PROTECTION BY THE FLUORIDE ION AGAINST PHTHALOCYANINE-INDUCED PHOTODYNAMIC KILLING OF CHINESE HAMSTER CELLS

When a dilute F- solution was added to a culture of Chinese hamster cells that had been preincubated with an aluminium phthalocyanine sensitizer derived from AlPcCl, the photosensitivity of the cells was markedly reduced compared to control cells not treated with F-. Under the same treatment conditions, the reduction in [3H]thymidine incorporation into cellular DNA caused by light and this sensitizer and the production of DNA-protein crosslinks caused by light and this sensitizer were also inhibited by F-. In contrast, the killing of Chinese hamster cells, the reduction of thymidine incorporation by the cells, and the production of DNA-protein crosslinks in the cells caused by the combination of light and either Photofrin II or the silicon phthalocyanine HOSiPcOSi(CH3)2(CH2)3-N(CH3)2 were not inhibited by F-. We conclude that the aluminium phthalocyanine sensitizer used is largely or completely AlPc(OH)(H2O), that it is converted to a fluoro complex by F-, and that this compound probably is a less efficient generator of photochemical damage at a critical cellular target(s) than is AlPc(OH)(H2O). The inhibition of thymidine incorporation and DNA-protein crosslink formation indicates that the effects of F- can be expressed at intracellular sites. It is further concluded that the silicon phthalocyanine sensitizer and Photofrin II do not interact significantly with F-.     

PROTECTIVE ACTION OF CAROTENOID PIGMENTS AGAINST PHOTODYNAMIC DAMAGE TO LIPOSOMES*

Abstract— When liposomes (a model membrane system) are subjected to a dye-sensitized photo-oxidation, lysis, as measured by glucose leakage or a change in light scattering, results. Before lysis occurs, the membrane lipids undergo peroxidative damage, as determined by the appearance of malondialdehyde. Carotenoids incorporated into the liposomal membrane protect against both lipid peroxidation and liposomal lysis. Other ¹O₂ quenchers and free radical absorbers also protect liposomes from photodynamic damage.     

PHOTODYNAMIC INACTIVATION OF LYSOZYME BY EOSIN

Abstract— It has been demonstrated that singlet oxygen is the major oxidizing entity in the photo-dynamic inactivation of hen egg white lysozyme by eosin, using D₂O to enhance the solvent-induced decay lifetime, and azide ion as a specific scavenger. Two regimes of inactivation can be distinguished depending on whether the sensitizer is free or complexed to the enzyme. The kinetic analysis for free dye sensitization, based on photostationary measurements and inactivation quantum yields, indicates that at least 1 in 15 singlet oxygen interactions with lysozyme leads to loss of lytic activity. The direct attack of triplet eosin makes a lesser overall contribution in air-saturated solutions, where 1 in 4 reactions induces inactivation. Lysozyme binds 1 eosin molecule from pH 4 to 12, leading to almost total quenching of the tryptophyl residue fluorescence without inhibition of the enzymic activity. The inactivation quantum yields indicate that singlet oxygen generated from the bound dye is the inactivating agent, but the dominant attack takes place with the complexed fraction of lysozyme molecules. The tryptophyl residue loss is the same or smaller in changing from H₂O to D₂O despite the 5–10 times increase in quantum yield, indicating that singlet oxygen inactivates also by reacting with residues other than tryptophan. The photochemical and fluorescence results are consistent with the the identification of tryptophyl site 108 with the eosin binding site and a reaction target for singlet oxygen. In a re-examination of earlier work on eosin-sensitized photo-oxidation of I", it has been found that singlet oxygen is the oxidizing agent in aerobic solutions.     

