SPECTROSCOPIC STUDIES OF CUTANEOUS PHOTOSENSITIZING AGENTS—II. SPIN TRAPPING OF PHOTOLYSIS PRODUCTS FROM SULFANILAMIDE AND 4-AMINOBENZOIC ACID USING 5,5-DIMETHYL-1-PYRROLINE-1-OXIDE

Abstract— The photodecomposition of sulfanilamide, 4-aminobenzoic acid and related analogs in aqueous solution has been studied with the aid of spin traps 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) and CH₃NO₂ as well as by direct electron spin resonance techniques. The NH₂ radical was trapped by DMPO during the photolysis of aqueous solutions of sulfanilamide with a Xe arc lamp. Studies with [¹⁵N¹]-sulfanilamide indicated that the NH₂ radical was generated by homolytic fission of the sulfur-nitrogen bond. Under the same conditions DMPO trapped the H and SO⁻₃ radicals during photolysis of sulfanic acid. Direct photolysis of sulfanilamide, sulfanilic acid and Na₂SO₃ in the absence of any spin trap yielded the SO⁻³ radical. Photolysis of 4-aminobenzoic acid at pH 7 gave the H radical which was trapped by DMPO. At low pH values OH and C₆H₄COOH radicals were generated during the photolysis of 4-aminobenzoic acid. No e⁻_aq were trapped by CH₃NO₂ when acid (pH 4) and neutral aqueous solutions of sulfanilamide or 4-aminobenzoic acid were photoirradiated. The mechanism of formation of known photoproducts from the free radicals generated by sulfanilamide and 4-aminobenzoic acid during irradiation are discussed. The free radicals generated by these agents may play an important role in their phototoxic and photoallergic effects.     

MECHANISMS OF PHOTOELIMINATION REACTIONS OF ALKYL BENZOHYDROXAMATES

Abstract— Photolysis at 254 nm of alkyl benzohydroxamates [C, H, CONHOR: R = CH₃ H₂CH₃ CH(CH₃)₂, CH₂C₆H₅ CH(CH₃)C₂H₅ CH(CH₃)- n -C₆H₁₃] in acetonitrile or hydrocarbon solvents gives benzamide. These reactions can be sensitized by benzophenone (at ca. 350 nm) and are quenched by cis-piperylene. Racemization occurred when 2-octyl (+)-benzohydroxamate was irradiated in cyclohexane. These results are consistent with a mechanism involving a triplet biradical. Photolysis of phenyl benzohydroxamate [C₆H₅CONHOC₆H₅] and benzyl N -methylbenzohydroxamate [C₆H₅CON-(CH₃)OCH₂Q₆H₅] cannot be quenched with ris-piperylene and appear to be singlet reactions.     

THE REACTION OF METHYLENE WITH HYDROGEN

Abstract— An investigation has been made of the reaction between methylene, formed by the photolysis of ketene, and hydrogen. Ethane, ethylene and methane are the major hydrocarbon products, and it has been shown that the formation of these products may be adequately described by the sequence of processes
CH₂CO + hv → CH₂+ CO (1)
CH₂+ H₂→ CH₃+H (2)
2CH₃→ C₂H₆ (3)
CH₃+ H₂+ CH₄+ H (4)
CH₂+ CH₂CO → C₂H₄+ CO (7)
In particular, the relative rates of ethane and methane formation are consistent with the known rate constants for reactions (3) and (4), and it is not therefore necessary to postulate the participation of an 'insertion' process
CH₂+ H₂→ CH₄ (6) to account for the formation of methane.
Decrease of the energy possessed by the methylene, either by increase of the wavelength of ketene photolysis, or by increase of gas pressure, is shown to result in an increase in the reactivity of the methylene towards ketene relative to its reactivity towards hydrogen (i.e. the ratio k₂/k₂ increases).     

PHOTOLYSIS OF AQUEOUS SOLUTIONS OF POTASSIUM CIS-DIAQUOBIS (OXALATO) CHROMATE (III)

Abstract— The photolysis of aqueous solutions of cis -[Cr(C₂O₄)₂(H₂O)₂]- at 254 nm and pH 4 produced CO₂ and H₂ in nearly equal yields. The quantum yield of hydrogen, φ², increased by 9% and the yield of carbon dioxide, φ, by 65% as the pH was lowered from 4 to I. The total gas yield, φ_gas, decreased in the presence of added oxalate or chromium (II) ions and when the light intensity was lowered. The gas yield in D₂O was appreciably higher than in H₂O. The variation of φ_gas with pH (D) and with added oxalate ion was roughly parallel in the two liquid media. The gas yield increased in the series:
A tentative mechanism is suggested to explain the results.     

