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On the Relaxation Mechanisms of 6-Azauracil Jo~ao Paulo Gobbo,† Antonio Carlos Borin,*,† and Luis Serrano-Andres‡ † ‡
Instituto de Química, Universidade de S~ao Paulo, Av. Prof. Lineu Prestes, 748, 05508-900, S~ao Paulo, SP, Brazil Instituto de Ciencia Molecular, Universitat de Valencia, Apartado 22085, ES-46071 Valencia, Spain
bS Supporting Information ABSTRACT: The nonadiabatic photochemistry of 6-azauracil has been studied by means of the CASPT2//CASSCF protocol and double-ζ plus polarization ANO basis sets. Minimum energy states, transition states, minimum energy paths, and surface intersections have been computed in order to obtain an accurate description of several potential energy hypersurfaces. It is concluded that, after absorption of ultraviolet radiation (248 nm), two main relaxation mechanisms may occur, via which the lowest 3 (ππ*) state can be populated. The first one takes place via a conical intersection involving the bright 1(ππ*) and the lowest 1(nπ*) states, (1ππ*/1nπ*)CI, from which a low-energy singlet triplet crossing, (1nπ*/3ππ*)STC, connecting the 1(nπ*) state to the lowest 3 (ππ*) triplet state is accessible. The second mechanism arises via a singlet triplet crossing, (1ππ*/3nπ*)STC, leading to a conical intersection in the triplet manifold, (3nπ*/3ππ*)CI, evolving to the lowest 3(ππ*) state. Further radiationless decay to the ground state is possible through a (gs/3ππ*)STC.
1. INTRODUCTION The nucleic acid bases are the fundamental chromophoric units of DNA and RNA absorbing UV radiation below 300 nm. A crucial property shared by the nucleic acid base monomers is their photostability after interaction with UV light, a characteristic that has probably been acquired by natural selection and evolution,1 preventing destructive photochemical reactions.2 Early experiments concerning the interaction of the nucleobases with radiation showed a very low fluorescence quantum yield.3 5 Recent timeresolved spectroscopic experiments have probed internal ultrafast events, confirming excited state lifetimes of the order of a picosecond or less in the gas phase and solution.1,6,7 Time-resolved experiments have also shown that the nucleobases share a common pattern of deactivation mechanisms,1 related to very fast radiationless decay processes that deactivate the nucleobases back to the electronic ground state after absorption of UV light.8 From the theoretical point of view, the ultrafast radiationless decay of the DNA and RNA nucleobases has been rationalized in terms of internal conversion (CI) and intersystem crossing (ISC) processes. The crossings behave like funnels, a region in which the transfer of population between states is highly favorable.9,10 Nonetheless, it has been shown9,11 16 that the efficiency of the decay mechanism is not only assured by the existence of a low-energy CI, but it is also necessary to guarantee that it is accessible. On the other hand, it is known that the photochemical properties of many nucleobase tautomers and derivatives are distinct from those of their parent nucleobase.1,17 19 For example, it is worth comparing the properties of 2-aminopurine and its close isomer adenine (6-aminopurine);11 while the former has a strong emission that enables its use as a fluorescent probe for detecting conformational changes in proteins, the latter has a very efficient quenching mechanism, resulting in quantum yields of the order of 10 4 in aqueous solution. r 2011 American Chemical Society
Figure 1. Schematic structures and labeling for uracil (U) and 6-azauracil (6-AU). The structures do not represent detailed bonding characteristics.
