Time-resolved Visible Absorption Research Articles

Excited State Dynamics of Geometrical Evolution of α-Substituted Dibenzoylmethanatoboron Difluoride Complex with Aggregation-Induced Emission Property.

Organic molecules with an aggregation-induced emission (AIE) property have been attracting much attention from the viewpoint of application to solid state emissive materials. For the AIE mechanism, quantum mechanical studies proposed the restriction of the intramolecular motion (RIM) model with the contribution of the conical intersection (CI) and deduced the importance of the restricted access to a conical intersection (RACI) in the potential energy surface (PES). Although these theoretical studies have contributed to the elucidation of AIE phenomena, direct detection of the reaction dynamics is indispensable to clarify the actual PES and the deactivation mechanism. Along this line, we investigated excited state dynamics of the AIE molecule with dibenzoylmethanatoboron difluoride complexes using time-resolved absorption spectroscopies in both visible and infrared (IR) regions. While the reference system of 1,3-bis(4-methoxyphenyl)methanatoboron difluoride (2aBF2) showed strong emission in solution, the methyl-substituted derivative at the α-position of the dioxaborine ring (2amBF2) led to the very weak fluorescence in solution but strong emission in the solid state. Time-resolved visible absorption measurements revealed a peak shift and broadening of the stimulated emission in the solution of 2amBF2, owing to the rapid change of the molecular geometry. With the temporal evolution of time-resolved IR absorption signals and density functional theory (DFT) calculation of these systems, it was deduced that 2amBF2 has two stable geometries, namely, planar and bending, in the S1 state and the bending geometry in the S1 state led to rapid conversion to the S0 state. These results support the RACI model in the aggregated states, leading to the AIE properties.

Contact-Mediated Retinal-Opsin Coupling Enables Proton Pumping in Gloeobacter Rhodopsin.

When a chromophore embedded in a photoreceptive protein undergoes a reaction upon photoexcitation, the photoreaction triggers structural changes in the protein moiety that are necessary for the function of the protein. It is thus essential to elucidate the coupling between the chromophore and protein moiety to understand the functional mechanism for photoreceptive proteins, but the mechanism by which this coupling occurs remains poorly understood. Here, we show that nonbonded atomic contacts play an essential role in driving functionally important structural changes following photoisomerization of the chromophore in Gloeobacter rhodopsin (GR). Time-resolved ultraviolet resonance Raman spectroscopy revealed that the substitution of Trp222, which contacts with methyl groups of the retinal chromophore, with a Phe residue reduced the extent of structural change. The proton-pumping activity of the GR mutant was as small as 9% of that of the wild type. Time-resolved visible absorption and resonance Raman spectra showed that the photocycle of the mutant proceeded to the L intermediate following the all-trans to 13-cis photoisomerization step but did not result in the deprotonation of the chromophore. The present results demonstrate that the atomic contacts between the chromophore and the Trp222 side chain induce the structural changes necessary for proton transfer. The requirement for dense atomic packing in a protein structure for the efficient propagation of structural changes through a coupling mechanism is discussed.

Timing of NO Binding and Protonation in the Catalytic Reaction of Bacterial Nitric Oxide Reductase as Established by Time-Resolved Spectroscopy

Abstract Membrane-integrated nitric oxide reductases (NOR) catalyze the formation of nitrous oxide (N2O) from two NO molecules using two protons and two electrons at a heme/non-heme iron binuclear center. Despite extensive efforts, the mechanism underlying the NOR-catalyzed reaction has been poorly understood due to the rapidity of the reaction. Here, we utilized a photosensitive caged NO compound as a trigger for the NOR reaction to characterize the NO reduction mechanism by time-resolved visible absorption spectroscopy. We showed that the NOR reaction consists of three steps. One NO molecule binds to the reduced binuclear center to form a non-heme Fe(II)-NO species in the 1st phase (microsecond timescale), followed by a migration of NO to form the other chemical species, possibly 5-coordinate heme b3-NO, in the 2nd phase (timescale of tens of microseconds). Then, the NO bound to heme reacts with a second NO molecule in the 3rd phase (millisecond timescale), in which protonation and electron transfer promote N-N bond formation and N-O bond cleavage to yield N2O. These findings led us to propose a revised trans mechanism for NO reduction by NOR.

Lipidic cubic phase injector is a viable crystal delivery system for time-resolved serial crystallography.

Serial femtosecond crystallography (SFX) using X-ray free-electron laser sources is an emerging method with considerable potential for time-resolved pump-probe experiments. Here we present a lipidic cubic phase SFX structure of the light-driven proton pump bacteriorhodopsin (bR) to 2.3 Å resolution and a method to investigate protein dynamics with modest sample requirement. Time-resolved SFX (TR-SFX) with a pump-probe delay of 1 ms yields difference Fourier maps compatible with the dark to M state transition of bR. Importantly, the method is very sample efficient and reduces sample consumption to about 1 mg per collected time point. Accumulation of M intermediate within the crystal lattice is confirmed by time-resolved visible absorption spectroscopy. This study provides an important step towards characterizing the complete photocycle dynamics of retinal proteins and demonstrates the feasibility of a sample efficient viscous medium jet for TR-SFX.