THE PRESENCE OF PHOTOREACTIVE β-CARBOLINE FLUOROPHORES IN WEATHERED WOOL KERATIN

Abstract Solvent-cleaned Merino wool, which was enzymatically digested with protease K, displayed a fluorescence maximum at 500 nm when excited at 430 nm. The yield of this emission was approximately 15 times greater for a fiber tips digest than for a digest of the mid-length region of the same fibers.
Separation of the components in the wool tip digest by silica gel thin-layer chromatography revealed the presence of several fluorescent species. The chromatographic mobility of these species was similar to the behavior observed in a preparation of authentic 1-methyltetrahydro-P-carboline, 1,3-dicarboxylic acid.
Mass spectroscopy of the fluorescent compounds in the wool tip digest displayed molecular ions, [M + H]⁺ with m/z = 273.0890 and 257.0560, and molecular weights that define the formulae C₁₄H₁₂N₂O₄ and C₁₁H₈O₄N₂, respectively, which correspond to β-carboline 1,3-dicarboxylic acids. Mass spectral evidence also indicates the presence of two other β-carbolines.     

STRUCTURE AND CONFORMATION OF PHOTOSYNTHETIC PIGMENTS AND RELATED COMPOUNDS 7. ON THE CONFORMATION OF THE METHYL ESTER OF (20-METHYLPHYTOCHLORINATO)NICKEL(II)-A BAaERIOCHLOROPHYLL c MODEL COMPOUND

Abstract The crystal and molecular structure of the title compound 4 have been determined by single crystal X-ray crystallography. The structure shows the typical S,-ruffled conformation observed for Ni(II) tetrapyrroles. Compared to the structure of the methyl ester of (13²-demethoxycarbonyl-pheophorbidato a)nickel(II), 4 shows a smaller C_a-C_m-C_m angle and a higher degree of conformational distortion at the methyl-substituted C20 position. This local distortion of the macrocycle might account for the bathochromic shifted absorption spectra of the bacteriochlorophylls c compared to the d-series. Crystal data: C₃₅H₃₈N₄NiO₃; tetragonal, P4₃2₁2, a = 15.335(7) Å, c = 25.11(2) Å, V = 5904, Z = 8, λ(Mo Kα) = 0.71069 Å, μ= 0.701 mm⁻¹, P(000) = 2624, 130 K, R = 0.058 for 5700 reflections with F > 4.0σ(F).     

EFFECT OF TRANSMEMBRANE POTENTIAL ON CHARGE PHOTOSEPARATION IN LIPOSOMES

Abstract— An unexpected transmembrane potential effect on the recombination rate of the pheophytin or bacteriopheophytin anion-radicals (dissolved in membrane) and ascorbic ion-radicals (dissolved in aqueous interior) has been established in liposomes. The influence of transmembrane potential on the recombination rate of Ru³⁺ (dissolved in inner volume) and (C₁₈H₃₇)V⁺ or (C₁₄H₂₉)V⁺ (dissolved in membrane) was observed. The potential was created by a potassium concentration gradient between inner and outer volumes of liposomes in the presence of valinomycin. The effect of the potential was considered on the bases of: (1) it was determined by the diffusional drift of the hydrophobic radicals in a radial direction in the membrane, according to the direction of the electric field; (2) the electric field changed the rate constant of the electron transfer, owing to the effects on the free energy and electronic coupling. Our results show the first explanation to be preferable.     

THE PHOTOREDUCTION OF METHYLENE BLUE BY ARYLAMINOMETHANE-SULFONATES

Abstract— The photoreduction of methylene blue in the presence of arylaminomethanesulfonates (RAMS = RC₆H₄NHCH₂SO₃Na) was studied by laser and conventional flash photolysis. These compounds quenched the methylene blue triplet deviating from a normal Stern-Volmer behaviour. For low quencher concentrations, a Rehm-Weller relationship was found between the k _q's and the DL G 's obtained for the electron transfer reactions. The lack of further quenching at higher [RAMS] is ascribed to the formation of a ground state ion pair between the dye and the anionic quencher which, on excitation, forms a triplet state unable to under go electron transfer for steric reasons. A second order decay rate constant was found for the semireduced species (MB') ( ca. 5 × 10⁹ M _-1 s_-1, independent of the RAMS used) and is attributed to a proton transfer from the radical zwitterion (RC₆H₄NH CH₂SO₃⁻) to MB. The overall dependence on the substituent of the bleaching observed by continuous irradiation follows the triplet behaviour.     

