10234
J. Phys. Chem. 1992,96, 10234-10240
(31) (a) Wicchmann. M.; Port, H.; Laermer, F.; Frey, W.; Elsaesser, T. Chem. Phys. Lett. 1990,165,28-34. (b) Wiechmann, M.; Port, H.;Frey, W.; Laermer, F.; Elsaesser, T. J. Phys. Chem. 1991, 95, 1918-1923. (32) W a n , D. F.; Huston, A. L.; Scott, G. W. J. Chem. Phys. 1983, 79, 5802-5807. (33) Huston, A. L.; Scott, G. W. J . Phys. Chem. 1987,91. 1408-1413. (34) Gustav, K.; Colditz, R. 2.Chem. 1988, 28, 309-315. (35) Gustav, K., private communication. (36) Stewart, J. M.; Machin, P.A.; Dickinson, C.; Ammon, H. L.; Heck, H. S.;Flack, H. XRAY System - Version 61976; Technical Report TR-446, Computer Science Center: University of Maryland, College Park MD, 1976.
(37) Allen, F. H.; Kennard, 0.; Watson, D. G.; Brammer, L.; Orpen, A. G.; Taylor, R.J. Chem. Sm., Perkin Trans. 2 1987, Sl-Sl9. (38) Pimentel, G. C.; McClellan, A. L. The Hydrogen Bond, W. H. Freeman: San Francisco, 1960. (39) Lemmert-Schmitt, E., Thesis, University of Stuttgart, 1990. (40)D u d s , J.; KarvaH, M.;Maiiisek, Z . Coll. Czech. Chem. Commun. 1973, 38, 215-223, 224-242, 243-250. (41) Note added in pmfi After submission of the manuscript we obtained information about the work of Greenwood et al. (Greenwood, R.J.; Mackny, M.F.; Wilshire, J. F. K. AWL J. Chem. 1992,45,965-968)describing X-ray analysis of HTBM-TIN. Their results are in good agreement with oun.
Resonance Raman Spectra of Chlorin and Chlorophyll Radical Anions J i i H u e i Perng and David F. h i a n * Department of Chemistry, University of California, Riverside, California 92521-0403 (Received: August 3, 1992; In Final Form: September 16, 1992)
Resonamx Raman (RR) spectra arc reported for the anions Zn(0EC); OEC (OEC = octaethykhlorin), CW (Chl = chlorophyU a), and Pheo- (Pheo = pheophytin a). The reduction of Zn(0EC) and OEC leads to relatively small shifts in the frequencies (most less than 5 cm-l) of the skeletal modes of the macrocycles. These results indicate that reduction of the chlorins does not result in large structural perturbations. In contrast, reduction of Chl and Pheo leads to large shifts in the frequencies of a number of the vibrational modes of the ring. The G-keto group on ring V appears to be the key structural element in facilitatingthese perturbations. The keto stretching modes of Chl and Pheo downshift by 26 and 41 cm-I, rarpeaivcly. The c b c b stretching vibrations of the two rings upshift by 10 cm-I. These observations indicate that reduction of the photosynthetic pigments results in significantstructural and/or electronic perturbationsof the macrocycle. The observation that the and c b c b stretching vibrations undergo relatively large shifts upon duction of the photosynthetic macrocycles suggests that these modes might serve as benchmarks for anion formation in vivo. This should facilitate time-resolved RR studies on photosynthetic proteins. Collectively, the RR studies suggest that the interaction of the C9-ketogroup with the protein matrix could provide a means of mediating conformationalchanges and hence biological redox activity.
