bluuebunny/crossref_metadata_2025 huggingface dataset:
VECTORIAL ELECTRON TRANSPORT ACROSS LIPID VESICLE MEMBRANES DRIVEN BY TWO PHOTOREACTIONS. I. ETHYLENEDIAMINETETRAACETIC ACID AS AN ELECTRON DONOR via FLAVIN TRIPLET STATE IN THE INNER COMPARTMENT AND CYTOCHROME c AS AN ELECTRON ACCEPTOR FROM CHLOROPHYLL TRIPLET STATE IN THE OUTER COMPARTMENT
Authors: Zhao Zhan‐Gong , Tollin Gordon
Published: January 2008

Negatively charged vesicle suspensions containing chlorophyll a (chl) dissolved in the lipid bilayer, flavin mononucleotide (FMN) and/or ethylenediaminetetraacetic acid (EDTA) enclosed in the inner compartment as electron sources and oxidized cytochrome c (cyt c[ox]) in the outer compartment as an electron acceptor have been studied using laser flash photolysis and steady‐state irradiation methods. Cytochrome c initially quenches the chl triplet state (3chl) generating the chlorophyll cation radical (chi+′) in the membrane. Reverse electron transfer from cyt c(red) to chl+. subsequently occurs in a kinetically biphasic reaction, with rate constants of 430 pT 30 and 21.9 pT 1.7 s−1 for the fast and slow phases, respectively. In the absence of FMN, reduction of chl+′ by EDTA in the inner compartment can be observed during steady‐state irradiation but not in a laser flash photolysis experiment. This is due to a low reaction yield, which is probably limited by the repulsive electrostatic interaction between EDTA and the negatively charged membrane. When FMN was enclosed together with EDTA in the inner Compartment, the reaction yield of vectorial electron transfer across the bilayer from EDTA to cyt c(oX) was increased by a factor of six during steadystate white light irradiation. Laser flash photolysis and steady‐state irradiation experiments using red and blue light excitation have demonstrated that the enhancement mechanism involves the formation of fully reduced FMN by blue light‐sensitized photooxidation of EDTA via the flavin triplet state, occumng simultaneously with red lightsensitized electron transfer to cyt c via the chlorophyll triplet state.
GENE EXPRESSION OF TEH B875 LIGHT‐HARVESTING PREPOLYPEPTIDES FROM RHODOSPIRILLUM RUBRUM IN ESCHERICHIA COLI
Authors: Ghosh Robin , Cornacchia Luigi , Bachofen Reinhard
Published: January 2008

The gene coding for the prepolypeptides of α and β, obtained as a 429 bp fragment from chromosomal DNA of Rhodospirillum rubrum S1 by polymerase chain reaction amplification, were cloned in tandem into the high‐level expression vector pOTSNco 12 for epxression in Escherichia coli. The vector pOTSNco 12 is a derivative of the pAS vector system, which contains the strong ΛPL promotor and is under tight control by the cI857 repressor encoded by the expression strain AR58. Induction of transcription from the ΛPL promotor is achieved by shifting the growth temperature from 32 to 42d̀C. Expression of the gene products was monitored by sodium dodecylsulfate polyacrylamide gel electrophoresis and western blotting. The expressed B875 light‐harvesting prepolypeptides were located in the E. coli inner membrane and could not be removed by washing with high salt. The amount of expressed B875 light‐harvesting prepolypeptides was estimated to be about 0.1% of the total soluble protein.
CHARACTERIZATION OF THE SPECTROSCOPIC PROPERTIES OF A TETRAHYDROCHRYSENE SYSTEM CONTAINING A RIGIDIFIED HYDROXYNITROSTILBENE CHROMOPHORE: AN INHERENTLY FLUORESCENT LIGAND DESIGNED FOR THE ESTROGEN RECEPTOR
Authors: Anstead Greorgy M. , Hwang Kwang‐Jin , Katzenellenbogen John A.
Published: January 2008

Abstract— A tetrahydrochrysene system that embodies a hydroxy‐ and nitro‐substituted stilbene chromophore held rigidly near planarity by the tetracyclic nature of the compound was prepared as a fluorescent ligand for the estrogen receptor. It shows strong solvent‐dependent fluorescence at long wavelengths. The solvent polarity dependence suggests that the fluorescence arises from an excited state with much nπ* character in cyclohexane; stronger emission comes from an intramolecular charge transfer state that has lower energy in more polar solvents, and finally progressive quenching of the charge transfer state occurs in solvents of higher polarity. The quenching effect is particularly evident in protic solvents. In water, however, the compound shows fluorescence of unusually high energy for an intramolecular charge transfer state, which suggests that photochemistry may be occurring. In solutions of gamma‐cyclodextrin, emission from the nitrotetrahydrochrysene is red shifted and intensified relative to water. Photobleaching occurs in H2O but not in ethanol or gamma‐cyclodextrin solution. The change in dipole moment between the ground and excited states for the nitrochrysene is 12.9 D, similar to our previous measurements for related nitrostilbenes. The compound displays red‐shifted emission in triethylamine, perhaps due to an excited state hydrogen‐bonded complex, The absorption and emission properties of the corresponding nitrophenolate were also studied. The nitrophenolate exhibits reverse solvatochromism in its absorption spectra. In conclusion, the high sensitivity of the emission energy and quantum yield of the title compound make it of potential utility as a fluorescent probe.
DIFFERENTIAL EXPRESSION OF PYRIMIDINE DIMER‐BINDING PROTEINS IN NORMAL AND UV LIGHT‐TREATED VERTEBRATE CELLS
Authors: McLenigan Mary , Levine Arthur S. , Protić Miroslava
Published: January 2008

