Solvent-dependent photoinduced tautomerization of 2-(2′-hydroxyphenyl)benzoxazole

Article abstract of DOI:10.1021/jp013915o

The solvent-dependent ground-state conformational equilibrium and excited-state dynamics of 2-(2′-hydroxyphenyl)benzoxazole have been characterized in several solvents on the femtosecond to nanosecond time scales. The only observable ground-state tautomer

Full text of DOI:10.1021/jp013915o

J. Phys. Chem. A 2002, 106, 3665-3672
3665
Solvent-Dependent Photoinduced Tautomerization of 2-(2′-Hydroxyphenyl)benzoxazole
Osama K. Abou-Zied,† Ralph Jimenez,† Elizabeth H. Z. Thompson,‡ David P. Millar,‡ and
Floyd E. Romesberg*,†
Departments of Chemistry and Molecular Biology, The Scripps Research Institute, 10550 N. Torrey Pines Rd.,
Maildrop CVN22, La Jolla, California 92037
ReceiVed: October 22, 2001; In Final Form: January 22, 2002
The solvent-dependent ground-state conformational equilibrium and excited-state dynamics of 2-(2′-
hydroxyphenyl)benzoxazole have been characterized in several solvents on the femtosecond to nanosecond
time scales. The only observable ground-state tautomer is the enol, which exists in equilibrium between the
syn- and anti-rotational isomers. In the anti-enol isomer, the phenyl hydroxyl group appears to not interact
strongly with solvent but rather forms a strong intramolecular hydrogen bond with the benzoxazole oxygen
atom. In the syn-enol isomer, the phenyl hydroxyl proton may interact with solvent or form an internal hydrogen
bond with the benzoxazole nitrogen atom. Upon excitation, the proton is transferred from the oxygen atom
to the nitrogen atom of the internally hydrogen bonded syn-enol isomer in 170 fs, regardless of the solvent.
The lifetime of the resulting excited keto tautomer is solvent dependent and on the order of picoseconds. In
addition to these dynamics, several additional dynamic processes are detected which may correspond to
relaxation of a distorted excited keto tautomer.
Introduction
Proton-transfer reactions are of fundamental importance in
chemistry and biology. A variety of molecules have intramo-
lecular hydrogen bonds (H-bonds) that may be photoinduced
to undergo proton transfer.1,2 This process is known as excited-
state intramolecular proton transfer, or ESIPT. Molecules that
can undergo ESIPT allow for the controlled initiation and
subsequent time-dependent characterization of both the H-bond
itself and the proton transfer process. We have recently proposed
one such molecule, 2-(2′-hydroxyphenyl)benzoxazole (HBO),
as a structural mimic of a DNA base pair for which tautomer-
ization may be initiated at a defined time and position within
duplex DNA (Figure 1).3 In contrast to previous model base
pairs, such as dimers of 7-azaindole,4 HBO may be incorporated
into duplex DNA.3 Therefore, HBO may be used to probe the
biologically relevant duplex environment. Experimental (UV/
vis, CD, duplex melting temperature) and theoretical studies
(molecular dynamics simulations) of DNA containing the HBO
model base pair demonstrate that, in a given DNA sequence
context, HBO packs within an A-form duplex and that the
resulting duplex has a structure and stability similar to a fully
native duplex. The conformational and tautomerization dynamics
of the model base pair are dominated by interbase packing
effects that are unique to the duplex environment.5 To fully
interpret the effects of the biological environment on the
dynamics of the HBO model base pair it is important to
understand the dynamics of HBO in a variety of simple solvent
environments.
Figure 1. (a) Tautomerization of a natural DNA base pair and ESIPT
in HBO. (b) Modeled structure of DNA duplex containing an HBO
model base pair.3,5
loxazole (HPMO).6-17 In the ground state, HBO, HBT, HPI,
and HPMO exist in a conformational equilibrium between the
syn- and anti-enols. In addition, the phenoxy group of each syn-
enol may form an internal H-bond or an intermolecular H-bond
with a solvent molecule (referred to as “solvated” or “open”).
Only the internally H-bonded syn-enol efficiently forms the keto
tautomer due to favorable electronic (∆pKa) and structural
properties (H-bond). Proton transfer does not occur in the anti-
enol species due to an insufficient increase in the basicity of
Several studies have documented the solvent-dependent con-
formational isomerism of HBO and several analogues, including
2-(2′-hydroxyphenyl)benzothiazole (HBT), 2-(2′-hydroxyphen-
yl)benzimidazole (HBI), and 2-(2′-hydroxyphenyl)-4-methy-
* Corresponding author. E-mail: floyd@scripps.edu. Fax: (858) 784
7472.