-
Introduction
Photosynthetic light-energy conversion involves the transient formation of a (bacterio)chlorophyll+/(bacterio)pheophytinradical pair. Accordingly, the structural and electronic characteristics of these radicals must influence the process of charge separation.I4 In order to gain a better understanding of the physical properties of tetrapyrrolic radicals, we and others have been using resonance Raman (RR) spectroscopy to examine t h e ~ p e c i e s . ~ Most - ~ ~ vibrational studies have focused on cation radicals and, as a consequence, the effects of oxidation are reasonably well characterized.524 In contrast, only a few RR studies have been reported for the anion radicals of tetrapyrr~les.~“’~ In order to assess the effects of reduction on the properties of tetrapyrrolic macrocycles,we have been performing a systematic RR investigation of the anion radicals of a variety of ring systems?”’ Recently, we reported the results of our studies on the porphyrinic anions O E P and Zn(0EP)- (OEP = octaethylporphyrin).” Herein, we extend these vibrational studies to the anions of the structurally related chlorin system OEC- and Zn(0EC)- (OEC = octaethylchlorin) and to the anions of the photosynthetic pigments Pheo- and Chl- (Chl = chlorophyll a; Pheo = pheophytin a ) . The structures of these macrocycles are shown in Figure 1. As far as we know, no RR studies have been reported for any of these anionic species. Collectively, our RR studies of OEC, Zn(OEC)-, Pheo-, and Chl- provide new insights into the structural and electronic properties of these macrocyclic anions. The RR data for the anions of the photosynthetic pigments also serve as benchmarks for the characterization of transient radicals in vivo via time-resolved RR spectroscopy.
Experhntrl Section tram-OEC was prepared from OEP (Midcentury Chemicals, Posen, IL)according to the method of Whitlock et al.33Zn(0EC) was prepared by reacting OEC with zinc(I1) acetate according 0022-3654/92/2096-10234303.00/0
to the method of Stolzenberg and Ster~hic.’~ Chl was isolated from spray-dried Spirulina maxima algae (Earthrise Co., San Rafael, CA) by the method of Smith et The crude product was dissolved in a minimal amount of dichloromethane and chromatographedon neutral alumina (Aldrich). The elution was initiated with n-hexane and then followed with dichloromethane to remove the yellow products. The major green band was collected by using 0.55% (v/v) propanol in dichloromethane as the eluting solvent. Normally, this procedure was repeated in order to obtain pure Chl. Pheo was prepared by systematically adding drops of concentrated hydrochloric acid to a solution of Chl in ethyl ether.36 The demetaiation was monitored by UV-vis absorption spectroscopy. Pheo was purified by using the same procedure as that used for Chl. The various anions were prepared in a Vacuum Atmospheres Model HE43 glovebox equipped with a Model 493 Dri-Train by using electrochemicalinstrumentation described elsewhere.29 The solvents for the reductions were butyronitrile (purified by the method of Van Duyne and Reilly”) and DMF (purified by the method of Fajer et with -0.1 M tetrabutylammonium hexafluorophosphate(Kodak, twice recrystallized from absolute ethanol and dried at 110 OC in vacuo) serving as the supporting electrolyte. The electrochemicalstudies on OEC and Zn(0EC) were perfonned in butyronitrile whereas the studies on Pheo were conducted in DMF. The studies on Chl were conducted in both solvents. The integrity of the anions was confirmed by cyclic voltammetry, coulometry, and UV-vis absorption spectroscopy. The RR spectra of the neutral and anionic species were obtained on instrumentation previously described.29 The spectra of the neutral species were acquired for samples in rigorously degassed dichloromethane (Fisher, freshly distilled from calcium hydride under argon) solutions. The samples were housed in NMR tubes and spun to prevent photodecomposition. The spectra of the anions were obtained in situ in an air-tight electrochemical cell. The laser 0 1992 American Chemical Society
Spectra of Chlorin and Chlorophyll Radical Anions
\
The Journal of Physical Chemistry, Vol. 96, No. 25, 1992 10235
/
gg
OEC
Pheo
300
nm
800 300
nm
800
300
nm
800 300
nm
800
\\
'
"/
A
H
i
H Y
coophvtvl k&* Chl Figure 1. Structures of OEC, Zn(OEC), Chl, and Pheo.
Zn(0EC)
power at the sample was was -4 cm-'.