Abstract— The expression of UV damage‐specific DNA‐binding proteins was examined in various phylogenetically distant species with differing DNA repair phenotypes. Two distinct constitutive DNA‐binding activities, one specific for cyclobutane pyrimidine dimers and the other for non‐cyclobutane dimer photoproducts, were detected. The expression of these binding activity was found to be variable throughout the animal kingdom: cold‐blooded vertebrastes show a constitutive cyclobutance dimer‐binding activity excusively, and primates reveal only non‐cyclobutane expression (rather than the constitutive presence)of these UV damage‐specific DNA‐binding activities after UV traeatment correlate with the cell's capacity for DNA repair. In addition, cyclobutane pyrimidine dimer‐binding activities could be detected only in cells with eestablished photoreactivating activity
A PULSED PHOTOLYSIS PROCEDURE FOR DETERMINING OXYGEN EQUILIBRIUM PARAMETERS OF LOW‐AFFINITY NONCOOPERATIVE HEMOGLOBINS
Authors: Astatke Mekbib , Parkhurst Lawrence J.
Published: January 2008

Abstract— It is often very difficult to obtain precise values for oxygen equilibrium constants for low‐affinity nonco‐operative hemoglobins owing to the ease with which they are oxidized and denatured with even the most gentle stirring or agitation. We have developed a procedure that eliminates this problem by maintaining the hemoglobin as HbCO except during the brief time it is present as Hb, and HbO2 forms following photolysis. Oxygen is continuously removed by an enzyme system. The solution is photolyzed to 100% every2–3 min during the deoxygenation process to obtain maximum absorbance changes at the Hb‐HbCO isosbestic. These maximum absorbance changes at known times during the deoxygenation provide the necessary data for obtaining the oxygen equilibrium constant. These absorbance changes are used with enzyme kinetics equations to obtain calculated times. The simple equations, which neglect heme concentration, will be satisfactory for nearly all conditions, but for generality the complete equations are given. The variance minimizations are with respect to observed and calculated photolysis times. The results for native carp Hb over a range of temperatures are in excellent agreement with results obtained with considerably more material and greater difficulty by other methods.
LASER‐INDUCED FLUORESCENCE IN MALIGNANT and NORMAL TISSUE OF RATS INJECTED WITH BENZOPORPHYRIN DERIVATIVE
Authors: Andersson‐Engels Stefan , Ankerst Jaro , Johansson Jonas , Svanberg Katarina , Svanberg Sune
Published: January 2008

Abstract— Laser‐induced fluorescence was used to characterize the localization of intravenously administered ben‐zoporphyrin derivative‐monoacid (BPD‐MA) 3 h postinjection in different rat tissue types, including an induced experimental malignant tumor. A comparison of the fluorescence properties and the demarcation potential between the newer sensitizer BPD‐MA and four other substances, hematoporphyrin (HP), polyhematoporphyrin ester (PHE), tetrasulfonated phthalocyanine (TSPc) and the commercially available Photofrin earlier investigated, is included. The fluorescence light was induced with a nitrogen laser, emitting at 337 nm. The fluorescence spectrum in the region380–750 nm was analyzed by a polychromator equipped with a diode array detector. The demarcation potential between tumor and surrounding tissue in terms of fluorescence signal for the tumor model used was 2:1 for BPD‐MA. In comparison with the other drugs, HP shows about the same demarcation potential, whereas Photofrin and PHE exhibit about 3 times better and TSPc about 1.5 times better demarcation. By also employing the endogenous tissue fluorescence signature the contrast was enhanced by a factor of about 2 for each of the five drugs.
PHOTODYNAMIC DESTRUCTION OF LYSOSOMES MEDIATED BY NILE BLUE PHOTOSENSITIZERS
Authors: Lin Chi‐Wei , Shulok Janine R. , Kirley Sandra D. , Bachelder Cindy M. , Flotte Thomas J. , Sherwood Margaret E. , Cincotta Louis , Foley James W.
Published: January 2008

Previous studies have established that a number of Nile blue derivatives are potent photosensitizers and that they are localized primarily in the lysosomes. The present study examines whether the lysosome is a main target of the photocytotoxic action mediated by these sensitizers. Chosen for this study were NBS‐61 and sat‐NBS, which represented, respectively, derivatives with high and moderate degrees of lysosomal selectivity. Overall results indicated that both derivatives are very effective in mediating a photodestruction of lysosomes. This is indicated by the light‐and drug‐dose‐dependent losses of acid phosphatase staining particles, reduction of hexosaminidase in the lysosomecontaining subcellular fraction, and impairment of the lysosomes to take up and sequester acndine orange. Ultrastructurally, swollen and ruptured lysosomes were seen as one of the first evidences of cell damage mediated by these photosensitizers. However, the study also showed that sat‐NBS, which is less lysosomal selective, was less effective in mediating lysosomal destruction. Also, the degree of lysosomal destruction mediated by sat‐NBS did not parallel the degree of cytotoxicity generated. This implies that for derivatives that are not exclusively localized in the lysosome, other subcellular sites may also be damaged by the photodynamic action and may play a role in the photocytotoxic process.
PHOTOSENSITIZATION WITH ETIOBENZOCHLORINS AND OCTAETHYLBENZOCHLORINS
Authors: Kessel David , Morgan Alan
Published: January 2008

The photophysical and photobiological properties of a series of etiobenzochlorins were evaluated in cell culture using murine leukemia L1210 cells. In the series of agents tested, the chlorin‐(mono)sulfonate was the most efficacious, the tin chlorin somewhat less so and the tin chlorin‐sulfonate much less active. The parent chlorin was essentially inactive at the limit of solubility. Photodamage was assessed by measuring alterations in surface hydro‐phobicity (via a two‐phase partitioning procedure), amino acid transport and membrane potential. Additional information was provided from fluorescence microscopy, which was used to identify sites of sensitizer binding and effects of photodamage on the binding patterns of fluorescent probes specific for mitochondria, lysosomes and plasma membranes. Effects of photodamage on fluorescence lifetime distribution of the membrane probe trimethylamino‐diphenyl hexatriene were examined. The data obtained were consistent with localization of the parent etiobenzochlorin and tin derivative at lysosomal loci. the chlorin‐sulfonate at plasma and mitochondrial membranes and tin‐sulfonate at the cell surface.
INHIBITION OF 80 kDa PROTEIN PHOSPHORYLATION BY SHORT‐WAVELENGTH UV LIGHT IN NIH 3T3 CELLS
Authors: Shin Incheol , Yoon Yoo Sik , Kang Kewon , Park Sang Dai , Joe Cheol O.
Published: January 2008

