CALCULATED RATES AND EQUILIBRIA OF THERMAL INTERCONVERSION OF TRIPLET (3g-) AND SINGLET (1Δg and g+) MOLECULAR OXYGEN

Abstract— From spectroscopic data and rate constants in the literature, equilibrium constants and rates of thermal formation of singlet oxygen (¹Δ_g and ¹Σ_g⁺) were calculated for a number of conditions. For the gas phase we estimate K _eq(¹Δ_g³Σ_g^-) = 1.67 exp(-94.31 KJ/RT) and K _eq(¹Σ_g⁺/³Σ_g^-) = 0.33 exp(-157.0 KJ/RT). The calculated rate constants for the ³Σ_g⁺→¹Δ_g transition of O₂ at 25°C varied from 2.5 × 10^-11 s^-1 in water to 4.8 × 10^-16 s^-1 in air, assuming equal solvent interactions with the ground and excited states. Physical quenchers for singlet oxygen are expected to be catalysts for its thermal formation. Equations are presented which allow one to estimate whether such catalysis by quenchers will result in a pro-oxidant effect.     

HO2 ELIMINATION FROM α-HYDROXYALKYLPEROXYL RADICALS IN AQUEOUS SOLUTION

Abstract— In aqueous solutions α-hydroxyalkylperoxyl radicals undergo a spontaneous and a base catalysed HO₂ elimination. From kinetic deuterium isotope effects, temperature dependence, and the influence of solvent polarity it was concluded that the spontaneous reaction occurs via an HO₂ elimination followed by the dissociation of the latter into H⁺ and O₂^-. The rate constant of the spontaneous HO₂ elimination increases with increasing methyl substitution in α-position ( k (CH₂(OH)O₂) < 10s^-1 k (CH₃CH(OH)O₂) = 52s^-1 k ((CH₃)₂C(OH)O₂) = 665 s^-1). The OH^- catalysed reaction is somewhat below diffusion controlled. The mixture of peroxyl radicals derived from polyhydric alcohols eliminate HO₂ at two different rates. Possible reasons for this behaviour are discussed. The mixture of the six peroxyl radicals derived from d -glucose are observed to eliminate HO₂ with at least three different rates. The fastest rate is attributed to the HO₂ elimination from the peroxyl radical at C-l ( k > 7000s^-1). Because of the HO₂ eliminations the peroxyl radicals derived from d -glucose do not undergo a chain reaction in contrast to peroxyl radicals not containing an α-OH group. In competition with the first order elimination reactions the α-hydroxylalkylperoxyl radicals undergo a bimolecular decay. These reactions are briefly discussed.     

PULSE RADIOLYSIS STUDIES IN MODEL LIPID SYSTEMS: FORMATION AND BEHAVIOR OF PEROXY RADICALS IN FATTY ACIDS

Abstract— Radiolytic formation and peroxidation of fatty acid radicals have been investigated by pulse radiolysis techniques in oleate, linoleate, linolenate and arachidonate systems. A strong absorption band at 280 nm associated with conjugated radicals, R_conj, formed in polyunsaturated fatty acid moieties has been used as a probe for kinetic processes occurring at doubly allylic sites in the hydrocarbon chain. Formation of R_conj by O^- has been found to be more efficient than the less selective OH radical. Peroxidation of R_conj is shown to be somewhat slower, ( k _R+ O₂˜ 3 × 10⁸ M ^-1 s^-1), than O₂ reactions with radicals in oleate ( k _R+ O₂= 1 × 10⁹ M ^-1 s^-1). Peroxy radicals generated in these reactions disappear slowly by essentially second order processes (2 k RO₁˜ 10⁷ M ^-1 s^-1). The superoxide radical, O^-₂, shows little if any reactivity towards 0.01 M linolenate or arachidonate over periods of 20 s.     

SEARCH FOR SINGLET OXYGEN LUMINESCENCE IN THE DISPROPORTIONATION OF HO2/O2

Abstract— During the reaction HO₂+ HO₂ (or O₂^-) = H₂O₂+ O₂ in aqueous solution, no luminescence in the region 620–720 nm, expected if the product O₂ were formed in a singlet state, could be detected. If any singlet O₂ is formed, its yield must be less than 10%. Faint luminescence, sometimes found at shorter wavelengths, was shown to arise from reaction of HO₂ with impurities in the reagents present.     

