Calcium as a modulator of photosensitized killing of H9c2 cardiac cells.

Illumination of H9c2 rat heart cells in the presence of Rose Bengal resulted in dose-dependent cell killing (assessed by trypan blue staining) and modification of ionic currents flowing through the heart cell membrane. Inhibitors of voltage-gated ionic currents were shown to have little effect on cell killing. Ionic current measurements were used to assess the increase in leak conductance of these cells, which has been suggested to be a causal factor in killing of other cell types (1). Inhibitors of voltage-gated ionic currents, including the sodium channel blocker tetrodotoxin (100 microM) and the calcium channel blocker lanthanum (10 microM) were shown to have little effect on cell killing. The potassium channel inhibitor tetraethylammonium (20 mM) inhibited cell killing, but the effect is viewed as being caused by an inhibition of leak current. The time course of block of voltage-activated ionic currents during illumination, in the presence of Rose Bengal, was rapid compared with that for induction of leak current and for cell killing. These observations are consistent with a role for leak current in photosensitized killing of cardiac cells. They are interpreted with respect to calcium influx through the leak current pathway as a trigger for the cellular response.     

Effect of dipole modifiers on the magnitude of the dipole potential of sterol-containing bilayers

The effects of various subclasses of flavonoids, Rose Bengal, and different styrylpyridinium dyes on the magnitude of the dipole potential of membranes composed of pure phospholipids and sterol-containing bilayers were investigated. Changes in the steady-state membrane conductance induced by cation-ionophore complexes were measured to examine the changes in the dipole potential of lipid bilayers. The characteristic parameters of the Langmuir adsorption isotherm for different flavonoids and Rose Bengal and the slope of the linear dependence of the dipole potential change on the aqueous concentrations of RH dyes were estimated. Chalcones (phloretin and phloridzin) and flavonols (quercetin and myricetin) strictly decrease the dipole potential of phospholipid- and sterol-containing membranes; the unsaturation of the C-ring and the hydrophobicity of the molecule contribute to the ability of the flavonoid to reduce the bilayer dipole potential. Rose Bengal decreases the magnitude of the bilayer dipole potential to a similar extent, but its affinity for membrane lipids is higher; the effects of RH dyes, chalcones, and phloroglucinol are determined by sterol concentration and type.     

Type I and type II sensitizers based on Rose Bengal onium salts

Abstract— New Rose Bengal oniurn salts containing one or two iodonium, sulfonium, phosphonium, and pyrylium ions have been prepared as part of a program to develop sensitizers which can function as Type I radical photoinitiators and Type II energy transfer donors depending on experimental conditions. The absorption spectra of the onium salts in different solvents indicate an equilibrium between tight and loose ion pairs which depends on the solvent polarity, the cation and concentration. Typical Rose Bengal photochemistry requires the structure be that of the loose ion pair in the solvent of choice. Similar factors also influence bleaching behavior, and bleaching is the result of electron transfer processes. The quantum yields of singlet oxygen production from the onium salts in polar solvents are similar to that of the parent, Rose Bengal disodium salt.     

Inhibition of the mitochondrial F1-ATPase by rose bengal mediated photooxidation. Interaction of the Fe2+ chelate of bathophenanthroline with the sensitizer

Rose Bengal mediated photooxidation of mitochondrial F1-ATPase and its beta-subunit resulted in inactivation and loss of about 50 and 60% of their histidine residues, respectively. The beta-subunit was not cleaved upon photooxidation. Photooxidation of histidine probably results in changes in the conformational stability of F1-ATPase leading to its inactivation. The participation of singlet molecular oxygen during the photooxidation process is suggested by the selective loss of histidine residues, while other amino acids, also sensitive to singlet oxygen attack, were not affected. Photochemical damage of F1-ATPase was prevented by various phenanthroline compounds, the order of efficiency being bathophenanthroline-Fe chelate greater than bathophenanthroline greater than orthophenanthroline-Fe chelate greater than bathophenanthroline-sulfonate-Fe chelate. The prevention by bathophenanthroline-Fe chelate of photochemical damage is interpreted on the basis of its interaction with the photosensitizer, Rose Bengal, probably implying a chemical reaction which decreases the actual concentration of the sensitizer and, thereby, the extent of photoinactivation.     