The exposure of NIH 3T3 fibroblast cells to 254 nm UV radiation resulted in a temporary depression of DNA synthesis and inhibition of 80 kDa protein phosphorylation. This inhibition of protein phosphorylation was correlated with decreased protein kinase C activity in the membrane fractions of UV‐damaged cells. The inositol tnphosphate contents measured, by the competitive binding assay using bovine adrenal binding protein, showed 80% reduction in the fibroblasts treated with 15 J/m2 of UV light. The intracellular diacylglycerol concentration was also markedly reduced in UV‐damaged cells. The results suggest that UV light causes acute reductions of inositol triphosphate and diacylglycerol contents in cells along with decreases in membrane protein kinase C activity, which leads to the inhibition of phosphorylation of an acidic protein of 80 kDa.
ADRIAMYCIN RESISTANCE IN CHINESE HAMSTER FIBROBLASTS FOLLOWING OXIDATIVE STRESS INDUCED BY PHOTODYNAMIC THERAPY
Authors: Fisher Anita M. R. , Ferrario Angela , Gomer Charles J.
Published: January 2008

Photodynamic therapy (PDT) generates reactive oxygen species that are responsible for the initial cytotoxic events produced by this treatment. An extended (16 h) porphyrin incubation prior to light irradiation increased expression of the 75, 78 and 94 kDa glucose‐regulated stress proteins (GRP), as well as the cognate form of the 70 kDa heat shock protein. However, these stress proteins were not induced following isoeffective PDT doses using a short (1 h) porphyrin incubation protocol. In the current study, Chinese hamster fibroblasts were used to examine sensitivity to adjunctive PDT and adriamycin as previous reports indicate a correlation between stress protein synthesis and a decrease in adriamycin cytotoxicity. Treatments that either induced GRP (i.e. PDT with an extended porphyrin incubation or exposure to the calcium ionophore A23187) or did not induce GRP (i.e. PDT with a short porphyrin incubation or UV irradiation) were followed at increasing time intervals with a 1 h adriamycin incubation. A time‐dependent decrease in adriamycin cytotoxicity was observed when cells were first exposed to either of the PDT protocols or to A23187. Alterations in intracellular drug levels did not account for the change in adriamycin sensitivity. Likewise, intracellular glutathione concentrations and antioxidant enzyme activities were not significantly altered following PDT or A23187. Parameters associated with altered adriamycin sensitivity included a decrease in the percentage of S phase cells following PDT and A23187 as well as a depletion of intracellular ATP after PDT using the extended porphyrin incubation. These results demonstrate that PDT can be added to the growing list of diverse stresses producing transient resistance to adriamycin and that stress protein induction is not universally associated with all oxidative treatments inducing this resistance.
PHOTOACTIVE METHYLENE BLUE DYE DERIVATIVES SUITABLE FOR COUPLING TO PROTEIN
Authors: Motsenbocker M. , Masuya H. , Shimazu H. , Miyawaki T. , Ichimori Y. , Sugawara T.
Published: January 2008

Methylene blue is a very strong photoactive dye that has an absorption peak (668 nm) that corresponds well to a popular low‐cost diode laser. However, it has not been used in photodynamic tumor therapy and immunodiagnostics because it cannot be covalently coupled to protein. Therefore, methylene blue derivatives having a succinimido or maleimido functional group were synthesized and coupled to antibody, serum albumin and transfemn proteins. Incorporation of dye into antibody protein at high ratios (more than three per molecule) caused precipitation and loss of antibody activity. Inclusion of one or more carboxylic acid residues in the methylene blue derivative before coupling to protein alleviated the precipitation problem, and up to 36 methylene blue dye molecules could be attached to an antibody fragment using bovine serum protein as a carrier. Methylene blue derivatives and protein complexes formed from them oxidized luminol when stimulated with red light. The new dye conjugates were used in an optically pumped chemiluminescence immunoassay for α‐fetoprotein. These compounds and techniques should also be useful for photodynamic tumor therapy where it is desired to attach a red‐absorbing photoactive dye to antibody protein.
In vitro PHOTODYNAMIC EFFECTS OF LYSYL CHLORIN p6: CELL SURVIVAL, LOCALIZATION AND ULTRASTRUCTURAL CHANGES
Authors: Leach Michael W. , Higgins Robert J. , Autry Susan A. , Boggan James E. , Lee Shwn‐Ji H. , Smith Kevin M.
Published: January 2008

The in vitro cell survival, localization and ultrastructural changes following irradiation were examined in 9L glioma cells sensitized with a new photosensitizer, lysyl chlorin p6 (LCP). In clonogenic assays, LCP was 10–100‐fold more phototoxic than photofrin II on a μg/mL basis. Lysyl chlorin p6 uptake was blocked when cells were incubated at 2°C. In view of the chemical properties of LCP, this finding indicates that uptake probably occurred through the endocytic pathway. Fluorescence studies showed LCP localized in a region of the endocytic compartment similar in size, shape and distribution to that labeled by lucifer yellow CH (LY), as well as localizing diffusely throughout the perinuclear cytoplasm. Cells stained with both LY and LCP, however, had distinctly separate regions of staining. Lysyl chlorin p6 localization differed from that of fluorescent probes labeling the mitochondria, Golgi apparatus and endoplasmic reticulum. Ultrastructural changes at both 2 and 30 min after laser irradiation were similar. Mitochondria were often condensed or swollen and also had constrictions and cytoplasmic invaginations. The Golgi apparatus, perinuclear space and rough endoplasmic reticulum (RER) were dilated. These data demonstrate that LCP localizes in a portion of the endosomal compartment, but that morphologic damage initially occurs in the mitochondria, Golgi apparatus and RER.
ROLE OF HYDROXYL‐BEARING AMINO ACIDS IN DIFFERENTIALLY TUNING THE ABSORPTION SPECTRA OF THE HUMAN RED AND GREEN CONE PIGMENTS
Authors: Merbs Shannath L. , Nathans Jeremy
Published: January 2008