-
15 mW, and the spectral slit width
A Ekctrocbemirrtry. The redox potentials measured for OEC, Zn(OEC), and Pheo were identical to those previously reportd.YJ9.40 In the case of Chl, the electrochemical behavior is highly solvent dependent" as has been found to be the case for bacteriochlorophyll.'2 In butyronitrile, Chl exhibits one reversible oxidation at 0.71 V and three irreversible reductions at -1.15, -1.36, and -1.57 V. However, in DMF, two different cyclic voltammograms can be obtained depending on the direction of scanning. If the initial scan is cathodic, two reversible reductions are observed at -1.06 and -1.50 V and one irreversible oxidation occurs at 0.7 V. Alternatively, if the initial scan is anodic, one irreversible oxidation occurs at 0.7 V and three reductions are observed, one irreversible at -0.85 V and two reversible at -1.06 and -1.50 V. B. AWrptioa Spcctn. The absorption spectra of OEC, Zn(OEC), Pheo,Chl, and their anions are shown in Figure 2. The general featobserved in the spectra of OEC- and Zn(0EC)-, which include low-intensity, split Soret bands accompanied by weaker shoulders on the low-energy side and bands in the visible region that are not observed in the parent neutrals, are similar to those observed in the absorption spectra of the OEP- and Zn(OEP)-31 (and other metalloporphyrin anim39.These general features are also characteristic of Chl- and Pheo-, although the multiple bands observed in the visible region are not well resolved for these For all the anions investigated here, new bands are also ohpemd in the 7WOCLnm region. These bands are quite pronounced for OEC- and Zn(0EC)- but less so for Chl- and Pheo-. These features are not due to phlorins which also exhibit absorptions in the 7WOCLnm The red absorptionbands of these latter species are significantly broader than those of the anions shown in Figure 2. The absorption spectrum of the electrochemical reduction product of Chl in butyronitrile is shown in Figure 3. The a b sorption spectrum obtained in this advent is considerablydifferent from that observed in DMF. The spectrum is also different from that of neutral Chl. In some respects, the absorption spectrum of the reduction product in bytyronitrile appears to be a composite spectrum of Chl and Chl-. However, the absorption spectrum of the former species exhibits at 584 and 542 nm whereas neither Chl nor Chl- exhibits bands at these wavelengths. This observation suggests that the reduction product generated in butyronitrile is different from either Chl or Chl-; consequently,
H g m 2. Rarm-temperatun absorption spectra of Zn(0EC). OEC, Chl,
Pheo,and their electrochemicallygenerated anions. The solvents for the neutral and anion complexes are dichloromethaneand butyronitrile, respectively, for both Zn(0EC) and OEC. The solvent is DMF for both the neutral and anion complexes of Chl and Pheo. The ordinates for the spectra of the anions are scaled by the factors indicated.
300
425
550
a
0
Wavelength (nm) Flgm 3. Room-temperatureabsorption spectra of Chl and compound X in butyronitrile.
this species was designated compound X. C. Rerolrrsce Rimrn Spectra and Vibmtiolul hignmeots. 1. =(ON), OEC, md 'Lbcir Adoar. The high-frequency regions of the B-8tate-cxcitation (x,= 406.7 nm) RR spectra of Zn(0Ec) and Zn(0EC)- are shown in Figure 4. The frequencies and assignmentsfor the prominent RR bands of these complexesare Summanzed ' in Table I. The assignmentsfor Zn(0EC) are taken from our previous work on this complex and CU(OEC).~The adgmlcnrs for the RR bands of zn(0EC)- were made by analogy to thaw of the neutral complex. These assignments are relatively straightforward because reduction results in frequency shihp that are in general less than 5 cm-'. The largest shifts are observed for modes in the 1350-1450-cm-' region. The vibrational eigenvectors for these modes amtain contributions from a number of internal COOrdinBtCO including C,N, C,Cb and C,C, stretching
10236 The Journal of Physical Chemistry, Vol. 96, No. 25, 1992
Perng and Bocian
OEC-
I Y 900
1100
1300
1500 1700 Raman Shift (cm- 1) Figure 4. High-frequency regions of the Estate-excitation (X, = 406.7 nm) RR spectra of Zn(0EC) and Zn(0EC)-. The solvents for the neutral and anion complexes are dichloromethane and butyronitrile, respectively. The anions were generated electrochemically with tetrabutylammonium hexafluorophosphate serving as the supporting electrolyte. The spectra of the anions were obtained in situ in an air-tight electrochemical cell (see text).
