Themodynamic and transport properties of intermediate states of the photocyclic reaction of photoactive yellow protein (PYP) were studied by a combination of the pulsed laser-induced transient grating (TG), transient lens (TrL), and photoacoustic (PA) spectroscopies from tens of nanoseconds to hundreds of milliseconds. The diffusion coefficients (D) of PYP in the ground state (pG) and of the second intermediate state (pB) were determined by the TG analysis, and it was found that D of pG is about 1.2 times larger than D of pB. At the same time, D at various denatured conditions were measured using guanidine hydrochloride as the denaturant. D of completely unfolded protein is about 0.4 times that of the native form. The enthalpy of pB is estimated to be 60 kJ/mol by the TrL method with an assumption that the volume change of pB is not sensitive to the temperature. Since the enthalpy of the first intermediate state (pR) is as high as 160 kJ/mol, it implies that most of the photon energy is stored as the strain of the protein in pR, and this may be the driving force for the successive reaction to pB. From the temperature dependence of the volume change, the difference in the thermal expansion coefficients between pG and pR was calculated. All of the characteristic features of PYP, the negative volume change, the larger thermal expansion coefficient, and the slower diffusion process, indicate that the intermediate pR and pB are reasonably interpreted in terms of the unfolded (loosened) protein structure.

The energetics, protein dynamics, and diffusion coefficients of three mutants of photoactive yellow protein, R52Q, P68A, and W119G, were studied by the transient grating and pulsed laser-induced photoacoustic method. We observed a new dynamics with a lifetime of approximately 1 micro s in the transient grating signal, which is silent by the light absorption technique. This fact indicates that, after the structure change around the chromophore is completed (pR(1)), the protein part located far from the chromophore is still moving to finally create another pR (pR(2)) species, which can transform to the next intermediate, pB. Although the kinetics of pR(2)-->pB-->pG are very different depending on the mutants, the enthalpies of the first long-lived (in micro seconds, 100-micro s range) intermediate species (pR(2)) are similar and very high for all mutants. The diffusion coefficients of the parent (pG) and pB species of the mutants are also similar to that of the wild-type photoactive yellow protein. From the temperature dependence of the volume change, the difference in the thermal expansion coefficients taken as indicator of the flexibility of the structure between pG and pR(2) is measured. They are also similar to that of the wild-type photoactive yellow protein. These results suggest that the protein structures of pR(2) and pB in these mutants are globally different from that of pG, and this structural change is not altered so much by the single amino acid residue mutation. This is consistent with the partially unfolded nature of these intermediate species. On the other hand, the volume changes during pR(1)-->pR(2) are sensitive to the mutations, which may suggest that the volume change reflects a rather local character of the structure, such as the chromophore-protein interaction.

The energetics and protein dynamics of photoactive yellow protein (PYP) were studied by transient grating (TG) and photoacoustic (PA) spectroscopies. The enthalpy difference (ΔH) between the ground state (pG) and the first intermediate (pR) at 20 °C was determined by the TG method as 160 kJ/mol, which is much larger than ΔH of the photoisomerized chromophore (p-coumaric acid) in water (50 kJ/mol). By simultaneous measurements of the TG and PA signals, the volume change (ΔV) for the pG → pR process is determined to be −7 cm3/mol (contraction) at 20 °C. Interestingly, the volume contraction increases with decreasing temperature. At 0 °C the volume change becomes ca.  −15 cm3/mol. This temperature-dependent volume change may indicate the structural fluctuation of PYP protein in the solution phase.

Sulfur-doped TiO 2 was prepared by two methods; one was simple oxidation annealing of TiS 2 , the other was mixing of titanium isopropoxide and thiourea. These two sulfur-doped TiO 2 preparations showed fairly different photocatalytic activity under visible light. The dynamics of photogenerated charge carriers were studied by the transient absorption measurement in the region of mid-IR. In both samples, excitation by 532 nm pulse led to photocarrier generation to the same extent. Nevertheless, the reactivity of the photocarriers was totally different. Photogenerated electrons and holes transferred to reactant gas in the latter sample, whereas they did not in the former sample. We attributed the different carrier behavior to the difference in the distribution of S atoms or particle size. These observations can explain the difference in capability of photocatalysis under visible light.

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Photophysics and electron dynamics in dye-sensitized semiconductor film were studied by transient mid-IR spectroscopy, in particular on the nano- to millisecond time scale. As sensitizers, a Ru complex and 9-phynol xanthane derivatives were used. We simultaneously observed electrons injected into the conduction band of a semiconductor and the change of vibration bands due to formation of a cation of a dye molecule. The transient absorption components derived from these two species decayed differently. From this observation, we found that injected electrons decay through two paths; one was back electron transfer, and the other was decay to deep trap sites that cannot be detected by mid-IR light. We could also estimate the amount of electron injection for different dye-sensitized TiO2. Our transient mid-IR spectroscopy in long time scale introduces a semi-new approach to the slow dynamics in dye-sensitized semiconductors including back electron transfer.

The present account describes how photochemical reactions over metal oxides are traced by time-resolved infrared (IR) absorption spectroscopy. The ac-coupled amplification of the IR signal allows detection of transient absorbance changes as small as 10 )6 with a time resolution of 50 ns. Band-gap excited electrons in TiO 2 and NaTaO 3 present a structureless absorption of IR light from 3000 to 1000 cm )1 . Reaction-perturbed decay of this absorption evidences the assignment to photoexcited electrons, not to holes. The efficiency of the water splitting reaction on NaTaO 3 -based catalysts correlates with the quantity of electrons detected by the IR absorption. A short-lived intermediate state of 2-propanol oxidation on TiO 2 is identified by its vibrational band at 1640 cm )1 superposed on the structureless absorption of electrons. Ru dye (N3) on a TiO 2 film is irradiated with a 532-nm light pulse to simulate dye-sensitized solar cells. The neutralization rate of dye cations and the decay rate of electrons injected in the film are quantified, leading to a three-state model which describes the relaxation of injected electrons. These results demonstrate the ability of this method in tracing photochemical kinetics over metal oxides.

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