitate the electron transfer reaction. Under reducing conditions, on the other hand, charge recombination takes place in the special pair to generate the triplet state, and the triplet energy is transferred to the accessory BChl, and then to the carotenoid (Car)2 to be dissipated. Here, the relevant pigment molecules must be arranged to maximize the overlap and mixing of both the HOMO and the LUMO of the donor and the acceptor molecules to facilitate the triplet-energy transfer reaction through the electron-exchange mechanism.3 Changes in the electronic structures upon electronic excitation of a pigment molecule can be probed indirectly by excitedstate Raman spectroscopy. When we measure the S0, S1, and * Corresponding author. Phone: 81-798-54-6389. Fax: 81-798-51-0914. E-mail:
[email protected]. † Kwansei Gakuin University. ‡ Satya Wacana Christian University.
T1 Raman spectra of its variously isotope-substituted homologues and determine the stretching force constants in the conjugated system, we can use each force constant as a scale of the π-bond order of the relevant bond. In a Hu¨ckel-type MO description, where the molecular orbitals are expressed as a linear combination of the 2p atomic orbitals, a pair of large coefficients for the orbitals of adjacent atoms is expected to give rise to a large change in bond order and vice versa (see the Discussion section for details). In this way, we can predict a bond that contributes most to the HOMO and the LUMO as a bond that gives rise to a large change in π-bond order upon electronic excitation. In order to obtain information concerning changes in π-bond order (hereafter, abbreviated as “bond order”) upon excitation, we first recorded the resonance-Raman spectra of BChl a having the bacteriochlorin skeleton in the T14,5 and S16 states, and tried to probe them by comparing the frequencies of Raman lines in the different electronic states; here, we temporarily correlated the Raman lines having similar intensity and frequency. The results suggested systematic changes in bond order, in the order S0, T1, and then S1 state.6 In order to empirically assign each Raman line to a particular stretching vibration, we recorded the Raman spectra of unlabeled and totally 15N- and 2H-labeled BChl a in the S0, T1, and S1 states.7-9 The empirical assignments lead us to the following
10.1021/jp000645l CCC: $19.00 © 2000 American Chemical Society Published on Web 08/04/2000
Macrocycles of BChl a and BPhe a conclusion concerning triplet excitation: The bond orders of the Ca-Cm and Ca-Cb bonds having a double-bond character decrease in bond order, whereas the bond orders of the Ca-N bonds having a single-bond character increase in bond order. The same technique was applied to BPhe a, and similar changes in bond order were found upon triplet excitation.9 However, in both BChl a and BPhe a, the S1 Raman spectrum was too complicated to make a reliable set of assignments. In order to elucidate changes in bond order upon triplet and singlet excitation of chlorophyll (Chl) a having the complicated chlorin skeleton, we had to apply a more sophisticated technique,10 because its ground- and excited-state Raman spectra were too complicated to try any empirical assignment. We recorded the S0, S1, and T1 Raman spectra for unlabeled and totally 15N-, 13C-, 13C +15N-, 2H-, and 2H + 15N-labeled Chl a to collect maximum information concerning the Raman-active normal modes in order to facilitate normal-coordinate analysis: After determining a set of force constants in the S0 state, we tried to determine the T1- and S1-state force constants of the carbon-carbon and carbon-nitrogen bonds in the 16-membered ring, assuming that the rest of the force constants remain unchanged. Extremely complicated changes in bond order were elucidated by the use of the set of stretching force constants as a probe.10 In the present investigation, we have attempted to determine the stretching force constants in the S0, T1, and S1 states for the carbon-carbon and the carbon-nitrogen bonds within the entire bacteriochlorin skeleton by the use of the unlabeled and the totally 15N-, 13C-, 2H-, and 13C + 15N-labeled species of both BChl a and BPhe a. First, we used changes in the stretching force constants to probe changes in bond order upon triplet and singlet excitation in various parts of the bacteriochlorin macrocycle; we tried to compare the region where the largest changes in bond order take place11 among cyclic conjugated systems including BChl a, BPhe a, and Chl a. Second, we tried to locate a bond where the largest change in stretching force constant (bond order) takes place upon electronic excitation in order to identify localization of the HOMO and the LUMO of BChl a. Third, we used the localized part to propose a possible interpretation of the arrangement of the special-pair BChls in the photoreaction center which facilitates charge separation. Finally, we proposed a possible interpretation of pigment arrangements facilitating the triplet-energy transfer from the special-pair to the accessory BChl, and then to spheroidene. Experimental Section Sample Preparation. (a) BChl a. The unlabeled BChl a having the natural-abundance isotopic composition (“the NA species”) and