TRIPLET STATES OF CAROTENOIDS FROM PHOTOSYNTHETIC BACTERIA STUDIED BY NANOSECOND ULTRAVIOLET AND ELECTRON PULSE IRRADIATION

Abstract— Absorptions of the triplet excited states of five carotenoids (15,15'-ds phytoene, all- trans phytoene, C-carotene, spheroidene and spirilloxanthin), extracted from the photosynthetic bacteria Rhodopseudomonas spheroides and Rhodospirillum rubrum, have been detected in solution using pulse radiolysis and laser flash photolysis. Triplet lifetimes, extinction coefficients, lowest energy levels and quantum efficiencies of formation have been determined. Comparison of the carotenoid triplet energy levels with that of O₂('Δ_g) suggests that spirilloxanthin, spheroidene and possibly alsoζ-carotene, would be expected to protect against photodynamic action caused by O₂ ('Δ_g), but not cis or trans phytoene. The S → T intersystem crossing efficiences of all five polyenes were found to be low, being a few per cent or less. In their protective role these triplet states can only therefore be effectively reached via energy transfer from another triplet, except in the case of O₂ ('Δ_g). The low crossover efficiencies also mean that light absorbed by such carotenoids in their possible role as accessory pigments would not be wasted in crossing over to the triplet state.     

INDICATION OF FAST TRIPLET FORMATION IN BIMOLECULAR QUENCHING OF EXCITED SINGLET STATES STUDIED BY CONVENTIONAL FLASH PHOTOLYSIS

The bimolecular quenching of the first excited singlet state of oxonine by allylthiourea leads to the formation of the triplet state of the dye. This has been proved by comparison with the triplet-triplet absorption spectrum of oxonine obtained by triplet-triplet energy transfer. The conventional flash experiments suggest that the dye triplet state is produced directly rather than by radical recombination.     

REACTIVITY OF ACRIDINE DYE TRIPLET STATES IN ELECTRON TRANSFER REACTIONS

Abstract—Rate constants, k _q , for the reaction of cationic and neutral acridine orange and 10-methylacridine orange triplet states (³AOH ⁺, ³AO, ³MAO⁺) with a series of electron donors have been measured. Two different protolytic forms of the semireduced dye radical are produced by acridine orange triplet quenching at various pH^M values in methanolic solution.
It is found that k ₄ decreases with increasing oxidation potential of the reducing agent for all triplet states in a manner which is expected for electron transfer reactions. The different reactivities of the cationic and neutral triplet forms can, therefore, be attributed to the difference in reduction potentials of these species. The difference in reduction potentials is related to the p K ^M values of triplet state, p K _T^M , and semireduced dye radical, p K ^M_S , by thermodynamic consideration. p K _T^M (³AOH⁺/³AO) is determined to be 11.2. From thisp K _S^M (AOH./AO;) is estimated to be 17–18. This is in striking contrast to the protolytic equilibrium of the semireduced dye radicals found to be pK^F= 4.1. We conclude that the last value represents the second protonation equilibrium (AOH⁺₂./AOH). This conclusion is confirmed by spectroscopic data.     

THYMINE HYDRATE FORMED BY ULTRAVIOLET AND GAMMA IRRADIATION OF AQUEOUS SOLUTIONS

Abstract— Evidence is presented for the formation of a thymine hydrate upon ultraviolet (UV) or gamma irradiation of aqueous solutions. The UV quantum efficiency exhibits a dependence on pH similar to that shown for uracil hydration, but the yield is three orders of magnitude smaller than for uracil. Hydration is not affected by wavelength, oxygen, or concentration of thymine. The reversal rate of the photohydrate to thymine is similar to the reversal rates of both isomers of the thymine hydrate formed by γ radiolysis, and depends on pH in the same way as the rate for the uracil photohydrate. The photohydrate of thymine is chromatographically identical to the cis isomer of 6-hydroxy-5, 6-dihydrothymine.     

ELECTRON SPIN RESONANCE STUDIES OF THE TRIPLET STATES OF MONOANIONIC THYMINE*

Abstract— Ultraviolet irradiation of the thymine anion tautomers 1-HT- and 3-HT^- at -196°C generates excited triplet states having δm=±2 resonances at 1067 and 1260 G respectively (with v= 9.0 GHz). Excitation at 270nm gave predominantly the low field signal while excitation at 300 nm produced only the high field resonance. These results when compared with the absorbance properties of the monoanions are consistent with the low field resonance being due to 1-HT^- and the high field resonance being due to 3-HT^-.     

