PHOTOFRAGMENTATION OF α-OXOCARBOXYLIC ACIDS IN AQUEOUS SOLUTION. AN EPR STUDY. FORMATION OF SEMIDIONE RADICALS BY DECARBOXYLATIVE SUBSTITUTION OF α-OXOCARBOXYLIC ACIDS BY ACYL RADICALS-I: GLYOXYLIC, PYRUVIC, α-OXOBUTYRIC, α-OXOGLUTARIC AND α-OXOISOCAPROlC ACIDS

Abstract— By means of in situ photolysis EPR of aqueous solutions of α-oxocarboxylic acids (RCO-CO₂H) at pH values above 5, semidione radical anions [RC(O^-)=C(O')R] and α-hydroxy-α-carboxy alkyl radicals [RC(OH)CO₂-] were detected. C0₂ was identified as a reaction product. On photolysis of mixtures of α-oxocarboxylic acids (RCOCO₂H and R'COCC₂H), "mixed" semidione radical anions [RC(O^->=C(O)R'] were observed in addition to RC(O^-)=C(O')R, R'C(O^-)=C(O')R', RC(OH)CO₂^- and R'C(OH)CO₂^-. The experimental results are explained in terms of photodecarboxylation (α-clea-vage) of electronically excited RCOCOJ to yield RCO and CO₂. The radicals RC(OH)CO₂^- are formed by reduction of RCOCO₂^- by CO₂^-. The semidione radicals are produced by addition of RCO to RCOCO₂^- followed by decarboxylation of the intermediate adduct. This mechanism was confirmed by generating acyl radicals independently and reacting them with α-oxocarboxylic acids. Selected product studies support the mechanism suggested.     

UPTAKE AND BIOCONVERSION OF α-TOCOPHERYL ACETATE TO α-TOCOPHEROL IN SKIN OF HAIRLESS MICE

Abstract— The photoprotective effect of topically applied α-tocopheryl acetate (vitamin E acetate), a stable derivative of α-tocopherol (vitamin E), and its possible bioconversion to the active antioxidant species (α-tocopherol) was examined in skin tissue of female hairless mice (HRS/J) exposed to UV-B irradiation. Our results indicate that topically applied α-tocopheryl acetate is absorbed into and retained by skin tissue. Furthermore, skin tissue from UV-B-irradiated animals that received daily topical α-tocopheryl acetate treatments contained significantly higher levels (P < 0.001) of α-tocopheryl acetate than non-UV-B-irradiated mice that received identical daily topical α-tocopheryl acetate treatments. Finally, free α-tocopherol levels in skin also were significantly increased (P < 0.00 1) by topical applications of α-tocopheryl acetate and skin levels of free α-tocopherol were significantly greater (P < 0.001) in UV-B-irradiated animals that received daily topical α-tocopheryl acetate treatments than in non-UV-Birradiated animals. These results suggest that UV-B irradiation enhances both the absorption of α-tocopheryl acetate and its bioconversion to free α-tocopherol.     

FORMATION OF REARRANGEMENT PRODUCT IN PHOTOLYSIS OF α,α-DIMETHYLDEOXYBENZOIN UPON CYCLODEXTRIN COMPLEXATION

Abstract Solid-state irradiation of cyclodextrin complexes of α,α-dimethyldeoxybenzoin results in the formation of a significant amount of rearrangement product, 4-isopropylbenzophenone, in addition to cage products. This behavior is not observed in the photolysis in solution or in micellar media.     

PHOTOPHYSICAL PROCESSES IN TROPOLONE, α-METHOXY-TROPONE AND COLCHICINE

Abstract— The spectral characteristics of the emission observed from tropolone, α-methoxy-tropone and colchicine in EPA at 77 K are reported. Luminescence polarization, lifetime, quantum yield and energy transfer experiments provide evidence that the luminescence observed is S₁→ S₀ fluorescence and that the lowest excited singlet state is π, π* The two compounds α-methoxy-tropone and colchicine have been found to exhibit dual luminescence. No phosphorescence could be detected in these three molecules. The photophysical processes for each molecule are discussed and their respective energy level diagrams are also suggested.     