CHLORIDE ENHANCEMENT OF QUENCHING OF TRIPLET METHYLENE BLUE BY Fe(II)

Abstract— The influence of chloride ion on the rate of decay of triplet methylene blue in 0.01 M acid in the absence and presence of ferrous ions was investigated by means of laser flash-photolysis monitored by kinetic spectrophotometry. Chloride weakly accelerates decay of ³MBH^2± in aqueous solution in the absence of Fe(II). Quenching of ³MBH²⁺ by Fe(II) is more strongly catalyzed by Cl^- in both water and 50 v/v% aq. CH₃CN. The uncatalyzed quenching constant, k ₅, is of the order of 1 × 10⁶ M ^-1 s^-1 while in 4.8 M aqueous chloride ( μ – 7.2 M ) k ₅= (37.2 ± 1.8) × 10⁶ M ^-1 s^-1. A possible role of chloride is as a bridging species in quenching via electron transfer between ³MBH²⁺ and Fe(II).     

THE ABSOLUTE VALUE OF THE REACTION RATE CONSTANT OF BILIRUBIN WITH SINGLET OXYGEN IN D2O

Abstract— The chemical reaction rate constant of bilirubin with singlet oxygen in basic aqueous solution has been redetermined to be 3.5 × 10⁸ M^-1 s^-1 by a competitive technique using a 1,3-diphenylisobenzofuran in sodium dodecyl sulfate micelles. Bilirubin also physically quenches a singlet oxygen with a rate constant of 9 × 10⁸ M ^-1 s^-1. The lifetime of singlet oxygen in D₂O solution has been determined to be 35 μ s . The absorption cross-section for the molecular oxygen ³∑_g-→¹δ _g + 1 v electronic transition at 1.06μn in aqueous solution is unexpectedly larger than the gas paase cross-section.     

THE INFLUENCE OF MOLECULAR CONFORMATION ON THE STABILITY OF ULTRAVIOLET STABILIZERS TOWARD DIRECT AND DYE-SENSITIZED PHOTOIRRADIATION: THE CASE OF 2-(2'-HYDROXY-5'-METHYPHENYL)BENZOTRIAZOLE (TIN P)

Abstract— The mechanism for photodegradation of the ultraviolet photostabilizer 2-(2'-hydroxy-5'-methylphenyl)benzotriazole (TIN P) upon direct and dye-sensitized (singlet molecular oxygen [O₂(1Δ_g)]-mediated) irradiation was studied. From the experimental TIN P photodegradation rate data, and low temperature (77 K) fluorescence and phosphorescence quantum yields, one can conclude that the photodegradative process involves phosphorescent states of TIN P. The open conformer of TIN P quenches O₂(¹Δ_g) by physical scavenging with a rate constant (k_q) in dimethylsulfoxide of 2.8 times 10⁶ M ^-1 s^-1. The intramolecular hydrogen-bonded conformer does not appreciably interact with O₂(¹Δ_g). In the presence of a relatively high concentration of OH- (either 5 times 10^-2 M KOH in ethanol or water at pH 13), the ionic form of TIN P (with an ionized phenol group) physically and chemically quenches O₂(¹Δ_g). The reaction rate constant ( k _r) is 1 times 10 ⁸ M ^-1 s^-1, and the ratio k _q/ k _r is approximately three in alkaline aqueous media.     

Dynamics of Flavin in Flavocytochrome b2: A Fluorescence Study

Abstract— The dynamics of the flavin bound to the flavocytochrome b₂ from Hansenula anomala were studied by fluorescence intensity quenching and quenching emission anisotropy with iodide. The fluorescence intensity of bound flavin is decreased 13-fold as compared to the free molecule. The remaining fluorescence decays with two lifetimes equal to 0.963 ± 0.040 and 4.635 ± 0.008 ns and fractional intensities of 0.036 ± 0.002 and 0.964 ± 0.002, respectively. The bimolecular diffusion constant was found to be 3.33 × 10⁹ M ^-1 s^-1 when the flavin is bound to the enzyme and 8.3 × 10⁹ Mv s^-1 when the flavin is free in solution. Thus, the flavin in flavocytochrome b₂ is accessible to the solvent, but the amino acid residues of the binding site inhibit the diffusion of iodide. The rotational correlation time of bound flavin was found to be 2.015 ± 0.365 ns, a value higher than that (155 ps) of free flavin in solution. Our results are discussed on the basis of local dynamics of the flavin.     

