5666
J . Phys. Chem. 1986, 90, 5666-5668
On the Electronic Spectrum of I-Methyluracil Joel S. Novrost and Leigh B. Clark* Department of Chemistry, B-014, University of California, Sun Diego, La Jolla, California 92093 (Received: June 2, 1986)
The electronic spectrum of 1-methyluracilhas been reexamined by measuring the polarized reflection spectra of two different faces of single crystals of the material. The results indicate that the lowest energy absorption region (275 nm) is not composite in nature and provide a transition moment direction of -9' (measured toward N3 from the N I X 4axis) for this band. The polarization direction of the second strong AT* transition at 213 nm is either -53O or +59' relative to the Nl-C4 axis. The +59' choice is found to be consistent with linear dichroism results for uracil and thymine, while the -53' possibility agrees more nearly with the results from a single-crystal study of thymine. Evidence for an nx* transition at 217 nm polarized perpendicularly to the molecular plane is presented.
Introduction The present paper is part of an ongoing effort to determine the spectroscopic properties of the pyrimidine and purine bases. Such monomer parameters are ultimately used in the interpretation of the optical properties of the various polymers that occur. 1Methyluracil was the object of an earlier single-crystal, direct absorption study by Eaton and Lewis.' The present work similarly yields polarized absorption spectra of single crystals of 1methyluracil; however, here the curves are obtained by Kramer-Kronig analyses of reflection data. Our results confirm the assignment of Eaton and Lewis for the polarization of the lowest energy absorption band (270 nm) and, in addition, provide evidence for the polarization of the second absorption band (205 nm). The derivation of transition moment directions from crystal spectra is obfuscated owing to various effects manifested by the intermolecular interactions in the crystal. The problem is particularly acute when molecules of low symmetry are studied. For pyrimidine bases, the planar symmetry restricts transition moment directions to two choices: (1) perpendicular to the molecular plane, n a* type, and (2) parallel to the molecular plane, x a* type. For the latter type one seeks the precise directions in the molecular plane of the various transition moments. The usual approach is to obtain absorption spectra with radiation incident normally to a crystal face and polarized along the principal directions of the face. The dichroic ratios obtained from such data are then interpreted in terms of the oriented gas model as originating from the projection onto that face of a unique, in-plane transition moment. A situation that appears to be not uncommon is that the spectral contours of a given band are found to be different along different crystal axes. Such situations when considered within the framework of the oriented gas model have led to the assignment of multiple transitions in order to account for the contour differences?4 The data reported in this paper show large band shape changes for the lowest energy band along different crystal axes but have sufficient internal redundancy so as to rule out the presence of multiple transitions. In this case the change in band shape must arise because of crystal-field-induced mixing.
-
-
Experiment and Results 1-Methyluracil forms orthorhombic crystals with eight molecules per unit The planes of the molecules lie strictly in the ab crystal plane. Slow evaporation of aqueous solutions yields large 5 X 5 X 20 mm crystals that are bounded on the ends by (001) and on the long sides by the form (110). We have obtained polarized spectra from both crystal planes so that the data include the orthogonal a, b, and c axes as well as the redundant I C direction of the (1 10) face. The latter axis lies in the plane of the molecules at 45' from both the a and b axes. An orthogonal projection of the eight molecules of the unit cell onto the (001) Present address: Clinical Chemistry Technical Center, Eastman Kodak Co., Rochester, NY 14650.
