Stepwise two-color laser photolysis studies of alpha-cleavage in highly excited triplet states of alpha-acyl-4-phenylphenols.

The photochemical properties of alpha-cleavage of C-O bond in highly excited triplet states (T(n) with n>2) of p-biphenyl acetate and p-biphenyl benzoate (Me-OBP and Ph-OBP) in solution were investigated in comparison with those in the lowest excited singlet and triplet states by using single laser and sequential two-color two-laser photolysis techniques. Upon 266 nm laser photolysis of Me-OBP and Ph-OBP, occurrence of C-O bond cleavage in the excited singlet state was recognized from the observation of the formation of p-phenylphenoxy radical (PPR) in the transient absorption. The quantum yields (Phi(rad)) of the PPR formation were determined to be 0.29 and 0.24 for Me-OBP and Ph-OBP, respectively. Triplet sensitization using acetone (Ac) provided efficient formation of the lowest triplet states (T(1)) of Me-OBP and Ph-OBP, and the molar absorption coefficients of the triplet-triplet absorption were determined. No photochemical reactions were found in the T(1) state. Upon 355 nm laser flash photolysis of the T(1) states of Me-OBP and Ph-OBP, formation of PPR accompanied with decomposition of the triplet state was confirmed in the transient absorption. This observation indicated that alpha-cleavage proceeds in the highly excited triplet state. The quantum yields (Phi(dec)) of the decomposition in the dissociative highly excited triplet state (T(R)) were determined to be 0.25 and 0.15 for Me-OBP and Ph-OBP, respectively. The reaction mechanism for alpha-bond cleavage in the T(R) state was discussed.     

Investigations on the mechanism of photodynamic action of different psoralens with DNA

Investigations on the photodynamic action of psoralens with DNA were performed, using experimental techniques of fluorescence lifetime and NMR-CIDNP, as well as SCF-MO and CNDO molecular orbital calculations. It has been shown that the formation of a biradical through the triplet state is the decisive step for psoralen dimer formation, as well as for cyclobutane addition with thymine, while singlet oxygen production is responsible for enzyme inactivation (e.g., lysozyme and trypsin). The molecular orbital calculations, in agreement with experimental results, indicate that the differences in biological effectivity of different psoralens are based on variations in triplet formation probability.     

Triplet states of bacteriochlorophyll and carotenoids in chromatophores of photosynthetic bacteria

Chromatophores from photosynthetic bacteria were excited with flashes lasting approx. 15 ns. Transient optical absorbance changes not associated with the photochemical electron-transfer reactions were interpreted as reflecting the conversion of bacteriochlorophyll or carotenoids into triplet states. Triplet states of various carotenoids were detected in five strains of bacteria; triplet states of bacteriochlorophyll, in two strains that lack carotenoids. Triplet states of antenna pigments could be distinguished from those of pigments specifically associated with the photochemical reaction centers. Antenna pigments were converted into their triplet states if the photochemical apparatus was oversaturated with light, if the primary photochemical reaction was blocked by prior chemical oxidation of P-870 or reduction of the primary electron acceptor, or if the bacteria were genetically devoid of reaction centers. Only the reduction of the electron acceptor appeared to lead to the formation of triplet states in the reaction centers.In the antenna bacteriochlorophyll, triplet states probably arise from excited singlet states by intersystem crossing. The antenna carotenoid triplets probably are formed by energy transfer from triplet antenna bacteriochlorophyll. The energy transfer process has a half time of approx. 20 ns, and is about 1 × 10³ times more rapid than the reaction of the bacteriochlorophyll triplet states with O₂. This is consistent with a role of carotenoids in preventing the formation of singlet O₂ in vivo. In the absence of carotenoids and O₂, the decay half times of the triplet states are 70 μs for the antenna bacteriochlorophyll and 6–10 μs for the reaction center bacteriochlorophyll. The carotenoid triplets decay with half times of 2–8 μs.With weak flashes, the quantum yields of the antenna triplet states are in the order of 0.02. The quantum yields decline severely after approximately one triplet state is formed per photosynthetic unit, so that even extremely strong flashes convert only a very small fraction of the antenna pigments into triplet states. The yield of fluorescence from the antenna bacteriochlorophyll declines similarly. These observations can be explained by the proposal that singlet-triplet fusion causes rapid quenching of excited singlet states in the antenna bacteriochlorophyll.     

