ARTICLE pubs.acs.org/JPCA
S0 and S1 State Structure, Methyl Torsional Barrier Heights, and Fast Intersystem Crossing Dynamics of 5-Methyl-2-hydroxypyrimidine Simon Lobsiger, Hans-Martin Frey, and Samuel Leutwyler* Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland
Philip Morgan and David Pratt Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, United States
bS Supporting Information ABSTRACT: We report the analysis of the S1 r S0 rotational band contours of jet-cooled 5-methyl-2-hydroxypyrimidine (5M2HP), the enol form of deoxythymine. Unlike thymine, which exhibits a structureless spectrum, the vibronic spectrum of 5M2HP is well structured, allowing us to determine the rotational constants and the methyl group torsional barriers in the S0 and S1 states. The 000, 6a01, 6b01, and 1401 band contours were measured at 900 MHz (0.03 cm 1 ) resolution using mass-specific two-color resonant two-photon ionization (2C-R2PI) spectroscopy. All four bands are polarized perpendicular to the pyrimidine plane (>90% c type), identifying the S1 r S0 excitation of 5M2HP as a 1nπ* transition. All contours exhibit two methyl rotor subbands that arise from the lowest 5-methyl torsional states 0A00 and 1E00 . The S0 and S1 state torsional barriers were extracted from fits to the torsional subbands. The 3-fold barriers are V300 = 13 cm1 and V30 = 51 cm1; the 6-fold barrier contributions V600 and V60 are in the range of 23 cm1 and are positive in both states. The changes of A, B, and C rotational constants upon S1 r S0 excitation were extracted from the contours and reflect an “anti-quinoidal” distortion. The 000 contour can only be simulated if a 3 GHz Lorentzian line shape is included, which implies that the S1(1nπ*) lifetime is ∼55 ps. For the 6a10 and 6b01 bands, the Lorentzian component increases to 5.5 GHz, reflecting a lifetime decrease to ∼30 ps. The short lifetimes are consistent with the absence of fluorescence from the 1nπ* state. Combining these measurements with the previous observation of efficient intersystem crossing (ISC) from the S1 state to a longlived T1 (3nπ*) state that lies ∼2200 cm1 below [S. Lobsiger, S. et al. Phys. Chem. Chem. Phys. 2010, 12, 5032] implies that the broadening arises from fast intersystem crossing with kISC ≈ 2 1010 s1. In comparison to 5-methylpyrimidine, the ISC rate is enhanced by at least 10 000 by the additional hydroxy group in position 2.
I. INTRODUCTION Their low fluorescence quantum yields make photophysical and dynamical studies of nucleobases and nucleic acids very difficult.1 To overcome this problem, a number of fluorescent nucleic acid base analogs have been investigated and incorporated into DNA and RNA.2 5-Methyl-2-pyrimidinone (5M2P), which closely mimics the pyrimidine bases thymine and cytosine, is known to fluoresce in aqueous solution3 with absorption and emission maxima at 309 and 380 nm, respectively. These are well separated from those of the canonical DNA bases, allowing 5M2P to be preferentially excited and monitored. 5-Methyl-2-hydroxypyrimidine is in tautomeric equilibrium with its keto form 5-methyl-2-pyrimidinone. Correlated quantum chemical calculations with large basis sets predict that in the gas phase the enol (5M2HP) tautomer is the dominant species.4 Kistler and Matsika58 performed detailed calculations of the excited-state energies and potential surfaces of gas-phase 5-methyl2-pyrimidinone using multireference configuration-interaction r 2011 American Chemical Society
