Influence of Arrestin on the Photodecay of Bovine Rhodopsin

Continued activation of the photocycle of the dim‐light receptor rhodopsin leads to the accumulation of all‐trans‐retinal in the rod outer segments (ROS). This accumulation can damage the photoreceptor cell. For retinal homeostasis, deactivation processes are initiated in which the release of retinal is delayed. One of these processes involves the binding of arrestin to rhodopsin. Here, the interaction of pre‐activated truncated bovine visual arrestin (Arr^Tr) with rhodopsin in 1,2‐diheptanoyl‐sn‐glycero‐3‐phosphocholine (DHPC) micelles is investigated by solution NMR techniques and flash photolysis spectroscopy. Our results show that formation of the rhodopsin–arrestin complex markedly influences partitioning in the decay kinetics of rhodopsin, which involves the simultaneous formation of a meta II and a meta III state from the meta I state. Binding of Arr^Tr leads to an increase in the population of the meta III state and consequently to an approximately twofold slower release of all‐trans‐retinal from rhodopsin.     

2-Naphthyl(carbomethoxy)carbene revisited: combination of ultrafast UV-vis and infrared spectroscopic study

Ultrafast laser flash photolysis (310 nm) of methyl 2-napthyldiazoacetate (2-NpCN2CO2CH3) in acetonitrile or cyclohexane produces a diazo excited state which absorbs broadly in the visible region (tau = 300 fs). The decay of the excited diazo compound is accompanied by growth of the vibrationally excited singlet 2-naphthyl(carbomethoxy)carbene ((1)NpCCO2CH3). The singlet carbene absorbs at 360 and 470 nm. In acetonitrile these bands do not decay over 3 ns, but they do decay by approximately 50% of their original intensity in cyclohexane in 3 ns. It is concluded that (1)NpCCO2CH3 has a singlet ground state in acetonitrile but a triplet ground state in cyclohexane. Related experiments reveal a singlet ground state in Freon-113 and chloroform. This interpretation is supported by ultrafast IR spectroscopy, which confirms that only (1)NpCCO2CH3 is formed within 50 ps of the laser pulse rather than a singlet-triplet equilibrium mixture of carbene. The planar singlet relaxes to the preferred perpendicular singlet over a few tens of picoseconds, as evidenced by a red shift of the carbonyl stretching vibration. Although our data agrees with previous studies, its interpretation is somewhat altered.     

The direct detection of an aryl azide excited state: an ultrafast study of the photochemistry of para- and ortho-biphenyl azide

Ultrafast laser flash photolysis (266 nm) of para- and ortho-biphenyl azide in acetonitrile produces azide excited states that have broad absorption bands centered at 480 nm. The para-biphenyl azide excited singlet state has a lifetime of 100 fs. The excited-state lifetime of the ortho-azide isomer is 450 +/- 150 fs. Decay of the azide excited states is accompanied by the formation of the corresponding known singlet nitrenes (para, lambdamax = 350 nm, ortho, lambdamax = 400 nm). Singlet para-biphenylnitrene is born with excess energy and undergoes vibrational cooling with a time constant of 11 ps to form the long-lived (tau approximately 9 ns) relaxed singlet nitrene. Singlet ortho-biphenylnitrene decays with a lifetime of 16 ps in acetonitrile at ambient temperature.     

Photo-deactivation pathways of a double H-bonded photochromic Schiff base investigated by combined theoretical calculations and experimental time-resolved studies