the totally 15N- and 2H-labeled BChl a (“the 15N and 2H species”) were obtained from the cells of Rhodobacter sphaeroides 2.4.1 that were grown in the 15N- and 2H-enriched media.7,8 The 15N enrichment factor was determined to be 88% by mass spectrometry, whereas the 2H enrichment factor was determined to be 97% by NMR spectroscopy.8 Two additional isotope species, one totally labeled with 13C (“the 13C species”) and the other totally labeled with both 13C and 15N (“the 13C + 15N species”) were obtained from the cells of Chromatium Vinosum grown in a medium modified from that of Bose.12 The medium consisted of 1.0 g of NH4Cl, 2.0 g of NaHCO3, 10 g of NaCl, 0.5 g of K2HPO4, 0.5 g of KH2PO4, 0.5 g of MgCl2‚ 6H2O, 0.05 g of CaCl2, 0.005 g of FeCl3‚6H2O, 2.0 g of Na2S2O3‚5H2O and 1.0 g of Na2S‚9H2O dissolved in 1 L of tap water; NaH13CO3 and 15NH4Cl were used as the 13C- and 15N-sources. The 13C enrichment factors of the 13C species and
J. Phys. Chem. B, Vol. 104, No. 34, 2000 8309 the 13C + 15N species were 95% and 98%, respectively, and the 15N enrichment factor of the 13C + 15N species was 93%, both being determined by mass spectrometry. (b) BPhe a. The NA and each isotope-substituted BPhe a were obtained from the relevant BChl a as follows: 2 mL of glacial acetic acid was added to 100 mg of dry BChl a, and the mixture was dried with nitrogen gas; then, 2 mL of toluene was added and dried again with nitrogen gas to completely remove the remaining acetic acid. The pigment was then purified by molecular-sieve chromatography using Sepharose CL-6B as the stationary phase and acetone/toluene (1:99 v/v) as the eluent. The BPhe a fraction thus obtained was analyzed by silica gel HPLC using a 4 mm i.d. × 30 mm column packed with LiChrosorb Si-60, 5 µm and an eluent acetone/n-hexane (1:4 v/v); it showed a single peak (detection at 355 and 420 nm). Since undeuterated acetic acid was used in this preparation, hydrogen atoms are attached to the pyrrole nitrogens in all the NA and isotope-substituted species. Raman Measurements. The S0 Raman spectra were recorded by the use of the 457.9 nm line of an Ar+ laser (Lexel 95); the T1 Raman spectra were recorded by one-color experiment using the 420 nm, 5 ns, and 10 Hz pulses from a dye laser (Lambda Physik FL-3003) which was pumped by the THG of a Nd:YAG laser (Lumonics HY-400); and the S1 Raman spectra were recorded by one-color experiment using the 351 nm, ∼50 ps, and 1 kHz pulses (THG) from a combination of a Nd:YLF laser (Quantronix 4216) and a Nd:YLF regenerative amplifier (Quantronix 4417). The details were described elsewhere.4,6 All the measurements were performed in acetone solution; the purity of each sample, after each Raman measurement, was above 94% as determined by HPLC analysis (detection at 420 nm). Normal-Coordinate Analysis. Figure 1a shows the chemical structure of BChl a and the typification of the five rings and the carbon atoms in the bacteriochlorin skeleton. The orientation of the acetyl group is based on NMR spectroscopy of BChl a in acetone solution which exhibited an NOE correlation between the methyl and the methine 1Hs (Mizoguchi, Furukawa, and Koyama, unpublished work). Figure 1c shows a simplified model of BChl a which was used in the normal-coordinate analysis. The model is based on the following assumptions: (1) All the side chains attached to rings II and IV are replaced by hydrogen atoms. (2) The bacteriochlorin skeleton has D2h symmetry. (3) The carbon-carbon and carbon-oxygen bond lengths were transferred from those determined by X-ray crystallography for methyl bacteriopheophorbide a,13 but those in the bacteriochlorin skeleton were modified by averaging based on the above assumption of D2h symmetry. (4) The aliphatic and olefinic C-H distances were set to be 1.095 and 1.08 Å, respectively. Figure 1b shows the chemical structure of BPhe a; the pair of hydrogen atoms attached to rings I and III has been confirmed by NMR spectroscopy (Sakamoto, Mizoguchi, and Koyama, unpublished). Figure 1d shows a model for BPhe a which is based on that of BChl a. The N-H distance was set to be 0.86 Å. The normal vibrations were calculated by Wilson’s GF-matrix method using Urey-Bradley-Shimanouchi (UBS) force field; non-UBS cross terms were also introduced. Programs BGLZ and LSMB, which had been originally written in Prof. Takehiko Shimanouchi’s laboratory,14 were run on a personal computer (NEC PC9821 Nr166). Results Normal-Coordinate Analysis of the S0, T1, and S1 Raman Spectra. (a) Characterization of Raman Lines. Figure 2a shows
8310 J. Phys. Chem. B, Vol. 104, No. 34, 2000
Figure 1. Chemical structures of (a) BChl a and (b) BPhe a, and the typifications of the five-membered rings and the carbon atoms in the bacteriochlorin skeleton. Models for (c) BChl a and (d) BPhe a which were used in the normal-coordinate analyses are also shown (see the Experimental Section for the details).
Sashima et al.
Figure 3. T1 Raman spectra of (a) BChl a and (b) BPhe a; the (1) NA, (2) 15N, (3) 13C, and (4) 2H species. Each spectrum was obtained by one-color experiment using the 420 nm, 5 ns, and 10 Hz pulses; a pair of spectra was recorded with high and low photon densities, and the difference spectrum was taken.