(MRCI) ab initio methods. However, their calculations refer to the less stable keto tautomer. We previously measured the UV spectrum of 5M2HP using mass-selective two-color resonant two-photon ionization (2CR2PI) spectroscopy, complemented by infrared depletion and holeburning experiments.4 The R2PI spectrum is well resolved Received: July 28, 2011 Revised: October 8, 2011 Published: October 09, 2011 13281
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Table 1. Calculated Rotational Constants of 5-Methyl-2-hydroxypyrimidine in the S0 and S1 States state S0
S1
S1S0
rotational constants
B3LYP/TZVP
RI-CC2/aug-cc-pVTZ
RI-MP2/aug-cc-pVTZ
SCS RI-MP2/aug-cc-pVTZ
A/MHz
5883.0
5813.6
5854.0
5847.4
B/MHz
1561.3
1561.7
1566.4
1562.0
C/MHz
1243.3
1240.5
1245.2
1242.2
A/MHz
6126.4
6057.2
B/MHz
1497.5
1491.6
C/MHz
1212.4
1205.8
A/MHz
243.4
243.6
B/MHz C/MHz
63.8 30.9
70.1 34.7
with many narrow bands, completely unlike those of uracil, thymine, and their methyl derivatives.912 Here, we investigate the electronic origin and lowest vibronic bands of the first excited state of 5M2HP using a laser resolution of 900 MHz (0.03 cm1) in the excitation step. The optically excited S1 state undergoes rapid intersystem crossing (ISC) leading to the lower lying T1 state, which exhibits a >5 μs lifetime.4 The ISC process is rapid enough to eliminate fluorescence but slow enough to obtain vibronically resolved excitation spectra over a range of ∼2000 cm1. The band contour analysis of 5M2HP is complemented by ab initio and density functional calculations of the structure and rotational constants of 5-methyl-2-hydroxypyrimidine and an analysis of the intersystem crossing rate.
II. COMPUTATIONAL METHODS AND RESULTS The electronic ground state of 5M2HP was optimized with the resolution of identity (RI) MøllerPlesset (MP2) method (using the aug-cc-pVXZ (X = D,T,Q) basis sets) and the spincomponent scaled (SCS) RI-MP2 method (using the aug-ccpVTZ basis set). Ground- and excited-state calculations were also performed with the second-order approximate coupledcluster (CC2) method with the resolution of identity (RI) approximation (using the aug-cc-pVXZ (X = D,T) basis sets). For the S1 state, structure optimization was performed starting from a nonsymmetric structure. It converged to a Cs symmetric structure with a planar 2-hydroxypyrimidine ring. Ground- and excited-state calculations were also performed using the B3LPY density functional (using the TZVP basis set). The thresholds for SCF and one-electron density convergence were set to 109 and 108 au, respectively. The convergence thresholds for all structure optimizations were set to 108 au for the energy change, 6 106 au for the maximum displacement element, 106 au for the maximum gradient element, 4 106 au for the rms displacement, and 106 au for the rms gradient. Calculations were performed using Turbomole 6.0.13 In addition, ground-state calculations with the B3LYP method (using the 6-31G(d,p) and 6-31++G(d,p) basis sets) and the MP2 method (using the 6-311++G(d,p) basis set) were performed using Gaussian03.14 The thresholds were set to 109 au for the SCF convergence and VERYTIGHT for geometry optimization. For B3LYP calculations the grid size was set to ultrafine. The rotational constants for the S0 and S1 states for the different methods using the largest respective basis sets are listed in Table 1. An overview of the results with all basis sets is given in the Supporting Information (Table 5). Methyl Group Internal Rotation. The methyl-hindered internal rotation potentials in the S0 and S1 states were calculated
Figure 1. Methyl group torsional potential energy curves for 5-methyl2-hydroxypyrimidine in the S0 and S1 states, calculated at the CC2/augcc-pVDZ, RI-MP2/aug-cc-pVTZ, and B3LYP/TZVP levels.