The human red and green cone pigments differ at either 15 or 16 amino acids, depending upon which polymorphic variants are compared. Seven of these amino acid differences involve the introduction or removal of a hydroxyl group. One of these differences, a substitution of alanine for senne at position 80, was found previously to produce a 5 nm blue shift. To determine the role of the remaining six hydroxyl group differences in tuning the absorption spectra of the human red and green pigments, we have studied six site‐directed mutants in which single amino acids from the green pigment have been substituted for the corresponding residues in the red pigment. Blue shifts of 7 and 14 nm were observed upon substitution of phenylalanine for tyrosine at position 277 and alanine for threonine at position 285, respectively. Single substitutions at positions 65, 230, 233, and 309 produced spectral shifts of 1 nm or less. These data are in good agreement with a model based upon sequence comparisons among primate pigments and with the properties of site‐directed mutants of bovine rhodopsin. Nonadditive effects observed in comparing the absorption spectra of red‐green hybrid pigments remain to be explained.
THE EFFECT OF ACCEPTOR GROUP VARIATION ON THE SOLVATOCHROMISM OF DONOR‐ACCEPTOR FLUOROPHORES
Authors: Anstead Gregory M. , Carlson Kathryn E. , Kym Philip R. , Hwang Kwang‐Jin , Katzenellenbogen John A.
Published: January 2008

Abstract The absorption and emission characteristics of five hydroxytetrahydrochrysenes substituted with acceptor groups (nitro, cyano, methylketone, 1° amide and methyl ester) (THC‐NO2, THC‐CN, THC‐COCH3, THC‐CONH2 and THC‐CO2CH3, respectively) were investigated in an extensive set of solvents. The order of absorption and fluorescence bathochromicity are: THC‐NO2 > THC‐COCH3 > THC‐CN ≥ THC‐CO2CH3 > THC‐CONH2 and THC‐NO2 >> THC‐COCH3 > THC‐CO2CH3 > THC‐CN > THC‐CONH2, respectively. The emission spectra of these compounds are sensitive to the solvent polarity (ET[30] scale) in the order: THC‐NO2 > THC‐COCH3 > THC‐CO2CH3 > THC‐CONH2 > THC‐CN. The response of the emission maxima of these compounds to the solvent polarity and hydrogen‐bond donor/acceptor properties (π*/α/β and acity/basity scales) was also determined. The emission energies of THC‐NO2 were most sensitive to π*, β, acity, and basity of the solvent; those of the amide were least sensitive to the solvent π*, β, and basity.

The ground‐ and excited‐state dipole moments were determined by semiempirical molecular orbital calculations and the absorption/fluorescence solvent‐shift method, respectively. THC‐NO2 had the largest ground‐ and excitedstate moments. The ester and amide had the smallest ground‐ and excited‐state moments, respectively. In general, unsatisfactory results were obtained for correlations of the emission and absorption energies, fluorescence solvatochromism and the ground‐ and excited‐state dipole moments with the Hammett substituent constants of the five acceptor groups. Acceptable correlations were obtained for the absorption and emission energies and the fluorescence solvatochromism with the substituent constants if the cyano compound was excluded.
PROTON, Mg2+ AND PROTEIN AS COMPETING LIGANDS FOR THE FLUORESCENT PROBE, MAG‐INDO‐1: A FIRST STEP TO THE QUANTIFICATION OF INTRACELLULAR Mg2+ CONCENTRATION
Authors: Morelle Bruno , Salmon Jean‐Marie , Vigo Jean , Viallet Pierre
Published: January 2008

Abstract Increasing evidence of the role of magnesium in various cellular mechanisms has led to the need to develop an accurate method for the evaluation of magnesium concentration in cells. 1H‐indole‐6‐carboxylic acid, 2–(4‐bis‐[carboxymethyl]amino‐3–[carboxy]ethoxy) (mag‐indo‐1) is used as a fluorescent indicator for ionized magnesium concentration. A physicochemical study of this probe has pointed out (1) that at concentrations higher than 10 μM, the presence of dimers can alter the different equilibria and (2) at concentrations, avoiding the dimer (≤ 10 μM), three fluorescent forms are in equilibrium with the deprotonated form of mag‐indo‐1 (L), which are the protonated form LH, the magnesium‐bound form LM and the protein‐bound form LP. A model is proposed that takes into account the equilibria between the four species. In a solution containing magnesium and protein, a complex fluorescence spectrum can be resolved by a combination of the three fluorescence spectra (L, LM, LP). However, under these conditions, the LH fluorescence spectrum is not taken into account for the spectral resolution. Finally, from the contribution of characteristic fluorescence spectra in the experimental fluorescence spectrum, the magnesium concentration can be estimated with accuracy. Such a method should be further applied to magnesium determination in different cell lines.
STEADY‐STATE OPTICAL POLARIZATION ANISOTROPY OF RODLIKE MOLECULES UNDERGOING TORSIONAL TWISTING MOTIONS
Authors: Ludescher Richard D. , Ludescher William H.
Published: January 2008

Abstract An analytical expression was derived for the steady‐state optical polarization anisotropy expected for a rodlike molecule undergoing torsional twisting motions about its long molecular axis. This expression is parameterized in terms of the ratio τ/φ where τ is the excited‐state lifetime of the chromophore and Φ is a characteristic time for rotational twisting (Yoshimura et al., J. Mol. Biol. 179, 453–467, 1984) that is proportional to the torsional rigidity of the flexible rodlike molecule; the amplitude at any given value of τ/φ is also determined by the angle between the chromophore transition dipole and the long molecular axis of the molecule.
EFFECT OF UVA AND BLUE LIGHT ON PORPHYRIN BIOSYNTHESIS IN EPIDERMAL CELLS*
Authors: Sassa Dan He Shigeru , Lim Henry W.
Published: January 2008