0022-3654/86/2090-5666$01 .50/0
plane is given in Figure 1. Details of the experimental procedures and the numerical analysis of the reflection data have been given earlier.69' The four reflection spectra obtained from the two crystal faces are presented in Figure 2 along with the corresponding absorption spectra. The spectra to about 41 000 cm-' of the a, b, and c axes have been reported by Eaton and Lewis,' and their results for the a axis are shown in Figure 2 for quantitative comparison of the two methods employed. The two experimental procedures give results that are coincident within the stated uncertainty ( i 1 6 % ) of the direct absorption measurements. The slight asymmetry apparent in the correspondence may originate in reflection losses not accounted for in the direct absorption work. Discussion of Spectra TransitionI. The transition centered at about 36 300 cm-' (275 nm) is strongly polarized along the a crystal axis. Eaton and Lewis report the dichroic ratio (coleb) of the 0,O component near 34 800 cm-' (287 cm-I) to be 250 f 100, and the two in-plane polarization directions consistent with this dichroic ratio are +7' and 0' according to the DeVoe-Tmm angle convention8(positive toward N3 from the NI-C4 reference axis). See Figure 3 for the numbering and angle conventions. The dichroic ratio for the 0,O obtained in the present work is 80 and carries an estimated uncertainty of about a factor of 3 arising from the relatively large percentage uncertainty in small calculated absorption coefficients from reflection data. A value of 80 is still large and corresponds to polarization angles of -3' or 10'. However, the dichroic ratio decreases as one moves through the band toward higher vibronic membranes. At 40000 cm-' this ratio becomes 6.4, and the corresponding in-plane polarization angles are -19' and + 2 Y , The question is how are such data to be interpreted? For a free molecule the polarization direction of the 0,Ois the direction of the pure electronic transition moment vector. The 0,O observed in crystal spectra cannot be interpreted in the same way. As discussed recently for crystal spectra of protonated 1-methylcytosine,electronic intensity changes owing to crystal-field mixing of crystal states arising from the vibronic levels of a given electronic band (Le., intraband mixing) can lead to serious changes in dichroic ratios in the 0,O region and all across the electronic band.g Such mixing appears to be a t work in crystal of 1methyluracil. The Lopolarized spectrum from the (1 10) crystal face exhibits a substantially different band contour from that observed for the a axis spectrum. Since we believe that there is (1) Eaton, W. A.; Lewis, T. P.J . Chem. Phys .1970,53, 2164. (2) Stewart, R. F.; Davidson, J. J. Chem. Phys. 1963, 39, 255. (3) Stewart, R. F.; Jensen, L .H. J. Chem. Phys. 1964, 40, 2071. (4) Anex, B. G.; Fucaloro, A. F.; Durra-Ahmed, A. J . Phys. Chem. 1975, 79, 2636. (5) Green, D. W.; Mathews, F. S.; Rich, A. J . Biol. Chem. 1962, 237, 3573. (6) Chen, H. H.; Clark, L. B. J. Chem. Phys. 1973, 58, 2593. (7) Zaloudek, F.; Novros, J. S.; Clark, L. B. J. Am. Chem. Soc. 1985,107, 7344. (8) DeVoe, H.; Tinoco, I., Jr. J . Mol. Biol. 1962, 4, 500. (9) Clark, L. B. J . Am. Chem. SOC.,in press.
0 1986 American Chemical Society
The Journal of Physical Chemistry, Vol. 90, No. 22, 1986 5667
On the Electronic Spectrum of 1-Methyluracil
I \e i
M
Figure 3. Angle and atom numbering conventions for 1-methyluracil. The distortion from hexagonal symmetry in the six-membered ring is real and taken from the crystal structure.
Figure 1. Projection of the eight molecules of the unit cell onto the (001) crystal plane.