MECHANISM OF THE PHOTOCHEMICAL REACTION OF DIPHENYLAMINES WITH CARBON TETRACHLORIDE

Abstract. The quantum yields of HCI (φ_HC1) formation have been measured for the photolysis of N -methyldiphenylamine (MeDPA), triphenylamine (TPA) and diphenylamine (DPA) in the presence of CCl₄ in polar solvents. The quantum yields of N-methylcarbazole formation (φ_mφca) have also been determined for the system MeDPA-CCl₄. With increasing CCl₄ concentration, φ_HCl increases as φ_MeCA decreases, and φ_HCl reaches maximum values 2.7 at 1 M CCl₄. Using laser photolysis, transient spectra have been recorded for MeDPA in the absence and presence of CCl₄ in polar and non-polar solvents, and for TPA. Transient absorption due to the triplet states and photocyclization products (without CCU), exciplexes, the (C₆H₅)₂ NCHi radical, the MeDPA⁺ cation radical, the (TPA⁺., CCl₄) ion pair, and the TPA⁺ cation radical have been identified. The mechanistic implications of these results are discussed.     

SPECTROSCOPIC STUDIES OF CUTANEOUS PHOTOSENSITIZING AGENTS–XII. SPIN TRAPPING STUDY OF THE FREE RADICALS GENERATED DURING THE PHOTOLYSIS OF PHOTO ALLERGENS BITHIONOL and FENTICHLOR

Free radicals were trapped and observed by ESR when photoallergens bithionol and fentichlor were irradiated in the presence of spin traps N- t -butyl-α-phenylnitrone (PBN) and 5,5-dimethyl-pyrroline-N-oxide (DMPO). In the absence of air, both PBN and DMPO trapped a carbon-centered radical. The carbon-centered radical, which was capable of abstracting a hydrogen atom from cysteine, glutathione, ethanol and formate, was identified as an aryl radical derived from the homolytic cleavage of the carbon-chlorine bond. In the presence of air, both carbon-centered radicals and hydroxyl radicals were trapped by DMPO. Under similar conditions, the yield of the hydroxyl radicals was greater from bithionol than from fentichlor. The presence of the hydroxyl radical was confirmed by kinetic experiments employing hydroxyl radical scavengers (ethanol, formate). Superoxide and H₂O₂ were not involved. Experiments with oxygen-¹⁷O indicated that the hydroxyl radicals came exclusively from dissolved oxygen. The precursor of the hydroxyl radical is postulated to be a peroxy intermediate (ArOO*) derived from the reaction of an aryl radical (Ar*) with molecular oxygen. Both bithionol and fentichlor photoionized only when excited in the UVC (<270 nm) region. Free radicals have long been postulated in the photodechlorination of bithionol and fentichlor and the present study provides supporting evidence for such a mechanism. Aryl and hydroxyl radicals are reactive chemical species which may trigger a series of events that culminate in photoallergy.     

TIME RESOLVED STUDIES OF 1.27 μm LUMINESCENCE FROM SINGLET OXYGEN GENERATED IN HOMOGENEOUS and MICROHETEROGENEOUS FLUIDS

-The luminescence at 1.27 μm from the ³→→¹δ_g transition of the oxygen molecule has been detected from a variety of liquid systems. A Q-switched laser delivering pulses of 532 nm light was the excitation source, a germanium photodiode was the detector and substituted porphyrins were used as photosensitizers. Protio- and deutero- forms of several solvents were studied and the singlet oxygen lifetimes determined directly agreed well with published values. T_δ in D₂O was found to be 55 μs and, by extrapolation from a series of H₂O - D₂O mixtures, a value of 3.3 μs was obtained for T_δ in H₂O. The technique was shown to be useful in measuring T_δ values in several microheterogeneous systems such as surfactant micelles, vesicles and human serum albumin.     