Figure 4. S1 Raman spectra of (a) BChl a and (b) BPhe a; the (1) NA, (2) 15N, (3) 13C, and (4) 2H species. Each spectrum was obtained by one-color experiment using the 351 nm, ∼50 ps, and 1 kHz pulses as a difference spectrum (see the caption of Figure 3). Figure 2. S0 Raman spectra of (a) BChl a and (b) BPhe a; the (1) NA, (2) 15N, (3) 13C, (4) 13C + 15N, and (5) 2H species. Each spectrum was recorded by the use of the 457.9 nm CW beam.
the S0 Raman spectra of the (1) NA, (2) 15N, (3) 13C, (4) 13C + 15N, and (5) 2H species of BChl a. Figures 3a and 4a show the T1 and S1 Raman spectra of the (1) NA, (2) 15N, (3) 13C, and (4) 2H species; the 13C + 15N species was found to be unstable against pulsed photoexcitation to the T1 and S1 states. The spectra of the NA, 15N, and 2H species were published and empirical assignments of the S0, T1, and S1 Raman lines were given by the use of the Raman spectra of the NA, 15N, and 2H
species; see Figure 5 of ref 8. The newly obtained Raman spectra of the 13C and 13C + 15N species supported this set of empirical assignments. Those assignments were based on the following considerations: (1) In-plane vibrations including carbon-carbon and carbon-nitrogen stretchings in the conjugated macrocycle as well as the methine and the methyl C-H in-plane deformation coupled with those skeletal modes can give rise to resonanceRaman lines. (The Raman intensity of out-of-plane vibrations is expected to be very small.) (2) On the basis of a set of resonance structures of the conjugated macrocycle, the intrinsic frequency of each stretching mode is expected to be in the following order: Ca′-Cm (2/3) > Cb-Cb ) Ca-Cb (1/2) > Ca-
Macrocycles of BChl a and BPhe a Cm ) Ca′-N (1/3) > Ca-N (1/6) > Ca′-Cb′ ) Cb′-Cb′, (0); the numbers in parentheses indicate the probability of finding a double bond in the set of resonance structures.4 (3) Carbonnitrogen stretching Raman lines should be affected by the 15N substitution, whereas the carbon-carbon stretching Raman lines should not. This classification of Raman lines applies not only to the set of the NA and 15N species but also to the set of the 13C and 13C + 15N species. (4) Raman lines due to those normal modes which are coupled with the methine and methyl C-H in-plane bending vibrations should be affected by the 2H substitution, because those vibrations shift to below 1000 cm-1. Sets of empirical assignments of the S0, T1, and S1 Raman lines for the NA species are given, and correlations of the Raman lines among the NA and the four (three) labeled species were made, as far as possible, before starting the normal-coordinate analysis. Figure 2b shows the S0 Raman spectra of the (1) NA, (2) 15N, (3) 13C, (4) 13C + 15N and (5) 2H species of BPhe a, and Figures 3b and 4b show the T1 and S1 Raman spectra of the (1) NA, (2) 15N, (3) 13C, and (4) 2H species. Here again, the doubly-labeled species was found to be unstable against pulsed-laser irradiation. The S0, T1, and S1 Raman spectra of the NA and 15N species have been published, and a set of empirical assignments of the Raman lines have been given.9 Before we started the normal-coordinate analysis, empirical assignments of the S0, T1, and S1 Raman lines were done by using additionally the S0 Raman spectra of the 13C, 13C + 15N, and 2H species and the T and S Raman spectra of 1 1 the 13C and 2H species. (b) Procedure of Normal-Coordinate Analysis. The final goal of the present study is to determine changes, upon triplet and singlet excitation, in the stretching force constants within the macrocycles of BChl a and BPhe a. Therefore, we focused our attention on the following four sets of stretching force constants (see Figure 1): (1) Ca′-Cm and Ca-Cm, (2) Cb-Cb and CaCb, (3) Ca′-N and Ca-N, and (4) Ca′-Cb′ and Cb′-Cb′. In fitting the calculated frequencies to the observed ones, we first assumed that the pair of force constants in each set was the same, and then, we split a particular set into two independent force constants when it became absolutely necessary. The reason for doing this was that we had a limited number of the S0, T1, and S1 Raman lines (as observables) in the 1650-1200 cm-1 region which we mainly focused on. The rest of the stretching force constants concerning the Cm-H, C-H, CdO, and C-C bonds in the peripherals as well as the central Mg-N bond were transferred from those of nickel(II) octaethylporphine (NiOEP)15 and Chl a.10 The bending and nonbonded repulsive force constants in the Urey-Bradley-Shimanouchi force field as well as the non-UBS, stretching-stretching cross terms were transferred from those of Ni-OEP15 and Chl a.10 Some of the cross terms were adjusted in the final refinement of the frequencies and the patterns of normal modes. An initial guess of the above four sets of S0-state force constants was made by two different methods: (a) A linear relation was deduced between the π-bond orders estimated by the resonance structures and the stretching force constants that were determined by previous normal-coordinate analyses for Ni-OEP,15 Chl a,10 β-carotene (Mukai et al., unpublished work), and retinal.16 Then, an initial set of stretching force constants was chosen, using the bond orders estimated by the use of resonance structures (vide supra). (b) A linear relation was deduced, for the same set of compounds listed above, between the carboncarbon bond lengths determined by X-ray crystallography and