ENERGY TRANSFER-ENHANCED CHEMILUMINESCENCE OF ADAMANTANONE (n,π*) AND ESTER (π,π*) SINGLET AND TRIPLET EXCITED STATES IN THE THERMOLYSIS OF SILYLOXYARYL-SUBSTITUTED SPIROADAMANTYL DIOXETANES†

Abstract— The thermal generation of singlet and triplet excited states from silyloxyaryl-substituted spiroadamantyl dioxetanes lab and the adamantylidineadamantane dioxetane (1c) was investigated by direct and enhanced chemiluminescence (CL). 9,10-Diphenylanthracene (DPA) and 9-fluorenone were used as energy acceptors in the singlet-singlet (S-S), naphthalene and europium chelate Eu(TTA)₃Phen (TTA = thenoyltrifluoroacetone, Phen = 1,10-phenanthroline) in the triplet-triplet (T-T) and 9,10-di-bromoanthracene (DBA) in triplet-singlet (T-S) energy transfer experiments. The direct chemiluminescence observed in the thermolysis of dioxetanes lab consisted of fluorescence derived from the singlet-excited adamantanones 2a,b. In the presence of naphthalene, selective T-S energy transfer with DBA (napthalene as quencher) displayed the adamantanone triplets 2a,b and with Eu(TTA)₃Phen (naphthalene as mediator) also the silyloxyaryl ester 3 triplets. From the Stern-Volmer constants (k_TNT_T⁰) the triplet lifetimes t⁰_t of these triplet state products were assessed. By using the Hastings-Weber standard, the total triplet excitation yield (φ^t) was estimated to be ca 20%. The energies of the first excited singlet and triplet states of the adamantanones 2a,b and the silyloxyaryl ester 3, the products of the thermally induced decomposition of dioxetanes la-c , were determined by semiempirical calculations (AMI-based configuration interaction), which included explicitly solvent effects on the excitation energies in terms of a self-consistent reaction field approach. The calculations revealed that the first excited singlet and triplet states of the adamantanones 2a,b are expectedly n,π*-type excitations while the silyloxyaryl ester 3 possesses π,π* character. The semiempirical computations suggest that excitation of the adamantanones 2a,b as well as the silyloxyaryl ester 3 is feasible in the thermolysis of the spiroadamantyl dioxetanes lab , which has been confirmed by the experimental energy transfer studies.     

EXCITED SINGLET AND TRIPLET PRODUCTS GENERATED FROM OXIDATION OF THE -CO-CH- CONTAINING MOLECULES

Abstract— In a previous study. we found indirect chemiluminescence from air oxidation of various simple ketones in alkaline aprotic solvents containing 9,10-diphenylanthracene and 9,10-dibromoanthracene. By summarizing the experimental results. it was concluded that excited products. mainly in the triplet state. were generated from air oxidation of those simple compounds having a -CO-CH-group via dioxetane intermediates.
Recently. we have succeeded in observing an intense direct chemiluminescence due to the generation of the excited singlet product by the air oxidation of such anthracene derivatives having a -CO-CH-group as 9,10-diisobutyrylanthracene and related compounds. The finding lends strong support to the hypothesis that dioxetane or dioxetanone may be the critical intermediate to give fluorescent singlet products.     

9, 10-DICYANOANTHRACENE-SENSITIZED PHOTOOXYGENATION OF α,α'-DIMETHYLSTILBENES. MECHANISM AND KINETICS OF THE COMPETING SINGLET OXYGEN AND ELECTRON TRANSFER PHOTOOXYGENATION REACTIONS

Abstract— The 9, lodicyanoanthracene-sensitized photooxygenation of 2-methyl-2-butene and (+)-limonene proceeds via the singlet oxygen pathway in carbon tetrachloride as well as in acetonitrile, although the fluorescence of the sensitizer in acetonitrile is quenched by these olefins in an electron transfer quenching mechanism. The 9, 10-dicyanoanthracene-sensitized photooxygenation of cis- and trans-ä, ä′-dimethylstilbenes occurs exclusively via the singlet oxygen pathway in carbon tetrachloride; in acetonitrile, however, singlet oxygen and electron transfer photooxygenation reactions compete with one another. Addition of tetra-n-butyl ammonium bromide and increasing oxygen concentrations favor the formation of the singlet oxygen product, whereas addition of anisole, increasing substrate concentrations and decreasing oxygen concentrations favor the electron transfer photooxygenation products. In carbon tetrachloride, exciplexes of the sensitizer and the dimethylstilbenes are formed which give rise to cidrrans-isomerization of the substrates. In acetonitrile, neither exciplex formation nor cisltrans-isomerization are observed. A mechanism is proposed which allows us to calculate product distributions of the competing singlet oxygen/electron transfer photooxygenation reactions and thus to determine the efficiencies with which encounters between the singlet excited sensitizer and the substrates finally result in electron transfer photooxygenation products. Using (I) these efficiencies, (2) the β-value obtained from singlet oxygen photooxygenation sensitized by rose bengal, and (3) the appropriate k-values determined from fluorescence quenching of 9, 10-dicyanoanthracene in MeCN by oxygen and the stilbene, allows the calculation of the quantum yield of oxygen consumption by this stilbene. The quantum yield thus calculated is strictly proportional to the rate of oxygen consumption experimentally obtained; this result is considered as convincing evidence for the mechanism proposed.     