AFFINITY FOR OXYGEN IN PHOTOREDUCTION OF MOLECULAR OXYGEN AND SCAVENGING OF HYDROGEN PEROXIDE IN SPINACH CHLOROPLASTS

Abstract— The apparent K _m for O₂ in the photoreduction of molecular oxygen by spinach class II chloroplasts and photosystem I subchloroplast fragments was determined. In both cases, a value of 2 ∼ 3 μ M O₂ was obtained. The reaction rate constant between O₂ and P-430, the primary electron acceptor of PS I, is estimated to be ∼ 1.5 × 10⁷ M ^-1 s^-1 and the factors affecting the production of superoxide by the photoreduction of O₂ in chloroplasts are discussed. Preliminary evidence is presented indicating the occurrence of an azide-insensitive scavenging system for H₂O₂ in chloroplast stroma.     

ACTINOMETRY IN MONOCHROMATIC FLASH PHOTOLYSIS: THE EXTINCTION COEFFICIENT OF TRIPLET BENZOPHENONE AND QUANTUM YIELD OF TRIPLET ZINC TETRAPHENYL PORPHYRIN

Abstract— The extinction coefficient ε_T, of triplet benzophenone in benzene has been directly determined by absolute measurements of absorbed energy and triplet absorbance, Δ D ⁰_T, under demonstrably linear conditions where incident excitation energy, E ₀, and ground state absorbance, A ₀, are both extrapolated to zero. The result, 7220 ± 320 M ^-1 cm^-1 at 530 nm, validates and slightly corrects many measurements relative to benzophenone of triplet extinction coefficients made by the energy transfer technique, and of triplet yields obtained by the comparative method.
As E ₀ and A ₀ both decrease, Δ D ⁰_T becomes proportional to their product. In this situation, the ratio R = (1/ A ₀)(dΔ D ⁰_T/d E ₀) = (ε_T - ε_G)φ_T. Measurements of R , referred to benzophenone, give (ε_T - ε_G)φ_T for any substance, without necessity for absolute energy calibration.
Both absolute and relative laser flash measurements on zinc tetraphenyl porphyrin (ε_T - ε_G at 470 nm = 7.3 × 10⁴ M ^-1 cm^-1) give φ_T= 0.83 ± 0.04.     

GAMMA AND PULSE RADIOLYSIS STUDIES OF THE REACTION BETWEEN SUPEROXIDE IONS AND OXYHEMOGLOBIN-METHEMOGLOBIN SYSTEM

Abstract— The radiolytic studies of oxyhemoglobin or methemoglobin in neutral aerated aqueous solutions with formate ions, lead to three conclusions:
The oxidation of oxyhemoglobin by O^-₂ is not important. The observed low oxidation yield is probably due to the slow reaction with hydrogen peroxide produced by O^-₂ disproportionation.
The reduction of methemoglobin in γ radiolysis reaches a plateau which could be explained by structural considerations.
The reduction of methemoglobin by O^-₂ ions, if it occurs, is relatively slow: k = 1.4 × 10³ M ^-1 s^-1. But a problem remains concerning the spectral characteristics of the product.     

RATE CONSTANTS FOR INTERACTION OF 1O21Δg) WITH AZIDE ION IN WATER

Abstract— The rate constant for quenching of ¹O₂ by azide ion in water was determined to be (5.0 ± 0.4) × 10⁸ M ⁻¹ s⁻¹ using a variety of sensitizers (including humic acids) and ¹O₂ acceptors. The apparent second-order rate constant decreases with pH below pH 5.5 in accordance with the protonation of azide ion to form hydrazoic acid (p K _a= 4.6). Quenching by hydrazoic acid is at least 2 orders of magnitude slower than by azide ion. Greater than 99% of all interactions between ¹O₂ and azide ion involve physical quenching rather than chemical reaction. Humic acid triplets are not significantly quenched by azide ion at concentrations less than 2 m M , allowing azide ion quenching to be used as a diagnostic test for the intermediacy of ¹O₂ in photosensitized oxidations in natural surface waters.     

ON THE REACTION OF SINGLET MOLECULAR OXYGEN WITH N-ALLYLTHIOUREA IN AQUEOUS SOLUTION

Abstract— Experiments on the photooxidation of N -allylthiourea, thiourea, and N-allylurea sensitized by the dye phenosafranine show that in N -allylthiourea the thiourea group is the site of singlet oxygen attack, while the allyl moiety neither reacts with nor quenches this metastable form of O₂ (in neutral aqueous solutions). Low concentrations of N^-₃ (a known quencher of singlet oxygen) strongly reduce the photooxidation of allylthiourea by a mechanism which apparently obeys simple competition kinetics. From these results the rate constant of the reaction between allylthiourea and singlet oxygen is obtained ( k = 4 × 10⁶ M ^-1 s^-1; pH = 7.1).     