Aza-substituted nucleic acid bases, although not detected in natural DNA or RNA, are interesting species due to their potential use as drugs for the treatment of several diseases,20 including cancer treatment when aza-substituted nucleic acid bases are employed simultaneously with chemotherapy and photodynamic therapy.21 Like other species derived from the natural nucleobases, aza-substituted analogues also have very distinct photochemical dynamics, as has been shown by Suzuki and co-workers.22 24 6-Azauracil (6-AU) (Figure 1) is an analogue of uracil, obtained by substituting the C6 atom by a nitrogen atom. Ab Received: January 11, 2011 Revised: March 25, 2011 Published: April 19, 2011 6243
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The Journal of Physical Chemistry B initio dynamics simulations25,26 showed a complex deactivation mechanism for uracil, which can be split into three possibilities, with the following two being the most relevant. The predominant mechanism (∼60% of the trajectories) starts off in the minimum of the 1(ππ* HL) (S2) state, where it stays for about 2.5 ps, then relaxes to the S1 1(nπ*) state, and finally to the ground state. The secondary (∼31% of the trajectories) indicates that the ultrafast deactivation mechanism8,25,26 can be explained by the barrierless character of the path from the 1(ππ* HL) state toward the ethene-like out-of-plane 1(gs/ππ*)CI conical intersection, involving combined stretching and twisting of the ethylenic bond, on the 1(ππ* HL) (HL: HOMO f LUMO) surface. Therefore, 6-azauracil and uracil differ in an important chemical bond from the photophysical point of view, that is, modifying the ethene-like chemical bond and introducing an isolated pair of electrons. Kobayashi et al.24 studied the excited states of 6-AU in acetonitrile (a polar aprotic solvent) employing experimental spectroscopic (absorption, transient-absorption, and time-resolved thermal lensing) and theoretical methods (time-dependent density functional theory (TD-DFT)). According to them, after 248 nm of irradiation, 6-AU has a high quantum yield of intersystem crossing (φISC = 1.00 ( 0.10), leading to the generation of a high amount of singlet O2 (1Δg), contrary to uracil. Another interesting feature of 6-AU is the reported low fluorescence quantum yield (φF = 10 4) after being irradiated with a 308 nm laser. The DFT calculations found several singlet and triplet states of different natures in a low-energy region; of special relevance is the existence of a dark 1(nπ*) state below the bright 1(ππ*) electronic state. Recently, Etinski and Marian27 have investigated the photophysical properties of 6-azauracil at the combined density functional theory and multireference configuration interaction (DFT/MRCI) level of theory. Linear interpolated energy profiles connecting the minima of the most relevant electronic states, spin orbit matrix elements, vibrational frequencies analysis, and intersystem crossing rates have been employed to propose a general, but not conclusive, mechanism for the experimental results reported by Kobayashi et al.24 There is a consensus that conical intersections28 provide a very efficient mechanism for population transfer among states of the same multiplicity (internal conversion, IC) of photoexcited molecules. If we are interested in the formation of triplet excited states, we have to keep in mind that they can also be populated by efficient intersystem crossing (ISC) from the initially excited singlet state, the efficiency of the interaction between singlet triplet states being described by the Fermi golden rule, which means that the extent of the interaction is ruled by the vibronic spin orbit coupling (SOC) factors and the Franck Condon (FC) weighted density of states.29 Nonetheless, besides determining the geometry and energetic position of the molecular structures related to the above-mentioned points, it is also necessary to investigate their accessibility from the initially populated states. A proper description of the potential energy hypersurfaces and related photophysical and photochemical properties requires nonadiabatic molecular dynamics computations. However, if the photophysical processes occur under conditions in which the excited state motion is slow, thermal equilibration is possible. If the excited state has a small amount of excess vibrational energy (i.e., with infinitesimal kinetic energy), the computation of minimum energy paths (MEPs) may represent a good source of information about the energetic profile. Indeed, it has been shown experimentally that energy transfer to the solvent
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molecules (vibrational cooling) occurs in a subpicosecond time scale.30,31 Therefore, if the excited state lifetime is greater than a few picoseconds, radiationless decay can be treated as occurring under conditions approaching thermal equilibrium, which means that state stationary points and minimum energy paths give important clues about the photophysical mechanisms.32,33 In this contribution, we investigate the photophysical relaxation mechanism of 6-azauracil with the CASPT2//CASSCF protocol. On the basis of a systematic search of relaxation paths connecting the relevant structures (such as state minima, intersystem crossings, and conical intersections), described with the computation of the minimum energy paths on the most important electronic hypersurface, a unified scheme of the singlet and triplet manifold, as well as a discussion of some questions raised recently,23,24,27 is presented. A comparison between the relaxation paths of 6-azauracil and uracil is also made.