TABLE I: Frequencies (em-') of the Promhat RR Biads of Zn(OEC), om llld Their Anions assimment" Zn(0EC) Zn(0EC)- b~ OEC OEC1629 1629 1620 -3 1602 1605 1617 1584 1582 -2 1572 1571 0 1569 1566 1571 1535 -2 1541 1546 1537 1491 -3 1505 1498 1488 1484 1482 1485 1455 -8 1464 1466 1463 -3 1395 1391 1382 1385 -3 1366 1368 1366 1369 -6 1354 1360 1347 -2 1349 1349 1334 1331 1322 2 1318 1320 1263 1 1259 1260 1262 0 1129 1130 1141 1141 1136 1115 1116
I 900
I!
OEC
1100
1300
1500
i 1700
Raman Shift (cm-l)
Figure 5. High-frequency regions of the B-state-excitation (X, = 406.7 nm) RR spectra of OEC and O E C . The solvents for the neutral and anion complexca are dichloromethane and buty-ronitrile, rcapcctively. The anions were generated and their RR spectra acquired as described in the legend of Figure 4.
b,3 -3 5 -7 3 2 -4 2 -3
1 1 1
OTaken from ref 44.
modes. Inspection of Figure 4 reveals that reduction of Zn(0EC) does not result in the appearance of any prominent new bands in the RR spectrum although some new features are observed. For example, bands due to both of the two highest frequency C,C, stretching modes are observed in the spectrum of Zn(0EC)- (1617 and 1629 cm-')whereas only a single prominent band is obstrved for Zn(0EC) (1620 cm-'). The 1620-cm-' band of the neutral complex does, however, exhibit a slight shoulder on the low-frequency side which suggests that the bands due to the two C,C, stretching modes of this species may be nearly coincident. The RR spectra of OEC and OEC-arehwnin F i i 5. The frequencies and assignments for the prominent RR bands of these macrocycles are summarized in Table I. To date, no detailed vibrational studies or normal-coordinate analysis have been reported for OEC. Consequently, the vibrational assignments for both this complex and OEC- were made by analogy to those of the Zn(1I) complexes. As is the case for Zn(OEC), reduction of OEC does not result in any large frequency shifts; most of the
900
1125
1350
1575
1800
Raman Shift ( c m - ' ) Figure 6. High-frequency regions of the B-state-excitation (h,= 406.7 nm)RR spectra of Chl and Chl-. The solvents for the neutral and anion complexca are dicblmmcthane and DMF,rcapcctively. The anions were generated and their RR spectra acquired as described in the legend of Figure 4. Solvent bands are marked with the letter S.
shifts are less than 5 cm-'.However, unlike Zn(OEC), the largest reduction-induced shifts observed for OEC involve modes above 1500 cm-', which contain predominantly C,C, and CbCb stretching character.& Finally, the RR spectrum of OEC- does not exhibit any prominent new bands with the exception of the intense feature at 1629 cm-l. This band is assigned as the highcat frequency C,C, stretching vibration. As is the case for Zn(OEC), this band is clearly observed in the RR spectrum of the anion but not the neutral species; however, its intensity is significantlylarger for OEC- than for Zn(0EC)-.