The photophysics of N,N'-bis(salicylidene)-p-phenylenediamine (BSP) is analyzed both theoretically and experimentally. The alternative intramolecular proton-transfer reactions lead to three different tautomers. We performed DFT and TDDFT calculations to analyze the topography of the reactions connecting the three tautomers. Deactivation paths through a Conical Intersection (CI) region are also analyzed to explain the low fluorescence quantum yield of the phototautomers. The complex molecular structure of BSP provides a large number of deactivation paths, almost all of them energetically available following the initial photoexcitation. Femtosecond (fs) time-resolved emission studies in solution and flash photolysis experiments (nano to millisecond regime) were performed to get detailed information on the time domain of the full photocycle. The picture that emerges by combining theoretical and experimental results shows a very fast (less than 100 fs) photoinduced single proton transfer process leading to a phototautomer where a single proton has moved. This species may deactivate through a low-energy CI leading in about 20 ps to a rotameric form in the ground state that has a lifetime of several tens of microseconds in solution. This process competes with another deactivation path taking place prior to the proton-transfer reaction which involves a low-energy CI leading to a rotamer of the enol structure. In the flash photolysis studies, the rotamer of the enol structure was directly identified by the positive transient absorption band in the 250-260 nm and its lifetime in n-hexane (10 ms) is almost 3 orders of magnitude longer than the lifetime of the photochrome (around 40 μs). Our findings do not exclude a double proton transfer reaction in the excited enol form to give a tautomer in less than 100 fs during the first (impulsive) phase of the reaction which reverts back to the photoproducts of the simple proton transfer in 1-3 ps.     

TIME-RESOLVED SPECTROSCOPIC STUDIES ON PHOTOPHYSICAL PROPERTIES OF BENOXAPROFEN IN MICELLAR SOLUTIONS: AN INTRAMICELLAR FLUORESCENCE QUENCHING

The micellar dependencies of the photophysical properties of benoxaprofen (BXP), a 2-phenyl benzoxazole derivative, have been investigated using fluorescence spectroscopy and laser flash photolysis techniques. The fluorescence of BXP in aqueous solution has been observed to be remarkably quenched upon addition of a surfactant, cetyltrimethyl ammonium bromide (CTAB) or Triton X-100, in contrast to its enhancement in sodium dodecyl sulfate (SDS) micellar solution. Time-resolved fluorescence measurements show that the fluorescence decays biexponentially in the micellar solution, indicating the relaxation of micellar environments surrounding the excited BXP. The major component of fluorescence lifetimes in CTAB or Triton X-100 micellar phase is even shorter (330–427ps) than in SDS micellar phase (731 ps). The nonradiative decay constants are significantly larger (ca 3.0 times 10⁹ s^?1) in the CTAB or Triton X-100 micellar phase than in SDS micelles by a factor of ca 10. The major nonradiative decay is interpreted to be the internal conversion due to nuclear geometric change of BXP in the first excited singlet state. This is consistent with the observation that the quantum yields of intersystem crossing are very low (less than 0.01) in the micellar solutions as determined by the laser flash photolysis technique. The laser-induced transient absorption spectrum of BXP in CTAB or Triton X-100 micellar solution shows that the decay kinetics of the transients in CTAB or Triton X-100 are significantly different from first order kinetics in SDS.     

The Role of Ligand‐Field States in the Ultrafast Photophysical Cycle of the Prototypical Iron(II) Spin‐Crossover Compound [Fe(ptz)6](BF4)2

Light‐induced excited spin‐state trapping (LIESST) in iron(II) spin‐crossover compounds, that is, the light‐induced population of the high‐spin (S=2) state below the thermal transition temperature, was discovered thirty years ago. For irradiation into metal–ligand charge transfer (MLCT) bands of the low‐spin (S=0) species the acknowledged sequence takes the system from the initially excited ¹MLCT to the high‐spin state via the ³MLCT state within ca. 150 fs, thereby bypassing low‐lying ligand‐field (LF) states. Nevertheless, these play a role, as borne out by the observation of LIESST and reverse‐LIESST on irradiation directly into the LF bands for systems with only high‐energy MLCT states. Herein we elucidate the ultrafast reverse‐LIESST pathway by identifying the lowest energy S=1 LF state as an intermediate state with a lifetime of 39 ps for the light‐induced high‐spin to low‐spin conversion on irradiation into the spin‐allowed LF transition of the high‐spin species in the NIR.     