2. METHODOLOGY The optimizations of minima, potential energy surface crossings, spin orbit coupling elements, transition states, and minimum energy paths (MEPs) were carried out at the multiconfigurational CASSCF34 level of theory. Cartesian coordinates for the relevant structures can be found in the Supporting Information. Dynamic correlation effects were taken into account with the second-order multiconfigurational perturbation theory approach,35 39 by recomputing the electronic energies at the relevant points described earlier. This approach, the so-called CASPT2//CASSCF protocol, has proved to be a general and accurate procedure for this purpose in previous studies.15,19 The active space comprises all π and π* orbitals plus three n orbitals, resulting in a 16 electrons in 11 orbitals calculation, CASSCF(16/11). The MEPs were obtained at the CASSCF(10/ 8) level, that is, with the π and π* orbitals in the active space, with the π electrons active. For each optimized structure along the MEP, a CASPT2(16/11) single point calculation was carried out with the standard zeroth-order Hamiltonian. An imaginary level shift40 of 0.2 au was applied to minimize the presence of intruder states. One-electron basis sets of ANO-L double-ζ plus polarization41 were employed, being represented by C, N, O (14s9p4d3f)/ H (8s4p3d) primitive gaussians contracted to C, O, N [3s2p1d]/ H [2s1p]. The spin orbit coupling (SOC) strength between selected states was computed within the AMFI framework, obtaining the length of the spin orbit coupling vector as described previously.42 No symmetry constraints were imposed, and all calculations were done with the MOLCAS-7.2 software.43 3. RESULTS AND DISCUSSION 3.1. Franck Condon Region and Excited States Minima. The optimized ground-state structure (Table 1 and Figure 2) is planar. In agreement with the results published by Kobayashi et al.,24 the C5 N6 bond distance (1.286 Å) is somewhat shorter than that of C5 C6 in uracil (1.342 Å,44 1.340 Å45). Our results are also comparable with those derived from the X-ray analysis of Singh and Hodgson (1.291 Å)46 and the minimum structure computed by Etinski and Marian27 at the RI-CC2 level (coupled cluster with approximate treatment of doubles) with Dunnings correlation-consistent basis sets cc-pVDZ. Computed properties for the low-lying singlet and triplet electronic states of 6-azauracil are compiled in Table 2, together with theoretical data reported by Kobayashi et al.24 and Etinski 6244
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Table 1. Computed and Experimental Geometrical Parameters of 6-Azauracil parameter state GS
N1 C2
C2 N3
N3 C4
C4 C5
C5 N6
N6 N1
C2 O2
C4 O4
ref
1.378
1.383
1.378
1.475
1.286
1.337
1.191
1.200
this work
1.38
1.39
1.39
1.47
1.29
1.35
1.22
1.23
24
1.40
1.40
1.41
1.48
1.31
1.35
1.23
1.23
27
1.37
1.38
1.36
1.45
1.29
1.35
1.22
1.22
46
1.373
1.378
1.382
1.360
1.354
1.357
1.195
1.361
this work
1.38
1.41
1.38
1.36
1.41
1.47
1.23
1.43
27
S2
1.312
1.309
1.456
1.401
1.349
1.384
1.315
1.219
this work
T1
1.33 1.382
1.32 1.369
1.53 1.391
1.41 1.424
1.39 1.456
1.46 1.331
1.34 1.197
1.24 1.220
27 this work
1.43
1.39
1.42
1.45
1.45
1.36
1.23
1.24
27
S1
and Marian.27 For the sake of comparison, related properties for uracil taken from some selected studies44,45,47,48 are also included in Table 2. It is known that the gas-phase absorption spectrum of uracil49 is composed of two bands. The lowest-energy and most relevant band from the photophysical point of view is centered at about 5.1 eV, being attributed to the transition to the S2 1(ππ* HL) excited state. The S1 excited state is derived from the excitation nO f π*, where nO is the lone electron pair from O4 (Figure 3), being located vertically at 4.93 eV above the ground state.44 The energetic order of the lowest-lying excited states of 6-azauracil can be compared to that of natural uracil base. As observed in uracil, the lowest excited state of 6-azauracil, S1 1(nπ*) (Table 2), is also derived from an n f π* excitation. The S1 1(nπ*) state is placed vertically at 4.36 eV above the ground state (4.10 eV at the TD-DFT level24 and 4.35 eV at the RI-CC2 level27), with μ = 2.18 D. In comparison to uracil, the 6-AU S1 1(nπ*) excited state is red-shifted by 0.57 eV. As can be seen in Figure 3, the n orbital of 6-AU has contributions from the lone pairs located on the O4 and N6 atoms (nO), that is, from the nitrogen atom inserted into the uracil natural base skeleton, and the π* (LUMO) has a C4 C5 bonding character and a C5 N6 and C4 O4 antibonding character. The optimized geometry of the S1 1(nπ*) state of 6-azauracil (Figure 2) shows some trends worth noting. The S1 1(nπ*) state is planar, with a C2 O2 carbonyl bond length (1.195 Å) almost the same as in the ground state (1.191 Å), but the C4 O4 bond distance is 0.161 Å longer than that of the ground state (1.361 Å vs 1.200 Å). For uracil, the same trend was observed by Climent et al.44 (C2 O2, 1.201 Å vs 1.200 Å for the ground state; C4 O4, 1.362 Å vs 1.203 Å for the ground state) and by Marian et al.,45 although their reported C4 O4 bond length is only slightly larger in the S1 1(nπ*) excited state (1.30 Å vs 1.20 Å for the ground state). An elongation of the C5 N6 (1.354 Å vs 1.286 Å for the ground state) and a shortening of the C5 C4 bond lengths (1.360 Å vs 1.475 Å for the ground state) is also computed for the S1 1(nπ*) of 6-AU, in line with the results reported for uracil by Climent et al.44 and Marian et al.45 The bond lengths computed by us are slightly shorter than those reported in ref 27. As for uracil, the bright state of 6-azauracil, the S2 1(ππ* HL) excited state, is computed vertically at 4.79 eV from the ground state (4.70 eV at the TD-DFT level24 and 4.93 eV at the RI-CC2 level27), in line with the experimental value (4.79 eV); it is associated with the largest oscillator strength up to 5 eV (f = 0.115). In comparison to uracil, for which the S2 1(ππ* HL) state is placed vertically at
Figure 2. Structure and main bond lengths (Å) for the optimized geometry of the lowest-lying electronic states of 6-azauracil. The structures do not represent detailed bonding characteristics.