I
3000
2500
I
I
A
(A)
2000 I
-
I METHYLURACIL (001)(110)
I
j
present case since polarized fluorescence studies of various thymines and uracils suggests great spectral purity across the whole band.1° Probably the best compromise interpretation of the data in order to obtain useful transition moment directions is to use oscillator strengths evaluated from the area under the entire electronic band envelope. For band I the overall dichroic ratio CfJfb) is 19.9, and the corresponding in-plane angle possibilities are -9' and 16'. As noted above, the differences between absorption band contours for radiation polarized along different crystal axes have lead to the presumption that more than a single transition is present. For the present case, if a second transition of different polarization occurred in this spectral region, then a strong component would be expected along the b or c axis. Such is not the case, and we conclude, therefore, that but one transition is present. We believe that band contour changes arise largely from intraband mixing, i.e., among vibronic members of a given electronic transition. Mixing of states depends strongly on lattice sums of dipoldipole interactions. These lattice sums are dependent on the direction of the wave vector k (normal to crystal face) through the so-called macroscopic term which involves k-d where d is the transtion moment vector.' 1~12 Different sets of mixing coefficients m u r not only for the two principal axes of each crystal face but also for different crystal faces. The result is that the mixing coefficients among the series of vibronic components of a given transition can be very different (even in sign) from axis to axis and from crystal face to crystal face, Along one axis intensity might shift always toward the lower energy members of the series, while the reverse might occur for some other crystal axis. Substantial changes in the co!tour or shape of diffuse bands can result. For 1-methyluracil the k-d term is close to zero for the lowest transitioqon the (001) face. However for ICof (110) the angle between k and d is 45', and the macroscopic term is appreciable. The observed band shapes are consistent with this interpretation. If the earlier data from thymine anhydrate crystal4 are reworked assuming but a single transition in the 36 000-cm-' region, the possible in-plane directions for thymine are -12' and +70'. For the three crystal systems so far studied, there is then overall consistency that the first electronic band of these systems is polarized a t small to modest negative angles: l-methylthymine2 (-20°), thymine (-12'), 1-methyluracil (-go). Transition ZZ. The analysis of the second strong absorption band, 11, a t about 47 000 cm-' (213 nm) is reasonably straightis 2.2 and corresponds to the forward. The dichroic ratio fb/f, possible in-plane transition moment directions of -53' or +59'. The experimental data of the earlier study of thymine terminated at 210 nm, and there was considerable stated uncertainty in the quantitative values of the absorption coefficients for band I1 owing to the nature of the Kramers-Kronig p r ~ c e d u r e .Nevertheless, ~ the estimated possible polarization angles for I1 were given as -31'
+
\
I
I
Figure 2. Reflection and absorption spectra of the (001) and (110) crystal faces of 1-methyluracil. The inset shows two representative molecules projected onto the (001) plane and the intersection of the (1 10) plane with the (001) plane. The spectrum obtained for radiation incident normal to (1 10) and polarized parallel to this dashed intersection line is labeled IC. The direct absorption result of ref 1 is shown for comparison.
but one electronic transition in this frequency region, the differences in band shape must arise from crystal-field effects or possibly from vibronically i n d u d depolarization inherent to the molecules. The latter possibility is probably unimportant in the
(10) Callis, P. R. Chem. Phys. Lett. 1979, 61, 563. (11) Fox, D.; Yatsiv, S.Phys. Rev. 1957, 108, 938. (12) Clark, L. B.;Philpott, M. R. J . Chem. Phys. 1970, 53, 3790.
5668 The Journal of Physical Chemistry, Vol. 90, No. 22, 1986 Transition
II
Novros and Clark TABLE I: Oscillator Strengths fa
fb fc
f,c(obsd)
101 thymine
single crystal
ibi thymine linear dichroism
( c ) UlOClI linear dlchrolsm
CH3 (d) 1-methyluracil Elngle crystal
Figure 4. Correlation of results for band 11. The wedges represent the uncertainties given for the stretched film work.