An ultrafast study of phenyl azide: the direct observation of phenylnitrenium ion

Ultrafast photolysis (lambda(ex) = 308 nm) of phenyl azide in 100% formic acid produces a broadly absorbing transient within the instrument time resolution (300 fs), which is assigned to an excited state of the azide. The azide excited state fragments within 300 fs to form singlet phenylnitrene. The decay of the nitrene (tau = 12.0 ps) produces a new species with absorption centered at 500 nm, which is assigned to phenylnitrenium ion. The lifetime of phenylnitrenium ion is 110 ps in 100% formic acid. This is the first spectroscopic observation of phenylnitrenium ion.     

State preparation and excited electronic and vibrational behavior in hemes

The temporally overlapping, ultrafast electronic and vibrational dynamics of a model five-coordinate, high-spin heme in a nominally isotropic solvent environment has been studied for the first time with three complementary ultrafast techniques: transient absorption, time-resolved resonance Raman Stokes, and time-resolved resonance Raman anti-Stokes spectroscopies. Vibrational dynamics associated with an evolving ground-state species dominate the observations. Excitation into the blue side of the Soret band led to very rapid S2 --> S1 decay (sub-100 fs), followed by somewhat slower (800 fs) S1 --> S0 nonradiative decay. The initial vibrationally excited, non-Boltzmann S0 state was modeled as shifted to lower energy by 300 cm(-1) and broadened by 20%. On a approximately 10 ps time scale, the S0 state evolved into its room-temperature, thermal distribution S0 profile largely through VER. Anti-Stokes signals disappear very rapidly, indicating that the vibrational energy redistributes internally in about 1-3 ps from the initial accepting modes associated with S1 --> S0 internal conversion to the rest of the macrocycle. Comparisons of anti-Stokes mode intensities and lifetimes from TRARRS studies in which the initial excited state was prepared by ligand photolysis [Mizutani, T.; Kitagawa, T. Science 1997, 278, 443, and Chem. Rec. 2001, 1, 258] suggest that, while transient absorption studies appear to be relatively insensitive to initial preparation of the electronic excited state, the subsequent vibrational dynamics are not. Direct, time-resolved evaluation of vibrational lifetimes provides insight into fast internal conversion in hemes and the pathways of subsequent vibrational energy flow in the ground state. The overall similarity of the model heme electronic dynamics to those of biological systems may be a sign that the protein's influence upon the dynamics of the heme active site is rather subtle.     

飞秒时间分辨的光电子影像对3-甲基吡啶分子超快动力学研究

利用飞秒时间分辨的光电子影像技术结合时间分辨的质谱技术,研究了3-甲基吡啶分子激发态的超快过程. 实时观察到了3-甲基吡啶分子S₂态向S₁态高振动能级的超快内转换过程,该内转换的时间大约为910fs. 二次布居的S₁态主要通过内转换衰减到基态S₀,该内转换的时间尺度为2.77 ps. 光电子能谱分布和光电子角分布显示,S₂态和S₁态在电离的过程中跟3p里德堡态发生偶然共振. 本次实验中还用400 nm两个光子吸收的方法布居了3-甲基吡啶的3s 里德堡态. 研究表明,3s 里德堡态的寿命为62 fs,并主要通过内转换快速衰减到基态.     

Ultrafast Photodynamics of the Indoline Dye D149 Adsorbed to Porous ZnO in Dye‐Sensitized Solar Cells

We investigate the ultrafast dynamics of the photoinduced electron transfer between surface‐adsorbed indoline D149 dye and porous ZnO as used in the working electrodes of dye‐sensitized solar cells. Transient absorption spectroscopy was conducted on the dye in solution, on solid state samples and for the latter in contact to a I^?/I₃^? redox electrolyte typical for dye‐sensitized solar cells to elucidate the effect of each component in the observed dynamics. D149 in a solution of 1:1 acetonitrile and tert‐butyl alcohol shows excited‐state lifetimes of 300±50 ps. This signature is severely quenched when D149 is adsorbed to ZnO, with the fastest component of the decay trace measured at 150±20 fs due to the charge‐transfer mechanism. Absorption bands of the oxidized dye molecule were investigated to determine regeneration times which are in excess of 1 ns. The addition of the redox electrolyte to the system results in faster regeneration times, of the order of 1 ns.     