5.18 eV44 above the ground state, the S2 state of 6-AU experiences a red shift. The excitation takes place from the π (HOMO) bonding orbital, with mainly C5 N6 character, to the π* orbital (LUMO) (Figure 3). The optimized geometry of the S2 1(ππ* HL) excited state of 6-azauracil (Figure 2) shows elongated C2 O2 (1.315 Å), C5 N6 (1.349 Å), and N6 N1 (1.384 Å) bond lengths as compared to the ground-state equilibrium geometry. The C4 O4 bond distance is 1.219 Å, similar to that computed for the ground state. As for the S1 1(nπ*) state, the C5 C4 bond length is shorter in the S2 1(ππ* HL) excited state (1.401 Å) than in the ground state. Again, the bond distances computed by us are slightly shorter than those computed at the RI-CC2 level (Table 1 and ref 27). The S2 1 (ππ* HL) state exhibits the largest dipole moment (μ = 6.03 D) of all states considered by us. 6245
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Table 2. Singlet and Triplet Low-Lying Electronic States of 6-Azauracil: Vertical Excitation Energy (EVA), Oscillator Strengths (f), and Dipole Moments (μ) 6-azauracil theoretical this work EVA (eV)
f
S1 1(nπ*)
4.36
0.0002
S2 1(ππ*) S3 1(nπ*)
4.79 5.06
0.115 0.0075
S4 1(ππ*)
5.54
0.145
T1 3(ππ*)
3.53
T2 3(nπ*)
3.92
T3 3(nπ*)
4.74
T4 3(ππ*)
4.76
state GS
experimental ref 24
μ (D)
ref 27
ref 24
EVA (eV)
f
EVA (eV)
f
EVA (eV)
2.18
4.10
0.000
4.35
0.0005
6.03 2.86
4.70 4.71
0.122 0.002
4.93 5.20
0.1529 0.0026
4.79
2.28
5.40
0.067
5.80
0.0540
5.42
1.61
3.00
3.54
1.55
3.56
4.03
2.37
4.32
4.87
1.38
4.22
5.00
1.56
uracil ref 44a
ref 47b
ref 48c μ (D)
state
EVA (eV)
f
EVA (eV)
f
S1 1(nOπ*) S2 1(ππ*)
4.93 5.18
0.0006 0.1955
4.81 5.27
0.000 0.181
S4 1(ππ*)
6.18
0.0733
T1 3(ππ*)
3.80
3.84
3.60
3.68
T2 3(nOπ*)
4.71
4.59
1.78
4.40
T3 3(nπ*)
5.33
GS
EVA (eV)
f
EVA (eV)
f
4.89 5.50
0.000 0.199
4.61 5.48
0.000 0.263
6.25
0.000
5.83
0.001
6.41
0.056
6.15
0.050
4.16 1.53 5.36
S3 1(nπ*)
a
ref 45d
5.73
CASSCF//CASPT2. b RI-CC2/aug-cc-pVDZ. c RI-CC2/TZVP. d DFT/MRCI/TZVP.
The triplet states of uracil, observed in solution,50 52 are characterized by a strong dependence of the intersystem crossing quantum yield (φISC) on the excitation energy in the lowestenergy absorption band, varying from 1.4 10 3 at 4.43 eV (280 nm) to 1.6 10 2 at 5.39 eV (230 nm).53,54 As mentioned in the Introduction, 6-AU has a φISC value close to 1. Our results for the triplet states of 6-azauracil (Table 2) show a reverse nature compared to the singlet excited states. Thus, the π f π* excitation corresponds to the T1 3(ππ* HL) excited triplet state, while the n f π* excitation gives rise to the T2 3(nπ*) excited state. The same energetic order is obtained for uracil in the Franck Condon (FC) region. The T1 3(ππ* HL) state is computed by us to lie 3.53 eV higher in energy than the ground state, still below the S2 1(ππ* HL). According to the DFT results of Kobayashi et al.,24 the T1 3 (ππ* HL) state would be 3.00 eV above the ground state. Following the T1 3(ππ* HL) state, we computed the T2 3(nπ*), T3 3(nπ*), and T4 3(ππ* HL) states at 3.92 eV (3.56 eV, ref 24), 4.74 eV (4.32 eV, ref 24), and 4.76 eV (4.22 eV, ref 24), respectively, above the ground state. With respect to uracil, our results suggest that the triplet states are stabilized in 6-AU. The optimization of the T1 3(ππ* HL) state geometry (Figure 2) gives a planar structure, in contrast with the butterfly shaped nuclear arrangement found for the T1 state of 6-AU by Etinski and Marian27 and for uracil.44,47,55 Apart from this, in comparison to the ground-state equilibrium structure, the
minimum energy structure of the T1 3(ππ* HL) excited state shows the same trends as obtained for the other excited states: an elongated C5 N6 bond (1.456 Å) and shorter C5 C4 bond (1.424 Å). The computed dipole moment for the T1 3(ππ* HL) state (1.61 D) is similar to that predicted for the ground state (μ = 1.56 D), indicating that polar solvents will not have much influence on the T1 3(ππ* HL) adiabatic excitation energy. In uracil, the dipole moment computed for the T1 state (3.60 D)47 is smaller than that for the S0 state (Δμ = 0.56 D), so the influence of polar solvents will be greater than that for the modified nucleobase. 3.2. Relaxation Mechanisms. 