Abstract— To study porphyrin biosynthesis in normal human keratinocytes and A431 cells derived from human epidermoid carcinoma, cultured cells were incubated with delta‐aminolevulinic acid (ALA), the precursor of porphyrin synthesis, and accumulation of porphyrins was measured spectrofluorometrically. Both human keratinocytes and A431 cells accumulated porphyrins in a time‐dependent and a dose‐dependent fashion. Protoporphyrin was the predominant porphyrin accumulated by both cell types. Porphyrin accumulation was enhanced by Ca Mg ethylene‐diaminetetraacetic acid, a ferrochelatase inhibitor, and the enhancement was reversed by the addition of iron, suggesting the utilization of iron by ferrochelatase. The effect of light on porphyrin accumulation was evaluated by exposing the ALA‐loaded A431 cells to ultraviolet‐A (UVA) and blue light radiation, followed by continued incubation with ALA for 2–48 h. There was an enhancement of porphyrin accumulation 2–48 h after the radiation as compared with nonirradiated controls. Consistent with this finding, ferrochelatase activity decreased in these cells at 24 h and 48 h. These data demonstrate that human keratinocytes and A431 cells are capable of porphyrin biosynthesis, and that exposure of porphyrin‐containing A431 cells to light, which includes the Soret band spectrum, decreases the ferrochelatase activity, which is responsible, at least in part, for the further increase in porphyrin level.
HEMOGLOBIN OXYGEN AFFINITY IS INCREASED IN ERYTHROPOIETIC PROTOPORPHYRIA
Authors: Hirsch Rhoda Elison , Lin Margaret J. , Pulakhandam Usha Rao , Nagel Ronald L. , Sandberg Sverre
Published: January 2008

Abstract— Whole blood and hemolysates from seven normal and three erythropoietic protoporphyria patients were compared in terms of their hemoglobin function. The oxygen affinity (P50) of the erythropoietic protoporphyria hemolysates compared to normals (13.1 ± 0.2 vs 17.5 ± 0.3 mmHg; P < 0.001) and erythropoietic protoporphyria erythrocytes compared to normals (23.4 ± 0.6 vs 27.1 ± 0.5 mmHg; P < 0.001) were increased while oxygen‐binding cooperativity (n‐value of the Hill equation) were similar. We conclude that hemoglobin function in erythropoietic protoporphyria patients is altered, but without pathophysiologic consequences. Because hemoglobin in which protoporphyrin IX substitutes for heme has a low oxygen affinity, our findings of a higher than normal affinity in erythropoietic protoporphyria red cells and hemolysates may indirectly support the findings by others that protoporphyrin IX binds to hemoglobin at non‐heme‐binding sites. In addition, based on the effect of other abnormal hemoglobins, this shift in P50 will decrease any tendency for anemia in erythropoietic protoporphyria patients.
UPCONVERSION OF THE SINGLET OXYGEN 1269 nm EMISSION TO THE VISIBLE
Authors: Allan D. , Gorman A. A. , Hamblett I. , Hodgson B. W. , Lambert C.
Published: January 2008

Abstract— Experiments are described that enable the kinetic behavior of singlet oxygen, O2(IΔg), to be monitored in the time‐resolved mode using a photomultiplier to detect deep orange light (γmax 660 nm). This orange light is a consequence of the upconversion of the natural emission of O2(IΔg) at 1269 nm.
THE EFFECTS OF PHOTODYNAMIC THERAPY USING DIFFERENTLY SUBSTITUTED ZINC PHTHALOCYANINES ON VESSEL CONSTRICTION, VESSEL LEAKAGE AND TUMOR RESPONSE
Authors: Fingar Victor H. , Wieman T. Jeffery , Karavolos Pamela S. , Doak Karola Weber , Ouellet René , van Lier Johan E.
Published: January 2008

Abstract— The effects of four different zinc phthalocyanines were studied during and after photodynamic therapy (PDT). Measurements of vessel constriction, vessel leakage, tumor interstitial pressure, eicosanoid release, and tumor response of chondrosarcoma were made in Sprague‐Dawley rats. Animals were injected intravenously with 1 μmol/ kg of mono‐, di‐, or tetrasulfonated zinc phthalocyanine, or 1 μmol/kg of a zinc phthalocyanine substituted with four tertiary butyl groups. Tissues were exposed to 400 J/cm2 670 nm light 24 h after photosensitizer injection. An additional group of animals was given indomethacin before treatment. The use of the monosulfonated and tertiary butyl substituted zinc phthalocyanines in PDT caused the release of specific eicosanoids, caused vessel constriction, and induced venule leakage and increases in tumor interstitial pressure. Tumor cures of 27% and 7% were observed. Photodynamic therapy using the disulfonated zinc phthalocyanine did not induce vessel constriction or the release ofeicosanoids, however; tumor cure was 43%. The use of thc tetrasulfonated zinc phthalocyanine caused intermediate effects between the mono‐ and disulfonated compounds. The administration of indornethacin to animals completely inhibited the effects of PDT using the monosulfonated compound but had minimal effects on treatment using the disulfonated compound. This suggests that the monosulfonated and disulfonated compounds act by different mechanisms of destruction.
LARGE MUTAGENIC LESIONS ARE INDUCED BY PHOTODYNAMIC THERAPY IN MURINE L5178Y LYMPHOBLASTS*
Authors: Deahl J. Thom , Oleinick Nancy L. , Evans Helen H.
Published: January 2008

Abstract— Mutagenic lesions at the thymidine kinase locus (tk) in mouse lymphoma L5178Y (LY) cells treated with red light and either Photofrin (PF) or chloroaluminurn phthalocyanine (AIPc) as the photosensitizer were compared in the relatively photodynamic therapy (PDT)‐sensitive strain LY‐R16 and the relatively resistant strains LY‐S1 and LY‐SR1. Southern blot analysis revealed that 92% (36/39) of the PDT‐induced thymidine kinase (TK −/‐) mutants of strains LY‐R16 and LY‐SR1 lost the entire active tk allele. (Strain LY‐S1 lacks a known tk polymorphism and has not been analyzed for loss of the active tk allele.) A decrease in galactokinase (GK) activity in the TK−/‐ mutants has been taken as an indication that the mutagenic lesion extends from the tk gene to the closely linked galactokinase gene (gk). Using PF as the photosensitizer, GK activity was decreased in 45% of the LY‐R16 mutants and in 22% of the LY‐S1 and LY‐SR1 mutants. With photoactivated AIPc, 59% of the TK −/‐ mutants of strains LY‐S1 and LY‐SR1 showed GK inactivation. (LY‐R16 mutants were not analyzed because of the low LY‐R16 mutant frequency induced by PDT with AlPc.) Thus, many of the TK−/‐ mutants of LY cells induced by PDT with either PF or AlPc harbor multilocus lesions.
PHOTOPHYSICAL PROPERTIES OF CHLOROPHYLL a LANGMUIR‐BLODGETT MULTILAYER FILMS
Authors: Tkachenko N. V. , Grandell D. , Ikonen M. , Jutila A. , Moritz V. , Lemmetylnen H.
Published: January 2008