Photodynamic action of Rose Bengal silica nanoparticle complex on breast and oral cancer cell lines

Rose Bengal, an anionic photosensitizer was conjugated to organically modified silica nanoparticles having 3-amino propyl groups by electrostatic or covalent interaction. The drug-nanoparticle complexes were characterized by FTIR, light scattering and zeta potential measurements. Significant changes were observed in the spectroscopic properties of the drug when it is conjugated with nanoparticles. The toxicity of the free drug and drug-nanoparticle complex was studied against oral (4451) and breast (MCF-7) cancer cell lines. Both complexes with nanoparticles were more phototoxic than free Rose Bengal, with the covalent complex being the more effective. Studies carried out on cellular uptake, photostability and singlet oxygen generation suggest that enhanced phototoxicity is primarily due to the enhanced uptake of the drug-nanoparticle complex.     

Metal-free visible-light-promoted intermolecular [2+2]-cycloaddition of 3-ylideneoxindoles

The Rose Bengal sensitized intermolecular [2 + 2]-cycloaddition of 3-ylideneoxindoles for the synthesis of spirocyclic oxindoles was developed successfully under visible light irradiation conditions. The cycloaddition products were obtained in good yields (up to 93%) with excellent diastereoselectivity and regioselectivity by using a low loading of Rose Bengal (0.125 mol%) as a triplet sensitizer. This work demonstrates the potential benefits of Rose Bengal in visible light catalysis.     

Microbubble-sonosensitiser conjugates as therapeutics in sonodynamic therapy

A Rose Bengal sonosensitiser has been covalently attached to a lipid microbubble and the resulting conjugate shown to produce higher levels of singlet oxygen, enhanced cytotoxicity in a cancer cell line and a greater reduction in tumour growth than the sonosensitiser alone.     

Site-directed photochemical disruption of the actin cytoskeleton by actin-binding Rose Bengal-conjugates

The in situ light-induced, non-enzymatic digestion of cytoskeletal actin by a xanthene dye conjugated to heavy meromyosin, anti-actin antibodies and/or anti-myosin antibodies is reported. The dye Rose Bengal was conjugated to either anti-actin antibodies, anti-myosin antibodies or heavy meromyosin. Under our experimental conditions, visible light induced the non-enzymatic breakdown of cytoskeletal actin when mammalian tissue culture cells were probed either with Rose Bengal-conjugated anti-actin and/or anti-myosin antibodies. Similar results were obtained when tissue culture cells were probed with Rose Bengal-conjugated heavy meromyosin before irradiation with visible light. The in situ photochemical reaction depended on the presence of actin-binding Rose Bengal-conjugates.     

An extraction method for the determination of the Rose Bengal content in the preparations of Rose Bengal labelled with131I

The paper describes an extraction radiometric method for the determination of Rose Bengal content in the radiopharmaceutical preparations of Rose Bengal labelled with¹³¹I. Two methods have been studied. The first one is based on the addition of an excess and substoichiometric amounts of Septonex /carbetoxypentadecyltrimethylammonium bromide/ to the sample of Rose Bengal-¹³¹I and on the activity measuring of the chloroform extracts. The second method is based on the extraction of Rose Bengal-¹³¹I with substoichiometric amount of Septonex from the sample of Rose Bengal-¹³¹I and the sample of Rose Bengal-¹³¹I diluted with a known amount of an inactive Rose Bengal preparation.     

PHOTOSENSITIZATION OF MELANIN: AN ELECTRON SPIN RESONANCE STUDY OF SENSITIZED RADICAL PRODUCTION AND OXYGEN CONSUMPTION

Abstract Rose Bengal is shown to photosensitize free-radical production and oxygen consumption in solutions of melanin from autooxidation of 3,4-dihydroxyphenylalanine (DOPA). In anaerobic solutions the sensitizer enhances rates of free-radical production by up to a factor of 20. In aerobic solutions, rates of oxygen consumption can be increased by a factor of several hundred. The reactions appear to involve the triplet state of the sensitizer. The effect of the sensitizer in increasing oxygen consumption is quenched by low concentrations of azide and enhanced by D₂O, suggesting that a singlet oxygen mechanism is involved.     