The Journal of Physical Chemistry, Vol. 96, No. 25, 1992 10237
Spectra of Chlorin and Chlorophyll Radical Anions
TABLE II: Freqoencics (em-') of the Promiaeet RR lkads of CM, WcqrilldTheirAnioar
assignmenta
Chl
Chl-
&,- Pheo
Pheo-
&/-
-41 -1665' -26 1706 -1665' 1691 0 1623 1614 -10 1623 1624 1604 -1 1602 4 1605 uic;cmj 1606 1577 (sh) -3 1577 (shy -4 1580 1581 u(C,C,) 1572 16 1560 8 1556 Y(CbCb) 1552 1554 (sh) 15 1541 1 1 1539 1530 v(cbcb) -2 1496 1502 6 1496 1498 (sh) u(C,C,) 1480 -5 -12 1485 1476 1488 u(C,C,) v(c,cb) 1376 1384 8 1376 1389 13 1358 -3 1354 -1 1361 1355 b'(c,cb) 1346 -2 0 1348 1342 1342 u(C,N) u(C,N) 1329 1334 5 1328 1225 1220 -5 1221 1223 2 b(C,H) 1188 9 U(C,C~O)1179 1156 1160 4 1158 1161 3 u(C,N) 1142 1139 -3 1149 1153 4 u(C,N) u(CpP0)
vIC.C,)
8
c,
#Taken from refs 45 and 46. 'Partially obscured by solvent peak. Csh= shoulder. 2. Chi, Pbeo, and Their Anions. The high-frequency regions of the B-state-excitation (kx= 406.7 nm) RR spectra of Chl, Pheo, and their respective anions are shown in Figures 6 and 7. The frequencies and assignmentsfor the prominent RR bands of these complexes are summarized in Table 11. The assignments for Chl were taken from our previous work on Ni(I1) Pheo model complexes4sand from the work of Lutz and co-workers on Chl and Pheo.24*46The assignments for the RR bands of Chl- and Pheo- were made by analogy to those of the neutral species. The assignments for the anions can be made with reasonable oonfidence despite the fact that certain RR bands are strongly affected by reduction. The keto stretchingmodes are the most strongly affected. These modes of Chl and Pheo lose RR intensity and downshift by -26 cm-'(1691 to -1665 cm-') and -41 cm-' (1706 to 1665 an-'), respectively. (The bands due to the stretching modes appear as weak shoulders on the 1660-cm-' solvent band.) The CbCb modes of both macrocycles also exhibit substantial reduction-induced shifts. The Chl modes at 1530 and 1552 cm-' upshift to 1541 and 1560 cm-', respectively. The analogous modes of Pheo at 1539 and 1556 cm-'upshift to 1554 and 1572 cm-', respectively. Other modes associated with both the ring skeleton and with the ring V also exhibit substantial reduction-induced shifts. Certain modes downshift, such as the C,C, vibrations at 1486 cm-',whereas others upshift, such as the c,cb vibrations at 1376 Cm-'. The c,,,clo(ring v) stretching vibration of Chl also upshifts upon anion formation (from 1179 to 1188 cm-').The analogue of this mode is not obsmed for either Pheo or Pheo-. 3. Compound X. The RR spectrum of compound X is shown in Figure 8. Although the RR spectrum of this species is qualitatively similar to that of neutral Chl, it is not identical. These difkenca cannot be attributed to solvent effects because the RR spectra of neutral Chl are essentially identical in solvents that yield fivacoordinate speCieS.4' In particular, the 1606- and 1530-cm-' RR bands of Chl are upshifted to 1611 and 1535 cm-' in compound x. spectral simulations indicatethat these features cannot be obtained by superimposing the RR spectra of Chl and Chl-. In addition, a very weak feature is observed in the RR spectrum of the latter species near 1734 CIX-'. Neither Chl nor Chl- exhibits a band in this region. Although the 1734-cm-' band is very weak and could be due to an overtone, it is also possible that the band is due to the carbonyl stretching vibration of the Cl(rcarbomethoxy group. This mode is not observed in the B-state-excitation RR spectrum of Ch124*46 but is observed as a very weak band in the Q,-excitation surface-enhanced RR spectrum.48