Abstract— Multilayer Langmuir‐Blodgett films of chlorophyll u and chlorophyll a in L‐α‐phosphatidylcholine dipal‐mitoyl matrix have been prepared. The films were protected by covering them with eight layers of stearic acid to ensure high film stability during the light illumination. Based on the time‐resolved fluorescence measurements a model with two types of chlorophyll arrangement was developed. One arrangement could be attributed to an aggregated state and the other to a homogeneously distributed state. Transient absorption in the wavelength range from 380 and 740 nm and the time range from 0.1 to 100 μs were measured. The chlorophyll a radicals and the triplet state were observed and their lifetimes estimated.
ULTRAVIOLET‐INDUCED DNA DAMAGE AND ITS PHOTOREPAIR IN TAIL FIN CELLS OF THE MEDAKA, Oryzias lutipes
Authors: Funayama Tomoo , Mitani Hiroshi , Shima Akihiro
Published: January 2008

Abstract— To study UV‐induced damage and its photorepair in vivo, endonuclease‐sensitive sites (ESS) induced by UV irradiation in the DNA of the tail fin of the Medaka, Oryzias latipes. were examined. Isolated fins were irradiated with UV‐B, but no ESS were detected after increasing doses of UV‐B up to 500 J/m2. Then fins were irradiated with UV‐B and fin cells were dissociated by treatment with trypsin and EDTA. Ultraviolet‐B‐induced ESS were detected in the epithelial cells near the tissue surface but not in the inner cells. The capacity for photo repair of both inner and outer epithelial cells and cells in fin ray was the same and was higher than that for a line of cultured cells, OL32.
BIOLOGICALLY EFFECTIVE DOSE OF SOLAR ULTRAVIOLET RADIATION ESTIMATED BY SPORE DOSIMETRY IN TOKYO SINCE 1980
Authors: Munakata Nobuo
Published: January 2008

Abstract— The biologically effective dose of solar UV radiation has been measured in Tokyo since 1980 using Bacillus subtills spores. To determine the cumulative dose in a half day, several samples of UV‐sensitive spores were exposed in successive intervals from the solar‐noon time. Because fluence‐survival curves were exponential, the number of lethal hits received by the spores was calculated for each interval and termed inactivation dose (ID). The total number of hits obtained in a half day (half‐day ID) was correlated with the amount of global insolation by a power‐function regression. The regression analyses were performed for the data collected on 35 days from 1980 to 1986 and for the data collected on 53 days from 1989 to 1991. The latter data set yielded significantly larger estimates of half‐day ID relative to the insolation than the former. These analyses suggested that the biologically effective dose relative to the insolation increased about 30% at some time in the later part of 1980s at this location. Changes of solar activity, air pollution and stratospheric ozone layer were considered as potentially responsible for this increase, but identification of the causative factors requires further efforts.
RELATIVE GROUND AND EXCITED STATE ENERGIES OF CH3(CH=CH)5CH=NC4H9, ITS HYDROGEN‐BONDED AND PROTON‐TRANSFERRED SPECIES, AND CHARGE PARTITIONING AND DISTRIBUTION IN THE PROTONATED SCHIFF BASE OF RETINAL*
Authors: Blatz Paul E. , Tompkins Jay A.
Published: January 2008

Abstract— CH3(CH=CH)5CH=NC4H9 (compound 1) is structurally related to the Schiff base of retinal, the prosthetic group in visual pigments. Dilute solutions of a weak acid (phenol) and 1 in a hydrocarbon solvent, when subjected to decreasing temperature, show striking changes in electronic absorption spectra. Initially only the spectrum of compound 1 is present, but as the temperature is lowered, the absorbance of 1 decreases, and the spectrum of the H‐bonded form of 1 appears and increases. Continued temperature lowering then causes a decrease in absorption of the H‐bonded form and an appearance and rise in absorption of the proton‐transfcrred form of 1. Concentrations of the various species are measured as a function of temperature, and by standard procedures, the thermodynamic constants for both reaction steps are computed. Values of Δ0 are taken as relative energies among the three ground states, and the λmax value of each species yields relative energies among excited states. By employing data from electronic absorption spectroscopy, nuclear magnetic resonance (NMR) and theoretical calculations for retinal Schiff base, charge partitioning between nitrogen and the polyene chain and charge distribution among the carbon atoms of the polyene chain are calculated.
EFFECT OF MELITTIN ON PHOTOCYCLE AND PHOTOPOTENTIAL OF PURPLE MEMBRANE: SITES OF INTERACTION BETWEEN BACTERIORHODOPSIN AND MELITTIN
Authors: Shi Hua , Hu Kun‐Sheng , Huang Ying , Dong Na‐Xin
Published: January 2008

Abstract— Purple membrane (PM) suspension and artificial bilayer lipid membranes (BLM) containing PM sheets were treated with melittin. Both the decaying of the photocycle intermediate M412 and proton translocation were inhibited by melittin: The yields and rate of the slow‐decaying component of M412 (M412s) together with the proton release and its uptake rate were significantly decreased, but the rate of the fast‐decaying component of M412 (M4120 had only slight changes. Relatively high concentrations of melittin could cause aggregation in PM suspensions. Addition of melittin to a BLM solution increased the continuous photopotential signal but decreased the transient signal. We suggest that there might exist strong interactions between melittin and bacteriorhodopsin in addition to the melittin–lipid action. On the other hand, the results also indicate that proton translocation was more likely to be coupled with M412s and both were more sensitive to the changes caused by the melittin–PM interaction than was M412f.
USE OF A PULSED LASER DIODE TO MEASURE PICOSECOND FLUORESCENCE LIFETIMES
Authors: Bruice Doljg , Miners Jennifer
Published: January 2008