INTRACELLULAR TARGET FOR α-TERTHIENYL PHOTOSENSITIZATION: INVOLVEMENT OF LYSOSOMAL MEMBRANE DAMAGE

Abstract— Intracellular targets for the photosensitizer α-terthienyl (αT) were examined by fluorescence microscopy and microfluorospectrometry using human nonkeratinized buccal cells. Intracellular distribution of αT was observed as fluorescent patches widely dispersed in the cytoplasm. The distribution of the fluorescent patches was compared with that of acid phosphatase activity visualized as an azo dye produced by the fast garnet 2-methyl-4-[(2-methyl-phenyl)azo]benzenediasonium sulfate reaction. Because both the distribution sites coincided, lysosomes were the likely sites of intracellular affinity of αT. However, because acid phosphatase is not a specific lysosomal marker, we tried to detect another lysosomal enzyme, β-galactosidase, to confirm if the fluorescent patches were lysosomes, using fluorescein-di-(β-D-galactopyranoside) (FDG) as a fluorogenic substrate. Without UV-A (320–400 nm) irradiation of the cells after uptake of αT and FDG, no significant fluorescence was observed. In contrast, with prior UV-A irradiation in the presence of αT and FDG, the bright yellow fluorescence of fluorescein, which is the digested product of FDG, was clearly detected in the cells by fluorescence microscopy. This observation implied that inflow of external FDG into the lysosomes is caused by lysosomal membrane damage on αT photosensitization. The present results indicated that lysosomes are the primary photosensitization site of αT.     

EXCITATION OF CHLOROPHYLL DURING THE PEROXIDASE-CATALYZED OXIDATION OF ETHYL α-FORMYLPHENYLACETATE

Abstract When micelle-solubilyzed chlorophyll is present during the horseradish peroxidase catalyzed aerobic oxidation of ethyl α-formylphenylacetate its fluorescence is observed. The excitation of chlorophyll may occur via energy transfer from the enzyme-generated triplet ethyl benzoylformate. These results imply that excited states may be generated in the roots of Datura innoxia .     

ON THE QUENCHING OF SINGLET OXYGEN BY α-TOCOPHEROL

Abstract —D-α-tocopherol was found to be an effective quencher of ¹O₂ molecules ( k = 2.5 times 10⁸→mol^-1 s^-1 in pyridine) by measuring its effect on the autosensitized photooxidation of rubrene. The quenching process was shown to be almost entirely 'physical', that is, α-tocopherol deactivated about 120 ¹O₂ molecules before being destroyed. The results suggest that this process may be a mechanism for the protective effect of α - tocopherol in photodynamic action.     

MODE OF ACTION OF α-TERTHIENYL ON ESCHERICHIA COLI: EVIDENCE FOR A PHOTODYNAMIC EFFECT ON MEMBRANES

The photodynamic effects of α-terthienyl (αT) in near-UV light (UV-A) on Escherichia coli showed close agreement with the light absorption of αT at different wavelengths suggesting that αT is the primary absorbing molecule responsible for the photosensitized reaction. Studies with DNA repair deficient mutants of E. coli indicated that the bactericidal action of αT/UV-A was not mediated by DNA damage, in direct contrast to the well-known photosensitizer, 8-methoxypsoralen. By using a closed borosilicate glass reaction vessel and various gas mixtures, it was demonstrated that photosensitization of both E. coli and a more resistant bacterium, Pseudomonas aeruginosa , was absolutely dependent on the presence of oxygen. The rate of killing by αT/UV-A showed a rather small dependence on preincubation temperatures, with quite rapid killing at 5°C, suggesting that the movement of αT across the cytoplasmic membrane of E. coli is not the rate limiting step in killing and perhaps is not even necessary for killing. Sodium dodecyl sulphate-polyacrylamide gels of cell membrane proteins after 15 and 30min of treatment with αT/UV-A showed substantial random crosslinking of these proteins. The results taken overall suggest that αT is a photodynamic photosensitizer which exerts its primary effect at the level of the cytoplasmic membrane.     