J. Phys. Chem. B, Vol. 104, No. 34, 2000 8311 the corresponding carbon-carbon stretching force constants determined by previous normal-coordinate analyses. Then, an initial set of carbon-carbon stretching force constants was chosen by the use of the bond lengths listed in Figure 1. The carbon-nitrogen stretching force constants were transferred from those of Ni-OEP and Chl a, whose values were more or less similar irrespective of bond lengths. The initial set of force constants “b” gave rise to much better results, a fact which lead us to a conclusion that the initial set “a” based on the resonance structures tends to overestimate difference in bond order among the carbon-carbon and carbonnitrogen bonds in the macrocycle, even though their order in bond order may be correct. In picking up those normal modes that are responsible for the observed resonance-Raman lines, out of a number of calculated normal modes, the following assumptions have been made: (i) The carbon-carbon and the carbon-nitrogen stretching modes in the conjugated bacteriochlorin skeleton, in general, can give rise to resonance-Raman intensity. Among them, those normal modes whose displacements of atoms exhibit the center of symmetry should give rise to the highest Raman intensity; actually, those normal modes gave high Raman intensity when probed in resonance with the Soret absorption.17 (ii) When the skeletal vibrations are coupled with each other to form a normal mode exhibiting C2 symmetry, in relation to either the axis connecting rings I and III or the axis connecting rings II and IV, can give rise to medium or weak Raman lines. (iii) A pair of localized symmetric modes, which overlap each other, can give rise to medium or weak Raman lines. (iv) The methine C-H in-plane bending vibrations [hereafter denoted as δ(Cm-H)] and the methyl (C-H) in-plane degenerate deformation and symmetric deformation vibrations (hereafter denoted “dd” and “sd”, respectively) can also appear in the resonance-Raman spectrum, when they are coupled with the skeletal stretching vibrations. Correlating such calculated “Raman-active” modes with the observed Raman lines, we tried to fit their frequencies by adjusting the carbon-carbon and the carbon-nitrogen stretching force constants in the bacteriochlorin skeleton. First, we have determined those stretching force constants of S0 BChl a; here, we could obtain reasonably good agreement between the observed and the calculated frequencies, keeping the pair of force constants in each set (1-4) the same (vide supra). Second, we adjusted the force constants of T1 BChl a starting from those of S0 BChl a. In this process, we had to split set 4 force constants into two independent parameters, and to shift the Ca′-Cb′ force constant to a higher value, leaving that of Cb′-Cb′ as in the S0 state. Finally, we adjusted the force constants in S1 BChl a starting from those of T1 BChl a. Here, it was necessary to make the Cb-Cb stretching force constant independent from the CaCb stretching force constant in set 2, and to shift the former to a very high value. The stretching force constants of BPhe a were determined in a similar way: First, we determined the force constants of S0 BPhe a by transferring those of S0 BChl a and by slightly modifying the set 2 force constants. Second, we determined the force constants of T1 BPhe a by modifying all the force constants of T1 BChl a. Finally, we determined the force constants of S1 BPhe a starting from those of S1 BChl a. Here, it was necessary to change the Cb-Cb stretching force constant to a much higher value, and to return the Ca′-Cb′ force constant to the initial value in the S0 state. We kept the Cb′-Cb′ stretching force constants unchanged throughout all the normal-coordinate analysis.
8312 J. Phys. Chem. B, Vol. 104, No. 34, 2000
Sashima et al.
Figure 5. A set of carbon-carbon and carbon-nitrogen stretching force constants determined for (a) BChl a and (b) BPhe a in the S0, T1, and S1 states (see Figure 1 for the typification of the carbon atoms in the bacteriochlorin skeleton). Four sets of force constants, i.e., (i) Ca′-Cm ) Ca-Cm, (ii) Cb-Cb ) Ca-Cb, (iii) Ca′-N ) Ca-N, and (iv) Ca′-Cb′ ) Cb′-Cb′ were assumed in the S0 state, and sets (ii) and (iv) were split into two independent force constants when it became necessary.
Assignments of Raman Lines in the S0, T1, and S1 States. (a) BChl a. Figure 5a depicts the sets of stretching force constants determined by the present normal-coordinate analyses of the S0, T1, and S1 Raman spectra, and Figure 6a compares the calculated frequencies (broken lines) with those of the Raman lines (solid lines) for the NA, 15N, 13C, and 2H species in the S0, T1, and S1 states. Agreement between the calculated and the observed frequencies is satisfactory for all the isotope species in the three different electronic states, when we consider the small number of stretching force constants used as fitting parameters. Some disagreement is seen when complicated coupling with vibrations outside of the bacteriochlorin skeleton is taking place. Larger disagreement is seen for the 2H species, where the pattern of coupling is completely changed because of the shifts of all the C-H in-plane bendings to below 1000 cm-1. Further refinement using a larger number of force constants is absolutely necessary to obtain better agreement between the observed and calculated frequencies. However, we believe that the present stage of normal-coordinate analysis suffices to assign the observed Raman lines in the 1650-1200 cm-1 region, and to characterize changes in bond order upon singlet and triplet excitation. Tables 1, 3a, and 4a list the assignments of the Raman lines in the S0, T1, and S1 states, respectively, for the NA and the variously isotope-substituted species of BChl a. Figure 7a depicts, for the NA species, typical normal modes giving rise to strong or medium Raman lines in the three electronic states. The Raman lines of S0 BChl a shown in Figure 2 can be assigned as follows (detailed description will be given only for the NA species with occasional reference to the 2H species): The strongest Raman line at 1611 cm-1 can be assigned to a