THE PHOTOINACTIVATION OF THE RESPIRATORY CHAIN IN SARCINA LUTEA (MICROCOCCUS LUTEUS) AND PROTECTION BY ENDOGENOUS CAROTENOID

Abstract. Irradiation of isolated membranes of Sarcina lutea (Micrococcus luteus) with blue light rapidly inactivated the respiratory malate oxidase system under aerobic but not anaerobic conditions. This inactivation was much faster than that seen in whole cells suggesting that the intact organism possesses protective mechanisms capable of preventing or repairing light damage. Three photosensitive sites have been detected by comparing the effect of blue light on membranes from the carotenoid-containing wild-type and a carotenoidless mutant. The sites have been identified as the initial malate dehydrogenase enzyme, a flavoprotein assayed by phenazine methosulphate reduction, a sulphydryl group associated with the dehydrogenase complex but not involved in phenazine methosulphate reduction and the respiratory quinone, menaquinone. Menaquinone was found to be sensitive only in carotenoidless membranes and not in membranes from the pigmented wild-type. Studies of the variation of photosensitivity with wavelength suggest that the three sites are sensitized by different chromophores and that the quinone acts as its own photosensitizer.     

PHOTODYNAMIC SENSITIZERS FROM CHLOROPHYLL:PURPURIN–18 AND CHLORINp6

Abstract— Two easily prepared derivatives of chlorophyll,purpurin–18 and chlorin p ₆, are potent sensitizers of cell killing by low-intensity red light. The internal anhydride group inpurpurin–18 provides the potential of covalently linking in one step the chlorin to cell targeting agents such as antibodies.     

CAROTENOID CHROMOPHORE LENGTH AND PROTECTION AGAINST PHOTOSENSITIZATION

Abstract— Carotenoid pigments were extracted and purified from wild-type and mutants 7 and 93a of Sarcina lutea , and tested for their ability to quench ¹O₂. The wild-type pigment (P-438, 9 conjugated double bonds) is as active in quenching ¹O₂ as is β-carotene. On the other hand, the pigment P-422 (8 conjugated double bonds) from mutant 7 is 2 or 3 times less efficient, whereas phytofluene and phytoene from S. lutea are 100 and 1000 times less efficient, respectively, than is β-carotene at quenching ¹O₂. It was also found that the broad EPR signal, induced by light in benzene solutions of chlorophyll a and hydroquinone, and related to chlorophyll oxidation, is efficiently quenched by P-438 and to a much smaller extent also by Sarcina phytoene.     

MULTIPLE MODES OF PHOTODYNAMIC ACTION BY CERCOSPORIN

Abstract— Cercosporin, one of a number of 4,9-dihydroxyperylene-3,10-quinones synthesized by plant pathogenic fungi, abundantly produces singlet oxygen when illuminated in solution. Singlet oxygen production is substantially decreased and superoxide production is greatly enhanced in the presence of the reducing agents ergothioneine (2-thiol-L-histidine betaine) or urate. Both ergothioneine and urate enhance superoxide production at a rate approximately 25-fold that elicited by an equimolar amount of methionine, the agent that is traditionally used in such assays. Such 'switching' in production of different active oxygen species under different environmental conditions may be of significance in biological processes, in this case host cell killing by the plant pathogen.     

QUENCHING OF CHLOROPHYLL FLUORESCENCE BY NITROBENZENE

Abstract—Nitrobenzene quenching of chlorophyll fluorescence in ethanol has been investigated. Steady state relative quantum yields have been measured and fluorescence decay rates were determined using both nanosecond photon counting and picosecond pulses from a mode-locked Nd³⁺ glass laser.
The fluorescence decay is described by
1( t )= I ₀ exp (- t/τ−At^1/2 )
the form predicted for decay governed by the kinetics of the continuum model of diffusion controlled reactions. From the parameters of the fluorescence decay, the encounter distance is 5–7 A° the mutual diffusion coefficient is 0.62 × 10^--⁵ cm2s^-1± 12%.
Some of the fluorescence quenching is also attributed to static quenching by a nitrobenzene-chlorophyll, ground-state complex. The equilibrium constant for formation of this ground-state complex was determined to be 4.1 M ^-1. The combined dynamic and static quenching model allows calculation of quantum yields of fluorescence in good agreement with the experimentally determined quantum yields.     