Photoinduced Electron Transfer between N,N, Nr,N' ‐Tetra‐(p‐methylphenyl)‐4, 4‐diamino‐1, 1'‐diphenyl Ether (TPDAE) and Fullerenes (C60/C70) by Laser Flash Photolysis

The photoinduced electron‐transfer reaction of N, N, N', N'‐tetra‐(p‐methylphenyl)‐4,4'‐diamino‐1,1'‐diphenyl ether (TPDAE) and fullerenes (C₆₀/C₇₀) by nanosecond laser flash photolysis occurred in benzonitrile. Transient absorption spectral measurements were carried out during 532 nm laser flash photolysis of a mixture of the fullerenes (C₆₀/C₇₀) and TPDAE. The electron transfer from the TPDAE to excited triplet state of the fullerenes (C₆₀/C₇₀) quantum yields and rate constants of electron transfer from TPDAE to excited triplet state of fullerenes (C₆₀/C₇₀) in benzonitrile have been evaluated by observing the transient absorption bands in the near‐IR region where the excited triplet state, radical anion of fullerenes (C₆₀/C₇₀) and radical cations of TPDAE are expected to appear.     

Excited state intramolecular proton transfer in Schiff bases. Decay of the locally excited enol state observed by femtosecond resolved fluorescence

Although the late (t>1 ps) photoisomerization steps in Schiff bases have been described in good detail, some aspects of the ultrafast (sub-100 fs) proton transfer process, including the possible existence of an energy barrier, still require experimental assessment. In this contribution we present femtosecond fluorescence up-conversion studies to characterize the excited state enol to cis-keto tautomerization through measurements of the transient molecular emission. Salicylideneaniline and salicylidene-1-naphthylamine were examined in acetonitrile solutions. We have resolved sub-100 fs and sub-0.5 ps emission components which are attributed to the decay of the locally excited enol form and to vibrationally excited states as they transit to the relaxed cis-keto species in the first electronically excited state. From the early spectral evolution, the lack of a deuterium isotope effect, and the kinetics measured with different amounts of excess vibrational energy, it is concluded that the intramolecular proton transfer in the S1 surface occurs as a barrierless process where the initial wave packet evolves in a repulsive potential toward the cis-keto form in a time scale of about 50 fs. The absence of an energy barrier suggests the participation of normal modes which modulate the donor to acceptor distance, thus reducing the potential energy during the intramolecular proton transfer.     

Wavepacket Splitting and Two‐Pathway Deactivation in the Photoexcited Visual Pigment Isorhodopsin

Isorhodopsin is the visual pigment analogue of rhodopsin. It shares the same opsin environment but it embeds 9‐cis retinal instead of 11‐cis. Its photoisomerization is three times slower and less effective. The mechanistic rationale behind this observation is revealed by combining high‐level quantum‐mechanical/molecular‐mechanical simulations with ultrafast optical spectroscopy with sub‐20 fs time resolution and spectral coverage extended to the near‐infrared. Whereas in rhodopsin the photoexcited wavepacket has ballistic motion through a single conical intersection seam region between the ground and excited states, in isorhodopsin it branches into two competitive deactivation pathways involving distinct conical intersection funnels. One is rapidly accessed but unreactive. The other is slower, as it features extended steric interactions with the environment, but it is productive as it follows forward bicycle pedal motion.     