† Department of Chemistry.
‡ Department of Molecular Biology.
10.1021/jp013915o CCC: $22.00 © 2002 American Chemical Society
Published on Web 03/26/2002

3666 J. Phys. Chem. A, Vol. 106, No. 15, 2002
Abou-Zied et al.
the oxygen, sulfur, and secondary amine groups in HBO, HBT,
and HBI, respectively, upon excitation. In the syn-enol ESIPT
results in diminished fluorescence in the normal Stokes-shifted
emission region, ascribable to the excited enol, and instead
fluorescence is observed at longer wavelengths, due to the
excited keto (ESIPT product). In non-H-bonding solvents, such
as hexane, which are unable to effectively compete for the
phenolic proton, excitation results in very efficient ESIPT and
long-wavelength fluorescence. However, in protic solvents,
which are more able to compete for the phenolic proton,
emission from both enol and keto are observed with intensities
that depend on the concentration of the syn-enol relative to the
solvated and anti-enols.
ond time-resolved fluorescence (TRF) spectroscopy, were
employed to access the wide range of time scales required to
follow all of the ESIPT dynamics and assign the molecular
origin of each signal.
Materials and Methods
Sample Preparation and Characterization. HBO was
purchased from Aldrich and purified by vacuum sublimation.
2-(4-Biphenylyl)-6-phenylbenzoxazotetrasulfonic acid potassium
salt (furan-2) was purchased from Lambda Physik. MBO was
synthesized by dissolving HBO in N,N-dimethylformamide
(DMF) and adding a slight excess of K2CO3 and methyl iodide
and purified by column chromotography. HPLC grade hexane
and methanol (Fischer Scientific) and DMSO (Aldrich) were
used without further purification.
Steady-State Absorption and Fluorescence. Steady-state
absorption spectra were measured using a Cary 300 BIO
spectrophotometer. Samples were contained in a 1 cm path
length quartz cell containing approximately 50 µM solutions,
and fluorescence spectra were measured on Spex Fluorolog
spectrometer in 1 cm path length quartz cell using a right angle
detection configuration.
Several time-resolved studies have characterized the HBO
proton-transfer dynamics and the lifetime of the enol and keto
tautomer excited states. The short wavelength fluorescence of
HBO was reported to have a decay time constant of 1080 (
360 ps in dimethyl sulfoxide (DMSO)8 In HBI, the excited-
state lifetime for the anti-enol species is reported in neutral
aqueous and ethanol solutions to be 370 ps and 1.5 ns,
respectively.9,10 Most of the published work on HBO has only
been able to assign an upper limit (∼10 ps) to the ESIPT time
constant.8 With femtosecond time resolution, Ernsting and co-
workers measured a gain in the transient absorption signal of
60 ( 30 fs, which was assigned to keto tautomer formation in
the excited state.11 However, the pulse duration used in the
experiment (170 fs) indicates that the measured time constant
for the HBO ESIPT process may not be accurate. Laermer and
co-workers measured an HBT ESIPT time constant in tetra-
chloroethylene of 170 ( 20 fs.12 The ESIPT reaction has also
been observed for HBT in polar solvents, but the proton-transfer
kinetics do not show simple monoexponential behavior, although
they do occur on a similar time scale as that observed in
tetrachloroethylene.13 Only an upper limit of 100 ps has been
determined for the proton-transfer time constant in HBI due to
insufficient time resolution.10 HPMO ESIPT is thought to
proceed by two distinct mechanisms.14 The first, a direct proton
transfer, was characterized by a 100 fs time constant, while the
second pathway involved twisting motion of the biaryl ring
system and was characterized by two time constants of 200 fs
and 3 ps. The HBO keto lifetime was found to be solvent
dependent, displaying an apparent inverse relationship with the
solvent dielectric constant and ranging from ∼70 to ∼ 400 ps.15
The lifetime of the keto tautomer of HBT in solution at room
temperature lies in the subnanosecond range.16-18 The decay
profile is biexponential and wavelength-dependent either as a
result of vibrational relaxation processes or due to emission from
both syn- and anti-keto species. In HBI, fluorescence lifetime
for the keto species is reported in neutral aqueous and ethanol
solutions to be 1.8 and 4.1 ns, respectively.9,10 With HPMO,
the planar excited-state ketone was found to have a 2-6 ps
lifetime in p-dioxane.14
Transient Absorption (TA). The transient absorption mea-
surements were performed using a pump-probe setup. A home-
built, diode laser (Spectra Physics Millenia) pumped, mode-
locked, titanium:sapphire oscillator produces sub-30-fs pulses
centered at 800 nm. The pulses are amplified by a commercial
chirped pulse amplifier system (Spectra Physics Spitfire/
Evolution). The amplifier produces 40-50 fs pulses centered
at 800 nm at a 1-5 kHz repetition rate. Frequency conversion
is accomplished with a home-built optical parametric amplifier
(OPA) and is based on the design of Yakovlev et al.19 The signal
beam from the OPA is subsequently doubled twice in two type
I BBO crystals to generate approximately 10-20 µJ energy in
the UV region (300-340 nm). The pulses are compressed by
double passing through a pair of fused silica prisms to
compensate for the group velocity dispersion.