CHLOROPHYLL a AND CAROTENOID AGGREGATES AND ENERGY MIGRATION IN MONOLAYERS AND THIN FILMS

Abstract— The spectra of absorption, fluorescence and excitation of monolayers and thin films containing chlorophyll a together with a carotenoid (cis-β-carotene, trans-β-carotene, fucoxanthin, or zeaxanthin), were measured at — 196°C. The concentration ratios used, (Chl)/(Car), were 6:1, 4:1, 3:1, 2:1, 1:1 and 1:3, and the area densities, 3·70, 2·55, 1·76, 0·71, 0·37 and 0·17 nm²/pigment molecule. In dilute monolayers, (3·70 nm²/molecule), with a constant concentration ratio (Chl)/(Car) = 3:1, evidence of three β-carotene forms, with absorption bands at 460, 500 and 520 nm (C₄₆₀, C₅₀₀ and C₅₂₀), and of a chlorophyll a form with an absorption band at 669–672 (Chl_669–672) was found. On increasing the density to 0·2–0·3 nm²/molecule, a conversion of C₄₆₀ and C₅₂₀ into C₅₀₀, was observed, and several more additional (probably more strongly aggregated) chlorophyll a forms appeared, with absorption bands at 672–733 nm. With excess carotene [(Chi)/(Car) = 1:3] the forms C₄₆₀, C₅₀₀, C₅₂₀ and Chl_669–672 were present even in the most dense films (0·2–0·3 nm²/molecule). The same was found with other carotenoids: if one of the pigments was in excess, aggregated forms of the other tended to disappear. In the transfer of energy from carotenoids to chlorophyll a, C₅₀₀ was found to be the main donor. In layers with a concentration ratio (Chl)/(Car) = 3:1, the efficiency of transfer was less than 10 per cent at the lowest density used (3·70 nm²/molecule); it increased to 50 per cent, as the density was increased to 0·20 nm²/molecule. When the relative concentration of the carotenoid was increased to (Chl)/(Car) = 1:1, the efficiency of energy transfer dropped to 25 per cent even at 0·20 nm²/molecule. It seems that the efficiency of energy transfer between carotene molecules (prior to its transfer to chlorophyll a) is low, and effective transfer occurs only between β-carotene and immediately adjacent chlorophyll a molecules.     

含肼甲酸钇热分解制超细氧化钇

以甲酰钇为前驱化合物并将肼添加到甲酸钇中，热分解此含肼甲酸钇可制得超细氧化钇，粒度<100 nm。     

PHOTODYNAMIC INACTIVATION OF YEAST CELLS SENSITIZED BY HEMATOPORPHYRIN

Abstract— Yeast cells are inactivated by treatment with hematoporphyrin and light. The inactivation is mainly mediated by singlet oxygen. The quantum yield of singlet oxygen increases with increasing pH, while the efficiency of cellular inactivation decreases with increasing pH. Cells in the stationary phase are much more resistant to the treatment than cells in exponential growth. Membrane damage seems to be the main determining step in the photoinactivation.     

PHOTODYNAMIC INACTIVATION OF YEAST SENSITIZED BY EOSIN Y

Abstract. Wild-type diploid yeast has been irradiated with visible light in the presence of eosin Y to investigate the photodynamic inactivation of this model eukaryote. Light, eosin Y and oxygen were all required for substantial inactivation, and no dark recovery was detected. Long periods of irradiation were required for greater than 90% inactivation, corresponding to a very small low-dose quantum yield. Neither binding nor uptake of the dye by yeast was detected. Corrections for the photooxidative bleaching of eosin Y during irradiation indicate that bleaching causes a significant reduction in the apparent rate of inactivation. The results suggest that eosin Y acts as an extracellular sensitizer where the likelihood of damage to the cell envelope is enhanced.     

CONTROL OF CHLOROPHYLL b BIOSYNTHESIS BY PHYTOCHROME

In the mustard seedling cotyledons, chlorophyll b appears from the very beginning in white light provided that a red light pulse pretreatment was given 12 h prior to the onset of white light. The red light pulses act through phytochrome. Without pretreatment no chlorophyll b is detectable at least during the first 60 min after the onset of white light (25°C). Biogenesis of chlorophyll b specifically depends on the action of phytochrome during the pre-steady state period as well as during the steady state period of chlorophyll accumulation. In light pulse experiments, it was found that formation of chlorophyll b takes place stoichiometrically at the cost of chlorophyll(ide) a.     