PHOTOREGULATION OF α-CHYMOTRYPSIN ACTIVITY IN ORGANIC MEDIA: EFFECTS OF BIOIMPRINTING

Abstract α-Chymotrypsin exhibits photoswitchable activities in an organic solvent after covalent modification of the protein backbone with thiophenefulgide active ester (2). The thiophenefulgide-modified α-chymotrypsin exhibits reversible photoisomerizable properties between states (3)-E and (3)-C. The modified α-chymotrypsin, where nine lysine residues are substituted by thiophenefulgide units, retains 60% of the activity of the native enzyme. The activities of thiophenefulgide-modified α-chymotrypsin toward esterification of N -acetyl-L-phenylalanine (4) by ethanol in cyclohexane are controlled by the configuration of the attached photoisomerizable component and by prior bioimprinting of the protein backbone with the reaction substrate (4). The esterification of (4) in cyclohexane using bioimprinted (3)-C is two-fold faster than in the presence of (3)-E. In the presence of a nonbioimprinted enzyme, esterification of (4) by (3)-C is five-fold faster than with (3)-E. The activity of bioimprinted (3)-E toward esterification of (4) is 4.5-fold higher than that of nonbioimprinted (3)-E. Switchable cyclic esterification of (4) is accomplished by sequential photoisomerization of the thiophenefulgide-modified α-chymotrypsin between states (3)-C and (3)-E.     

ANTIVIRAL ACTIVITY OF THE PHOTOACTIVE THIOPHENE α-TERTHIENYL

The naturally occurring thiophene, α-terthienyl, was investigated for phototoxicity against several viruses and a line of mouse cells. The compound was extremely phototoxic to the two-membrane-containing animal viruses, murine cytomegalovirus (MCMV) and Sindbis virus (SV). Antiviral activity was detected at 10⁵μg/m in the presence of UVA. However, no effect was seen in the absence of UV-A, even at 0.1 μg/m of αT. Mouse cells were much more resistant to αT, as was the bacterial virus T4, which does not contain a membrane. Murine CMV, which had been inactivated by αT and UVA, penetrated mouse cells efficiently; but the viral DNA could not replicate, and late viral proteins were not made. Thus viral gene expression was inhibited in the photoinactivated virus. In order to account for all these data we suggest that αT may interact with viral proteins in addition to membrane lipids.     

Thermolysis of 3‐alkyl‐3‐methyl‐1,2‐dioxetanes: Activation parameters and chemiexcitation yields

3‐Methyl‐3‐(3‐pentyl)‐1,2‐dioxetane 1 and 3‐methyl‐3‐(2,2‐dimethyl‐1‐propyl)‐1,2‐dioxetane 2 were synthesized in low yield by the α‐bromohydroperoxide method. The activation parameters were determined by the chemiluminescence method (for 1 ΔH‡ = 25.0 ± 0.3 kcal/mol, ΔS‡ = −1.0 entropy unit (e.u.), ΔG‡ = 25.3 kcal/mol, k₁ (60°C) = 4.6 × 10⁻⁴s⁻¹; for 2 ΔH‡ = 24.2 ± 0.2 kcal/mol, ΔS‡ = −2.0 e.u., ΔG‡ = 24.9 kcal/mol, k₁ (60°C) = 9.2 × 10⁻⁴s⁻¹. Thermolysis of 1–2 produced excited carbonyl fragments (direct production of high yields of triplets relative to excited singlets) (chemiexcitation yields for 1: ϕ^T = 0.02, ϕ ≤ 0.0005; for 2: ϕ^T = 0.02, ϕ^S ≤ 0.0004). The results are discussed in relation to a diradical‐like mechanism. © 2001 John Wiley & Sons, Inc. Heteroatom Chem 12:459–462, 2001     

NEW PHOTO-CONVERTIBLE REACTIONS OF BLUE-FLUORESCENT CALF α-CRYSTALLIN

Abstract— Both native blue fluorescent α-crystallin from calf lenses and UV (300 nm)-irradiated blue-fluorescent α-crystallin, when further irradiated with 365 nm-UV light, produce photo-products capable of emitting a new fluorescence at 455 nm. Illumination of the photo-products with 420 nm visible light regenerates the original fluorescence at 420–425 nm. In addition, another fluorescence at 400 nm has also been found in UV (300 nm)-irradiated blue-fluorescent α-crystallin, when exposed to 365 nm-UV light.     

SENSITIZED PHOTOOXIDATION OF PINENES BY 9,10-DICYANOANTHRACENE

Abstract —The products of 9,10-dicyanoanthracene sensitized photooxidation of α- and β-pinene were separated and identified. The mechanism of this reaction was investigated by fluorescence spectroscopy, solvent dependence and determination of oxidation potentials of olefins. The formation of an exciplex between excited dicyanoanthracene and pinenes was observed in non-polar solvents. On the bases of these observations, an electron transfer mechanism is proposed for the initial step of this reaction.     