-
-
Diacdoa The reduction-induced changes observed in the vibrational spectra of Zn(0EC) for the most part parallel those observed for the Zn(OEP).3' In the case of OEC, there are also a number of
900
1125
1350
1575
1800
Raman Shift (cm-1) Figure 7. High-frequencyregions of the B-stateexcitation (X, = 406.7 nm) RR spcctra of Pheo and Pheo-. The solvents for the neutral and
anion complexes are dichloromethane and DMF, respectively. The anions were generated and their RR spectra acquired as described in the legend of Figure 4. Solvent bands are marked with the letter S. parallels with OEP, although these parallels are not as close as for the Zn(1I) complexes. For all four ring systems, anion formation results in relatively small shifts in the vibrational frequencies. These observations suggest that the structural changes that occur upon anion formation in both the free bases and Zn(1I) complexes of the chlorins and porphyrins are relatively small. In the cast of Zn(OEC), the largest frequency shifts occur in modes whose eigenvectors contain substantial CONstretching contributions whereas, for OEC, modes with predominantly C,C, and c b c b stretching character are more affected by reduction. This same trend is observed for Zn(0EP) versus OEP." The fact that different types of normal modes are affected upon reduction of the Zn(0EP) vmus OEP was suggested to be due to the influence of the positive charge at the center of the former macrocycle. In particular, the presence of the charge would tend to draw electron density away from the periphery and toward the center of the macrocycle. This in turn would result in larger perturbations on modes containing contributions from internal coordinates near the core, such as the CONstretches, than on vibrations containing contributions from internal coordinates located in the periphery Of the such 88 the CbCb stretches. These Same Charge effects appear to be operative in the chlorin system. In the case of the Zn(OEC), reduction results in downshifts of modes containing C,C,, CbCb, and CONstretching character (Table I). This shift pattern is also observed for Zn(OEP).3' These types of vibrations are expected to downshift upon reduction based on the nodal pattern of the e#+orbital (or e +-likeorbital, in the case of the chlorin) in which the unpaired electron resides in the anions. This orbital is predominantly antibonding with respect to the C,C,, CbCb, and CaN bonds.49*50In the cast of OEC, the reduction-induced shifts for certain C,C, and CbCb modes follow the trends expected on the basis of the nodal pattern Of the %*-I&C orbid. HWCVCI, the SbiftS Of OthCI CaC, and CbCb vibrations are anomalous. In particular, the 1602-, 1541- and 1482-, and 1464-cm-' modes upshift upon anion formation rather than downshift. In contrast, the C,C, and c b c b modes of OEP downshift upon ring reduction3' as expected on the basis of the nodal pattern of the e * 0rbital.4~9~ The origin of the anomalous shift pattern observe! for OEC is not clear, although it is most likely due to changes in the composition of the vibrational eigenvectors that occur upon ring reduction.
w,
Pemg and Bocian
10238 The Journal of Physical Chemistry, Vo1. 96, No. 25, 1992
900
11'25
1350
1575
1800
Raman Shift (cm-3
Figure 8. High-frequency regions of the Bstate-excitation (X, = 406.7 nm) RR spectra of Chl and compound X. The solvents for the two complexar are dichloromcthaneand butyronitrile, reapectively. The anions were generated and their RR spectra acquired as described in the legend of Figure 4. Solvent bands are marked with the letter S.