3.2.1. Singlet Manifold. The photophysical properties of DNA and RNA nucleobases depend on the radiationless pathway involving two intermediate electronic states: the bright 1(ππ*) and dark 1(nπ*) excited states. As mentioned before, the photophysics of 6-azauracil differs from that exhibited by uracil, being characterized by a high quantum yield for intersystem crossing. According to Kobayashi et al.,24 this distinct behavior can be rationalized by taking into account the energy gap between the 1(ππ*) and 1(nπ*) excited states. After irradiation with UV light, most of the energy absorbed by 6-AU is carried by the bright S2 1(ππ* HL) excited state, computed vertically at 4.79 eV with an oscillator strength of 0.115 (Table 2). The minimum energy path (MEP) (Figure 4) on the S2 hypersurface, starting in the Franck Condon region, along with the evolution of other singlet and triplet states, shows 6246
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Figure 3. Most important n and π molecular orbitals of 6-azauracil at the ground state equilibrium structure.
Figure 5. Proposed photochemical scheme for the 6-azauracil molecule. The ground state potential energy surface is represented in black, the singlet states in blue, T1 3(ππ*) in red, and T2 3(nπ*) in wine. Radiative processes (fluorescence and phosphorescence, respectively) are presented by blue and green arrows.
Figure 4. Evolution of the ground and lowest singlet and triplet excited states of 6-azauracil, along the minimum energy path (MEP) on the hypersurface of the S2 1(ππ* HL) state from the Franck Condon region (point 1) toward the S2 1(ππ* HL)min, computed at the CASPT2//CASSCF level.
that the S2 1(ππ* HL) state evolves directly toward its relaxed planar minimum, S2 1(ππ* HL)min, placed adiabatically from the S0 state at 4.35 eV, corroborating the results reported by Etinski and Marian.27 In the S2 1(ππ* HL)min region, the system could potentially release the excess of energy by emitting radiation. Nonetheless, according to Kobayashi et al.,24 there is no emission from this state. Therefore, another efficient relaxation mechanism must take place. A schematic view of the most relevant photophysical pathways computed by us for 6-AU is displayed in Figure 5, to be discussed in the following paragraphs. The first plausible event to be considered would be via a conical intersection connecting the S2 1(ππ* HL) and ground states, 1(gs/ππ*)CI, computed to be adiabatically 4.35 eV above the ground state (see Figure 5).
Although S2 1(ππ* HL)min and 1(gs/ππ*)CI stationary points are computed to be isoenergetic, it is expected that they are linked through a barrier as linear interpolation in internal coordinates (LIIC) suggests. This energetic barrier is high enough to prevent the evolution of the S2 excited state from its minimum energy structure to the conical intersection with S0. In comparison to the S2 1(ππ* HL)min (Figure 2) geometry, the intermediate transition state structure, S2 1(ππ* HL)TS (Figure 6), exhibits a shorter C2 O2 (1.255 Å) and an elongated C5 N6 (1.400 Å) and C4 C5 (1.457 Å) bond distance. Nonetheless, the most striking feature of the S2 1(ππ* HL)TS geometry is the out-of-plane deformation of the H5 C5 N6 N1 dihedral angle involving the H5 atom (Figure 6). From the computed data (Figure 5), one notices that two other deactivation mechanisms are possible, the first via the conical intersection (1nπ*/1ππ*)CI, involving the S1 1(nπ*) and S2 1(ππ* HL) excited states, and the other through the intersystem crossing (1ππ*/3nπ*)STC between the S2 1(ππ* HL) and T2 3(nπ* HL) excited states. These possibilities will be analyzed in the following paragraphs. Direct population of the S1 1(nπ*) state, located vertically 4.36 eV above the ground state, is not very likely, as the computed oscillator strength (f = 0.0002) suggests. However, we found a conical intersection between the S1 1(nπ*) and S2 1 (ππ* HL) states, (1nπ*/1ππ*)CI, located adiabatically 0.2 eV above S2 1(ππ* HL)min. The existence of this conical intersection was proposed recently.27 The geometry of the (1nπ*/1ππ*)CI structure (Figure 6) does not differ much from that of S2 1 (ππ*)min (Figure 2), and the planarity exhibited by the initial 6247
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Figure 6. Selected CASSCF optimized conical intersections and transition state structures. The structures do not present detailed bonding characteristics.