In the one-color TA experiments, the pulses were split into
a pump pulse (2/3) and a probe pulse (1/3). The pump pulse
traveled a constant distance to the sample cell while the probe
pulse was delayed in time before being directed into the cell.
In the two-color TA experiments, light from the idler beam of
the OPA was doubled twice to obtain a probe pulse with a
wavelength in the blue region (420-460 nm). A beam splitter
was used to split a small portion of the probe beam to be used
as a reference beam. The excitation, probe, and reference beams
were focused into the sample with a fused silica lens (f ≈ 10
cm), but only the excitation and signal beams were overlapped
in the sample. The sample was contained in a 0.2 mm quartz
flow cell. After passing through the sample, both signal and
reference beams were detected using silicon photodiodes. The
signal levels were recorded by lock-in amplifiers referenced to
a phase-locked chopper (synchronized to the amplifier Q-
switch), which blocks half the pulses in the excitation beam.
The delay stage and data acquisition were controlled with a PC.
It was typically possible to measure samples with optical density
on the order of 10-4, after averaging the signal from ∼5000
pulses for each delay position. All experiments were carried
out at room temperature. The finite response time of the system
was determined by recording the absorption change of furan-2
in water. The data were fit by numerical iterative convolution
using a variable Gaussian instrument response function to obtain
the best fit with the observed decay, as judged by the distribution
of weighted residuals. The calculated full-width at half-
No comprehensive characterization of conformation equilib-
rium and excited-state dynamics of HBO has been reported.
Therefore, to develop a more thorough understanding of the
environmental factors effecting the tautomerization of HBO, a
steady-state and time-resolved characterization of HBO was
carried out in solvents of differing polarities and H-bonding
abilities. Three solvents were chosen, hexane, in which virtually
only low-energy keto HBO fluorescence is detected, methanol
(MeOH), where enol and keto fluorescence are both detected,
and DMSO, in which most of the detected fluorescence is from
the enol species. Femtosecond one-color and two-color transient
absorption (TA) pump-probe experiments, as well as picosec-

Photoinduced Tautomerization
J. Phys. Chem. A, Vol. 106, No. 15, 2002 3667
Figure 3. Steady-state emission spectra of HBO in (a) hexane, (b)
methanol, and (c) DMSO. The spectra were measured at different
excitation wavelengths as indicated in the figure and the text.
and IR spectroscopy.8 The presence of an HBO anion is
excluded on the basis of the work of Krishnamurthy and Dogra7
as well as Mosquera et al.10 HBO shows two, low-energy ππ*
absorptions. Following the assignments of Do¨rr et al., an
absorption at ca. 320 nm is assigned to the anti-enol and an
absorption at ca. 330-334 nm corresponds to the syn-enol.8
Both transitions are at the same wavelength in hexane and
DMSO (321 and 334 nm) but slightly blue-shifted in methanol
(319 and 330 nm). This result is not consistent with simple
polarity effects and may derive from specific H-bonding between
the methanol donor and either the benzoxazole nitrogen or
oxygen acceptors. A weak absorption band at ∼370 nm was
observed in DMSO. This absorption was assigned by Do¨rr et
al. to the anti-keto tautomer. The fluorescence spectrum consists
of two strong emission bands at ca. 362-370 nm and 474-
500 nm, whose peak frequencies and relative intensities are
solvent dependent. The UV emission arises from the enol
tautomer, while the lower energy emission arises from the keto
tautomer after ultrafast ESIPT from the initially prepared enol
tautomer. These assignments are supported by absence of the
low-energy emission band in the fluorescence spectrum of 2-(2′-
methoxyphenyl)benzoxazole (MBO, Figure 2), which, due to
the methoxy group, serves as a non-proton-transfer model. The
keto emission band is dominant in hexane (at 500 nm), keto
and enol (at 365 nm) emission bands are of equal intensity in
methanol, and the enol emission band is dominant in DMSO.