Direct Evidence for Transient Pair Formation between a Solvated Electron and H3O(+) Observed by Picosecond Pulse Radiolysis.

The reaction between the solvated electron and hydronium cation H3O(+) in water constitutes a fundamental reaction in chemistry. Due to significant rearrangement of solvent molecules around both the electron and H3O(+), the reaction rate of this process is not controlled by diffusion. The presence of a reaction barrier suggests the formation of an intermediate that has so far not been observed. Here, the time-resolved visible absorption spectra in three concentrated acid solutions, perchloric, sulfuric, and phosphoric, at various concentrations are recorded by the picosecond pulse radiolysis method. In contrast to previous reports, a strong blue shift of the absorption band of the solvated electron in acidic solutions compared to neat water is clearly observed, consistent with formation of a pair between the solvated electron and hydronium cation.

Two-step photoionization of trans-stilbene in acetonitrile via an ion-pair precursor studied with picosecond time-resolved absorption and Raman spectroscopies

Picosecond time-resolved visible absorption and Raman spectra of trans-stilbene in acetonitrile have been measured. A new transient absorption band is observed at 440nm, whose decay time constant coincides with that of the rise of the radical cation. In addition to the known Raman bands of the radical cation, a new Raman feature showing a fast decay is found at 1599cm−1. We assign the 440nm absorption band and the 1599cm−1 Raman band to a precursor of the radical cation. The precursor may well be an ion pair that holds an ejected electron still around the cation.

Direct Observation of Interfacial Charge Recombination to the Excited-Triplet State in All-trans-Retinoic Acid Sensitized TiO2 Nanoparticles by Femtosecond Time-Resolved Difference Absorption Spectroscopy

The excited-state dynamics of all-trans-retinoic acid (ATRA), both free in n-hexanol and bound to a TiO2 nanoparticle have been studied by use of femtosecond time-resolved visible absorption spectroscopy excited at 400 nm. Three excited singlet states, S3, S2, and S1, have been observed for the ATRA molecule, which can be assigned as 1Bu+ (π, π*), (n, π*), and 2Ag- (π, π*) singlet excited manifolds, respectively. Photoinduced electron injection from the excited singlet state S3 to the conduction band of the TiO2 nanoparticle is observed in an ATRA-sensitized TiO2 colloidal solution in hexanol; the subsequent interfacial charge recombination between the injected electron and the ATRA cation is investigated. It is found that the charge recombination is mainly via the triplet state, and the branching ratio for the population on the excited triplet state and the ground state during the charge recombination is about 6.0. The observed rate constants for the charge recombination to the triplet state and the grou...

Ultrafast Bimolecular Reaction Kinetics between S1 trans-Stilbene and Carbon Tetrachloride Studied by Sub-Picosecond Time-Resolved Visible Absorption Spectroscopy

Abstract Ultrafast bimolecular reaction kinetics between S1 trans-stilbene and carbon tetrachloride is monitored by sub-picosecond time-resolved visible absorption spectroscopy. The observed kinetics is well explained either by Smoluchowski’s theory or by Collins-Kimball’s theory of diffusion-controlled reactions. These results suggest that the molecular motion in solution for a few picosecond time period is well described by diffusion.

Ultrafast Dynamics of Photochemical Radical Formation from [Re(R)(CO)3(dmb)] (R=Me, Et; dmb=4,4′-dimethyl-2,2′-bipyridine): A Femtosecond Time-Resolved Visible Absorption Study