Structures of polypyridine mononuclear Ir complexes in the ground state and the lowest triplet state

The structures of eight Ir^III centered polypyridine complexes were determined by density-functional-theory calculations. The differences in the optimized geometries between the ground state and the lowest excited triplet state were mainly considered. A crystal structure of [IrCl(bpy)(terpy)](PF₆)₂ was also obtained by the X-ray diffraction study, where bpy is 2,2′-bipyridine and terpy is 2,2′:6′,2″-terpyridine. The computed geometries are in good agreement with the experimental ones. Those in the triplet biradical states were determined to evaluate the energy difference between the triplet and the ground states. The resulted values correlate well with the observed emission energies. To investigate the nature of the electronic transition involving the ground and the first excited triplet states, a Mulliken population analysis of the spin densities on the eight complexes was performed. The geometric changes from free tterpy ligand {tterpy = 4′-(4-tolyl)-2,2′:6′,2″-terpyridine} to the Ir^III complexed ligand, and then to triplet biradical were examined. The planarity enhanced the π-π^∗ excitation in the ligand and consequently gave the stable triplet biradical of the complex. It was found that efficient phosphorescence should be impacted by the presence of one coplanar polypyridine ligand.     

Photochemical reactions of triplet p-phenylbenzyl derivatives studied by using laser flash triplet-sensitization techniques.

Photochemical properties in the triplet states of p-phenylbenzyl derivatives (PBX; X = H, Cl, Br, OH or SH) were investigated by using laser photolysis techniques of triplet-acetone sensitization. Beta-bond dissociation of triplet phenylbenzyl mercaptan (X = SH) was shown for the first time while formation of triplet states were seen for phenyl toluene and phenylbenzyl alcohol by triplet energy transfer. Beta-bond cleavage was absent for halide compounds (X = Cl and Br) upon sensitization because of the short triplet lifetimes due to a heavy atom effect. The reactivity for beta-bond cleavage was interpreted in terms of difference in the bond enthalpy, D(C-X), relative to the triplet energies.     

Triplet state of the primary donor in reaction centers of the phototrophic bacterium Rhodobacter sphaeroides R26 with active photoinduced electron transfer

EPR characteristics of transient paramagnetic states photoinduced in isolated reaction centers of Rhodobacter sphaeroides R26 with intact electron transfer have been studied. It was demonstrated that the detected weak triplet state EPR signal belongs to the primary donor molecule and is populated via the conventional mechanism of radical pair S-T0 mixing. The distortion of the spectral shape of this signal is explained by the triplet quantum yield anisotropy brought about by the short lifetime of precursor radical pairs. The angular dependence of the anisotropy was evaluated. It was shown that the spectral shape of the triplet state of photosystem II reaction center observed in the case of singly-reduced primary quinone acceptor can also be described by the anisotropic quantum yield of the triplet, with practically the same angular dependence. These properties confirm the conclusions on the mechanism of photoinduced electron transfer in photosystem II, made in previous publications. The peculiarities in the functioning of photosystem II reaction centers are probably determined by strict limitations on the triplet state generation.     

Luminescence and photoreactivity of Ag(8-quinolinol)(8-quinolinolate)

The complex Ag(8-quinolinol)(8-quinolinolate) shows an IL fluorescence while an IL phosphorescence does not appear. The IL triplet is deactivated to a reactive LMCT state. A subsequent photoredox reaction leads to generation of elemental silver.     

EPR characterisation of the triplet state in photosystem II reaction centers with singly reduced primary acceptor Q(A)

The triplet states of photosystem II core particles from spinach were studied using time-resolved cw EPR technique at different reduction states of the iron--quinone complex of the reaction center primary electron acceptor. With doubly reduced primary acceptor, the well-known photosystem II triplet state characterised by zero-field splitting parameters |D|=0.0286 cm(-1), |E|=0.0044 cm(-1) was detected. When the primary acceptor was singly reduced either chemically or photochemically, a triplet state of a different spectral shape was observed, bearing the same D and E values and characteristic spin polarization pattern arising from RC radical pair recombination. The latter triplet state was strongly temperature dependent disappearing at T=100 K, and had a much faster decay than the former one. Based on its properties, this triplet state was also ascribed to the photosystem II reaction center. A sequence of electron-transfer events in the reaction centers is proposed that explains the dependence of the triplet state properties on the reduction state of the iron--quinone primary acceptor complex.     