The enol emission spectrum is shifted to lower energy in
methanol and DMSO (365 and 370 nm, respectively) relative
to that in hexane (362 nm). The opposite trend is observed for
keto emission where the emission wavelength is shifted to higher
energy in methanol and DMSO (474 and 463 nm, respectively)
relative to that in hexane (500 nm).
Figure 2. Absorption (solid lines) and emission (dashed lines) spectra
of HBO dissolved in hexane, methanol, and DMSO. The spectra of
MBO dissolved in hexane is also shown. The excitation wavelength
was 320 nm.
maximum (fwhm) was ∼89 fs in both the one- and two-color
experiments. The fit also serves to locate time zero. Due to the
use of a single OPA, a change in the pump wavelength required
a change in the probe wavelength. Therefore, to examine the
wavelength dependence of the TA signals, three two-color
experiments were conducted with pump/probe wavelengths of
320/430, 333/445, and 338/453 nm.
Time-Resolved Fluorescence (TRF). The time-dependent
emission was measured using time-correlated single photon
counting which has been described elsewhere.20 Briefly, samples
contained in quartz cells (1 cm path length) were excited at
333 nm using the frequency-doubled output from a synchro-
nously pumped, mode-locked, and cavity-dumped dye laser
(Coherent 702). Sample emission was collected at right angles
to the excitation beam, collimated by a lens, passed through a
motor-controlled polarizer, and focused on the entrance slit of
a monochromator (Jobin-Yvon H-10). Emission at 362-370 and
480-500 nm was examined. The instrument response function
of the system was obtained by recording the scatter of the second
harmonic of the excitation pulse and determined to have a width
of 100 ps (fwhm).
Fluorescence emission spectra measured at various excitation
wavelengths in different solvents (Figure 3) revealed the relative
concentrations of the solvated syn- and anti-enols that contribute
to the emission band at 362-370 nm. In hexane, fluorescence
in this region is negligible after excitation in the 333 nm region
(syn-enol λmax), while fluorescence at these wavelengths is
maximal following excitation at 318 nm (anti-enol λmax).
Therefore, we assign the majority of HBO emission in hexane
in this region to fluorescence of the excited anti-conformer. In
methanol and DMSO, the fluorescence emission spectra mea-
Results
Absorption and Fluorescence Steady-State Spectra. To
characterize the effects of solvation on the steady-state spec-
troscopic properties of HBO, the absorption and emission spectra
were recorded in hexane, MeOH, and DMSO (Figure 2).
Features in the absorption spectra of HBO have previously been
assigned by Do¨rr et al. on the basis of a comparison with model
compounds where H-bonding is not possible, as well as 1H NMR

3668 J. Phys. Chem. A, Vol. 106, No. 15, 2002
Abou-Zied et al.
TABLE 1: Time Constants (ps) Measured by TA and TRFa
and Their Assignments
λdetection
,
solvated
enol
b
solvent
hexane
nm
anti-enol
τn
keto
Transient Absorption
12 (0.50)c
20 (0.12)d
295 (0.50)
295 (0.88)
0.56 (rise)e,f 295
methanol
DMSO
12 (0.67)c
68 (0.33)
0.56 (rise)e 68
12 (0.15)c
57 (0.85)
0.56 (rise)e 57
Time-Resolved Fluorescence
1370 (0.50)
hexane
362
500
295 (0.50)
295
methanol 365 510 (0.64) 1560 (0.36)
474 1560 (<1%)
370 970 (0.35)h 1380 (0.65)h
463 1380 (<1%)
68g (>99%)
57g (>99%)
DMSO
a Unless noted, time constants represent decays. Relative contribu-
tions are listed in parentheses. b Unassigned intermediate time scale
dynamics. See text for details. c One-color TA (333/333 nm). d Two-
f
color TA (320/430 nm). e Two-color TA (333/445 nm). Two-color TA
(338/453 nm). g Fit based on TA data. h See text for assignment.
Figure 4. Femtosecond transient absorption of HBO in hexane (top)
and furan-2 in water (bottom). Excitation and probe wavelengths were
333 and 445 nm, respectively. The data are plotted as a function of the
delay time between pump and probe pulses. The rise in furan-2 reflects
the time-integrated cross-correlation function of the two pulses. The
rise time onset of the signal in HBO reveals the formation of the keto
species by ESIPT. The solid lines represent the best fits from a rate
equation model as explained in the text.
sured at various excitation wavelengths showed a considerable
contribution from the excited states of both solvated syn- and
anti-conformers to the emission in this region. Therefore, relative
to hexane, MeOH and DMSO increase the stability of the
solvated syn-enol relative to the anti-enol of HBO, presumably
due to stronger H-bonding between the solvent and the HBO
hydroxyl group in the syn conformation.