Abstract— The use of an inexpensive pulsed laser diode (Hamamatsu picosecond light pulser PLP‐01) as the excitation source for a single photon timing spectrolluorimeter with microchannel plate photomultiplier detection was dem‐onstrated. The performance of the instrument was tested with two very short‐lived fluorescent dyes and two pho‐tosynthetic systems with wcll‐defined decay characteristics. Individual fluorescence decays were analyzed by modeling with a convolution of the instrument response function to a sum of exponential decay components. Accurate fluorcscence lifetimcs of the dyes cryptocyanine (55 ps in acetone and 83 ps in ethanol) and 1,1‘‐diethyl‐2,2′‐dicarbocyanine iodide (13 ps in acetone and 26 ps in ethanol) were obtained by analysis of the decay kinetics with a single exponential component. Fits to the fluorescence decay kinetics of isolated photosystem I particles and intact cyanobacterial cells required three and four decay components. respectively. The decay kinetics of the isolated photosystem I preparation were dominated (99%) by a very fast 9 ps lifetime, reflecting the preparation's small antenna size of approximately 30 chlorophyll a. The cyanobackria showed decay components of 35 ps, 160 ps, 400 ps and 1.95 ns similar to those described previously by Mullincaux and Holzwarth (Rinchim. Biophys. Acfa1098, 68–78, 1991). The performance of the pulsed laser diode as an excitation source for single photon timing is discussed in comparison with conventional sources of picosecond light pulses.
STRUCTURE‐SPECIFIC BINDING and PHOTOSENSITIZED CLEAVAGE OF BRANCHED DNA THREE‐WAY JUNCTION COMPLEXES BY CATIONIC PORPHYRINS
Authors: Nussbaum Jean M. , Newport Mary Ellen A. , Mackie Madelyn , Leontis Neocles B.
Published: January 2008

Abstract— The interactions of cationic porphyrins with DNA oligonucleotides that form branched, three‐way junction complexes (TWJ) were investigated using native gel electrophoresis, absorption spectroscopy and photochemical probing using DNA sequencing techniques. Meso‐tetra(pa ra‐N‐trimethylaniliniumyrjporphine (TMAP), meso‐tetra(4‐JV‐methylpyridiniumyl)porphine (T4MPyP) and meso‐tetra(4‐N‐methylpyridiniumytyporphine (T3MPyP) were found to bind more tightly to DNA TWJ than to DNA duplexes. The binding to the junction DNA persists at high ionic strength, conditions that greatly decrease porphyrin binding affinity to duplex DNA. The TWJ DNA binding sites of TMAP and T4MPyP were localized to the junction region based on the observation of site‐ and structure‐specific, porphyrin‐sensitized photodamage to guanosine residues flanking the junction region.
OXIDATION OF INDOLE‐3‐ACETIC ACID BY SONICATED NEUTROPHILS. PITFALLS IN CORRELATING EPR SIGNALS AND CHEMILUMINESCENCE
Authors: Escobar Jorge A. , Vásquez‐Vivar Jeannette , Cilento Giuseppe
Published: January 2008

Abstract –The oxidation‐peroxidation of indole‐3‐acetic acid by sonicated neutrophils, unlike the horseradish peroxidase‐catalyzed process, features not only the skatole radical, but also a tertiary carbon‐centered radical in the electron paramagnetic resonance (EPR) spectrum. Moreover, emission occurs only in the red.

From the present and earlier data it is concluded that the observation of the skatole radical does not necessarily indicate its involvement in the 1ΔgO2 generating process. Also unlikely is a correlation between the tertiary carbon‐centered radical and red emission.

Our results indicate that in general great care must be taken in attempting to correlate EPR signals with chemi‐luminescence and with oxygen consumption.
In vitro PHOTODEGRADATION OF CHLORPROMAZINE
Authors: Broeke L. T. van den , Ouijja E. H. , Bojarski J. , Henegouwen G. M. J. Beyersbergen VAN
Published: January 2008

The in vitro photodecomposition of chlorpromazine (CPZ) was investigated with the aim to evaluate possible reactive determinants that could play a role in the occurrence of the in vivo‐observed photosensitivity. In view of the in vivo situation, CPZ was dissolved in low concentration in buffered aqueous solution (pH 7.4) or in dilute human serum and irradiated with low intensity (5–7 W m‐2) UV‐A and UV‐B. No distinct difference was found between UV‐A or UV‐B irradiation as far as photoproduct formation is concerned. This suggests the same degradation mechanism at both wavelength ranges. In buffered aqueous solution, irradiation of CPZ resulted in 65 and 90% 2‐hydroxypromazine (PZOH), 5 and 7% promazine (PZH) and 2 and 0% chlorpromazinesulfoxide (CPZSO) under aerobic and anaerobic conditions, respectively. In dilute human serum, there was only a shift in the PZH/PZOH ratio, probably as a result of H‐atom or electron donation by sulfur containing groups present in proteins. The results demonstrate that photodegradation of CPZ in vitro, under conditions relevant to the in vivo situation, proceeds almost entirely by dechlorination rather than by radical cation formation (the essential pathway of CPZSO production). Thus we conclude that the thiyl radical cation probably does not play a major role in the in vivo‐observed phototoxic reactions.
PHOTOPHYSICAL AND PHOTOCHEMICAL PROPERTIES OF TRITON X‐165 IN AQUEOUS AND METHANOLIC SOLUTIONS
Authors: Ghosh H. N. , Palit D. K. , Sapre A. V. , Ramarao K. V. S. , Mittal J. P.
Published: January 2008