LIGHT ADAPTATION OF DARK-ADAPTED BACTERIORHODOPSIN STUDIED BY NANOSECOND TIME-RESOLVED ABSORPTION SPECTROSCOPY

Abstract— The aim of the study is to clarify the mechanism of light adaptation of dark-adapted bacteriorhodopsin. Double-pulse experiment was carried out at room temperature in aqueous suspension of dark- and light-adapted fragments of the purple membranes for different excitation laser light intensities. It is demonstrated that the route of light adaptation of the dark-adapted bacteriorhodopsin depends on laser light intensity used.     

THE INTERPRETATION OF THE ELECTRON SPIN RESONANCE SPECTRUM OBTAINED BY ULTRAVIOLET IRRADIATION OF THE PHOTO ALLERGEN BITHIONOL

Abstract— The initial radical produced on UV irradiation of the photoallergen bis (2-hydroxy-3,5-dichlorophenyl) sulphide (bithionol) in aqueous solution (pH 8) has been characterised involving chlorine isotope interaction (³⁵Cl 0.059, ³⁷Cl 0.049 mT) in the interpretation of the ESR spectrum. This is the first time that this effect has been observed in solution in these systems.     

QUENCHING OF OXONINE EXCITED STATES BY EDTA VIA ELECTRON TRANSFER AND DEACTIVATION PROCESSES. A COMPARATIVE STUDY OF EXCITED SINGLET AND TRIPLET STATE REACTIVITY

The quenching of excited singlet oxonine by EDTA in aqueous solution leads mainly to deactivation of the dye to the ground state and, to a lesser extent, to electron abstraction. The rate constants for these processes have been measured and compared to those for the same reactions involving the oxonine triplet state. The rate constant of electron abstraction is about ten times greater via the singlet state than via the triplet state. However, the rate constant of deactivation to the ground state is 10³-10⁴ times greater for the excited singlet state than for the triplet state, so that the efficiency of electron transfer is much smaller for the singlet state.     

TRIPLET STATE STUDIES OF FLAVlNS BY ELECTRON PARAMAGNETTC RESONANCE-III

Abstract— A continuation of studies is presented on the excited triplet state of flavins using EPR techniques. Detailed experiments are reported on the triplet state of flavin-mono-nucleotide (FMN) and flavin-adenine-dinucleotide (FAD). Action spectra of triplet yield are explained in terms of the optical absorption for FMN and FAD. Effects of light saturation, concentration quenching and oxygen on the triplet state are discussed. It is suggested that the rate constant k ₃ for the intersystem crossover from the excited singlet to the triplet state is increased by oxygen and quenchers such as KI. Detailed kinetic studies are presented on the formation of the triplet state.     

TRIPLET EXCITED STATES OF PYRIMIDINE NUCLEOSIDES AND NUCLEOTIDES

Abstract— The triplet absorption spectra, lifetimes, extinction coefficients, e_TT, and intersystem crossing quantum yields to the lowest triplet T₁, ø_T1, of thymidine, thymidine monophosphate, uridine and uridine monophosphate, have been determined in various solvents at 300 K.
The effect of H-bonding on ø_T1, of these nucleosides and nucleotides and also of uracil has been determined and discussed. This effect allows, an ordering of ^l,3 n, π* and ^1,3 π, π* states in protic and aprotic solvents.     

MECHANISM OF PHOTOSENSITIZATION BY PHEOPHORBIDE a STUDIED BY PHOTOHEMOLYSIS OF ERYTHROCYTES AND ELECTRON spIN RESONANCE SPECTROSCOPY

Abstract Phcophorbide a (PPa), a causal substance of food intoxication, when excited by exposure to light wavelengths of over 600 nm, caused the photohemolysis of goat erythrocytes in proportion to the incubation time of the cells. The addition of N-₃, an effective scavenger of ¹O₂, to the medium markedly inhibited the hemolysis of erythrocytes in a concentration-dependent manner, whereas the addition of superoxide dismutase (SOD) and catalase, inhibitors of O^-₂ and H₂O₂ generation, respectively, to the medium had little effect on it.
Methods for converting ¹O₂ to a nitroxide radical by 2,2,6,6-tetramethyl-4-piperidone (TMPD) and for trapping O^-₂ and OH by 5,5-dimethyl-l-pyrroline-A'-oxide (DMPO) were employed to observe directly these activated oxygens by electron spin resonance (ESR). The methods provided evidence that only ¹O₂, was produced by PPa, which was excited by light wavelengths of over 600 nm. Both the addition of N₃ to the solution and the removal of oxygen from the solution inhibited the generation of ¹O₂.
These results led us to conclude that ¹O₂ was mainly responsible for the hemolysis of erythrocytes by photoexcited PPa.     