The excited-state dynamics and photochemistry of [Re(R)(CO)3(dmb)] (R=Me, Et); dmb=4,4′-dimethyl-2,2′-bipyridine) in CH2Cl2 have been studied by time-resolved visible absorption spectroscopy on a broad time scale ranging from approximately 400 fs to a few microseconds, with emphasis on the femtosecond and picosecond dynamics. It was found that the optically prepared Franck-Condon 1MLCT (singlet metal-to-ligand charge transfer) excited state of [Re(R)(CO)3(dmb)] undergoes femtosecond branching between two pathways (≤400 fs for RMe; approximately 800 fs for REt). For both methyl and ethyl complexes, evolution along one pathway leads to homolysis of the Re-R bond via a 3SBLCT (triplet σ-bond-to-ligand charge transfer) excited state, from which [Re(S)(CO)3(dmb)]. and R. radicals are formed. The other pathway leads to an inherently unreactive 3MLCT state. For [Re(Me)(CO)3(dmb)], the 3MLCT state lies lowest in energy and decays exclusively to the ground state with a lifetime of approximately 35 ns, thereby acting as an excitation energy trap. The reactive 3SBLCT state is higher in energy. The quantum yield (0.4 at 293 K) of the radical formation is determined by the branching ratio between the two pathways. [Re(Et)(CO)3(dmb)] behaves differently: branching of the Franck-Condon state between two pathways still occurs, but the 3MLCT excited state lies above the dissociative 3SBLCT state and can decay into it. This shortens the 3MLCT lifetime to 213 ps in CH2Cl2 or 83 ps in CH3CN. Once populated, the 3SBLCT state evolves toward radical photoproducts [Re(S)(CO)3(dmb)]. and Et.. Thus, population of the 3MLCT excited state of [Re(Et)(CO)3(dmb)] provides a second, delayed pathway to homolysis. Hence, the quantum yield is unity. The photochemistry and excited-state dynamics of [Re(R)(CO)3(dmb)] (R=Me, Et) complexes are explained in terms of the relative ordering of the Franck-Condon, 3MLCT, and 3SBLCT states in the region of vertical excitation and along the Re-R reaction coordinate. A qualitative potential energy diagram is proposed.

Ultrafast dynamics of photochemical radical formation from

The excited-state dynamics and photochemistry of [Re(R)(CO)3(dmb)] (R=Me, Et); dmb=4,4'-dimethyl-2,2'-bipyridine) in CH2Cl2 have been studied by time-resolved visible absorption spectroscopy on a broad time scale ranging from approximately 400 fs to a few microseconds, with emphasis on the femtosecond and picosecond dynamics. It was found that the optically prepared Franck-Condon 1MLCT (singlet metal-to-ligand charge transfer) excited state of [Re(R)(CO)3(dmb)] undergoes femtosecond branching between two pathways (< or =400 fs for R=Me; approximately 800 fs for R=Et). For both methyl and ethyl complexes, evolution along one pathway leads to homolysis of the Re-R bond via a 3SBLCT (triplet sigma-bond-to-ligand charge transfer) excited state, from which [Re(S)(CO)3(dmb)]* and R* radicals are formed. The other pathway leads to an inherently unreactive 3MLCT state. For [Re(Me)(CO)3(dmb)], the 3MLCT state lies lowest in energy and decays exclusively to the ground state with a lifetime of approximately 35 ns, thereby acting as an excitation energy trap. The reactive 3SBLCT state is higher in energy. The quantum yield (0.4 at 293 K) of the radical formation is determined by the branching ratio between the two pathways. [Re(Et)(CO)3(dmb)] behaves differently: branching of the Franck-Condon state between two pathways still occurs, but the 3MLCT excited state lies above the dissociative 3SBLCT state and can decay into it. This shortens the 3MLCT lifetime to 213 ps in CH2Cl2 or 83 ps in CH3CN. Once populated, the 3SBLCT state evolves toward radical photoproducts [Re(S)(CO)3(dmb)]* and Et*. Thus, population of the 3MLCT excited state of [Re(Et)(CO)3(dmb)] provides a second, delayed pathway to homolysis. Hence, the quantum yield is unity. The photochemistry and excited-state dynamics of [Re(R)(CO)3(dmb)] (R=Me, Et) complexes are explained in terms of the relative ordering of the Franck-Condon, 3MLCT, and 3SBLCT states in the region of vertical excitation and along the Re-R reaction coordinate. A qualitative potential energy diagram is proposed.

Ultrafast Electronic Relaxation and Hydrogen-Bond-Formation/Dissociation Dynamics of Photoexcited All-trans Retinal in Protic Solvents

The ultrafast electronic relaxation and the hydrogen-bond-formation/dissociation dynamics of photoexcited all-trans retinal in 1-butanol/cyclohexane mixed solvents have been studied by femtosecond time-resolved visible absorption spectroscopy. Four transient absorption bands, which can be assigned to the S3, S2, S1, and T1 states, were observed in neat cyclohexane. The shapes and the dynamics of these absorption bands agree very well with those reported previously for all-trans retinal in hexane. In contrast, only three transient absorption bands, which can be assigned to the S3, S2, and T1 states, were identified in the mixed solvents. The band assigned to the S2 state showed a time-dependent peak shift, which is attributed to solvent reorganization on a picosecond time scale. A kinetic analysis of the three transient absorption bands has led to the conclusion that no state-ordering change of the (n, π*) and (π, π*) states takes place in the excited singlet manifold upon hydrogen-bond formation. The 1-butanol concentration dependence of the absorption spectra shows that the free and hydrogen-bonded species coexist in the S3 and T1 states, but that all of the retinal molecules are hydrogen-bonded in the S2 state. These observations indicate that an ultrafast hydrogen-bond-formation reaction takes place during or just after the S3 → S2 internal conversion and is complete within a time scale much shorter than the S2 lifetime. Dissociation of the hydrogen bonding is most likely to take place during or after the S1 → T1 intersystem crossing and is complete within a time scale much shorter than the T1 lifetime. The observed longer lifetime of the hydrogen-bonded S2 state is consistent with the higher isomerization quantum yield in protic solvents than in aprotic nonpolar solvents.