IMPROVED SURVIVAL FROM INTRACAVITARY PHOTODYNAMIC THERAPY OF RAT GLIOMA

The effectiveness of intratumoral photoradiation in photodynamic therapy (PDT) using a polyporphyrin photosensitizer was studied in the RT-2 rat glioma model. One week after intracerebral implantation of RT-2 cells, experimental rats received a single i.p. injection of 2 mg/kg of Photofrin. After administration of the photosensitizer (48 h), the tumors were partially resected and the exposed cavity was irradiated with 15 J of laser light at a wavelength of 630 nm. Further treatment with a large craniectomy significantly enhanced rat survival. Control rats which received no photosensitizer but were treated with surgery, alone or in combination with laser irradiation, succumbed from early tumor recurrence. Photodynamic therapy without decompressive surgery resulted in hemorrhagic infarction of residual tumor and adjacent brain with focal cerebral edema which resulted in cerebral herniation and early death. Our results indicate that photodynamic therapy is effective in treating residual brain tumor but at the expense of brain tissue surrounding the tumor. Unless relieved, intracranial pressure from photodynamic therapy-associated cerebral edema in this animal model resulted in shortened survival.     

由氯化钠制硝酸钠的转化法研究

我国硝石矿很少,硝酸钠主要靠合成法生产,即用纯碱(或烧碱)溶液吸收硝酸“尾气”而得:     

PHOTODYNAMIC DESTRUCTION OF LYSOSOMES MEDIATED BY NILE BLUE PHOTOSENSITIZERS

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.     

PHOTOPRODUCTION OF IONS FROM CHLOROPHYLL α IN SOLUTION STUDIED BY FLASH PHOTOLYSIS and PHOTOCONDUCTIVITY

Abstract— Ionic species were detected electrically on flash excitation of chlorophyll α following rapid production of the triplet state of the pigment. The action spectrum of photocurrent agreed well with the absorption bands of the pigment in the visible region; the threshold for the formation of ionic species in acetonitrile was 1.8 eV. Oxygen gas introduced into the solution completely prevented the appearance of the triplet and ionic species. The latter are suggested to result from electron-transfer between the triplet pigments. Decay processes of the ionic species varied markedly with changes in the solvent polarity. Direct photo-ionization of the pigment could be observed by UV excitation in some non-polar solvents.     

ELICITATION OF SPREADING DEPRESSION BY ROSE BENGAL PHOTODYNAMIC ACTION

Spreading depression refers to a slowly propagating depression of the ordinary electrical activity of the nervous tissue. It can be elicited by different types of physical or chemical non-specific stimuli. Various evidences suggest that transient alterations of cell membranes are involved. For this reason, and considering the action of free radicals on cell membranes, the elicitation of the reaction by dye photoactivation has been investigated. Isolated chick retina superfused in the dark with Ringer solution was able to regularly exhibit spreading depression when submitted to 1 microM rose bengal pulse of 5 min in duration, followed by 2.1 x 10(4) to 4.2 x 10(4) Jm-2 light pulse. The phenomenon was monitored either by visual inspection of the light-scattering milky wave that accompanies the reaction or by recording its characteristic slow voltage variation. The reaction was not triggered if the retina, superfused with the dye, was (a) maintained in the dark; (b) illuminated with red light (3.75 x 10(2) to 2.25 x 10(4) Jm-2), or (c) stimulated by white light but superfused with nitrogen-saturated solutions. It is concluded that, under the present conditions, the elicitation of spreading depression is contingent on the photoactivation of rose bengal in the presence of oxygen.     

PREVENTION BY PHYTOCHROME OF PHOTODELAY IN CHLOROPHYLL ACCUMULATION

A photodelay in chlorophyll- a and -b accumulation is observed in mustard ( Sinapis alba L.) cotyledons during the first 3 h after the onset of white light even at medium light fluxes (3500 lx). A pretreatment of two red light pulses or a 12 h far-red light pretreatment, both operating through phytochrome. prevent the photodelay completely. This is a specific phytochrome effect since'it can be separated from the effects of phytochrome on the rates during pre-steady state and steady state phases of chlorophyll accumulation in saturating white light. Thus, photostability of chlorophyll in nature is a photoresponse mediated through phytochrome.