arrangement is maintained. Therefore, due to the structural and energetic resemblance between S2 1(ππ*)min and the (1nπ*/1ππ*)CI, the latter may lead to a partial population transfer from the S2 1(ππ* HL) state toward the S1 1(nπ*) state and then to its minimum, S1 1(nπ*)min, located adiabatically at 3.76 eV above the ground state in the FC region. It is also worth noting that the MEP from the Franck Condon region on the S1 1 (nπ*) hypersurface leads directly to the S1 state minimum, 1 (nπ* HL)min. Once the S1 1(nπ*)min is reached, three other pathways are possible: (i) via the (gs/1nπ*)CI conical intersection between the S1 and S0 hypersurfaces, leading to a radiationless decay to the ground state; (ii) emission from the S1 1 (nπ*)min to the S0 state; (iii) a singlet triplet crossing (STC) with a low-lying triplet state, to be discussed in the next section. The (gs/1nπ*)CI conical intersection was found adiabatically 4.8 eV above the ground state, too high to be accessible from 1 (nπ*)min. Therefore, a radiationless decay of the S1 1(nπ*) state via the high-lying (gs/1nπ*)CI is not an effective deactivation mechanism. The relaxation pathway involving fluorescence from S1 1(nπ*)min is more likely. Kobayashi et al.24 did not observe fluorescence after excitation at 248 nm (∼5.00 eV), close to the vertical excitation energy of the S2 1(ππ* HL) state computed by us at 4.79 eV (Table 2). A very low fluorescence (φF = 10 4), however, was observed after excitation at 308 nm (4.02 eV). The computed vertical emission energy from the S1 1(nπ*)min state is 2.83 eV, in line with the experimental findings. The dependence of the fluorescence quantum yield on the excitation wavelength
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can be better understood after considering the relaxation mechanisms related to the triplet excited state, which will be done in the next section. As emphasized in the Introduction, based on recent ab initio dynamics simulations,25,26 the ultrafast deactivation mechanism of uracil can be understood by considering two pathways: the predominant mechanism (∼60% of the trajectories) starting in the minimum of the 1(ππ* HL) (S2) state, followed by population transfer to the S1 1(nπ*) state, and finally to the ground state; the secondary (∼31% of the trajectories) mechanism8,25,26 related to a barrierless path from the 1(ππ* HL) minimum toward the ethene-like out-of-plane 1(gs/ππ*)CI conical intersection, involving combined stretching and twisting of the ethylenic bond, on the 1(ππ* HL) (HL: HOMO f LUMO) surface. For 6-azauracil, the calculations indicate that the MEP along the hypersurface of the S2 1(ππ* HL) state leads in a barrierless way to the S2 1(ππ* HL)min state, from which a barrier must be surmounted in order to reach the 1(gs/ππ*)CI structure; the corresponding transition structure, S2 1(ππ* HL)TS, displays an out-of-plane deformation of the H5 C5 N6 N1 dihedral angle, involving the H5 atom. The substitution of the C6 atom by a nitrogen atom prevents the stretching and twisting of the C5 N6 bond, which is the relevant deformation necessary to allow the formation of the CI structure with an ethene-like out-of-plane deformation. Therefore, the primary deactivation mechanism observed in uracil (via the S1 1(nπ*) state) is also the most relevant for 6-azauracil. The absence of an emission band for the S2 1(ππ* HL) state, as claimed by the experimentalists,24 can be explained by the population transfer to the S1 state and to a triplet state, as discussed below. 3.2.2. Triplet Manifold. The MEP on the hypersurface of S2 1 (ππ* HL) (Figure 4) from the Franck Condon region toward the S2 minimum, 1(ππ* HL)min, indicates that the lowest-lying triplet state of 6-AU can be populated via the mechanisms described below (Figure 5). The results reported here were obtained in a vacuum. Because in polar solvents such as acetonitrile, employed by Kobayashi et al.,24 the 3(nπ*) states are blue-shifted,56 we will consider only the lowest 3(nπ*) state. Near the S2 1(ππ* HL)min region, a singlet triplet crossing (STC) between the S2 1(ππ* HL) and T2 3(nπ*) excited states ((3nπ*/1ππ*)STC) is computed at the CASSCF level, with a large spin orbit coupling (SOC) of about 17 cm 1. At the CASPT2 level, the singlet triplet splitting (ΔEST) is computed to be 0.16 eV (3.7 kcal 3 mol 1). Considering that an efficient population transfer from singlet to triplet states requires small energy gaps (in this work, ΔEST = 3.7 kcal 3 mol 1), large spin orbit couplings (SOC)57 (in this work, SOC = 16 cm 1), and a large transition dipole moment between the S2 1(ππ* HL) excited state and ground state45 (in this work, f = 0.115), the (3nπ*/1ππ*)STC mentioned above is expected to be a viable mechanism for populating the T2 3(nπ*) state. Starting from the (3nπ*/1ππ*)STC structure, the MEP on the T2 3(nπ*) hypersurface leads to the T2 3(nπ*)min state minimum, placed adiabatically 3.61 eV above the ground state. A conical intersection, (3nπ*/3ππ*)CI, connecting the T1 3 (ππ*) and T2 3(nπ*) triplet states was computed to be 0.08 eV (1.8 kcal 3 mol 1) adiabatically above T2 3(nπ*)min, favoring the population switch toward the T1 1(ππ*) state. From the (3nπ*/3ππ*)CI, the MEP on the T1 3(ππ*) hypersurface leads to the T1 3(ππ*)min state minimum, placed adiabatically 2.77 eV above the ground state, with a computed vertical 6248
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Figure 7. Selected CASSCF optimized conical intersections and singlet triplet crossing structures. The structures do not present detailed bonding characteristics. The notation (1nπ/3ππ 3nπ)STC represents the structures (1nπ/3ππ)STC and (1nπ/3nπ)STC.