pair of modes consisting of the Ca-Cm asymmetric and the CaCb symmetric stretchings taking place on both the ring I and ring III sides; the pair of modes is coupled in-phase to form an ag-type normal mode (Figure 7a). This mode has been referred to as “the ring-breathing mode”, whose frequency reduces on going from the penta- to the hexa-coordinated state, due to the expansion of the 16 -membered ring (see ref 9 for a review). Thus, the basis for such naming is now established. The normalmode analysis predicts the presence of another mode, in which the Ca-Cm asymmetric stretching is coupled with the Ca-Cb asymmetric stretching, instead; this mode showing a C2 symmetry is ascribable to the shoulder on the low-frequency side of the above ring-breathing Raman line. The second strongest Raman line at 1533 cm-1 can be assigned to a pair of localized Cb-Cb stretchings overlapping each other. A series of very weak Raman lines in the 1500-1400 cm-1 region can be associated with the Ca-Cm symmetric stretching, the Ca-Cb asymmetric and symmetric stretchings, and the methyl degenerate and symmetric deformations, as listed in the table. A pair of weak Raman lines at 1361 and 1340 cm-1 can be assigned to the Ca′-N symmetric and asymmetric vibrations, respectively, whereas a medium Raman line at 1288 cm-1 can be assigned to the Ca-N symmetric stretchings coupled in-phase (Figure 7). All those modes are coupled further with δ(Cm-H). The isotope substitutions affect the normal modes as follows: The 15N substitution little affects the pattern of normal mode, but selectively shifts, to the lower frequencies, those normal modes that are associated with the carbon-nitrogen stretchings. The 13C-substitution pushes, to large amounts, all the carbon-carbon and the carbon-nitrogen stretchings to the lower frequencies, but the patterns of the normal modes are
Macrocycles of BChl a and BPhe a
J. Phys. Chem. B, Vol. 104, No. 34, 2000 8313 lines. All those changes upon those isotope substitutions can be seen for all the S0, T1, and S1 Raman lines of both BChl a and BPhe a. The Raman lines of T1 BChl a can be assigned as follows (see Figure 3a, Table 3a, and Figure 7a): The strongest Raman line at 1587 cm-1 can be assigned, as in the case of the S0 state, to the ag-type, ring-breathing mode consisting of the Ca-Cm asymmetric stretchings and the Ca-Cb symmetric stretchings. The weak 1533 cm-1 Raman line, at the foot, can be assigned to a pair of localized Cb-Cb stretchings. The very weak 1477 cm-1 Raman line and a stronger 1409 cm-1 Raman line can be assigned to the Ca-Cm symmetric stretchings coupled with the Ca-Cb asymmetric stretchings. The broad 1324 cm-1 Raman line with medium intensity can be assigned to a pair of localized Ca-N symmetric stretchings which overlap each other. A very weak 1263 cm-1 Raman line can be assigned to the Ca′-Cb′ symmetric stretching coupled with δ(Cm-H). The Raman lines of S1 BChl a can be assigned as follows (see Figure 4a, Table 4a, and Figure 7a): The Raman line at 1568 cm-1 can be definitely assigned to the ag-type, ringbreathing mode because of its high Raman intensity. The Raman line at 1629 cm-1 on the high-frequency side can be assigned to an overlap of the localized Cb-Cb stretchings; its much lower Raman intensity can be explained in terms of such localized vibrations. The weak 1406 cm-1 Raman line is ascribable to a coupled mode consisting of the Ca-Cm symmetric and the CaCb asymmetric stretchings, whereas the strong 1318 cm-1 Raman line to a pair of localized Ca-N symmetric stretchings. The δ(Cm-H) mode is coupled with both of those normal modes. (b) BPhe a. Figure 5b depicts the sets of force constants determined, and Figure 6b compares the calculated and the observed frequencies for the NA, 15N, 13C, and 2H species of S0, T1, and S1 BPhe a. Agreement between the calculated and the observed frequencies for the Raman-active normal modes in the 1650-1200 cm-1 region is more or less similar to the case of BChl a. Tables 2, 3b, and 4b list the assignments of the Raman lines of S0, T1, and S1 BPhe a, respectively. Figure 7b depicts typical normal modes giving rise to high or medium Raman intensity. The Raman lines of S0 BPhe a shown in Figure 2b can be assigned as follows: The strongest Raman line at 1612 cm-1 can be assigned to the ag-type ring-breathing mode, i.e., the CaCm asymmetric stretching coupled with the Ca-Cb symmetric stretching, a pair of which is coupled further in-phase. A medium Raman line at 1586 cm-1, on the shoulder, can be assigned to the Ca-Cm asymmetric stretching coupled with the Ca-Cb asymmetric stretching, a pair of which is coupled in-phase (this mode now appears as a split Raman line). A weak, broad Raman line at 1542 cm-1 can be assigned to a pair of localized Cb-Cb
Figure 6. Comparison of the observed and the calculated frequencies of the Raman lines of (a) BChl a and (b) BPhe a in the 1650-1200 cm-1 region for the NA, 15N, 13C, and 2H species in the S0, T1, and S1 states. The solid lines indicate the observed Raman lines (the length of each line is proportional to peak intensity), whereas the broken lines indicate the calculated frequencies (their lengths are adjusted to those of the observed). In some cases, a single, broad observed Raman line is ascribed to a pair of independent modes.