or +91'. A comparison of the results for the two crystal systems does not provide a clear choice in removing the ambiguity in 8. Our value of -53' is 22' from -31°, while our value of +59' is only a bit further away (32') from their +91° value. See Figure 4. Correlation with Linear Dichroism Spectra. Linear dichroism (LD) studies of stretched poly(viny1 alcohol) films doped with planar chromaphores result in pairs of possible in-plane transition moments for each transition. The actual values of these angles relative to the N I X 4references axis are dependent on the correct identification of the orientation axis of the molecules along the stretching direction of the film; however, the relative angular differences between successive transitions should be independent of the choice of orientation axis. Matsuoka and Norden have studied both uracil and thymine in such stretched films.13 The LD for uracil is appreciably constant over the first absorption band but decreases at the red edge of the second band. The polarization angles for I work out to be 7' f 2' or -1 1' f 3'. The latter is nearly the value we report in this study. However, Matsuoka and Norden report that the drop in L D in the vicinity of I1 is consistent with an angle of 70-90° between I and 11. The angular differences between -9' for I and our -53O and +59' possibilities for 111 are 44' and 6 8 O , respectively, so that the +59O choice for I1 harmonizes the results of the two studies. These considerations are represented in Figure 4. The spectrum reported by Matsuoka and Norden for thymine does not show the drop in LD as I1 is approached, and this result indicates that I and I1 make equal angles with the orientation axis. The orientation axis for thymine was identified as +98' f 12, so that I1 for thymine should be expcted at either -31' or +47'. The latter value agrees within experimental uncertainty with the +59' choice for I1 of 1-methyluracil that harmonized the present crystal study and the LD results for uracil. Our alternate choice of -53' for I1 of 1-methyluracil would destroy that harmony. See Figure 4. Thus, the present work when coupled with the stretched film work support the +59' choice for 11. However, it must be noted that the corresponding choice given by Anex et al. for thymine is +91', and this value seems to be too far removed from the +59' value chosen above. It is difficult to see how experimental uncertainty in the thymine crystal work could account for the dis(13) Matsuoka, Y.;Norden, B. J . Phys. Chem. 1982,86, 1378
fd c a l c d ) ' ASO(crYst)b hal/
I
I1
0.027 0.544 0.007 0.295 0.286 0.193 0.195
0.552 0.251 ? 0.400 0.402 0.268 0.260
"Calculated from the (001) data for a and b. 'Aqueous solution, neutral pH.
bfiso= cf, +fb +fJ
parity, for a full inversion of the observed dichroic ratio of band I1 would be required. We do not know how to resolve the present disparity for this band with the existing data. Oscillator Strengths. Component oscillator strengths are internally consistent throughout as shown in Table I. The intensity expected along the ICaxis can be calcuated from the data of the (001) face as can the isotropic oscillator strengthAs, = 1/3(fo + fb +f,) for comparison to values obtained from an aqueous solution spectrum. In all cases agreement is within the estimated experimental uncertainty. This agreement supports the notion that interband intensity shifts are minimal. nr* State. Turning now to the IIc spectrum of the (110) crystal face, we see absorption that is polarized normal to the molecular planes. Any features observed here are candidates for nr* transitions. The very weak absorption in the region of band I was tentatively assigned by Eaton and Lewis to an nr* transition. Probably this absorption originates from the lowest rr*,in-plane band through crystal imperfections, the use of convergent radiation, etc. The fact that the contour of Z, reported by Eaton and Lewis is very nearly the same as that for I in the ICspectrum supports this conclusion. Callis has reached a similar c o n c l ~ s i o n . ~ ~ The next feature appearing at 46 000 cm-l in the Ilc spectrum and showing a crystal oscillator strengthf, = 0.06 is, therefore, the first observable candidate for the lowest nr* transition, for it cannot be accounted for in a similar fashion. The peak position of the second in-plane band shifts considerably from axis to axis of the crystal, so that the assignment of the absorption maximum at 46000 cm-' in the [IC spectrum as the exciton shifted the component of I1 cannot be ruled out although the origin of its intensity would be somewhat mysterious. In the absence of any calculations in regard to crystal energy shifts we think that an nr* assignment is a reasonable choice at the present time. The theoretical calculations of the transition energies by Hug and T i n o c ~ 'predict ~ the lowest energy nr* transition to occur somewhat to the blue of the first rr* transition. It should be noted that an "extra" band at about this position (-215 nm) has been observed in CD16 and MCD" spectra. Registry No. 1-Methyluracil, 615-77-0. (14) Callis, P. R. Ann. Rev. Phys. Chem. 1983,34, 329. (15) Hug,W.; Tinoco, I., Jr. J . Am. Chem. Sac. 1974, 96, 665. (16) Sprecher, C. A.; Johnson, W. C. Biopolymers 1977, 16, 2243. (17) Voelter, W.; Records, R.; Bunnengerg, E.; Djerassi, C. J. Am. Chem. SOC.1968, 90, 6163.