The methoxy derivative of HBO (MBO) shows absorption
maxima in hexane (313 and 324 nm) that are slightly blue-
shifted relative to those of HBO (Figure 2), which may result
from steric effects induced by the larger methoxy group of MBO
relative to the smaller hydroxy group of HBO. The larger
methoxy group may interfere with coplanarity of the two
aromatic rings and results in reduced delocalization of electron
density through the conjugated biaryl system. The emission of
MBO is more structured and blue-shifted relative to HBO. The
nearly mirror image relationship to the absorption spectrum is
consistent with the absence of ESIPT-mediated relaxation in
the excited state.
Time-Resolved Absorption and Fluorescence. Time-
resolved experiments were performed to characterize the
dynamics of HBO in each solvent (Table 1). One- and two-
color TA was employed to measure the fast dynamics (100 fs
to 300 ps), and TRF was used to measure slower decays (100
ps to 20 ns). The one-color TA experiments may be complicated
by probe pulse (333 nm) absorption in either enol or keto excited
states in addition to ground-state absorption. Two-color TA
experiments were performed with pump/probe wavelngths of
320/430, 333/445, and 338/453 nm to examine the wavelength
dependence. The TRF detection wavelength was chosen on the
basis of the steady-state emission maxima, described above, to
probe specific emitting species. The average lifetime values are
reported for cases where the same decay component was
measured with both methods or for cases where the component
was detected at two different fluorescence detection wave-
lengths.
Figure 5. Femtosecond transient absorption of HBO in hexane in the
intermediate time frame from 0 to 5 ps. Pump-probe wavelengths
employed are indicated in each case. The solid lines are the best fits.
The analysis gives a rise time of 170 fs in all the transients followed
by a decay of 12 ps in the one-color experiment, a decay of 20 ps in
the 320/430 nm two-color experiment, and a rise time of 560 fs in the
333/345 and 338/453 nm experiments. The fit is calculated from time
origin in the case where an absorption decrease around zero delay is
observed.
color experiment (333/445 nm) in Figure 4. For comparison,
the presumably response limited rise time of furan-2 is also
shown. The pump pulse prepares the system in the enol excited
state, from which the syn-tautomer promptly tautomerizes by
ESIPT to the keto tautomer. The fast time scale dynamics of
HBO lead to a signal rise after time zero as shown in Figure 4,
which reflects proton transfer. The rise time is apparent and
identical in each experiment. The rise time is attributed to
absorption of the excited-state keto. The data are fit with a single
Transient Absorption Spectroscopy. The TA data for HBO
in hexane are shown in Figures 4, 5, and 6a. The small decrease
in intensity around zero delay may be due to absorption from
the S1 state of the enol, as suggested previously.21 The data
from the one- and two-color experiments are collected in Table
1. The fastest time scale dynamics are illustrated for the two-

Photoinduced Tautomerization
J. Phys. Chem. A, Vol. 106, No. 15, 2002 3669
Figure 7. Single-color (333 nm) femtosecond transient absorption of
HBO and MBO in hexane. Solid lines represent the best fit to the data
calculated from time zero.
time of 560 fs was observed in the 333/445 and 338/453 nm
TA data. The one-color (333/333 nm) and the two-color (333/
445 nm) experiments were also performed with HBO in MeOH
and DMSO, which showed identical time constants (a 12 ps
decay and a 560 fs rise, for the one- and two-color experiments,
respectively). The molecular origin(s) of this intermediate time
scale decay or rise component is unclear (vide infra). Oscillations
which persist for at least 5 ps are observed in almost all of the
transients (Figure 5). These oscillations may be due to a vibronic
wave packet that is created in the keto S1 state after ESIPT.
However, the presence of such oscillations in MBO indicates
that this wave packet motion may be associated with vibrational
modes in the S0 or S1 state of the enol tautomer.
HBO also possesses longer time scale dynamics in hexane.
A 295 ps decay is observed in the one- and two-color (333/
445, 320/430 nm) TA experiments. This time constant is
consistent with that observed in cyclohexane for the excited-
state keto lifetime (257 ps).15 The observed 295 ps decay is
therefore assigned to excited-state keto lifetime. One color and
two-color (333/445 nm) TA data for HBO in MeOH and DMSO
showed a 68 and 57 ps decays, respectively, again assigned to
the keto excited-state lifetime, in agreement with previous
studies.8,15
Time-Resolved Fluorescence Studies. The TRF transients
of HBO in hexane monitored in either the enol emission re-
gion (362 nm) or keto emission region (500 nm) are shown in
Figure 6b. With 362 nm detection, a biexponential decay was
observed with short and long components of 295 and 1370 ps,
respectively. The 295 ps decay matches the TA decay and is
assigned to the excited keto lifetime (detected in the very blue
edge of the emission band), and the 1370 ps decay is assigned
to lifetime of the excited anti-enol, based on the excitation
dependent study described above. With detection at 500 nm,
only a 295 ps decay was observed, which is consistent with
excited keto emission.