The reduction-induced changes observed in the vibrational spectra of Chl and Pheo are in general similar to one another. These include large down&& of the C p O keto stretching modes and substantial upshifts of the CbCb stretching vibrations. A
numberofotherringskektalmodesofthephotasynthttic~ts also undergo sigdkant reduction-induced shifts (Table 11). The magnitudes of these shifts are in general larger than those of the corresponding modes of the chlorin model systems. Collectively, these mults suggest that much larger structural and/or electronic changes occur upon reduction of Chl or Pheo than do upon reduction of Zn(0EC) or OEC. The principal structural difference between the photosynthetic pigments and the chlwin model systems is the additional isocyclic ring with its conjugating C p O group (Figure 1). The fact that the l a r w d d o n - i n d u d frequency M t sOCCUT for the stretching modes suggests that this group is the key structural feature that differentiates the response of the photosynthetic pigments from that of the chlorin model systems upon addition of an electron. The lower frequencies for the C g 4stretching vibrations of Chl- and Pheo- indicate that the r bond order for this moiety is lower in the anions. This could be due to the contribution of resonance forms such as those shown in Figure 9. In this regard, electronic structure calculations of Chl- and Pheo- predict that substantial electron density (-0.1 of an unpaired spin) resides on the oxygen atom of the Cg-keto group of these two a n i ~ n s . ~This ' ~ ~amount ~ of electron density in the antibonding orbital would substantially lower the frequency of the Cp=O stretching vibration. The frequency shifts of the skeletal vibrations lend additional insight into the structural and electronic perturbations that occur upon reduction of Chl and Pheo. In this regard, the C,C, stretching modes of both rings downshift (with the exception of the 1496-cm-' mode of Pheo) upon reduction. This is consistent with the nodal pattem of the LUMO for the photosynthetic pigments.*52 In addition, the CmCIO stretching mode of Chl upshifts upon ring reduction (Table 11). An upshift in this mode is consistent with an increase in 'Ibond order which is expected from the contribution of the resonance structures for the anions shown in F w r e 9. Despite the consistency in the reduction-induced shift patterns observed for the C 9 4 , C,C,, and CmCIO stretching modes of
Figure 9. Resonance structures of Chl-.
chl and Pheo, the Shifts 0-4 for the c b c b vibrations Of both rings are anomalous. These latter modes undergo sisnificant upshifts whereas they would be predicted to downshift on the baais of the nodal pattern of the LUMO of the ph-ynthctic pigments and the resonance structure3shown in Figure 9. One explanation for this anomalous behavior is that induction due to the i n d negative charge on the oxygen atom of the C+to group removes electron density from the al&e orbital of the anion. This orbital, which is the HOMO of the neutral ring, is antibonding with respect to the CbCb bond~;~O,~' consequently, a loss of electron density Would be CXpCCtCd to Upshift the cbcb Stretches. HOWCVCr, it seems unlikely that this effect would outweigh the effects of the increase in antibonding electron density due to population of the es+-likeorbital. In addition, lass of electron density from the alu-likcorbital would be expected to result in upshiftsfor the C,C, stretches (rather than the observed downshifts). A second possibility is that the relatively law fresucncy for the cpoetrdchiag vibration in the anions results in increased mixing between this vibration and the CbCb Stretches. This Would tend to k l W S C the f r c q d i e s of the latter modc9. This possibility also seems unlikely because the highest frequency C,C, modes (whose frequencies arc closer to those of the C g 4stretches than are those of the CbCb modes) downshift 88 CXpaded. It is possible that the c b c b modes of the anions are more strongly mixed with the C g - 0 vibrations than are the C,C, modes. However, in the case of neutral species, normal-coordinate calculations clearly indicate that this is not the A third possible explanation for the reduction-inducedUpshifts O w e d for the c b c b Stretches is that the structures of the anions are sienifcantly different than thw of the neutral pigments. Large structural changes could significantly alter the forms of the vibrational eigenvectors. If these structural changes lead to greater mixing between the CbCb vibrations and the higher frequency C,Cm stretches, the former modes would be expected to upshift while the latter would be expected to dawnshift. This picwe is qualitativelycoBshnt with the rcduction-induced shifts observed for the CbCb and ClCm stretching vibrations of Chl but less satisfactory for Pheo (Table 11). The fact that none of the above scenarios a n n p l d y explains the reduction-induced shifts observed for both Chl and Pheo suggests that a number of factors may contribute to the shift patterns. F i i y , it is interestingto speculateon the identity of compound X. Coulometry indicates that a reduced species is formed. However, both the absorption and RR spectra of this species arc different from those of Chl-. These spectra are also not simple
