Asahi Suzuki

Rate Constants for Quenching Singlet Oxygen and Activities for Inhibiting Lipid Peroxidation of Carotenoids and α-Tocopherol in Liposomes

The 1O2 quenching rate constants (kQ) of α-tocopherol (α-Toc) and carotenoids such as β-carotene, astaxanthin, canthaxanthin, and lycopene in liposomes were determined in light of the localization of their active sites in membranes and the micropolarity of the membrane regions, and compared with those in ethanol solution. The activities of α-Toc and carotenoids in inhibiting 1O2-dependent lipid peroxidation (reciprocal of the concentration required for 50% inhibition of lipid peroxidation: [IC50]−1) were also measured in liposomes and ethanol solution and compared with their kQ values. The kQ and [IC50]−1 values were also compared in two photosensitizing systems containing Rose bengal (RB) and pyrenedodecanoic acid (PDA), respectively, which generate 1O2 at different sites in membranes. The kQ values of α-Toc were 2.9×108M−1s−1 in ethanol solution and 1.4×107 M−1s−1 (RB system) or 2.5×106 M−1s−1 (PDA system) in liposomes. The relative [IC50]−1 value of α-Toc in liposomes was also five times higher in the RB system than in the PDA-system. In consideration of the local concentration of the OH-group of α-Toc in membranes, the kQ value of α-Toc in liposomes was recalculated as 3.3×106 M−1s−1 in both the RB and PDA systems. The kQ values of all the carotenoids tested in two photosensitizing systems were almost the same. The kQ value of α-Toc in liposomes was 88 times less than in ethanol solution, but those of carotenoids in liposomes were 600–1200 times less than those in ethanol solution. The [IC50]−1 value of α-Toc in liposomes was 19 times less than that in ethanol solution, whereas those of carotenoids in liposomes were 60–170 times less those in ethanol solution. There were no great differences (less than twice) in the kq and [IC50]−1 values of any carotenoids. The kQ values of all carotenoids were 40–80 times higher than that of α-Toc in ethanol solution but only six times higher that of α-Toc in liposomes. The [IC50]−1 values of carotenoid were also higher than that of α-Toc in ethanol solution than in liposomes, and these correlated well with the kQ values.

Kinetics and Dynamics of Singlet Oxygen Scavenging by α-Tocopherol in Phospholipid Model Membranes

Scavenging of singlet oxygen (1O2) by alpha-tocopherol (alpha-Toc) was investigated in liposomes. 1O2 was generated by photoirradiation in the presence of two photosensitizers, water-soluble methylene blue (MB) and lipid-soluble 12-(1-pyrene)dodecanoic acid (PDA). The rates of oxidation of alpha-Toc differed depending on the photosensitizing dye and the membrane charge: in the MB-system, alpha-Toc was oxidized fast in negatively charged dimyristoylphosphatidylcholine (DMPC) liposomes containing dicetylphosphate (DCP) and slowly in neutrally charged DMPC liposomes and positively charged DMPC liposomes containing stearylamine (SA), but in the PDA-system, the oxidation rate was independent of the membrane charge. The charge-dependent difference in the MB-system would be due to the site of 1O2 generation depending on the charge-dependent distribution of MB, because positively charged MB increased the zeta-potential of DCP-DMPC liposomes by its interaction with DCP at the membrane surface, but changed the zeta-potentials of DMPC and SA-DMPC liposomes less because of its location in the bulk water phase. The oxidation rate of alpha-Toc in liposomes was different from that in EtOH solution: in the MB system, the oxidation rate was faster in EtOH solution than in DMPC or SA-DMPC liposomes but the same as that in DCP-DMPC liposomes. However, in the PDA system, the oxidation rate was slower in EtOH solution than in DMPC liposomes with or without a charge. Membrane fluidity changed the rate of alpha-Toc oxidation in liposomes, the rate being higher in the liquid crystalline phase than the gel phase, as judged by the higher rate in DMPC liposomes than in dipalmitoylphosphatidylcholine (DPPC) liposomes at 30 degrees C. The rate constants of alpha-Toc for scavenging, the chemical reaction and physical quenching of 1O2 were determined in membranes using DCP-DMPC liposomes labeled with 1,3-diphenyl-isobenzofuran (DPBF), which traps 1O2. These constants differed in the two photosensitizing systems, being higher in the MB-system than in the PDA-system, and were lower than those in EtOH solution.

Effects of α-tocopherol analogs on lysosome membranes and fatty acid monolayers

Abstract The surface pressures of α-tocopherol analogs, fatty acids, and their mixtures were measured in their spread monolayers at an air—water interface. The surface pressure—area isotherms for the mixed monolayers of α-tocopherol and either stearic acid, oleic acid or linoleic acid deviated positively from those calculated on the basis of the additivity rule, and the magnitude depended on the length of the phytyl side chain in α-tocopherol and on the degree of unsaturation of the fatty acid chains. Lysosome membranes of mouse liver were stabilized by addition of α-tocopherol. A decrease in the length of the phytyl side chain in α-tocopherol reduced its ability to stabilize lysosome membranes. A good correlation was obtained between the extent of stabilizing activity of α-tocopherol analogs on lysosome membranes and the degree of positive deviation of the surface pressure for their mixtures with fatty acids.