Static and time-resolved absorption spectroscopy of the bacteriorhodopsin mutant Tyr-185-->Phe: evidence for an equilibrium between bR570 and an O-like species.

The light-dark adaptation, photocycle kinetics, and acid-induced blue formation of the bacteriorhodopsin (bR) mutant Tyr-185-->Phe (Y185F) expressed in Halobacterium halobium have been investigated by both static and time-resolved visible absorption spectroscopy. Evidence is presented that a pH-dependent equilibrium exists between a bR570-like form (bRY185F570) and a red-shifted species in the light-adapted form of Y185F. In two related papers, we show that this species has vibrational features similar to the O intermediate. Key findings are that light adaptation causes formation of a purple species similar to bR570 and a second long-lived red-shifted species with a lambda max near 630 nm, well above the pH for the acid-induced blue transition. The concentration of the red-shifted species is pH- and salt-dependent, decreasing reversibly at high pH and high ionic strength. The dark-adapted state of Y185F also contains a small amount of the red-shifted species which is reversibly titratable. Dark adaptation is much slower than wild-type bR and causes a parallel decay of light-adapted bR and the red-shifted species. Time-resolved visible absorption spectroscopy reveals that the purple and the red-shifted species undergo separate photocycles. The purple species exhibits a relatively normal photocycle except for an increased rate of M formation kinetics. The red-shifted species has a photocycle involving a red-shifted K intermediate and a second longer lived intermediate possibly similar to N. The apparent absence of an O intermediate in the late photocycle of Y185F is attributed to cancellation by depletion bands due to the photoreacting red-shifted species.(ABSTRACT TRUNCATED AT 250 WORDS)

Vibrational spectroscopy of bacteriorhodopsin mutants. Evidence that Thr-46 and Thr-89 form part of a transient network of hydrogen bonds.

The role of Thr-46 and Thr-89 in the bacteriorhodopsin photocycle has been investigated by Fourier transform infrared difference spectroscopy and time-resolved visible absorption spectroscopy of site-directed mutants. Substitutions of Thr-46 and Thr-89 reveal alterations in the chromophore and protein structure during the photocycle, relative to wild-type bacteriorhodopsin. The mutants T89D and to a lesser extent T89A display red shifts in the visible lambda max of the light-adapted states compared with wild type. During the photocycle, T89A exhibits an increased decay rate of the K intermediate, while a K intermediate is not detected in the photocycle of T89D at room temperature. In the carboxyl stretch region of the Fourier transform infrared difference spectra of T89D, a new band appears as early as K formation which is attributed to the deprotonation of Asp-89. Along with this band, an intensity increase occurs in the band assigned to the protonation of Asp-212. In the mutant T46V, a perturbation in the environment of Asp-96 is detected in the L and M intermediates which corresponds to a drop in its pK alpha. These data indicate that Thr-89 is located close to the chromophore, exerts steric constraints on it during all-trans to 13-cis isomerization, and is likely to participate in a hydrogen-bonding network that extends to Asp-212. In addition, a transient interaction between Thr-46 and Asp-96 occurs early in the photocycle. In order to explain these results, a previously proposed model of proton transport is extended to include the existence of a transient network of hydrogen-bonded residues. This model can account for the protonation changes of key amino acid residues during the photocycle of bacteriorhodopsin.

Replacement of leucine-93 by alanine or threonine slows down the decay of the N and O intermediates in the photocycle of bacteriorhodopsin: implications for proton uptake and 13-cis-retinal----all-trans-retinal reisomerization.

We report that the replacement of Leu-93 in bacteriorhodopsin by Ala (L93A) or Thr (L93T) slows down the photocycle by approximately 100-fold relative to wild-type bacteriorhodopsin. Time-resolved visible absorption spectroscopy and resonance Raman experiments, respectively, show the presence of long-lived O-like and N-like intermediates in the photocycles of the above mutants. We infer the existence of an equilibrium between the N and O intermediates in the photocycles of these mutants. The L93A and L93T mutants exhibit normal proton pumping under continuous illumination, suggesting that the decay of the N and/or O intermediate, and consequently, proton translocation, can be accelerated by the absorption of a second photon. Since the 13-cis----all-trans reisomerization of retinal is completed during the decay of the N and O intermediates, we conclude that the interaction of Leu-93 with retinal is important in this phase of the photocycle. This conclusion is supported by a recent structural model of bacteriorhodopsin that suggests that Leu-93 is near the C-13 methyl group of retinal.