The TRF of HBO in methanol was monitored at either 365
or 474 nm. At 365 nm, two decays were apparent with time
constants of 510 and 1560 ps, corresponding to the two enol
conformers, as shown in the excitation-dependent experiments
described above. The 1560 ps decay may be assigned to the
excited anti-enol lifetime on the basis of the similarity to the
long time scale observed in hexane. By elimination, the 510 ps
Figure 6. Time-resolved (a) transient absorption and (b) fluorescence
of HBO in hexane (0-20 ns). Transient absorption is shown for the
one-color experiment (closed circles) and the 333/445 nm two-color
experiment (open circles). The excitation wavelength in the TRF
experiment was 333 nm, and the detection wavelength is shown for
each case. The data from the TRF experiment were fit to a biexponential
decay convoluted with the instrument response function shown in the
figure as a dashed line.
rise exponential function convoluted with an 89 fs pulse width.
A time constant for the proton transfer of 170 ( 20 fs was
determined. This conclusion is confirmed by measuring the TA
of MBO, which is incapable of tautomerization and shows no
rise time (Figure 7). This time constant may not directly
represent the ESIPT rate constant. The use of a time constant
determined from monitoring a single wavelength requires two
assumptions. The transition moment must be wavelength
independent (Condon approximation), and the time constant
must not reflect vibrational relaxation within the product
manifold. It is unclear to what extent these approximations are
justified in this study. Nevertheless, it is clear by comparison
with MBO that the 170 fs time scale is associated with the
proton transfer and, therefore, represents an upper limit to the
transfer time. Within experimental error, the same time constant
was observed in MeOH, MeOD, and DMSO, indicating that
the proton transfer is solvent and isotope independent. These
results agree with those obtained previously for the thiazole
analogue HBT.12,13
The TA experiments also exhibited a 0.5-20 ps time scale
process. The observed process was strongly dependent upon
the pump and probe wavelengths (Figure 5). The single color
data (333/333 nm) showed a decay with a time constant of 12
ps, while the two-color data (320/430 nm) showed a similar
decay with a slower time constant of 20 ps. However, a rise

3670 J. Phys. Chem. A, Vol. 106, No. 15, 2002
Abou-Zied et al.
Figure 8. Ground-state equilibria and excited-state dynamics of HBO.
decay is assigned to the solvated syn-enol excited state. With
474 nm detection an instrument-limited decay was detected that
must correspond to the decay of the keto excited state (measured
above to be 68 ps). A small amplitude component (<1%) of
the 1560 ps decay was also present due to detection of the low-
energy tail of the anti-enol fluorescence.
enol relative to the solvated and anti-enols. This ratio is large
(>20:1) in hexane, demonstrating the almost exclusive presence
of the internally H-bonded syn-enol, but drops to ∼1:1 and ∼1:6
in methanol and DMSO, respectively, due to the increased
H-bonding capacity of these solvents (Figure 2).
The equilibrium between the syn- and anti-conformers is also
solvent dependent. On the basis of the excitation-dependent
emission experiments and the time-resolved studies reported in
this work, as well as the calculations by Do¨rr and co-workers,8
we assign the 318 nm absorption to the anti-enol and the 333
nm absorption to the syn isomer. In hexane HBO exists primarily
as the syn-enol in equilibrium with a small concentration of
the anti-enol. In MeOH and DMSO, the fluorescence excitation
spectra showed a large contribution from the excited states of
both solvated syn- and anti-conformers to the emission at 370
nm. Relative to hexane, DMSO and MeOH each selectively
increases the stability of the solvated syn-enol of HBO relative
to the anti-enol. This stabilization may result from specific
H-bonding interactions or from polarity effects.