MEMBRANE POTENTIAL CAN INFLUENCE THE RATE OF MEMBRANE PHOTOMODIFICATION

Abstract— Though cellular photomodification has been shown to change cellular resting membrane potential, an effect of membrane potential on the rate of photomodification has never been reported. Here we demonstrate that the rate of photomodification of potassium channels in frog atrial cells is voltage dependent. The rate of potassium channel photomodification using negatively charged Rose Bengal as the photosensitizer is about 2.5 times greater at the resting membrane potential of -70 m V compared to +40 mV. Similar results are obtained using the positively charged photosensitizer methylene blue. On the other hand, the rate of photomodified increase of leak current in the same cells does not significantly change in this voltage range with Rose Bengal as photosensitizer, but demonstrates a voltage dependence like that of potassium current when methylene blue is the photosensitizer. These observations cannot be explained based on voltage-dependent partitioning of the sensitizer, as similar effects on potassium current were obtained using either a positively charged or negatively charged sensitizer.     

LYSIS OF EGG PHOSPHATIDYLCHOLINE LIPOSOMES BY SINGLET OXYGEN GENERATED IN THE GAS PHASE

Abstract Singlet molecular oxygen was generated in the gas phase by heterogeneous energy transfer from a film of dry Rose Bengal and bubbled into suspensions of egg phosphatidylcholine liposomes, leading to membrane lysis and lipid peroxidation. The analysis based on large target diffusion kinetics indicates that approximately 3000 interactions were required for lysis of 1.5 μm liposomes. This experimental method makes it possible to measure the reactivity of gas phase singlet oxygen with aqueous biological systems.     

Structure-based prediction of the conductance properties of ion channels

The HOLE procedure allows the prediction of the absolute conductance of an ion channel model from its structure. The original prediction method uses an empirically corrected Ohmic method. It is most successful, with predictions being reliable to within a factor of two. A new modification of the procedure is presented in which the self-diffusion coefficients of water molecules from molecular dynamics simulation are used to replace the empirical correction factor. A "prediction" of the conductance for the porin OmpF by the new method is made and shown to be very close to the experimental value. HOLE also allows the prediction of the effect that the addition of non-electrolyte polymers will have on channel conductance. The method has great potential to yield structural information from data provided by single channel recordings but needs further validation by making measurements on channels of known structure. Preliminary results are given of single channel records establishing the effects of non-electrolytes on the conductance of gramicidin D channels. As an example of the potential uses of the procedure application is made to examine the oligomerization of alpha-toxin (alpha-hemolysin) channels. A model for the alpha-toxin hexamer, based on the crystal structure for the heptamer, is generated using molecular mechanics methods. The compatibility of the structures with single channel conductance data is assessed using HOLE.     

THE PHOTODYNAMIC EFFECT OF ROSE BENGAL ON PROTEINS OF THE MITOCHONDRIAL INNER MEMBRANE

Photodynamic action promoted by Rose Bengal was evaluated in solutions of unsaturated fatty acids or histidine, and on beef heart submitochondrial particles. Rose Bengal-promoted photooxidation of histidine was mainly due to the opening up of the imidazole ring by singlet oxygen. Photosensitization of polyunsaturated fatty acids (PUFA) resulted in oxygen consumption and thiobarbituric acid-reactive substances (TBARS) formation, the extent of which was linearly related to the increasing degree of unsaturation. Photosensitization of submitochondrial particles caused oxygen consumption and TBARS production. These processes involved two different reaction components: during the first, most of the mitochondrial proteins were inactivated, the most sensitive being succinate dehydrogenase and cytochrome c. The values for the rate ratios of [TBARS] formation/[O2] consumption for the first and second phase were 0.36 and 1.32%, respectively, pointing to a larger contribution of lipid peroxidation during the second phase. The calculation of the rate constants for reaction of singlet oxygen with mitochondrial proteins suggests that singlet oxygen is more reactive towards proteins than to PUFA. The biological role of this selectivity is discussed in terms of the mitochondria as one of the first targets for photosensitized reactions.     