ON THE PHOTOCHEMISTRY OF X-RAY PRODUCED RADICALS TRAPPED IN FROZEN ETHANOL MATRICES

Abstract— Ethanol and ethanol-water matrices were exposed to X-rays at 77K and the photochemistry and paths of radical conversion were investigated by EPR methods. The main X-ray induced radical, CH3ĊHOH, is probably photoionized by 254 nm light. The following radicals are produced during prolonged UV-irradiation of CH₃ĊHOH radicals: ĊH₃, ĊHO, H and 2 types of radicals giving singlet EPR spectra. One of these radicals (d) is bleachable with 580 nm light, ĊH₃ and ĊH₃ĊHOH being formed during the bleaching, the other one (e) is unbleachable and the most stable radical in the matrix during annealing. The CH₃ radicals decay at 77 K (τ∽ 10 min) and produce CH₃-CHOH radicals and the unbleachable radical (e). Stable H-atom signals were seen in X-irradiated ethanol-water mixtures (volume ratio 2:1) at 77 K. The H-atom signals increased during photobleaching of the trapped electrons in the matrix and during UV-photolysis of CH₃CHOH radicals.     

PHOTOREDUCTION OF IRON(III)PORPHYRINS

Abstract— Ferrideuteroporphyrin in benzene, water or micelle solutions containing primary or secondary alcohols as well as in pure or basic 2-propanol solutions is clearly reduced to the ferrous state by continuous light irradiation in the Soret region. Quantum yields range between 4 × 10⁻⁴ and 3 × 10⁻² depending on the solvents used and on the coordination state of the ferric porphyrin. As inferred from laser pulse photolysis experiments, the primary chemical step appears to be the homolytic cleavage of the bond between the ferric ion and a coordinated alcoholate anion leading to the ferrous porphyrin and the alkoxy radical. This cleavage is found to occur within less than 50 ns. The alkoxy radical rearranges leading to the α-hydroxyalkyl radical which reacts with excess ferric porphyrin leading to further reduction. The reaction rate constant for the reaction of α-hydroxyisopropyl radicals is found to be k = (2.1 ± 0.3) × 10⁸ M ⁻¹ s⁻¹ in pure 2-propanol. As expected, this rate is greatly increased in basic 2-propanol where α-hydroxyisopropyl radicals deprotonate.     

PRIMARY PRODUCTS OF LIQUID WATER PHOTOLYSIS AT 1236, 1470 AND 1849 Å

Abstract— The yield of the primary products of the liquid water photolysis at 1236 and 1470 Å is reported. It was found that besides the dissociation of the excited water molecules into H and OH radicals probably e⁻_aq is also formed. The H and OH radicals were scavenged by means of formate, and the e⁻_aq together with a part of H₂O* by adding carbon dioxide. The quantum yields determined at 1236 Å, are: Φ(H, OH) = 1.03 & 0.02, 0.06 <Φ( e⁻_aq , H₂O*) < 0.12 and at 1470 Å,: Φ(H, OH)=0.72±0.02, 0.037 < Φ( e⁻_aq , H₂O*) <0.075. The quantum yield of high purity water at 1849 Å in the absence of any scavengers is Φ(H, OH)=0.022. Previously published data by us for 1849 Å are also given: Φ(H, OH)=0.33 ± 0.01 and 0.02 < Φ ( e⁻_aq H₂O*) < 0.04. Reaction mechanisms are proposed.     

A METHOD FOR THE CORRECTION OF QUANTUM YIELDS OF FLUORESCENCE IN SOLUTIONS

Abstract— The correction factors due to the overlap of absorption and emission spectra have been calculated for various concentrations of fluorescein dye at pH 12 by using the experimentally obtained value of apparent molar absorptivity, ε₂, for the fluorescence radiation. The observed fluorescence intensities for the same-surface detection for a given geometry have been corrected for the secondary emission effect thus calculated, and also for refraction and dilution effects, to give corrected relative quantum yields for the solutions. These have been converted to the absolute values by comparison of the integrated fluorescence quantum spectrum of a dilute solution of fluorescein with that for quinine sulphate in 0.1 N H₂SO₄. The corrected quantum yields compare well with values given in the literature.     