Frameshift mutations in bacteria produced in the dark by several furocoumarins; absence of activity of 4,5',8-trimethylpsoralen.

4 furocoumarins, namely psoralen (P), 8-methoxypsoralen (8-MOP), 4,5',8-trimethylpsoralen (TMP) and angelicin (A) were tested for dark mutagenesis in E. coli lac-. Three compounds; P, 8-MOP and A were shown to be weak frame-shift mutagens. TMP, surprisingly in view of its very active photosensitizing action, was found to be non-mutagenic. These results are discussed in relation to the photosensitizing action of the furocoumarins.     

Triplet state of the primary donor in reaction centers of the phototrophic bacterium <Emphasis Type="Italic">Rhodobacter sphaeroides</Emphasis> R26 with active photoinduced electron transfer

EPR characteristics of transient paramagnetic states photoinduced in isolated reaction centers of Rhodobacter sphaeroides R26 with intact electron transfer have been studied. It was demonstrated that the detected weak triplet state EPR signal belongs to the primary donor molecule and is populated via the conventional mechanism of radical pair S-T₀ mixing. The distortion of the spectral shape of this signal is explained by the triplet quantum yield anisotropy brought about by the short lifetime of precursor radical pairs. The angular dependence of the anisotropy was evaluated. It was shown that the spectral shape of the triplet state of photosystem II reaction center observed in the case of singly-reduced primary quinone acceptor can also be described by the anisotropic quantum yield of the triplet, with practically the same angular dependence. These properties confirm the conclusions on the mechanism of photoinduced electron transfer in photosystem II, made in previous publications. The peculiarities in the functioning of photosystem II reaction centers are probably determined by strict limitations on the triplet state generation.     

The pH dependence of the reactions of flavin triplet states with amino acids: A laser flash photolysis study

The pH dependence of the rate constants of reaction of several amino acids with the triplet states of flavin mononucleotide in aqueous solution has been determined. In addition, the relative contributions of hydrogen atom transfer, electron transfer and physical deactivation to the overall process of triplet quenching by amino acids have been estimated.Analogous experiments to those with amino acids were carried out with EDTA as the substrate. The results indicate that the flavin triplet state abstracts an electron from EDTA but does not form an excited state flavin-EDTA complex as suggested in a previous study.     

Magnesium (Mg2+) cofactor in a triplet state reduces the proton of a coordinated water molecule to the hydrogen atom and pushes it out of the complex at a high speed

Quantum chemistry methods [RHF/UHF + MP4(FULL), DFT:B3LYP] with the 6-311+ +G**(p,d) basis set were used to elucidate the properties of six coordinated Mg2+ complexes with water, glutamic acid and ATP/GTP in singlet (S) and triplet (T) states. In the triplet state, the magnesium complex concentrates its spin density on a coordinated water molecule (inner or outer coordination shell). Within the molecule, a redox reaction occurs, and one of the hydrogen atoms is pushed out of the complex at a speed of approximately 125 m/s. In water solution, the energy of the triplet state is higher than that of the singlet state. In a mixed environment composed of water, amino acids and ATP/GTR, the energy of the magnesium complex in the triplet state is lower than that in the singlet state by 1.5-2.0 kcal/mol. A little difference in T and S states allows the Mg(2+)-ATP/GTP complex to switch easily between two reaction mechanisms.     