on a 10 grid starting from the corresponding minimum energy structures. During structure optimization the dihedral angle of one methyl H atom with respect to the pyrimidine ring plane was used as a driving coordinate, while the other structure parameters were optimized at each step. The S0 potentials were calculated at the MP2/aug-cc-pVTZ, CC2/aug-cc-pVDZ, and B3LYP/TZVP levels and the S1 state potentials at the CC2 and time-dependent B3LYP (TD-B3LYP) levels. The calculated energies were leastsquares fitted to a periodic 3-fold (V3) plus 6-fold (V6) potential form15 V ðθÞ ¼
V3 V6 ð1 cos 3θÞ þ ð1 cos 6θÞ 2 2
ð1Þ
The potentials are shown in Figure 1, and the fitted V3 and V6 barrier heights for the different computational methods are summarized in Table 2. All three methods predict the S0 state V3 barrier to lie in the range 1113 cm1. The magnitude of the V6 contribution is close to 4 cm1 for all methods. However, the CC2 method predicts a positive V600 contribution, while the MP2 and B3LYP methods predict it to be negative. The CC2 and TD-B3LYP methods both predict the V3 barrier to increase by 13282
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Table 2. Methyl Torsional Barriers in the S0 and S1 States (in cm1) Calculated at the MP2/aug-cc-pVTZ, CC2/aug-ccpVDZ, and B3LYP/TZVP Levels RI-CC2/
RI-MP2/ aug-cc-pVTZ
barriers
S0
V300
12.8
13.2
11.0
V600
3.8
3.9
3.4
S1
B3LYP/TZVP
aug-cc-pVDZ
state
V300
67.7
69.2
V600
10.2
11.6
about five times (to 6869 cm1) upon S1 r S0 excitation. Again, the CC2 method predicts a positive and the TD-B3LYP method a negative V60 contribution. Hydroxy Group Internal Rotation. The rotation potentials and barrier heights for the OH group internal rotation were calculated at the CC2/aug-cc-pVDZ level. Potential barrier heights were obtained from least-squares fits to a cyclic 2-fold (V2) plus 4-fold (V4) potential. The barriers are high, of the order of 25003000 cm1. Details are given in the Supporting Information. The resulting tunneling splittings will be far smaller than our experimental resolution; we therefore neglect contributions from the hydroxy rotor in the further discussion.
III. EXPERIMENTAL METHODS AND RESULTS III.A. Experimental Methods. 5-Methyl-2-hydroxypyrimidine was synthesized according to ref 16. The molecular-beam/ mass spectrometer experimental setup has been described elsewhere.4 Neon carrier gas (Linde, g99.995%) at ∼1.5 bar backing pressure was passed through a pulsed nozzle (0.4 mm diameter) containing the 5M2HP heated to 170 °C. 2C-R2PI spectra at vibrational (0.4 cm1) resolution were measured with a frequencydoubled Lambda-Physik FL3002 dye laser. The frequency-doubled output of a Radiant Dyes NarrowScan D-R dye laser was employed for the rotational band contour measurements. The bandwidth of this laser is typically 0.025 cm1 (750 MHz) in the visible range, as measured with a HighFinesse Angstrom WS6 high precision wavemeter. The UV bandwidth after frequency doubling is expected to be (2)1/2 wider (0.035 cm1 or 1050 MHz). Because the exact bandwidth in the UV is not known, the Gaussian width to simulate the contours was set to 0.03 cm1 (900 MHz), which is intermediate between the measured frequency width in the visible and the projected width in the UV after frequency doubling. Spectra were recorded at a 0.012 cm1 step size. The wavelength was calibrated by measuring the DCM dye laser fundamental frequency with the WS6 Wavemeter. A laser pulse energy of 100 μJ was used for the high-resolution scans, although spectra taken at pulse energies up to 300 μJ show no signs of broadening. We conclude that the spectra are well below intensities that would lead to optical saturation. Prompt ionization, e.g., without time delay between excitation and ionization laser, was performed using an Ekspla NT342B optical parametric oscillator (UV-OPO). III.B. Resonant Two-Photon Ionization Spectra. Figure 2a shows the vibrationally resolved 2C-R2PI spectrum of supersonic jet-cooled 5M2HP between 31 200 and 33 980 cm1.4 The spectrum exhibits a sharp 000 band at 31 529 cm1. This is 749 cm1 to the blue of the electronic origin of 5-methylpyrimidine17 and 476 cm1 to the blue of the 000 band of
Figure 2. (a) S1 r S0 two-color resonant two-photon ionization spectrum of 5-methyl-2-hydroxypyrimidine with vibronic assignments (ionization at 215 nm). The frequency scale is relative to the electronic origin at 31 529 cm1. (b) Low-energy part, magnified 15, with methyl torsional band structure.