emission energy of 2.30 eV, in agreement with the experimental phosphorescence observed at 2.42 eV.58 Just slightly above T1 3 (ππ*)min (0.12 eV), we found an intersystem crossing connecting T1 3(ππ*) and the ground state, (gs/3ππ*)STC, with a computed SOC of 2.7 cm 1. Due to the small value of the spin orbit coupling, no fast population transfer from T13(ππ*)min to the ground state is expected. Therefore, we predict a longer lifetime for the T1 3(ππ* HL) state, suggesting that it would be possible to observe the sensitization of 6-azauracil in the presence of molecular oxygen, giving rise to the formation of O2 (1Δg) with a high quantum yield, as shown experimentally by Kobayashi et al.24 Although the 3(ππ)* state and the (gs/3ππ*)STC are close in energy, their geometries are different. The (gs/3ππ*)STC geometry (Figure 7) is characterized by a distorted H5C5C4O dihedral angle of about 67° and a change of the N6N1C2N3 angle of about 16°. There is another possibility to populate the T1 3(ππ*) state. As mentioned in the Singlet Manifold subsection, there is a population transfer from the bright S2 1(ππ* HL) state to the dark S1 1 (nπ*) state via the (1nπ*/1ππ*)CI conical intersection. The MEP on the S1 1(nπ*) hypersurface from the (1nπ*/1ππ*)CI region leads to the (1nπ*/3ππ*)STC crossing region, located near S1 1(nπ*)min. The computed SOC is 64.7 cm 1, large enough to suggest this route as an efficient decay mechanism to the lowest T1 3(ππ*) state. The geometry of the (1nπ*/3ππ*)STC structure (Figure 7) differs from that of the ground state in the increase of the C4 O4 and the decrease of the C4 C5 bond distances, as well as a closure of the O4C4N3 angle by more than 15° and an out-of-plane deformation of the N3H bond by about 47°. Besides, the higher 3(nπ*) state is located in the same energetic region at 3.82 eV, complicating even more an accurate determination of the triplet relaxation. Both paths, singlet and triplet, ultimately lead to an energy transfer to the T1 excited state. With respect to the wavelength dependence of φF,24 it is possible to make some considerations. As stated by Kobayashi et al.,24 after irradiation at 308 nm, the S1 1(nπ*) state can be populated directly, evolving to the S1 1(nπ*)min state minimum, from which an emission can be observed. The weakness of the emission is due to a highly efficient singlet triplet crossing, denoted here as (1nπ*/3ππ*)STC, that transfers most of the energy to the triplet state. The situation is quite different when
6-azauracil is irradiated at 248 nm, because, under this condition, it is possible to access the S2 1(ππ* HL) state directly and, according to our results (Figure 5), although the S1 1(nπ*) may be populated as well, one more channel is open due to another STC, (3nπ*/1ππ*)STC. This bifurcation may cause a more severe quenching of the fluorescence. For uracil, according to Climent et al.,44 there are two different paths to populate the lowest triplet state, T1 3(ππ*). The first one starts with a STC between the dark 3(nOπ*) and bright 1(ππ*) states, (3nOπ*/1ππ*)STC, at 4.6 eV, for which the SOC is computed to be 25 cm 1. Then, following the 3(nOπ*) state hypersurface, the population can be transferred to the T1 3(ππ*) state via a conical intersection, (3nOπ*/3ππ*)CI, located slightly above (0.01 eV) the 3(ππ*)min. Following the T1 3(ππ*) hypersurface from the (3nOπ*/3ππ*)CI point, the system reaches the 3 (ππ*)min. This energy relaxation is very similar to the first one proposed in this subsection, but for 6-AU, it must be more efficient due to the large SOC computed. It is also interesting to note that this mechanism has been reported to occur not only in nucleobases but also in other chromophores such as isoalloxazine59 and psoralen.60 The other path proposed for uracil by Climent et al.44 involves a direct population transfer from the bright 1(ππ*) state to the T1 state via a STC, (1ππ*/3ππ*)STC, located at 4.2 eV adiabatically from the S0 state. Nonetheless, the computed SOC amounts for only 1 cm 1, which makes it a very unlikely mechanism.