Photothermal Spectra of Thylakoids Isolated from Cucumber Cotyledons at Various Stages of Greening

The character of interaction between carotenoids (Cars) and chlorophylls (Chls) in thylakoids isolated from cucumber cotyledons at three stages of greening (3, 6, and 24 h of irradiation with 120 µmol m^–2 s^–1) was studied. The shapes of the steady state photoacoustic spectra were changed with the change in time of greening and with the frequency of radiation modulation. The shapes show that changes not only in the contents of various pigments but also in pigment interactions with surrounding occur and that processes of thermal deactivation characterised by different kinetics take place. Slow processes of thermal deactivation are in most cases due to deactivation of triplet states. Long living triplet states are very often engaged in photochemical reactions that can destroy the tissue. Analysis of the time-resolved photothermal spectra shows that at later stage of greening, the chlorophyll (Chl) molecules are better shielded against photo-destruction because Cars more efficiently quench their triplet states. The yield of formation of the pigment triplet states measured by the time resolved photothermal method, always at the same energy absorbed by pigment mixture, declined during sample greening. The decay time of the slow component of pigment thermal deactivation, due predominantly to deactivation of the triplet state of Chl, decreases with the increase of time of greening from 6.2 µs for the 3-h sample to 1.5 µs for the 24 h sample. The energy taken by Cars from Chls is dissipated into heat, therefore the steady state and quick thermal deactivation values increased during the greening process. The Cars/Chls ratio in the thylakoids decreased during greening approximately 2 fold. Hence at a later phase of greening the Cars can quench the triplet states of Chls more efficiently than at an earlier phase of greening.     

Triplet-triplet energy transfer in Peridinin-Chlorophyll a-protein reconstituted with Chl a and Chl d as revealed by optically detected magnetic resonance and pulse EPR: Comparison with the native PCP complex from Amphidinium carterae

The triplet state of the carotenoid peridinin, populated by triplet-triplet energy transfer from photoexcited chlorophyll triplet state, in the reconstituted Peridinin-Chlorophyll a-protein, has been investigated by ODMR (Optically detected magnetic resonance), and pulse EPR spectroscopies. The properties of peridinins associated with the triplet state formation in complexes reconstituted with Chl a and Chl d have been compared to those of the main-form peridinin-chlorophyll protein (MFPCP) isolated from Amphidinium carterae. In the reconstituted samples no signals due to the presence of chlorophyll triplet states have been detected, during either steady state illumination or laser-pulse excitation. This demonstrates that reconstituted complexes conserve total quenching of chlorophyll triplet states, despite the biochemical treatment and reconstitution with the non-native Chl d pigment. Zero field splitting parameters of the peridinin triplet states are the same in the two reconstituted samples and slightly smaller than in native MFPCP. Analysis of the initial polarization of the photoinduced Electron-Spin-Echo detected spectra and their time evolution, shows that, in the reconstituted complexes, the triplet state is probably localized on the same peridinin as in native MFPCP although, when Chl d replaces Chl a, a local rearrangement of the pigments is likely to occur. Substitution of Chl d for Chl a identifies previously unassigned bands at ∼ 620 and ∼ 640 nm in the Triplet-minus-Singlet (T − S) spectrum of PCP detected at cryogenic temperature, as belonging to peridinin.     

Reaction mechanisms of riboflavin triplet state with nucleic acid bases.

ESR and laser flash photolysis studies have determined a reasonable order of reactivity of nucleotides with triplet riboflavin (3Rb*) for the first time. ESR detection of triplet state reactivity of Rb with nucleoside, polynucleotide and DNA has been obtained simultaneously. In addition, ESR spin elimination measurement of the reactivity of 3Rb* with nucleotides in good accord with laser flash photolysis determination of the corresponding rate constants offers a simple and reliable method to detect the reactivities of nucleic acids and its components with photoexcited flavins. Kinetic, ESR and thermodynamic studies have demonstrated that Rb should be a strong endogenous photosensitizer capable of oxidizing all nucleic acid bases, and preferentially two purine nucleotides with high rate constants.     