The TRF of HBO in DMSO was monitored at either 370 or
463 nm. At 370 nm, two decays were apparent with time
constants of 970 and 1380 ps. The difference between these
time constants is not as large as those in MeOH, and therefore,
their assignment is more difficult. In analogy to the experiments
described above, the 970 and 1380 ps decays may be assigned
to the solvated syn- and anti-enols, respectively. However, we
note that the relative amplitudes of the two decays are more
consistent with the opposite assignments considering excitation
at 333 nm (maxima of the syn-enol absorption). With 463 nm
detection a pulse limited decay was also detected and is assigned
to keto excited state (on the basis of the TA experiments
described above, the lifetime is 57 ps). As with methanol, a
small contribution (<1%) from the anti-enol fluorescence decay
was detected at 463 nm. The assignments of the two enol
lifetimes are consistent with previous studies.8,15
Solvent-Dependent Dynamics of HBO. There are at least
three components to HBO tautomerization: enol dynamics;
proton-transfer dynamics; keto dynamics. The solvent-dependent
dynamics of each component has been characterized. After
excitation, the solvated syn- and anti-enols fluoresce back to
their respective ground states, while the internally H-bonded
syn-enol efficiently tautomerizes to the keto form. The lifetime
of the anti-enol, ca. 1400 ps, showed no solvent dependence.
Since it is expected that an intermolecular H-bond with solvent
would contribute to the structural and electronic properties of
the anti-enol, the solvent independence implies the existence
of an intramolecular H-bond. As expected, the solvated syn-
enol shows a solvent-dependent lifetime. Although the TRF
signal was only detectable in methanol and DMSO, the lifetime
was twice as long in DMSO compared to methanol. The
increased solvent sensitivity of the solvated syn-enol is expected,
relative to an intramolecularly H-bonded anti-enol, due to the
intimate involvement of the solvent in this structure. An
equilibrium between an unsolvated anti-enol and a solvated syn-
enol is consistent with the observation that the equilibrium is
Discussion
Solvent-Dependent Conformational Equilibrium of HBO.
The solvent-dependent conformational equilibrium of HBO
is depicted in Figure 8. Two rotational conformers of HBO exist
in the ground state, the syn- and anti-enols. Moreover, two syn-
enol isomers exist in a solvent-dependent equilibrium. Of the
two isomers, one syn-enol has an internal O-H---N bond and
one “open” or “solvated” syn-enol possesses intermolecular
H-bonds with solvent. These syn-enols may be differentiated
experimentally. Only the “closed” syn-enol has an intramolecular
O-H---N bond that is required for ESIPT. Therefore, photo-
excitation results in efficient ESIPT and strongly red-shifted
fluorescence at approximately 500 nm. Excitation of the solvated
syn- or anti-enols results in fluorescence at ∼370 nm, as no
ESIPT is possible. The ratio of these two steady-state emission
bands may be used to quantitate the concentration of the syn-

Photoinduced Tautomerization
J. Phys. Chem. A, Vol. 106, No. 15, 2002 3671
shifted toward the syn-enol with the stronger interacting solvents
MeOH and DMSO, relative to hexane.
the total decay relative to the 295 ps component (1:9 relative
amplitudes). The lower energy probe experiments (333/445 and
338/453 nm) each detect a 560 fs rise time. Unfortunately the
uncertainty in the relative amplitudes of the rise times prevents
an assessment of their relative contributions.
The closed syn-enol, which has an intramolecular H-bond,
undergoes efficient proton transfer upon excitation. We observed
a 170 fs rise time in the TA measurements, which we assign to
the excited-state proton transfer. This rise time is independent
of solvent, as well as H to D isotope substitution, consistent
with a strongly exothermic proton transfer with weak coupling
to solvent degrees of freedom. The isotope independence has
often been interpreted as resulting from a barrier-free ESIPT,
where the rate is controlled by C-O-H bending vibrations that
deliver the proton to the acceptor. However, H to D isotope
substitution should decrease the frequency of such bending
vibrations by a factor of approximately 1.4, predicting a proton-
transfer time constant of ∼240 fs for the heavier isotope, which
would be easily detected by the TA experiment. It is therefore
possible that the ESIPT occurs adiabatically with a small
activation barrier involving HBO vibrations that do not include
the H or D atoms, such as torsional motion within the biaryl
ring system.
These decay and rise components may derive from multiple
processes whose relative contribution depends on the pump and
probe wavelengths. It is possible that the pump pulse excites
different ground states or that the probe pulse observes different
combinations of ground and excited states. However, it is
possible to speculate on the dynamics that might give rise to
these data. For example, the 12 ps decays might represent
rethermalization of the ground-state molecules disturbed by the
probe pulse, which is consistent with the observation of a similar
time scale process (7 ps) in MBO. The rise time might
correspond to vibrational cooling within the excited keto. Chou
et al. reported a 8-10 ps rise time during fluorescence
upconversion studies of ESIPT in 10-hydroxybenzo[h]quino-
line.26 These authors ascribed this dynamics to vibrational
cooling of the excited keto that is initially prepared in a
vibrationally hot state after ESIPT, as is also likely with HBO.