DESIGN AND PERFORMANCE OF THE OKAZAKI LARGE SPECTROGRAPH FOR PHOTOBIOLOGICAL RESEARCH

A computer-operated spectrograph was recently built at Okazaki, Japan. Different specimens can be placed on a horseshoe-shaped focal curve (10 m long) covering a wavelength range of 250 to 1000 nm so they can be irradiated simultaneously. The linear dispersion is about 0.8 nm/cm. The photon fluence rate on the focal curve is 5 x 10¹⁵. photons x cm^-2 x s^-1 at 300nm and 1 x 10¹⁶ photons x cm^-2 x s^-1 at 600 and at 900 nm. The spectral half width is 5.5 nm or less on the focal curve. The stray light content is about 10^-5 of the main peak at the peak wavelength ± 100 nm. Specimens are set in microcomputer-controlled threshold boxes so that wavelengths, photon fluence rates, photon fluences and timing of irradiations are controlled automatically according to a pre-programmed schedule. An optical fiber system is also provided for remote irradiations.     

THE PHOTOCHEMICAL INTERACTION BETWEEN THE TRIPLET STATE OF 8-METHOXYPSORALEN AND THE MELANIN PRECURSORL–3, 4 DIHYDROXYPHENYLALANINE

Abstract— The photochemical interaction between 8-methoxypsoralen (8-MOP) and the melanin precursorL–3,4-dihydroxyphenylalanine(dopaH₂) has been studied using laser flash photolysis. Triplet excited 8-MOP was thus found to abstract electrons from dopaH₂ ( k ∼ 2 × 10⁹ dm³ mol^-1 s^-1) to form semireduced 8-MOP and semioxidised dopaH₂.The technique of pulse radiolysis was used to establish separately the spectra of (a) the semi-reduced form of 8-MOP at pH 6.5 and (b) the semioxidised forms of dopaH₂ at pH 6.5, 5.8, 4.6 and 3.3. The corresponding λ_max and extinction coefficients found were: for 8-MOP at pH 6.5, λ_max= 350 nm (= 9050 dm³ mol^-1 cm^-1); for dopa at pH 6.5, λ_max= 305 nm (ε= 12000 dm³ mol^-1 cm^-1) and for dopaH at pH 3.3, λ= 305 nm (ε= 5900 dm³ mol^-1 cm^-1).     

AMPHIPHILIC COPOLYMERS AS MEDIA FOR LIGHT-INDUCED ELECTRON TRANSFER-II. ELECTROSTATIC EFFECT ON THE BACK ELECTRON TRANSFER

Abstract— Photosensitized reduction of zwitterionic viologen (SPV) and methyl viologen (MV²⁺) was investigated using an amphiliphilic copolymer having phenanthryl and sulfonate groups (APh) as photosensitizer in aqueous solutions. In the presence of triethanolamine the accumulation of SPV * (photoproduct) was found to be faster than that of MV⁺. This attributed to the electrostatic repulsion between SPV. and anionic segments of APh. Such difference between SPV and MV²⁺ was minimized in the case of the related monomer model. Retardation of the back reaction for the APh-SPV system was also demonstrated by laser photolysis, k _b= 8.7 × 10⁷ M ^-1 s^-1 for the polymer system as compared to k _b= 2.8 × 10⁹ M ^-1 s^-1 for the monomer model system. Strong salt-effects on the yield of the photoreduction and the rate of back reaction confirm the strong electrostatic interaction between the photoproducts and polyanions. This remarkable electrostatic effect of the polyanions was simulated by electrochemical redox reactions by using a graphite electrode coated with APh.     

TRANSIENT PHOTOCHEMISTRY OF NEUTRAL RED

Abstract— Flash photolysis of neutral red between pH 1.3 and pH 11 yields the triplet species ³DH₂⁺²³DH⁺ and ³D. Both ³DH₂⁺² and ³D exhibit first order decay with rate constants of 1.6 ± 0.3 × 10⁴ s^-1 but ³DH⁺ decays within the lifetime of the flash. Over the entire pH range, ascorbic acid quenches the triplet, forming the semireduced radicals DH₃⁺² DH₂⁺ and DH, all of which exhibit second order decay with k = 1.8 ± 0.4 ± 10⁸ M ^-1s^-1 most probably by recombination with semioxidized ascorbic acid. The dependence of the rate of decay of radical neutral red on the identity of reversible reductants supports the back-electron transfer mechanism, as does digital simulation of complex radical disproportionation schemes. In contrast to the efficient reduction of triplet neutral red by ascorbic acid, its reduction by EDTA is quite inefficient.     