Extending Metal‐to‐Polyoxometalate Charge Transfer Lifetimes: The Effect of Heterometal Location

In an effort to develop robust molecular sensitizers for solar fuel production, the electronic structure and photodynamics of transition‐metal‐substituted polyoxometalates (POMs), a novel class of compound in this context, was examined. Experimental and computational techniques including femtosecond (fs) transient absorption spectroscopy have been used to study the cobalt‐containing Keggin POMs, [Co^IIW₁₂O₄₀]^6? ( 1 a ), [Co^IIIW₁₂O₄₀]^5? ( 2 a ), [SiCo^II(H₂O)W₁₁O₃₉]^6? ( 3 a ), and [SiCo^III(H₂O)W₁₁O₃₉]^5? ( 4 a ), finding the longest lived charge transfer excited state so far observed in a POM and elucidating the electronic structures and excited‐state dynamics of these compounds at an unprecedented level. All species exhibit a bi‐exponential decay in which early dynamic processes with time constants in the fs domain yield longer lived excited states which decay with time constants in the ps to ns domain. The initially formed states of 1 a and 3 a are considered to result from metal‐to‐polyoxometalate charge transfer (MPCT) from Co^II to W, while the longer‐lived excited state of 1 a is tentatively assigned to a localized intermediate MPCT state. The excited state formed by the tetrahedral cobalt(II) centered heteropolyanion ( 1 a ) is far longer‐lived (τ=420 ps in H₂O; τ=1700 ps in MeCN) than that of 3 a (τ=1.3 ps), in which the single Co^II atom is located in a pseudo‐octahedral addendum site. Short‐lived states are observed for the two Co^III‐containing heteropolyanions 2 a (τ=4.4 ps) and 4 a (τ=6.3 ps) and assigned solely to O→Co^III charge transfer. The dramatically extended lifetime for 1 a versus 3 a is ascribed to a structural change permitted by the coordinatively flexible central site, weak orbital overlap of the central Co with the polytungstate framework, and putative transient valence trapping of the excited electron on a single W atom, a phenomenon not noted previously in POMs.     

Photochemistry of iron(iii)-lactic acid complex in aqueous solutions

Photochemistry of a 1: 1 Fe^III-lactic acid complex, [Fe(Lact)]⁺, in aqueous solutions was studied by stationary photolysis, nanosecond laser flash photolysis (355 nm, 6 ns), and femtosecond pump-probe spectroscopy (400 nm, 200 fs). The quantum yield of photolysis of [Fe(Lact)]⁺ upon excitation at 355 nm is 0.4 and 0.22 in the deoxygenated and air-saturated solutions, respectively. Weak transient absorption in the range 500–750 nm was observed in the nanosecond experiments. It was assigned to a [Fe^II...-O-CH(Me)-COO^·]⁺ radical complex. The spectral properties of the ligand-to-metal charge transfer excited state and the characteristic time of formation of the radical complex (1.5 ps) were determined in the femtosecond spectroscopy experiments. A reaction mechanism was proposed, which involves inner-sphere electron transfer in the excited complex with the formation of a radical complex [Fe^II...-O-CH(Me)-COO^·]⁺ and its subsequent transformation to the end product of the photochemical reaction.     

Two Consecutive Polar Amino Acids at the End of Helix E are Important for Fast Turnover of the Archaerhodopsin Photocycle

Archaerhodopsins (ARs) is one of the members of microbial rhodopsins. Threonine 164 (T164) and serine 165 (S165) residues of the AR from Halorubrum sp. ejinoor (HeAR) are fully conserved in ARs, although they are far from the proton transfer channel and the retinal Schiff base, and are likely involved in a hydrogen‐bonding network at the end of the Helix E where most microbial rhodopsins assume a “bent structure”. In the present work, T164 and/or S165 were replaced with an alanine (A), and the photocycles of the mutants were analyzed with flash photolysis. The amino acid replacements caused profound changes to the photocycle of HeAR including prolonged photocycle, accelerated decay of M intermediate and appearance of additional two intermediates which were evident in T164A‐ and T164A/S165A‐HeAR photocyles. These results suggest that although T164 and S165 are located at the far end of the photoactive center, these two amino acid residues are important for maintaining the fast turnover of the HeAR photocycle. The underlying molecular mechanisms are discussed in relation to hydrogen‐bonding networks involving these two amino acids. Present study may arouse our interests to explore the functional role of the well‐conserved “bent structure” in different types of microbial rhodopsin.     