conserved except for that of the 1445 cm-1 Raman line. Here again, the (13C + 15N) substitution selectively shifts, further to the lower frequencies, those normal modes which are related to the carbon-nitrogen stretchings. The 2H substitution decouples all the skeletal stretchings from the methine and the methyl C-H in-plane deformations shifting to below 1000 cm-1. As a result, the normal modes become less crowded, and the localized or delocalized skeletal modes that are coupled either in-phase or out-of-phase appear as independent Raman lines. The pair of modes consisting of the Ca-Cm asymmetric stretching that is coupled with the Cb-Cb symmetric and asymmetric stretching, for example, now appears as split Raman
TABLE 1: Assignments of the S0 Raman Lines for the Unlabeled (NA) and Totally 15N-, 13C-, 13C + 15N-, and 2H-Labeled BChl a NA
15N
1611
1611
1533 1493 1463 1450 1424 1361 1340 1288
assignment
13C
(0)
Ca-Cm asym + Ca-Cb sym
1557
1531 1494 1461
(2) (+1) (2)
1426 1391
(+2)
Cb-Cb Ca-Cm sym Ca-Cm sym + Ca-Cb asym Ca-Cm sym + Ca-Cb sym Ca-Cm sym + Me dd Me sd
1355 1330 1278
(6) (10) (10)
Ca′-N sym + δ(Cm-H) Ca′-N asym + δ(Cm-H) Ca-N sym + δ(Cm-H)
1331 1306 1262
(∆)
13C
+ 15N
(∆)
assignment
2H
assignment
1559
(+2)
Ca-Cm asym + Ca-Cb sym
1603
1491
1491
(0)
Cb-Cb
1445 1416
1442
(3)
Me dd + Ca-Cm sym Ca-Cm sym + Ca-Cb sym
1385
1380 1347 1316 1294 1250
(5)
Me sd Ca-N asym Ca′-N sym + δ(Cm-H) Ca′-N asym + δ(Cm-H) Ca-N sym + δ(Cm-H)
1581 1533 1498 1442 1364 1337
Ca-Cm asym + Ca-Cb sym (in-phase) Ca-Cm asym + Ca-Cb asym Cb-Cb Cb-Cb Ca-Cm sym + Ca-Cb asym Ca′-N sym (in-phase) Ca′-N sym (out-of-phase)
1318 1263 1234
Ca-N sym (out-of-phase) Ca-N sym (in-phase) Ca′-N asym
(15) (12) (12)
8314 J. Phys. Chem. B, Vol. 104, No. 34, 2000
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TABLE 2: Assignments of the S0 Raman Lines for the Unlabeled (NA) and Totally 15N-, 13C-, 13C + 15N-, and 2H-Labeled BPhe a 13C
+15N
NA
15N
(∆)
assignment
13C
1612 1586 1542 1492 1465 1448 1426
1610 1582 1541 1490 1462 1441 1415
(2) (4) (1) (2) (3) (7) (11)
Ca-Cm asym + Ca-Cb sym Ca-Cm asym + Ca-Cb asym Cb-Cb Ca-Cm sym Ca-Cm sym + Ca-Cb sym Ca-Cm sym + Me dd Ca-N asym
1557 1493 1449 1431 1393 1334
1557 1535 1487 (1450) 1431 1388 1326
1343 1337 (6)
Ca′-N asym + δ(Cm-H)
1312
1294
(18)
Ca′-N asym + δ(Cm-H)
1298 1291 (7) 1218 1212 (6)
Ca-N sym + δ(Cm-H) δ(N-H)
1270 1200
1259 1196
(11) (4)
Ca-N sym + δ(Cm-H) δ(N-H)
(∆)
assignment
Ca-Cm asym + Ca-Cb sym Ca-Cm asym + Ca-Cb asym (6) Cb-Cb (+1) Ca-Cm sym + Me dd (0) Me dd (5) Ca-Cm sym + Ca-Cb asym (8) Ca′-N sym + δ(Cm-H) (0)
2H
assignment
Ca-Cm asym + Ca-Cb sym Ca-Cm asym + Ca-Cb asym Cb-Cb Ca-Cm sym + Ca-Cb asym Ca-Cm sym + Ca-Cb asym Ca-N asym Ca-N sym + Ca′-N sym (out-of-phase) 1291 Ca-N sym + Ca′-N sym (in-phase) 1182 δ(N-H)
1599 1574 1541 1480 1453 1432 1312
TABLE 3: Assignments of the T1 Raman Lines for the Unlabeled (NA) and Totally 15N-, 13C-, and 2H-Labeled (a) BChl a and (b) BPhe a NA
15N
(∆)
assignment
1587 1533 1477 1409 1324 1263
1587 1534 1478 1410 1313
(0) (+1) (+1) (+1) (11)
Ca-Cm asym + Ca-Cb sym Cb-Cb Ca-Cm sym + Ca-Cb asym Ca-Cm sym + Ca-Cb asym + δ(Cm-H) Ca-N sym + δ(Cm-H) Ca′-Cb′ sym + δ(Cm-H)
1587 1551 1522 1489 1451 1365 1235 1198
1587 1550 1522 1489 1451 1360 1230 1195
(0) (1) (0) (0) (0) (5) (5) (3)
Ca-Cm asym + Ca-Cb sym Ca-Cm asym + Ca-Cb asym Cb-Cb Ca-Cm sym Ca-Cm sym + Ca-Cb asym Ca′-N sym + δ(Cm-H) δ(N-H) δ(N-H)
13C
assignment
(a) BChl a 1537 Ca-Cm asym + Ca-Cb sym 1432
Me dd + Ca-Cm sym + Ca-Cb asym
1292
Ca-N sym + δ(Cm-H)
(b) BPhe 1542 Ca-Cm asym + Ca-Cb sym (1476) Cb-Cb 1428 Me dd 1398 Ca-Cm sym + Ca-Cb asym 1370 Ca-N asym 1338 Ca′-N sym 1261 Ca-N sym 1212 δ(N-H) 1181 δ(N-H)