Sensitization of aluminum chloride adsorbed tin(IV) oxide nanocrystalline films with Rose Bengal

The anionic dye Rose Bengal was found to surface chelate more strongly to SnO2 nanocrystalline films previously kept immersed in a solution of washed and dried AlCl3. Dye-sensitized photoelectrochemical cells made from such films exhibit enhanced quantum and energy conversion efficiencies. The result is explained as caused by binding of AlCl3 to SnO2 surface by elimination of Cl atoms and stronger bonding of Rose Bengal to Al, enhancing dye adsorption and suppression of back electron transfer by bridging of an Al atom between Sn and the dye molecule.     

Metal-enhanced phosphorescence: interpretation in terms of triplet-coupled radiating plasmons

We report our detailed metal-enhanced phosphorescence (MEP) findings using Rose Bengal at low temperature. Silver Island Films (SiFs) in close proximity to Rose Bengal significantly enhance the phosphorescence emission intensity. In this regard, a 5-fold brighter phosphorescence intensity of Rose Bengal was observed from SiFs as compared to a glass control sample at 77 K. In addition, several factors affecting MEP, such as distance dependence and silver film morphology, were also investigated. Our findings suggest that both singlet and triplet states can couple to surface plasmons and enhance both fluorescence and phosphorescence yields. This finding suggests that MEP can be used to promote triplet-based assays, such as those used in photodynamic therapy.     

PHOTOSENSITIZATION OF MELANINS: A COMPARATIVE STUDY

Abstract— Measurements of photosensitized free-radical production and oxygen consumption have been made in several melanin systems. The melanins studied were cysteinyldopa melanin (a model for pheomelanin), natural pheomelanins extracted from red hair and from red chicken feathers, and eumelanin extracted from bovine eyes. Rose Bengal was used as photosensitizer. A significant enhancement in the rates of free radical production and oxygen consumption was found in all systems when sensitizer was present. The largest effects were found with cysteinyldopa melanin and effects on pheomelanins were shown to be greater (by factors of4–6) than on eumelanins.     

Spectroscopic and electrochemical study of Rose Bengal in aqueous solutions of cyclodextrins

The interaction of Rose Bengal (RB) in aqueous solution of LiClO4 0.1 M with alpha-cyclodextrin (alpha-CD), hydroxypropyl-beta-cyclodextrins (HP-beta-CD) and hydroxypropyl-gamma-cyclodextrins (HP-gamma-CD) were studied by spectrophotometric measurements. The presence of Induced Circular Signals and the results of the analysis of the modifications in the absorbance spectra of RB produced by the presence of CDs in solution indicate that RB forms inclusion complexes only with HP-beta-CD and with HP-gamma-CD.     

Preparation of Porous Polymer Networks of <Emphasis Type="Italic">N</Emphasis>-Vinylpyrrolidone with Triethylene Glycol Dimethacrylate and Determination of Their Specific Surface Area Using Rose Bengal Dye

Porous polymer networks of N-vinylpyrrolidone with triethylene glycol dimethacrylate were prepared by three-dimensional radical copolymerization using template approach based on incorporation of macromolecular nanoobjects with porogen function. The adsorption of Rose Bengal dye from aqueous solutions onto the copolymers prepared and its desorption were studied by electronic absorption spectroscopy. The specific surface area of the porous polymer networks was estimated from the Rose Bengal adsorption data. The porous polymer networks adsorb riboflavin from aqueous solution. The riboflavin desorption is characterized by complex pattern due to washout from pores of different size.     

The extraction behaviour of the Rose Bengal labelled with131I in the presence of cationic surfactants

The Rose Bengal forms with cationic surfactants /1-carbethoxypentadecyltrimethyl-ammonium bromide, cetyltrimethylammonium bromide and cetylpyridinium bromide/ the extractable ion associates with composition 11 or 12. The extraction constants of the ion associates of the Rose Bengal with cationic surfactants were determined radiometrically with the aid of Rose Bengal labelled with¹³¹I.