Abstract Laser flash photolysis of the nonionic surfactant Triton X‐165 was carried out at 248 nm in methanolic and aqueous solutions at different pHs. Cation radical, triplet, phenoxy radical and solvated electron were observed as the transient absorbing species. Various characteristics of these transient species and their decay constants are reported. Photoionization was found to be a major process and it was monophotonic. The results obtained are compared with the photolysis of a small molecule p‐methoxytoluene (PMT). Fluorescence and phosphorescence properties of the surfactants Triton X‐100, Triton X‐165 and PMT are also studied. A suitable reaction scheme is proposed to account for the observed results.
DETECTION OF PYRIMIDINE DIMERS AND MONOADDUCTSINDUCED BY 7‐METHYLPYRIDO(3,4‐c) PSORALEN AND UVA IN CHINESE HAMSTER V79 CELLS BY ENZYMATIC CLEAVAGE AND HIGH‐PRESSURE LIQUID CHROMATOGRAPHY
Authors: Dardalhon M. , Guillo L. A. , Moysan A. , Vigny P. , Averbeck D.
Published: January 2008

Abstract The photochemotherapeutically active psoralen derivative 7‐methylpyrido(3,4‐c) psoralen (MePyPs) has been recently shown to be able to photoinduce monoadducts of the C4‐cycloaddition type as well as pyrimidine dimers in DNA in vitro. In the present study, we report on the induction of these two types of photolesions in mammalian cells in culture. The MePyPs photocycloadducts were quantified in V79 Chinese hamster cells after treatment with MePyPs plus UVA following enzymatic hydrolysis of the DNA by DNase I, S1 nuclease and acidic phosphatase treatments. Concomitantly induced pyrimidine dimers were determined by two methods, high‐pressure liquid chromatography and alkaline gel electrophoresis after dimer‐specific endonucleolytic cleavage. The results show that, in Chinese hamster cells treated with MePyPs plus UVA, the yield of pyrimidine dimers is approximately 5‐10% that of MePyPs‐DNA photocycloadducts. Because psoralen monoadditions to DNA alone are generally not considered as being very phototoxic, a synergistic interaction of monoadditions with pyrimidine dimers may be expected to occur in order to explain the high photobiological effectiveness of this psoralen derivative.
BIODISTRIBUTION AND PDT EFFICACY OF A KETOCHLORIN PHOTOSENSITIZER AS A FUNCTION OF THE DELIVERY VEHICLE
Authors: Woodburn Kathryn , Chang C. K. , Lee Sangwan , Henderson Barbara , Kessel David
Published: January 2008

Abstract C8KC is a new ketochlorin photosensitizer that must be formulated with an emulsifier because of its poor water solubility. In this report, we compare properties of Cremophor EL (CRM) and Tween 80 as delivery vehicles for C8KC. Unlike Tween 80, CRM altered the physical properties of both human and mouse plasma lipoproteins, resulting in decreased electrophoretic mobility of the individual lipoproteins along with the formation of a l ipoprotein degradation product: a phospholipid fraction of low buoyant density. In human plasma, where there was sufficient low‐density lipoprotein (LDL) for a distinction to be made, CRM caused a shift in binding of a ketochlorin from albumin to LDL and the degraded lipoprotein fraction. In mice bearing the RIF tumor, the use of CRM for drug formulation was associated with longer plasma and tissue persistence of C8KC, and enhanced photodynamic therapy (PDT) efficacy. These results indicate the importance of both sensitizer and vehicle as determinants of PDT efficacy.
INTERACTIONS OF BOTH MELITTIN AND ITS SITE‐SPECIFIC MUTANTS WITH BACTERIORHODOPSIN OF Halobacterium halobium: SITES OF ELECI'ROSTATIC INTERACI'ION ON MELITI'IN
Authors: Jiang Qiu‐Xing , Hu Kun‐Sheng , Shi Hua
Published: January 2008

Abstract Melittin and its site‐specific mutants differentially delay the slow‐decaying component of the photocycle intermediate M412 of bacteriorhodopsin in the purple membrane and the acetylated purple membrane whose several lysine residues are modified. This effect is attributed to the interaction of the total positive charges of melittin or its mutants with the total negative charges of bacteriorhodopsin. The effects of melittin and its mutants on the Triton X‐100–solubilized bacteriorhodopsin monomers are somewhat complicated but are associated with their charges. These results show that there is electrostatic interaction between bacteriorhodopsin and melittin and that both N‐and C‐termini of melittin function as sites of the interaction, with Arg 22 and Arg 24 making a prominent contribution to the effective surface charge of melittin. Melittin, at certain concentrations, partially restores the decreased photoactivity of the bacteriorhodopsin monomers trapped in the Triton‐lipid‐protein mixed micelles, which suggests that melittin may compete with Triton X‐100 for the binding sites on the bacteriorhodopsin monomers. Other kinds of interactions between bacteriorhodopsin and melittin are also indicated. The possible states of melittin in membranes are discussed.
MEROCYANINE DYES: EFFECT OF STRUCTURAL MODIFICATIONS ON PHOTOPHYSICAL PROPERTIES AND BIOLOGICAL ACTIVITY
Authors: Redmond Robert W. , Srichai Monvadi B. , Bilitz James M. , Schlomer David D. , Krieg Marianne
Published: January 2008

AbstractMerocyanine derivatives were prepared by structural alterations at the barbituric acid or chalcogenazole moieties. The photophysical properties of the dyes were markedly influenced by the presence of selenium rather than sulfur as a substituent at position 2 of the barbiturate. In methanol, quantum yields of both triplet state (φτ) and singlet oxygen sensitization (φΔ) were increased by over an order of magnitude, with a concomitant decrease in fluorescence, when selenium was present in the molecule. Photoisomerization, one of the dominant deactivation pathways in the sulfur‐ or oxygen‐containing analogues, was completely absent in the selenium‐containing derivatives. Efficient triplet state formation was observed for selenium‐containing derivatives incorporated into L1210 cells by diffuse reflectance laser flash photolysis. Cytotoxicity studies, camed out using clonogenic assays on L1210 leukemia cells, showed a good correlation with φτand φΔ, measured in solution. Experimental evidence provided by this paper supports a triplet state‐, and probably singlet oxygen‐, mediated phototoxic mechanism. Photoisomerization or singlet state mechanisms can be discounted.