INVOLVEMENT OF SINGLET OXYGEN IN THE PHOTOTOXICITY MECHANISM FOR A METABOLITE OF PIROXICAM

It has been previously shown that a metabolite of piroxicam but not piroxicam itself causes phototoxicity to cells in vitro after exposure to UVA (320–400 nm) radiation. The phototoxicity mechanism for this metabolite, 2-methyl-4-oxo-2H-l,2-benzothiazine-l,l-dioxide (Compound I), was investigated. In vitro phototoxicity to human mononuclear cells was assayed using 0.5 m M Compound I and UVA radiation. The UVA fluence required for phototoxicity of Compound I was lower by a factor of 2-3 in D₂O buffer compared to H₂O buffer. Superoxide dismutase and mannitol, which remove O₂^- and OH", respectively, do not decrease the phototoxicity. The photodecomposition of Compound I was inhibited by sodium azide, enhanced by human serum albumin and unaffected by mannitol. Stable photoproducts of Compound I were not toxic to the cells. The quantum yield of singlet oxygen based on its emission at 1270 nm was 0.19 and 0.35 for Compound I and s2 ± 10^-3 and 10^-2 for piroxicam in D₂O and C₆H₆, respectively. While the extremely low quantum yield for singlet oxygen from piroxicam appears to account for its lack of phototoxicity, the phototoxicity mechanism for its metabolite, Compound I, most likely does involve singlet oxygen.     

SPECTROSCOPY AND PHOTOCHEMISTRY OF 8-SUBSTITUTED 5-DEAZAFLAVINS

Abstract— Absorption, fluorescence and phosphorescence spectra as well as fluorescence and phosphorescence quantum yields of 8-X-5-deazaflavins (X = C1, NO₂, p -NO₂-C₆H₄, N(CH₃)₂, NH₂, p -NH₂-C₆H₄, p -N(CH₃)₂-C₆H₄-N=N) were determined. It was found that all these data are highly influenced by the substituent at position 8 of the 5-deazaisoalloxazine skeleton. Also the photoreduction of 8-X-5-deazaflavins in the presence of electron donors was studied. It was established that the photoreduction leads to the formation of a 5,5'-dimer and/or a 6,7-dihydro compound. Reduction of the C(6)-C(7) bond is promoted by strong electron-donating substituents and bulky electron donors. 5-Deazaftavins with a reducible substituent at position 8 exhibit reduction of the substituent prior to the reduction of the 5-deazaisoalloxazine skeleton.     

PHOTODYNAMIC INACTIVATION OF E. COLI BY ROSE BENGAL IMMOBILIZED ON POLYSTYRENE BEADS

Abstract. The photodynamic inactivation of E. coli by visible light and O₂ was found to occur in the presence of the sensitizer rose bengal, immobilized by covalent bonding to polystyrene beads. The demonstrated absence of significant amounts of dissolved rose bengal indicated that an inactivation mechanism based on penetration of sensitizer molecules into the cell's interior could not be operating. Survival curves typically exhibited induction periods followed by rapid exponential death, with 99.99% kill requiring 1–2 h depending on conditions. A mechanism involving the participation of photo-generated singlet excited oxygen O₂(¹δ) in inactivation of E. coli is proposed. The photodynamic inactivation rate increased significantly in H₂O compared with H₂O, which is evidence supporting singlet oxygen as an active intermediate, since O₂(¹δ) has a much longer lifetime in H₂O than in H₂O. H₂O did not act as a short term poison in the absence of sensitizer.     

A PULSE RADIOLYSIS STUDY OF THE ONE-ELECTRON OXIDATION AND REDUCTION OF 5, 10-METHENYLTETRAHYDROFOLATE

Abstract— The one-electron reduction and oxidation of 5,10-methenyltetrahydrofolate has been studied in aqueous solution in the acidity range H₀= -1 to pH = 7 using the reducing species CO^-₂ and (CH₃)₂-COH and oxidising species Br^-₂, and H₂SeO+₃. The spectral and other properties of the radicals so formed were found to be indcpendent of the reductant/oxidant used. Two protolytic forms of both the oxidised and reduced radicals were observed with approximate p K , values of 0.5 ± 0.3 being determined. Both the bridged form (5.10-methenyltetrahydrofolate) and the unbridged form (5- formyltetrahydrofolate) were found to be easily oxidised, whereas only the former could be reduced.     

PRODUCTS AND RELATIVE REACTION RATES OF THE OXIDATION OF TOCOPHEROLS WITH SINGLET MOLECULAR OXYGEN

Abstract— The relative reactivity of singlet molecular oxygen, 0₂(¹Δ_g), α-,β-,Γ-and δ with -tocopherol (vitamin E) was investigated using microwave discharge generation as a uniquely clean source of singlet oxygen and using a hydrocarbon solvent to approximate the membrane environment. The relative efficiencies of the tocopherols for O₂(¹Δg) were found to decrease in the order: D-α-tocopherol > D-β-tocopheroI > D–Γ-tocopherol > D-δ-tocopherol. The reaction products in all cases were found to be mixtures of quinone and quinone epoxides apparently resulting from decomposition of the primary product, the hydroperoxydienone.