Hot Branching Dynamics in a Light‐Harvesting Iron Carbene Complex Revealed by Ultrafast X‐ray Emission Spectroscopy

Iron N‐heterocyclic carbene (NHC) complexes have received a great deal of attention recently because of their growing potential as light sensitizers or photocatalysts. We present a sub‐ps X‐ray spectroscopy study of an Fe^IINHC complex that identifies and quantifies the states involved in the deactivation cascade after light absorption. Excited molecules relax back to the ground state along two pathways: After population of a hot ³MLCT state, from the initially excited ¹MLCT state, 30 % of the molecules undergo ultrafast (150 fs) relaxation to the ³MC state, in competition with vibrational relaxation and cooling to the relaxed ³MLCT state. The relaxed ³MLCT state then decays much more slowly (7.6 ps) to the ³MC state. The ³MC state is rapidly (2.2 ps) deactivated to the ground state. The ⁵MC state is not involved in the deactivation pathway. The ultrafast partial deactivation of the ³MLCT state constitutes a loss channel from the point of view of photochemical efficiency and highlights the necessity to screen transition‐metal complexes for similar ultrafast decays to optimize photochemical performance.     

STUDIES ON CHROMOPHORE COUPLING IN ISOLATED PHYCOBILIPROTEINS. IV. FEMTOSECOND TRANSIENT ABSORPTION STUDY OF ULTRAFAST EXCITED STATE DYNAMICS IN TRIMERIC PHYCOERYTHROCYANIN COMPLEXES*

The excited state dynamics of trimeric phycoerythrocyanin has been studied by two-color femtosecond transient absorption spectroscopy with a time-resolution better than 200 fs. Upon selective excitation of the short-wavelength phycobiliviolin chromophore at 575 nm absorption bleachings are observed. An isotropic ultrafast decay of the initial bleaching of 585 ± 40 fs has been resolved at short detection wavelength (578 nm). Upon stepwise increase of the detection wavelength up to 617 nm, the bleaching showed a delayed rise above 593 nm with r_rice=380–580 fs. All other isotropic kinetic components in this wavelength range were longer than 100 ps. The ultrafast component is discussed in terms of an energy transfer process from the α-84 phycobiliviolin chromophore to the β-84 chromophore in adjacent monomer subunits of the trimer. It is concluded that the β-155 chromophore is the longest-wavelength chromophore. Exciton relaxation between closely spaced chromophore pairs is discussed as an alternative interpretation for the ultrafast component.     

没食子酸的激光光解研究

The transient species of gallic acid(GA)have been studied by 266 nm nanosecond laser flash photolysis inaqueous solution and acetonitrile.The intermediate with absorption at 320 nm was identified as excited triplet state(~3GA~*),the decay rates of which were obtained in aqueous solution and acetonitrile respectively.Energy transferfrom ~3GA~* to β-carotene was observed and the energy transfer rate constant k_(ent)was determined to be 2.2×10~9mol~(-1)·L·s~(-1).GA underwent photoionization during photolysis and the quantum yield of photoionization was de-termined to be 0.12 at room temperature with KI as a reference.     

Primary Reaction Dynamics of Proteorhodopsin Mutant D97N Observed by Femtosecond Infrared and Visible Spectroscopy†

Femtosecond time‐resolved spectroscopy in the visible and IR range was utilized to study the primary reaction dynamics of the proteorhodopsin (PR) D97N mutant in comparison with wild type PR at different pH values. The analysis of the data obtained in the mid‐IR closely resembles the results for wild type PR. The observation of the first ground state intermediate K is initially obscured by a complex reaction scheme of vibrational relaxation and heating effects, but its spectral signature clearly emerges at long delay times. In the visible range, a biexponential decay of the excited state within 30 ps and the formation of the K photoproduct is observed. The decay time constants derived for the D97N mutant in D₂O are slightly larger than in H₂O due to H/D exchange. This kinetic isotope effect is even less pronounced than for wild type PR at pH 6. These results support the current notion of a pH dependent hydrogen bonding network in the retinal binding pocket of PR and a weaker interaction between the retinal Schiff base and the counter ion complex compared to bacteriorhodopsin.