Effects of pressure on the phase transition of bilayers in liposomes influence of cholesterol and α-tocopherol

The effects of pressure on the gel-to-liquid crystalline phase transition temperature of dimyristoylphosphatidylcholine (DMPC) bilayers containing cholesterol, alpha-tocopherol, and alpha-tocopheryl acetate were studied by fluorescence depolarization. The transition temperature of cholesterol mixtures (greater than 7.5 mol%) was lower than that of 100% DMPC at atmospheric pressure, but it became higher than the latter on increase in pressure. The thermodynamic parameters of the transition (delta V, delta S, delta H) were estimated and the functions of cholesterol and alpha-tocopherols in the bilayers are discussed.

Effects of pressure on the mobility of the hydrocarbon chains in liposomes

Abstract The mobility of the hydrocarbon chains of dimyristoylphosphatidylcholine (DMPC) liposomes containing cholesterol and α-tocopheryl acetate (α-T acetate) was studied by the fluorescence depolarization method at high pressures up to 980 bars (1 bar=10 5 Pa). Plots of fluorescence polarization vs. pressure showed maxima in both the gel and liquid-crystalline state for cholesterol and α-T acetate mixtures. However, in the pure DMPC systems, the liquid-cystalline state did not have a maximum, although the gel state had. The pressures at which the maxima were observed in the gel and liquid-crystalline states shifted in different directions with increase in concentration of added substances. The results are discussed on the basis of the functions of cholesterol and α-T acetate in bilayer lipid membranes.

Effects of pressure and potassium chloride on the aggregation of poly(γ-benzyl-L-glutamate) in liposomal bilayers

Fluorescence depolarization studies were made on dimyristoylphosphatidylcholine liposomes containing four kinds of dansylated poly(gamma-benzyl-L-glutamate) with different degrees of polymerization or hydrocarbon chain lengths under high pressure at up to 981 bar (1 bar = 10 MPa). Potassium chloride promoted the aggregation of the synthetic peptides in liposomal bilayers at both atmospheric and high pressure. The chain lengths of the hydrocarbons of the peptides had more influence than their degrees of polymerization on aggregation.

Surface films of protein and a protein-lipopolysaccharide complex extracted from Pseudomonas aeruginosa

Protein and a protein-lipopolysaccharide complex were extracted from the cell envelopes of Pseudomonas aeruginosa as fractions of endotoxin and spread on aqueous subphases as monomolecular layers. The surface properties of these materials, i.e. surface pressure, surface viscosity and surface potential, were measured as a function of the surface area. The protein molecule was partially unfolded at the air water interface and had a limiting area of 0.54 m2/mg protein in the state of minimum compressibility. Lipopolysaccharide alone did not form a surface film on water. As a complex with protein, lipopolysaccharide did not dissolve completely in the aqueous phase but was attached to the protein film. This protein aggregates or dissociates in the bulk phase as a function of the pH of the solution so the effect of pH on its surface properties was examined. The effects of NaCl and KCl in the subphase on the protein film differed from their effects on a film of the complex. The film of the complex is mainly formed from protein and this protein film is stiffened and stabilized by lipopolysaccharide attached to the lower surface of the film.

Semiconductive Coordination Polymers: Dithiooxamides Copper Compounds

A number of coordination polymers have been synthesized in the last decade. Some of them are semiconductors1. One of the authors has studied the semiconductive properties of 1,6-dihydroxyphenazinato-Cu(II), 2,5-dihydroxy-p-benzoquinonato-Cu(II) and rubeanato-Cu(II)2,3. In this short note, we would like to briefly review the last one and its derivatives, to discuss on some problems in correlating the properties with the molecular structures, and to introduce the attempts to solve the problems.

Studies of fluorescence depolarization at high pressures of dimyristoylphosphatidylcholine liposomes containing poly(γ-benzyl-l-glutamate)

The behaviors of four kinds of poly(gamma-benzyl-L-glutamate) (PBLG) molecules and their locations in dimyristoyl-phophatidylcholine (DMPC) liposomes were studied by fluorescence techniques at high pressures of up to 981 bar. The fluorescent substances 1,6-diphenyl-1,3,5-hexatriene (DPH), 1-aminonaphthalene-8-sulfonate (ANS) and PBLGs labeled with a dansyl group were used as probes of hydrophobic and hydrophilic spheres and the motions of polypeptides, respectively. The changes in mobilities of the PBLG molecules depended on their concentrations, degrees of polymerization and the lengths of hydrocarbon chains attached to their terminals. The molecular motions of the PBLGs were altered by increase in their contents, caused by squeezing the PBLG molecules out of the membranes. Models of the membranes and the behavior of polypeptides in the membranes at high pressures are discussed.