Receptor-mediated photo-cytotoxicity: synthesis of a photoactivatable psoralen derivative conjugated to insulin

4'-Aminomethyl-4,5',8-trimethylpsoralen has been chemically conjugated to insulin using a carbodiimide derivative. The psoralen moiety retains its photochemical reactivity as evidenced by its ability to crosslink DNA after exposure to long wavelength ultraviolet light (UVA, 320-400 nm). This chimeric molecule has been used to selectively kill a population of lymphocytes whose expression of insulin receptors has been stimulated with phytohemagglutinin. Insulin carries the psoralen into the cell via receptor-mediated endocytosis, where it is subsequently activated by exposure to UVA light. The UVA induced activity of AMT-insulin can be blocked by the presence of native insulin. The viability of unstimulated lymphocytes was not affected by AMT-insulin and UVA light. The hybrid insulin-psoralen molecule may be a prototype for a family of phototoxic drugs which can be selectively delivered to subsets of lymphocytes.     

Effects of boron doping on structural,electronic, elastic,and optical properties of energetic crystal 2,6-diamino-3,5-dinitropyrazine-1-oxide: a theoretical study using the first principles calculation and Hirshfeld surface analysis

The activation or functionalization of the saturated C-H is an extremely active field at present. We have explored the triplet state thioxanthone in reactivity of the hydrogen transfer reaction between donors and acceptors. In our works, two donors with quasi-inert sp3 C-H of skipped diene (3,6-nonadiene) and cyclic acetals (benzodioxole) reacted with type II photoinitiators (triplet state of thioxanthone series, TXs) as acceptors are investigated. The excited energies of TXs were obtained by time-dependent density functional theory (TD-DFT). TXs show obvious photosensibility based on their low reorganization energies (< 60 kcal mol−1). The isoentropy reactions had linear geometries of transition state (TS). The distortion/interaction model was used to probe the existence of interaction between acceptors and donors in saddle point. The distortion energy and activation barrier of benzodioxole are much higher than those of the corresponding 3,6-nonadiene. The lower bond dissociation energy noticeably affect the transition state. The reaction of triplet state of TXs with skipped dienes were found to have an anomalous low tunneling factors by using Wigner correction and early transition state by using the bond-energy–bond-order method. The triplet state of TXs photoinitiator can induced the hydrogen abstraction from saturated cyclic acetals and the skipped alkadienes. The hydrogen abstraction experiment are confirmed by UV and real-time FTIR.     

Evidence for uptake of 8-methoxypsoralen and 5-methoxypsoralen by cellular nuclei

The fluorescent appearance of oral mucosa cells treated with 8-methoxypsoralen (8-MOP) and 5-methoxypsoralen (5-MOP) was observed by means of fluorescence microscopy. Fluorescence at the nuclei was weakened in 8-MOP-treated cells, while it was intensified in 5-MOP-treated cells. These findings were consistent with changes in the fluorescence intensities on association of the psoralen derivatives with DNA in aqueous solution. This intensity change of fluorescence and also the blue shift of the fluorescence maximum of the derivatives on association suggested that the environment around the psoralen molecules is as little polar as methanol. From the results of these fluorescence microscopic observations and spectroscopic analysis of fluorescence of derivatives interacting with DNA during equilibrium dialysis, we concluded that 8-MOP, as well as 5-MOP, is incorporated by nuclei of human cells.     

Variable reactivity of an engineered cysteine at position 338 in cystic fibrosis transmembrane conductance regulator reflects different chemical states of the thiol

In a previous study of T338C CFTR (cystic fibrosis transmembrane conductance regulator) we found that protons and thiol-directed reagents modified channel properties in a manner consistent with the hypothesis that this residue lies within the conduction path, but the observed reactivity was not consistent with the presence of a single thiolate species in the pore. Here we report results consistent with the notion that the thiol moiety can exist in at least three chemical states, the simple thiol, and two altered states. One of the altered states displays reactivity toward thiols like dithiothreitol and 2-mercaptoethanol as well as reagents: mixed disulfides (methanethiosulfonate reagents: MTSET+, MTSES-) and an alkylating agent (iodoacetamide). The other altered state is unreactive. The phenotype associated with the reactive, altered state could be replicated by exposing oocytes expressing T338C CFTR to CuCl2, but not by glutathionylation or nitrosylation of the thiol or by oxidation with hydrogen peroxide. The results are consistent with the hypothesis that substituting a cysteine at 338 can create an adventitious metal binding site. Metal liganding alters thiol reactivity and may, in some cases, catalyze oxidation of the thiol to an unreactive form such as a sulfinic or sulfonic acid.