While 560 fs is considerably faster than 10 ps, it is not
unreasonable compared to other vibrational redistribution
processes. Zewail et al. observed a decay for HPMO that was
fit with time constants of approximately 200 fs and 3 ps and
was ascribed to a proton transfer involving initial stretching of
the O-H bond followed by torsional motion across a low
barrier.14 It is also possible that the observed dynamics with
HBO reflect a complex superposition of several of the above-
mentioned processes. Additional experiments, particularly fem-
tosecond fluorescence upconversion, are required to further
address these issues.
This model of proton transfer is supported by the conclusions
of previous studies of related ESIPT systems. For example,
Chudoba et al. proposed that ESIPT in 2-(2′-hydroxy-5′-
methylphenyl)benzotriazole is modulated by a highly anhar-
monic in-plane vibration which modulates the separation
between proton donor and acceptor atoms and leads the system
to a barrierless channel on the excited-state potential.22 Ab initio
study of the ground- and excited-state proton transfer in HBO
at the SCF and CIS levels showed that in the course of excited-
state nuclear motion the O and N atoms are brought into
proximity by in-plane bending.23 The calculations predicted a
decreasing barrier with decreasing O---N distance. In HPMO,
a molecule very similar to HBO, two mechanisms were proposed
for the proton-transfer dynamics.14 A direct and barierless proton
transfer takes place in ∼100 fs that conserves the system
planarity. A second proton transfer route results from a
bifurcation of the initially excited wave packet in which some
trajectories evolve to the keto state with concomitant rotation
of the biaryl bond. Tautomerization along this twisting reaction
coordinate involves an energy barrier, as predicted theoreti-
cally.24 For similar systems involving internally H-bonded six-
membered rings, Sobolewski et al. predicted barrierless proton-
transfer on the basis of CASSCF and CIS electronic-structure
calculations.25
Conclusions
In the ground state, HBO exists as an equilibrium between
the syn- and anti- enols. The solvent-independent lifetime of
the anti-enol implies that this rotamer has an internal H-bond
between the phenolic hydroxyl group and the benzoxazole
oxygen atom. The syn-conformer exists in equilibrium between
an internally H-bonded enol and the solvated syn-enol. The
lifetime of the solvated syn-enol is longer in the H-bond
accepting solvent (DMSO), relative to the H-bond donating
solvent (MeOH). The syn-enol efficiently undergoes ESIPT
upon photoexcitation. The lifetime of the keto proton transfer
product is significantly longer in the weakly solvating environ-
ment of hexane relative to MeOH and DMSO.
The proton transfer occurs in 170 fs and is independent of
solvent or isotope. This implies that the proton transfer occurs
over a low barrier in the excited state and involves motions of
the molecule that are not strongly sensitive to O-H motion,
perhaps intra-aryl ring torsional motion, as is thought to play
an important role in related derivatives.
The lifetime of the excited keto shows a stronger solvent
dependence than did the enol isomers described above. The
decays varied by a factor of 5 in the different solvents. The
lifetime was the shortest in DMSO (57 ps), slightly longer in
MeOH (68 ps), and significantly longer in hexane (295 ps).
These data are consistent with that reported by Do¨rr and co-
workers, who measured the lifetime of the excited HBO keto
state in a large number of solvents.15 They measured lifetimes
that ranged from 69 ps in DMSO to 413 ps in carbon
tetrachloride and generally decreased with increasing solvent
polarity.
A process which occurred on a 500 fs to 20 ps time scale
that is intermediate between the ESIPT and keto lifetimes was
also observed. This process may correspond to torsional motion
of an alternative ketone product that arises from a second proton-
transfer pathway, as has been suggested for the analogous
compound HPMO.14 Additional experiments are currently in
progress to further define the molecular nature of this process.
In addition to the minimal number of dynamic processes (enol
lifetime, proton transfer rate, and keto lifetime) additional time
scale dynamics are reflected in the data. In each TA experiment,
one additional component is detected, which has a strong
wavelength dependence. With the one-color (333/333 nm) TA
this process appears as a 12 ps decay that is a major component
of the relaxation; the relative amplitude of the decay is equal
to that of the 295 ps decay. The two-color (320/430 nm) TA
detects a slower 20 ps decay which is a smaller component of
Acknowledgment. We thank Professor Julius Rebek for use
of the Spex Fluorolog spectrometer. Funding was provided by
the Skaggs Institute for Chemical Biology.
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