pyrimidine.19 The vibronic bands up to 1200 cm1 have comparable band widths, whereas above 1200 cm1 the rotational contours are perceptibly broadened.4 Here, we will concentrate on the rotational contours of the four lowest intensity bands in this spectrum as well as the associated torsional band structure. Figure 2b shows the low-frequency part of the spectrum on a 15 vertically expanded scale. Weak features 31 and 62 cm1 above the 000 band are associated with methyl torsional excitations that will be discussed in detail below. The comparative weakness of these excitations already indicates that the preferred methyl group orientation is the same in both electronic states. Similar torsional excitations are also observed at 486 and 504 cm1, 14 and 31 cm1 above the intense 473 cm1 band (assigned to 6a10), and at 755, 773, and 804 cm1, i.e., 11, 29, and 60 cm1 above the intense 744 cm1 band (assigned to 1410). Interestingly, however, no methyl torsional bands are observed in conjunction with the band at 532 cm1, assigned to 6b10. The weak band at 397 cm1 has been assigned to either the inplane fundamental 710 or the out-of-plane “butterfly” overtone 320.4 The weak excitations at 427 and 434 cm1, i.e., 30 and 37 cm1 above the 397 cm1 band, might also be methyl torsional bands, 13283
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Figure 3. (Black) Experimental 2C-R2PI rotational contours of the (a) 000, (b) 473 cm1, (c) 532 cm1, and (d) 744 cm1 bands at 0.03 cm1 resolution. (Red) Rotational simulations (including the A and E torsional subbands) using PGOPHER20 and a rigid-rotor Hamiltonian with parameters given in Table 1, Supporting Information.
Figure 4. Experimental contours and perturbed rigid-rotor contour simulations for three bands of 5-methyl-2-hydroxypyrimidine. (a) The 000 band: (top) experimental spectrum, (bottom) contour calculations of the torsional subbands 1E0 r 1E00 (red) and 0A0 r 0A00 (blue), (middle) total calculated spectrum. (b) Same for the +473 cm1 (6a10) band. (c) Same for the +532 cm1 (6b10) band. The respective simulated rotational parameters are given in Table 3, Supporting Information.
but due to their comparative weakness we do not discuss them further. III.C. Rotational Contours. Rigid-Rotor Simulations. Figure 3 shows the rotational contours of the 000, +473 cm1, +532 cm1, and +744 cm1 bands, measured at 0.03 cm1 resolution. Visual inspection reveals two prominent peaks in each band. The broader 1E0 r 1E00 subband lies to the red and the narrower
0A0 r 0A00 subband to the blue, as expected for a molecule that has a larger torsional barrier in the excited state than the ground state. Preliminary simulations of these contours using the rigidrotor program PGOPHER20 (red traces in Figure 3) show that all contours are 100% c type, analogous to the cases of 2- and 5-methylpyrimidine.21 This implies that the transition dipole moment (TDM) is perpendicular to the aromatic ring plane and 13284
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Table 3. Simulated S0 and S1 State Rotational Constants of 5-Methyl-2-hydroxypyrimidine (uncertainties in parentheses), Transition Dipole Moment Orientation, and Gaussian (ΔGauss) and Lorentzian (ΔLor) Broadening Contributions +473 cm1
origin band A subband
parameter
E subband
A subband
+532 cm1
E subband
A subband
E subband
calculation B3LYP/TZVP
A00 /MHz
5850 ((300)
5850 ((300)
5850 ((300)
5850 ((300)
5850 ((300)
5850 ((300)
B00 /MHz
1576 ((60)
1576 ((60)
1576 ((60)
1576 ((60)
1576 ((60)
1576 ((60)
1561.3
C00 /MHz A0 /MHz
1248 ((25) 6291 ((350)
1248 ((25) 6291 ((350)
1248 ((25) 6291 ((350)
1248 ((25) 6291 ((350)
1248 ((25) 6291 ((350)
1248 ((25) 6291 ((350)
1243.3 6126.4
B0 /MHz
1517 ((70)
1517 ((70)
1517 ((70)
1517 ((70)
1517 ((70)
1517 ((70)
1497.5
C0 /MHz
1220 ((30)
1220 ((30)
1220 ((30)
1220 ((30)
1220 ((30)
1220 ((30)
1212.4
Da00 /MHz
10 500 ((800)
10 500 ((800)
10 500 ((800)
Db00 /MHz
276
276
276
Da0 /MHz
9100 ((2000)
9100 ((2000)
9 100 ((2000)
Db0 /MHz
80
80
80
ΔI00 /amu Å2 ΔI0 /amuÅ2
2.1 0.8
2.1 0.8
2.1 0.8
a/b/c character
0/0/100
0/0/100
0/0/100
V300 /cm1
10
V30 /cm1
2.1 0.8
2.1 0.8
0/0/100
0/0/100
10
52
2.1 0.8 0/0/100 10
52
22
ΔGauss/MHz
900
900
900
900
900
900
ΔLor/MHz
3000
3000
5500
5500
5500
5500
Trot/K
6
6
6
6
6
6