stretchings. The following three weak Raman lines at 1492, 1465, and 1448 cm-1 can be associated with the Ca-Cm symmetric stretchings and the methyl degenerate deformation as shown in the table. The weak Raman lines at 1426, 1343, and 1298 cm-1 can be assigned to the Ca-N asymmetric, Ca′-N asymmetric, and Ca-N symmetric stretchings, respectively; the latter two modes are coupled with δ(Cm-H). The medium, broad Raman line around 1218 cm-1 can be assigned to an overlap of the N-H in-plane deformations [δ(N-H)], unique vibrations of this BPhe a molecule. The 15N, 13C, (13C + 15N), and 2H substitutions exhibit isotope effects on the S0 Raman spectra as in the case of S0 BChl a. In some cases, interpretation of shifts was not straightforward because of complicated couplings among various types of vibrations. The Raman lines of T1 BPhe a can be assigned as follows (see Figure 3b, Table 3b, and Figure 7b): A very strong 1587 cm-1 Raman line and a weak 1551 cm-1 Raman line can be assigned to the Ca-Cm asymmetric stretching coupled with the Ca-Cb symmetric (the ag-type, ring-breathing mode) and that coupled with the asymmetric stretchings (a mode showing C2 symmetry), respectively. The 1522 cm-1 Raman line is due to the localized Cb-Cb stretchings. Both the 1489 and 1451 cm-1 Raman lines are associated with the Ca-Cm symmetric stretchings, whereas the 1365 cm-1 weak profile with the Ca′-N symmetric stretching. The two split medium Raman lines at 1235 and 1198 cm-1 can be assigned to a pair of δ(N-H). Here, the 15N-substitution clearly distinguishes the last three normal modes which are associated with the carbon-nitrogen stretchings and the N-H in-plane bendings. The 13C and 2H substitutions drastically change the patterns of normal modes in the
2H
assignment
1575 1522
Ca-Cm asym + Ca-Cb sym Cb-Cb
(1353) Ca′-N sym 1323 Ca-N sym + Ca′-N sym 1577 1514
Ca-Cm asym + Ca-Cb sym Cb-Cb
1446
Ca-Cm sym + Ca-Cb asym
1365 1298 1219 1188
Ca′-N sym Ca-N sym δ(N-H) δ(N-H)
low-frequency region as shown in the table. Note that the δ(N-H) Raman lines are conserved, because BPhe a was prepared from BChl a by the use of acetic acid (vide supra). The Raman lines of S1 BPhe a can be assigned as follows (see Figure 4b, Table 4b, and Figure 7b): The medium Raman at 1642 cm-1 can be assigned to a pair of localized Cb-Cb stretchings as in the case of S1 BChl a. Its frequency is further increased in this molecule. The sharp and strong Raman line at 1587 cm-1 and a shoulder at 1565 cm-1 can be assigned, respectively, to the Ca-Cm asymmetric stretchings coupled with the Ca-Cb symmetric stretching (the ag-type ring-breathing mode) and those with the Ca-Cb asymmetric stretching (a mode showing C2 symmetry), respectively. The weak 1398 cm-1 Raman line is tentatively associated with the methyl symmetric deformation because no other candidates were found in this region. A pair of medium Raman lines at 1337 and 1294 cm-1 can be assigned to the Ca′-N asymmetric and the Ca-N symmetric stretchings, both being coupled with δ(Cm-H). A pair of weak Raman lines at 1228 and 1187 cm-1 are assigned to δ(N-H). Typical isotope effects are seen in this case. Table 5 compares four sets of assignments of the S0 Raman lines of BChl a given by Lutz,18 Donohoe et al.,17 Hu et al.,19 and the present work. The frequencies of the S0 Raman lines determined in the present investigation are used in the list; the assignment of normal mode to each Raman line is given not in terms of ped (potential energy distribution) but in terms of Lx matrix (displacements of atoms). Those vibrations that are found in common in all the sets of assignments are shown in italic in Table 5. When one takes into account the fact that the three sets of assignments have been obtained on completely different bases, the agreement can be considered to be satisfactory.
Macrocycles of BChl a and BPhe a
J. Phys. Chem. B, Vol. 104, No. 34, 2000 8315
Figure 7. Typical normal modes of (a) BChl a and (b) BPhe a giving rise to high or medium Raman intensity in the S0, T1, and S1 states. The frequencies of the observed Raman lines and their assignments in abbreviated forms are shown (see Tables 1-4 for the details).
Changes in the Stretching Force Constants upon Triplet and Singlet Excitation of BChl a and BPhe a. Now, we are going to compare the sets of force constants of (a) BChl a and (b) BPhe a among the S0, T1 and S1 states (see Figure 1 and Figure 5). In the process of normal-coordinate analyses, it was necessary to split, not set 1 (Ca′-Cm and Ca-Cm) or set 3 (Ca′-N
and Ca-N) force constant, but set 2 (Cb-Cb and Ca-Cb) and set 4 (Ca′-Cb′ and Cb′-Cb′) force constants (vide supra). Therefore, in the following discussion, we will refer to set 1 (set 3) force constants inclusively as Ca-Cm (Ca-N), and to set 2 (set 4) force constants separately as Cb-Cb and Ca-Cb (Cb′-Cb′ and Ca′-Cb′).