TRIPLET ENERGIES AND ELECTRON AFFINITIES OF METHYL-ACRYLATE AND METHYL-METHACRYLATE

Abstract— The gas phase electron affinities and triplet and singlet excitation energies of the title compounds have been measured by electron transmission and electron-energy-loss spectroscopies. The implications of the results to understanding the mechanism of electron-beam curing of materials containing the acrylate and methacrylate moieties are discussed.     

KINETIC AND SPECTROSCOPIC FEATURES OF SOME CAROTENOID TRIPLET STATES: SENSITIZATION BY SINGLET OXYGEN

Abstract— Triplet-triplet absorption spectra of a series of carotenoid pigments in benzene solution have been determined by pulse radiolysis experiments. The natural lifetimes in deaerated solution have also been measured. They fall in the range 2–10 µ s as found for other carotenoids under similar conditions. Pulsed laser (337 nm) excitation of benzene solutions containing oxygen, carotenoid and a photosensitized molecule (anthracene) showed the generation of absorption spectra of the triplet states. These absorptions decayed by first order kinetics in such a way as to indicate that they were formed in reactions with singlet oxygen, itself generated by interaction with the anthracene triplet state. Bimolecular rate constants for energy transfer from O*₂ (¹△_g), to carotenoid have been evaluated.     

FORMATION OF SEMIQUINONE RADICALS BY ULTRAVIOLET IRRADIATION OF PYRIMIDINES IN METHANOL

Abstract —Ultraviolet irradiation of pyrimidine and 4- and 5-methylpyrimidine in methanol at 113 K gave the corresponding semiquinone radicals in a monophotonic process. It is likely that this process involves an n * triplet state of the photoexcited pyrimidines.     

PHOTOCONVERSION OF PHYTOCHROME IN VIVO STUDIED BY DOUBLE FLASH IRRADIATION IN Mougeotia AND Avena

Abstract— The kinetics of phytochrome phototransformation from the red-absorbing form (P_r) to the far-red-absorbing form (P_fr) in vivo at 22°C were studied using a double flash apparatus with 1-ms flashes. Photoconversion by simultaneous flashes of red light saturates at a low P_fr level, indicating the possible attainment of a photoequilibrium between the excitation of P_r and the photoreversion of intermediates in the course of the I-ms flashes. At saturation energy, simultaneous flashes resulted in about 50% as much P_fr as was produced by saturating irradiation with 5 s red light. Intermediates of the phototransformation pathway were analysed by separating two red or a red and a far-red flash by variable dark intervals. In both plants phototransformation intermediates with half-lives < 1 ms occur, but they are too short-lived to characterize by our method. The subsequent intermediates have half-lives of about 7 ms and 150 ms in A vena , 2 ms and 10 ms in Mougeotia. The conversion from P_r to P_fr seems to be completed 1 s after the red flash in Avena. In the alga Mougeotia , P_fr formation seems to be finished within only 50 ms after the inducing red flash. The kinetics obtained from physiological and spectrophotometric experiments with Avena mesocotyls are almost identical. These observations indicate that the physiological response corresponds directly to the amount of P_fr produced and not to phototransformation intermediates or "cycling" between P_r and P_fr.     

MECHANISM OF PHOTOREDUCTION OF THIAZINE DYES BY EDTA STUDIED BY FLASH PHOTOLYSIS-II. pH DEPENDENCE OF ELECTRON ABSTRACTION RATE CONSTANT OF THE DYES IN THEIR TRIPLET STATE

Abstract— The pH dependence of the apparent reactivity of thiazine dyes in their triplet states has been studied in aqueous solutions, using as electron donor HY^-3, the trianionic species of ethylene diamine tetraacetic acid (EDTA), in the pH range 4–8. The pH dependence is found to be related to a change in the degree of protonation of the triplet excited dye. The apparent reactivity and lifetime of two differently protonated forms of thionine, azur B and methylene blue were determined by classical and dye-laser flash techniques, making it possible to evaluate the rate constant for electron abstraction of these molecules in their triplet states. It is found that: (a) protonation on the ring nitrogen increases the electron-abstraction rate constant of the triplet-state species about twenty-fold, and (b) methylation on the side amino groups decreases it.