Spectra of Chlorin and Chlorophyll Radical Anions
The Journal of Physical Chemistry, Vol. 96, No. 25, 1992 10239
composites of the spectra of the anionic and neutral species, although a number of the features observed in the RR spectra of compound X are more similar to those observed for Chl than for Chl-. Most notably, a band attributable to the C9=0 keto stretching mode is observed near 1691 cm-'whereas there is no evidence of a band near 1665 cm-'.Even a very weak band should be discernible in this latter region because the RR spectrum is uncongested. These seemingly incongruous observations could be explained if compound X were a dimeric species containing Chl and Chl- in which the oxygen atom of the C9-ketogroup of the anion is ligated to the Mg atom of the neutral species. This type of dimer is formed between neutral Chls in nonpolar solv e n t ~ The . ~ ~absorption spectrum of a dimeric species of this type would be expected to be similar to a composite spectrum of the neutral and anionic species but not identical, owing to the electronic interactions between the two macrocy~les.~~ Dimer formation could also ascount for the features observed in the RR spectrum, particularly the absence of a band due to the C9=0 stretching mode of the anion. Coordination of the (partially) negatively charged oxygen atom of the &-keto group of the anion to the Mg atom of the neutral species should upshift the frequency of the C 9 4 stretching vibration due to the redistribution of negative charge onto the metal ion. Indeed, it is possible that the weak shoulder observed at 1676 cm-' in the spectrum of compound X is the upshifted C9=0 stretching mode of the anion (Figure 8). The structures of both Chl and Chl- in the dimer rmght also be expected to be different from those of the monomeric species. This could ascount for the fact that the frequencies of certain C,C, and c&, stretching modes are different from those of either Chl or Chl-. The absence of RR bands assignable to the individual constituents of the dimer could be due either to a near coincidence of the frequencies of analogous modes of Chl and Chl- or to a lower RR intensity enhancement of one constituent (presumably Chl-) versus the other.
-
Conclusions The RR spectra reported herein for Zn(0EC)- and OECsuggest that reduction of these macrocycles does not result in large structural perturbations. In contrast, the data obtained for Chland Pheo- suggest that significant structural and/or electronic changes occur upon formation of these anions. The %-keto group on ring V appears to be the key structural element in facilitating these perturbations. The observation that the C 9 4 and cbc!, stretching vibrations undergo relatively large shifts upon reduction of the macrocycle suggests that these modes might serve as benchmarks for anion formation in vivo. This would facilitate timeresolved RR studies on photosyntheticproteins. The utility of these vibrations as anion marker bands in vivo assumes that these RR bands can be readily identified for both the neutral and anionic species. This could be compromised if the protein matrix exerts a significantly different influence on the vibrations in the neutral versus reduced species. The observation that the C9-keto group modulates the response of the ring to reduction is commensurate with the results of earlier RR studies on Chl' which indicate that this moiety is also the principal determinant of the response of the ring to o x i d a t i ~ n . ' ~In- ~this ~ connection, it has been suggested that conformational changes of chlorophylls in vivo might serve to modulate the electron-transfer p r o c e s ~ .The ~~~~ interaction of the C9-keto group with the protein matrix could provide a means of mediating these conformational changes and hence the redox a ~ t i v i t y . ~ ~ . ~ ~
Acknowledgment. This work was supported by grant GM36243 (D.F.B.) from the National Institute of General Medical Sciences. Referencea .ad Notes (1) Woodbury, N. W.; Becker, M.; Middendorf, D.; Parson, W. W. Bicchemistry 1985, 24, 7516. (2) Martin, J.-L.; Breton, J.; Hoff, A. J.; Mingus, S.;Antonetti, A. Proc. Nail. Acad. Sei. U.S.A. 1986, 83, 951.
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