8316 J. Phys. Chem. B, Vol. 104, No. 34, 2000
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TABLE 4: Assignments of the S1 Raman Lines for the Unlabeled (NA) and Totally 15N-, 13C-, and 2H-Labeled (a) BChl a and (b) BPhe a NA
15N
1629 1568 1406 1318
1628 1568 1405 1312
1642 1587 1565 1398 1337 1294 1228 1187
1642 1586 1563 1398 1332 1289 1235 (1182)
13C
(∆) (1) (0) (1) (6)
Cb-Cb Ca-Cm asym + Ca-Cb sym Ca-Cm sym + Ca-Cb asym + δ(Cm-H) Ca-N sym + δ(Cm-H)
(0) (1) (2) (0) (5) (5) (+7) (5)
Cb-Cb Ca-Cm asym + Ca-Cb sym Ca-Cm asym + Ca-Cb asym Me sd Ca′-N asym + δ(Cm-H) Ca-N sym + δ(Cm-H) (N-H) + δ(Cm-H) δ(N-H)
2H
assignment
(a) BChl a 1597 Cb-Cb 1519 Ca-Cm asym + Ca-Cb sym 1350 Ca-Cm sym + Ca-Cb asym + δ(Cm-H) 1271 Ca-N sym + δ(Cm-H)
1631 1559 1428 1318
Cb-Cb Ca-Cm asym + Ca-Cb sym Ca-Cm sym + Ca-Cb asym Ca-N sym + Ca′-N sym
1630 1575 1553 1427 1349 1271 1197 1156
Cb-Cb Ca-Cm asym + Ca-Cb sym Ca-Cm asym + Ca-Cb asym Ca-Cm sym + Ca-Cb asym Ca-N asym + Ca-Cm sym Ca-N sym δ(N-H) δ(N-H)
(b) BPhe a 1541
Ca-Cm asym + Ca-Cb sym
1386 1312 1258 1230 1164
Ca-Cm sym + Me sd + Ca-Cb asym Ca′-N asym + Ca-N asym Ca-N sym + δ(Cm-H) δ(N-H) + δ(Cm-H) δ(N-H)
assignment
TABLE 5: Comparison with the Results of Previous Normal-Coordinate Analyses for NA BChl a in the S0 Statea Raman freqb
Lutzc
Donohoe et al.d
Hu et al.e
present work
1611 1533 1493 1463 1450 1424 1361 1340 1288
Ca-Cm + [Cb-Cb, δ(Cm-H)] Cb-Cb C-C C-C C-C C-C C-N δ(Cm-H) C-N
Ca-Cm asym Cb-Cb + Ca-Cm Ca-Cb asym + Ca-Cm sym Ca′-N asym Ca-Cm sym Ca-Cb asym + Ca-Cm Ca′-N + Ca-Cb Ca′-N + δ(Cm-H) Ca-N sym + δ(Cm-H)
Ca-Cm asym + Cb-Cb Cb-Cb + Ca-Cm asym Ca-Cb asym + Ca-N asym Cb-Cb sym + Ca-Cm sym Ca-Cm sym + Ca′-N sym Ca-Cm sym Ca′-N sym + δ(Cm-H) Ca′-N asym + δ(Cm-H) Ca-Cb sym + δ(Cm-H)
Ca-Cm asym + Ca-Cb sym Cb-Cb Ca-Cm sym Ca-Cm sym + Ca-Cb asym Ca-Cm sym + Ca-Cb sym Ca-Cm sym + Me dd Ca′-N sym + δ(Cm-H) Ca′-N asym + (Cm-H) Ca-N sym + δ(Cm-H)
a Normal modes are described in terms of L matrics (displacements of atoms). b Frequencies determined in the present ivestigation. c Reference x 18. d Reference 17. e Reference 19.
(a) BChl a. In the S0 state, the stretching force constants are in the order, Ca-Cm > Cb-Cb ) Ca-Cb > Ca-N > Ca′-Cb′ ) Cb′-Cb′. On going from the S0 to the T1 state, both the CaCm and the Cb-Cb ) Ca-Cb stretching force constants decrease, whereas the Ca-N stretching force constant increases. A large increase in the Ca′-Cb′ stretching force constant also takes place. On going further to the S1 state, all the Ca-Cm, the Ca-Cb and the Ca-N stretching force constants decrease, whereas only the Cb-Cb stretching force constant increases drastically. In those changes in the values of stretching force constants, a compensation effect between increase and decrease is expected to take place, because triplet and singlet excitation is caused by the transition of a single π-electron from the HOMO to the LUMO out of 18 π- and 30 σ-valence electrons in the conjugated system that remain unchanged. Actually, an average value of the eight stretching force constants of this molecule (28 bonds) is 3.94, 4.08, and 4.03 in the S0, T1, and S1 states, respectively. The maximum difference, 3% (0.14/4.02), is probably within the limit of error in the process of fitting those force constants. (b) BPhe a. In the S0 state, the values of stretching force constants are exactly the same as those of BChl a except for the Cb-Cb ) Ca-Cb stretching force constants that are slightly increased. On going to the T1 state, changes in the stretching force constants similar to those in BChl a take place, although their values are varied; the Ca-Cm and the Cb-Cb ) Ca-Cb stretching force constants decrease, whereas the Ca-N and the Ca′-Cb′ stretching force constants increase. On going further to the S1 state, basically similar but more drastic changes in the force constants take place, when compared to the case of BChl a. The Ca-Cm stretching force constant is not changed at all; the Cb-Cb stretching and the Ca-Cb stretching force constants increase in a large and a small amount, respectively; and the Ca-N and the Ca′-Cb′ stretching force constants decrease substantially (the latter returns to the value in the S0 state). An average of the stretching force constants is 3.96, 3.99, and 3.93 in the S0, T1, and S1 states, respectively, in this
molecule. The maximum difference,
