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.     

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.     

GLOBAL ANALYSIS OF THE TIME-RESOL VED FLUORESCENCE OF α-CHYMOTRYPSINOGEN A AND α-CHYMOTRYPSIN POWDERS AS A FUNCTION OF HYDRATION

The time-resolved tryptophyl fluorescence of alpha-chymotrypsin A and alpha-chymotrypsin in the crystalline state and in buffer solution at room temperature was analyzed globally. Triple-exponential decay functions are necessary to adequately describe the tryptophyl fluorescence decay surfaces of the protein powders as a function of hydration and in solution. The fluorescence lifetimes of alpha-chymotrypsinogen A (tau 1 = 0.32, tau 2 = 1.30 ns, tau 3 = 3.98 ns) and alpha-chymotrypsin(tau 1 = 0.66 n s, tau 2 = 2.26 ns, tau 3 = 5.40 ns) are constant over the entire hydration range. The spectral positions of the decay-associated spectra of the hydrated powders do not shift as a function of hydration. This indicates that the structures of the zymogen and the active enzyme are unaffected by hydration. The lifetimes of alpha-chymotrypsinogen A in phosphate buffer pH 7.4 are tau 1 = 0.37 ns, tau 2 = 1.17 ns and tau 3 = 3.44 ns while the respective values of alpha-chymotrypsin are tau 1 = 0.47 ns, tau 2 = 1.40 and tau 1 = 3.89 ns.     

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.     

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.     

ON THE LUMINESCENCE OF α-DIKETONES OF THE STEROID SERIES

Abstract—The absorption and luminescent properties of two steroids [16-ketoestrone (I), 5α-androstane-3β-ol-16,17-dione (II)] are compared. The behaviour of these compounds which differ only in their A-ring is significantly different with regard to solvent dependence. The transfer of excitation energy in compound (I) from the phenolic to the diketonic chromophore is studied and compared with data from analogous estrogens. The results obtained in this study indicate a significant long range (presumably conformational) influence of the steroid A and D rings on each other.     

THERMAL GENERATION OF ELECTRONICALLY EXCITED STATES BY α-PEROXYLACTONES: ACTIVATION PARAMETERS AND EXCITATION YIELDS AS A FUNCTION OF α-DEUTERATION*

Abstract— The kinetics and the direct, the 9,10-dibromo- (DBA) and the 9,10-diphenylanthracene- (DPA) enhanced chemiluminescence of tert-butyl α-peroxylactone 1, with and without α-deuteration, was investigated in order to probe the mechanism of direct chemiexcitation by these hyperenergetic molecules. The small secondary isotope effect on the rates and activation parameters suggests that a diradical mechanism is obtained. The partitioning of the diradical intermediate into excited vs ground state product and the yield of triplet excited product are moderately (ca. 5-fold) increased by α-deuteration, but the singlet yield is unaffected. A convenient and useful chemiluminescence method for the determination of fluorescence quantum yields has been developed.     

MODIFICATION OF α-CHYMOTRYPSIN USING A WATER-SOLUBLE PHOTO-FENTON REAGENT

Abstract— -Modification of an enzyme, α-chymotrypsin, was examined by using a water-soluble photo-Fenton reagent. By photoirradiation of the enzyme with the reagent, which can occupy a binding site of the enzyme, a tryptophan residue in the vicinity of the active site was oxidized to N -formylkynurenine. Concurrently, the catalytic properties of the enzyme were largely changed: the K _m was increased and the k _catwas decreased. The decrease in k _cat for a specific amide substrate was the most significant among the esters and amides examined. The water-soluble photo-Fenton reagent would be useful to chemically modify relatively limited regions in biomolecules.     

PHOSPHORESCENCE SPECTRA OF α-DIKETONES IN LOW TEMPERATURE GLASSES

Abstract— The emission properties of o -anisil, p -anisil, benzil, and biacetyl in glasses containing varying proportions of isopropanol have been studied. The different kinds of phosphorescence spectra observed are proposed to be due to triplet states of different conformations and to triplet states of solvated species or other compounds formed by a reaction of the α-diketones with isopropanol.     

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]。     

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.     

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 .     

α-OXIDASE ACTIVITY IN PLANTS AS PROMOTOR OF ELECTRONIC ENERGY

Abstract The a-oxidase activity of higher plants acting on long chain fatty acids generates the lower aldehyde in the ground state; however if chlorophyll or chioroplasts are present the chlorophylls are excited most likely by a chemically initiated electron exchange (CIEEL) luminescence process with the putative a-peroxylactone intermediate. When the aldehyde is substituted for the acid, the lower aldehyde appears in the triplet state. The chiral discrimination observed in the quenching by D- and L-tryptophan of the chlorophyll sensitized emission indicates that the triplet aldehyde is generated within the enzymatic preparation and transfers energy while still bound to the enzyme.
Chlorophylls in chioroplasts are excited by addition of a long chain fatty acid or aldehyde. The mechanism, however, is unknown.     

TIME RESOLVED QUENCHING STUDY OF SERINE-195 LABELED α-CHYMOTRYPSIN

Abstract— Pyrenebutylmethylphosphonofiuoridate or pyrenepropylmethylphosphonofluoridate reacts with the serine 195 residue of α-chymotrypsin to yield a stoichiometric fluorescent derivative. The fluorescence decay of the modified enzyme was studied using a frequency-doubled, synchronously pumped picosecond rhodamine-6G laser excitation source with time-correlated single-photon-counting detection. Upon excitation above 330 nm, it could only be described adequately by a triple exponential decaying function. The exact reason for complexity of the emission has not yet been understood. Heterogeneous labeling seems irrelevant. The presence of different enzyme conformations on the single-photon-counting time-scale seems more plausible. The enzyme conformations where the pyrene label is situated close to the side groups of certain residues such as tryptophan and lysine show a much shorter fluorescence decay time resulting in non-exponential fluorescence behaviour. Dynamic fluorescence quenching experiments indicate that the pyrene label is located in a more polar region of the enzyme characterized by a negative charge. It seems that the pyrene group is too large to fit well in the hydrophobic pocket of the enzyme but is bent more towards the entrance of the cavity which is characterized by a higher polarity. Upon excitation at 296 nm, the excitation energy is transferred from the tryptophan chromophores towards the pyrene label.     

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.     

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.