SINGLET AND TRIPLET ENERGY TRANSFER IN α-DIKETONE DERIVATIVES OF URACIL and THYMINE

Irradiation of 1-(3,4-dioxopentyl)uracil (UPD) and 1-(3.4-dioxopentyl)thymine (TPD) in acetonitrile solution at 25°C, at the wavelength (280 nm) where only the pyrimidine absorbs the light, sensitizes both fluorescence and phosphorescence of the diketone chromophore in the sidechain. From comparison of the intensity in the corrected excitation spectra with the absorption spectra in acetonitrile solution, it was estimated that the yield of singlet energy transfer in UPD was 0.17 and in TPD was 0.44. It was also observed that the ratio of phosphorescence to fluorescence was greater in the sensitized emission than in that from direct excitation of the diketone chromophore. The yield of triplet energy transfer thus measured corresponds to minimum values for the yields of intersystem crossing from singlet excited state to triplet excited state of 0.075 in the uracil chromophore of UPD and of 0.14 in the thymine chromophore of TPD. These are in agreement with other recent values for these quantities. The value of this type of system as an intramolecular triplet counter is discussed.     

Thermolysis of 3‐alkyl‐3‐methyl‐1,2‐dioxetanes: activation parameters and chemiexcitation yields

3‐Methyl‐3‐(3‐pentyl)‐1,2‐dioxetane 1 and 3‐methyl‐3‐(2,2‐dimethyl‐1‐propyl)‐1,2‐dioxetane 2 were synthesized in low yield by the α‐bromohydroperoxide method. The activation parameters were determined by the chemiluminescence method (for 1 ΔH‡ = 25.0 ± 0.3 kcal/mol, ΔS‡ = −1.0 entropy unit (e.u.), ΔG‡ = 25.3 kcal/mol, k₁ (60°C) = 4.6 × 10⁻⁴s⁻¹; for 2 ΔH‡ = 24.2 ± 0.2 kcal/mol, ΔS‡ = −2.0 e.u., ΔG‡ = 24.9 kcal/mol, k₁ (60°C) = 9.2 × 10⁻⁴s⁻¹. Thermolysis of 1–2 produced excited carbonyl fragments (direct production of high yields of triplets relative to excited singlets) (chemiexcitation yields for 1: ϕ^T = 0.02, ϕ^S ≤ 0.0005; for 2: ϕ^T = 0.02, ϕ^S ≤ 0.0004). The results are discussed in relation to a diradical‐like mechanism. © 2001 John Wiley & Sons, Inc. Heteroatom Chem 12:176–179, 2001     

Radiolysis-Induced Oxidation of Bovine α-Crystallin

Abstract— Radiolysis of water by ionizing radiation results in the production of pure hydroxyl radicals. This technique, combined with analysis by tandem mass spectrometry (MS/MS), has been used to study the effect of hydroxyl radicals on the intact bovine α-crystallin protein. After exposure to -γ-irradiation, the oxidized α-crystallin was digested with trypsin and the resulting peptides were fractionated by reverse-phase HPLC. The isolated fractions were analyzed by matrix-assisted laser desorption ionization and by MS/MS to determine the locations and identities of the modifications. Structural analysis revealed that methionine 1 of αA- and αB-crystallin and methionine 68 of αB-crystallin were oxidized to methionine sulfoxide. Hydroxytryptophan was formed from each tryptophan residue in α-crystallin, although only tryptophan 9 of αA-crystallin was converted into N-for-mylkynurenine. This study has, for the first time, identified the sites of modification and the structures produced in the intact α-crystallin protein by exposure to hydroxyl radicals. By determining the consequences of in vitro exposure of α-crystallin to pure hydroxyl radicals, the in vivo contribution of this reactive oxygen species to the overall oxidative stress of the lens will be achieved from the identification of the modifications to α-crystallin purified from intact human lenses.     

α-氯代脂肪酸对某些稀土的萃取研究

Гавриловаа等^[1]曾报导用α-溴代己酸的氯仿溶液对钐、钕、钆和镥的萃取,其萃取曲线与无取代基的比较向酸性区移动了约1个pH单位。Preston报导了α-溴癸酸的二甲苯溶液对一系列金属离子的萃取,认为所试验的8个稀土金属与α-溴癸酸形成了不溶性的金属络合物^[2]。