OXODIPYRROMETHENES AS REACTIVE SINGLET OXYGEN ACCEPTORS. MEASUREMENT OF THEIR CHEMICAL REACTION RATES BY A LASER FLASH PHOTOLYSIS TECHNIQUE

Abstract— Singlet oxygen has been generated directly in 1,1,2-trichloro, 1,2,2-trifluoroethane solution by irradiation of the oxygen dimol 2³∑-_g→2¹δ_g transition with a pulsed dye laser and the ³∑_g^-→¹δ_g+ lv transition with a continuous Nd-YAG laser. The rates of chemical reaction and physical quenching of singlet oxygen so generated has been measured for a series of substituted oxodipyrro-methenes. The results show that the oxodipyrromethenes react with singlet oxygen at rates comparable to that for 1,3-diphenylisobenzofuran. The rate of quenching of singlet oxygen by ground state oxygen has been measured to be 2.5±0.3 × 10³ M^-l s^-1.     

THE QUANTUM EFFICIENCY FOR THE INTERPHOTOCONVERSION OF THE BLUE and PINK FORMS OF PURPLE MEMBRANE

When the cations bound to purple membrane are removed it turns blue, and when this blue membrane is irradiated its color changes to pink. Irradiation of pink membrane leads to the reformation of blue membrane. We have determined that the quantum efficiency for the formation of pink membrane from deionized blue membrane is 1.6 ± 0.6 ± 10 ⁴ at 0^oC, pH 5.0. We also found that the quantum efficiency for the back photoconversion, i.e. the formation of blue membrane from pink membrane, is 8.8 ± 1.6 ± 10^-3 at 0^oC, 55 times greater than that of the forward photoconversion reaction. The extinction coefficients of the pink membrane and blue membrane were determined to be 44 500 ± 670 cm^-1 M^-1 at 491 nm and 54 760 ± 830 cm^-1 M ^-1 at 603 nm, respectively, assuming light-adapted purple membrane is 63 000 cm^-1 M ^-1 at 568 nm. The quantum efficiency for forming pink membrane from blue membrane is much lower than that for forming the photointermediate of the blue membrane's photocycle. Their relationship is similar to that of light-adaptation and photocycle of the dark-adapted purple membrane.     

PICOSECOND LASER SPECTROSCOPY AND OPTICALLY DETECTED MAGNETIC RESONANCE ON A MODEL PHOTOSYNTHETIC SYSTEM

Fluorescence-detected magnetic resonance of triplets in zero magnetic field (FDMR), fluorescence fading (FF) due to triplet-formation, both at 4.2 K, and prompt fluorescence decay kinetics (FDK) at room temperature have been measured for free pheophorbide- a (f-Pheo) and bound (b-Pheo) to a synthetic polypeptide (L-L ys -L-A la -L-A la )_n, dissolved in dimethylformamide (DMF). Fluorescence decay kinetics measurements of f-Pheo in DMF yielded 1-5 ns lifetimes, for b-Pheo in DMF a ~ 50 ps decay-component was found emitting at 730–750 nm. Zero-field splitting parameters |D| and |E| of the lowest triplet state T₁ were determined from FDMR spectra as (337 and 24) 10^-4 cm^-1 for f-Pheo and (359 and 25) 10^-4 cm^-1 for b-Pheo, both in DMF. Decay rate constants of the three spin levels of T¹ of b-Pheo ( K _x= 1200 50 s^-1, k _y= 440 25 s^-1, k _z= 80 5 s^-1) and relative steady-state populations (N_x= 28 2%, N_y= 47 2%, N_z= 26 2%) determined from FF curves predict a fluorescence decrease at the D–E and D + E FDMR transitions, whereas experimentally a fluorescence increase is observed. The FDMR sign-inversion results from singlet-singlet energy transfer from b-Pheo monomers to their aggregates, followed by fast intersystem crossing to T₁. These results indicate that aggregates are formed by two or more b-Pheo molecules at different positions on the folded polypeptide chain. This situation resembles that in chlorophyll-proteins, containing low-lying traps, resulting from interaction of chromophores with other chromophores and with the protein environment.