the electronic transition is of nπ* type. The 0A0 r 0A00 and 1E0 r 1E00 torsional subbands were simulated separately using the rotational constants given in Table 1. Perturbed Rigid-Rotor Simulations. Previous studies have shown that perturbed rigid-rotor Hamiltonians18 are necessary to adequately reproduce the rotationally resolved spectra of methylated aromatics because of the large-amplitude excursions along the methyl torsional coordinate that are a consequence of low torsional barriers.2123 We used a similar procedure to analyze the rotational contours of 5M2HP. First, simulation of the 0A0 r 0A00 subband was started with the S0 rotational constants from the B3LYP calculation (Table 1). Once the pattern of lines was reproduced, the ground-state rotational constants were adjusted to improve the simulation. Since the calculations predict a small ground-state torsional barrier of ∼12 cm1 and a small inertial defect (which is confirmed by the simulations), we mainly adjusted the A00 constant, since it is most affected by the torsional motion. The excited-state rotational constants were adjusted to reflect the expected smaller inertial defect value in the excited state and varied until a satisfactory agreement with the subband contour was obtained, see Figure 4. Next, the 1E0 r 1E00 subband was simulated by adding torsionrotation terms Da and Db to the Hamiltonian, see ref 18 for details. Their magnitudes were initially set to the values determined for 5-methylpyrimidine.21 Then, both sets of parameters were adjusted for a best visual simulation of the experimental band contour, shown in Figure 4. Of the two torsionrotation constants, Da and Db, the simulation was found to be primarily sensitive to Da and much less sensitive to Db (the quoted values for Db are only estimates). This is a reasonable result sice the rotor axis is essentially parallel to a.18 In the final simulation, the Gaussian contribution to the line shape was estimated to be 900 MHz and the Lorentzian contribution was estimated to be 3000 MHz. The Lorentzian contribution
5883.0
corresponds to a fluorescence lifetime of 53 ps. The rotational temperature is most sensitive to the wings of the contour and best reproduced with a Trot = 6 ( 1 K. In simulating the +473 cm1 vibronic bands (see Figure 4), a similar procedure was followed. Since both vibronic bands originate from the same ground state as the origin, the S0 state rotational constants were kept fixed. The excited-state rotational constants were then varied for the A subbands until satisfactory contour simulations were obtained. The E subbands were then simulated using the A subband excited-state rotational constants and the same torsion-rotation terms as used in the simulation of the 000 E subband. The same Gaussian line shape contribution and Trot was employed as for the simulation of the origin. However, both the 473 and the 532 cm1 bands required substantially larger Lorentzian contributions of about 5500 MHz. This corresponds to fluorescence lifetimes of ∼29 ps. Table 3 lists the values of the parameters that were derived from these simulations and compares them with theory. Uncertainties in the rotational constants and torsionrotation constants were obtained by varying them until noticeable differences occurred in the simulations. Uncertainties in the excited-state parameters were assumed to be slightly larger than those for the ground state. Notably, the facile methyl group motion reduces the measured inertial defect from its “static” value of about 3.3 amu 3 Å2 to values of 2.1 amu 3 Å2 in the ground state and 0.8 amu 3 Å2 in the excited state. III.D. Experimental Methyl Group Torsional Barriers. For calculation of the methyl torsional energy levels we employed the internal axis method (IAM) torsional Hamiltonian15 including V3 and V6 potential terms, as given in eq 1. The methyl rotor axis is taken as the C7C5 bond between the methyl and ring carbon atoms. The reduced internal rotation constants15 Fred00 = 5.505 cm1 and Fred0 = 5.492 cm1 were calculated at the S0 and S1 state RI-CC2-optimized geometries. The one-dimensional torsional 13285
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Figure 6. S0 and S1 state methyl group 1-D torsional potentials for 5-methyl-2-hydroxypyrimidine. The torsional barriers are fitted to the torsional transitions accompanying the 000 band (see the text).