4. CONCLUSION In this contribution, the CASPT2//CASSCF protocol and double-ζ plus polarization ANO-L basis sets were used to investigate the photophysics of 6-azauracil, by means of geometry optimizations of the ground and excited states, as well as the computation of minimum energy paths, singlet triplet crossings, and conical intersections. The results presented here provide reliable clues about the deactivation mechanism of 6-azauracil. After population of the bright S2 1(ππ* HL) state, the MEP on its hypersurface from the Franck Condon region shows that the system evolves barrierless to the S2 1(ππ* HL)min state minimum, where crossings with singlet ((1nπ*/1ππ*)CI) and triplet ((3nπ*/1ππ*)STC) states are likely to occur. Both paths lead to an efficient energy transfer to the long-lived triplet excited 6249
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The Journal of Physical Chemistry B electronic state, T1 3(ππ*), which explains the experimental detection of oxygen singlet (1Δg). The possibility of another relaxation mechanism related to the radiationless decay to the ground state via a conical intersection ((gs/ππ*)CI), similar to that observed in uracil, was also studied; our results show that this mechanism is not likely because a high energetic barrier must be surmounted, decreasing drastically its efficiency. Another aspect addressed in this contribution concerns the wavelength dependence of the quantum yield for intersystem crossing (ΦISC). According to our results, after being exposed to electromagnetic radiation with a wavelength of 308 nm, the S1 (1nπ*) state is populated directly and most of its energy is transferred to T1 3(ππ*) via the low-energy singlet triplet crossing, (3nπ*/1ππ*)STC, as proposed in ref 24, but a small fraction of the excess energy can be emitted as fluorescence (φF = 10 4). However, if the system is irradiated with a more energetic light, able to populate the S2 excited state directly, the excess of energy will be redistributed between the singlet and triplet manifold, weakening even more the already low fluorescence.
’ ASSOCIATED CONTENT
bS
Supporting Information. Cartesian coordinates of the optimized structures. This material is available free of charge via the Internet at http://pubs.acs.org.
’ AUTHOR INFORMATION Corresponding Author
*E-mail:
[email protected].
’ ACKNOWLEDGMENT J.P.G. acknowledges CAPES (Coordenac-~ao de Amparo ao Pessoal de Ensino Superior, Project 2272/09-1) for a postdoctoral fellowship. L.S.-A. acknowledges financial support from Project CTQ2010-14892 and CSD2007-010 Consolider-Ingenio in Molecular Nanoscience of the Spanish MEC/FEDER. A.C.B. acknowledges continuous academic support from CNPq (Conselho Nacional de Desenvolvimento Cientfico e Tecnologico) and FAPESP (Fundac-~ao de Amparo a Pesquisa do Estado de S~ao Paulo). The services and computer time at the Laboratorio de Computac-~ao Científica Avanc-ada (LCCA) of the Universidade de S~ao Paulo are also acknowledged. The authors are also thankful to Prof. Manuela Merchan for stimulating discussions and careful reading of the manuscript. Unfortunately, Luis Serrano-Andres passed away unexpectedly. Luis, we miss you as a dear friend and scientist. ’ REFERENCES (1) Crespo-Hernandez, C. E.; Cohen, B.; Hare, P. M.; Kohler, B. Chem. Rev. 2004, 104, 1977. (2) Radiation Induced Molecular Phenomena in Nucleic Acids; Shukla, M., Leszczynski, J., Eds.; Springer: Berlin, 2008. (3) Daniels, M.; Hauswirth, W. W. Science 1971, 171, 675. (4) Morgan, J. P.; Daniels, M. Chem. Phys. Lett. 1979, 67, 533. (5) Callis, P. R. Chem. Phys. Lett. 1979, 61, 568. (6) Canuel, C.; Mons, M.; Pluzzi, F.; Tardivel, B.; Dimicoli, I.; Elhanine, M. J. Chem. Phys. 2005, 122, 074316. (7) Hare, P.; Crespo-Hernandez, C. E.; Kohler, B. Proc. Natl. Acad. Sci. U.S.A. 2007, 104, 435.
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