Figure 5. Experimental R2PI spectra showing the methyl rotor subbands (in black) which are compared to the calculated transitions (in red) for the (a) 000, (b) 473 cm1, (c) 532 cm1, and (d) 744 cm1 vibronic bands.
Schr€odinger equation was solved using a discrete variable representation which produced torsional eigenvalues, eigenfunctions for the ground and excited states, as well as the FranckCondon factors (FCFs) associated with the methyl rotor excitations.24 The V300 , V30 , V600 , and V60 parameters were determined iteratively by fitting the positions and intensities of the methyl torsional
subbands associated with the 000, 6a10, 6b10, and 1410 vibronic transitions, see Figure 5. The fits were performed using a genetic algorithm (GA) method:25 The lower/upper limits of the V3 terms were set to 5/20 cm1 for the S0 state and to 50/80 cm1 for the S1 state. The V6 potential terms were initially set to zero. An initial GA population of 30 randomly generated members (chromosomes) was created, where each member is defined by the S0 and S1 state V3 potentials (genes). The fitness of the members was defined with regard to a minimal χ2. Seventy percent of the fittest members were selected as parents. Eighty percent (24 members) of the daughter generation was generated by genetically mixing (uniform crossing over) of randomly selected parameters of two random parent members. Twenty percent of the daughter generation was generated by identical reproduction of the six fittest parent members (elitism). In a mutation step 2% of the parameters were randomly chosen and randomly changed (mutation) within the limits given above. The GA was iterated until χ2 reached a stable minimum. When fitting only the 3-fold terms, we obtained V300 = 13.1 cm1 and V30 = 50.7 cm1. Figure 6 shows the corresponding S0 and S1 state torsional potentials and associated excitations. The calculated torsional spectrum associated with the 000 band is shown in Figure 5a. Due to the increase of the barrier upon S1 r S0 electronic excitation, the 0A10 r 0A100 subband is shifted 2.3 cm1 to the blue of the 1E0 r 1E00 transition, see also Figures 4 and 6. The torsional population in the S0 state is assumed to conform to a Boltzmann distribution with Tvib = 10 K. At this temperature, the intensity of the 1E0 r 1E00 subband becomes approximately equal to the 0A10 r 0A100 subband; this 13286
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Table 4. Experimental and Calculated Methyl Rotor Transition Frequencies, FranckCondon Factors (FCF), and Barrier Heights of 5-Methyl-2-hydroxypyrimidine +473 cm1
origin band transition
exp
FCF calcd calcd
fcfa
exp
+532 cm1
FCF calcd calcd FCF exp FCF calcd calcd
+744 cm1 FCF
exp
FCF calcd calcd FCF
1E0 r 1E00 (cm1)
0.0 0.96
0.0
0.0
0.92
0.0 0.81
0.0
0.0
0.92
0.0
1.00
0.0
0.0
0.95
0.0 0.89
0.0
0.0
0.92
0A10 r 0A100 (cm1)
2.2 1.00
2.3
2.3
1.00
2.1 1.00
2.6
2.5
1.00
0.4
1.00
0.4
0.4
1.00
2.1 1.00
2.2
2.3
1.00
12.5 0.04 30.5 0.06
12.4 30.1
12.5 30.5
0.07 14.8 0.05 14.5 0.08 32.1 0.09 32.3
14.4 32.5
0.07 0.09
1.7c 1.7 c 0.07 12.5 0.10 19.4d 19.7d
