RESUMO
Internal stimuli-responsive controlled release from liposomal vesicles is an innovative approach for site-specific delivery of therapeutic drugs. In this study, to enhance the endosomal pH control of drug release from liposomes, a series of histidine-modified pH-sensitive Cn-His (n = 8, 12, 18) agents were designed and utilized as triggers for liposomal content release. The pH-dependent properties of Cn-His-incorporated liposomes were characterized using dynamic light scattering, ζ-potential, and fluorescence spectroscopy. The liposomes maintained a relatively uniform size across all pH conditions. However, the ζ-potential exhibited positive values at endosomal acidic pH levels and neutral or negative values at physiological pH levels. Furthermore, acidic pH-dependent release of both polar content (carboxyfluorescein) and nonpolar content (Nile red) was observed from the Cn-His-incorporated liposomes. Notably, with C12-His as the pH sensitizer, the pH dependence of liposomal content release was significantly evident. This work establishes endosomal pH-controllable liposome platforms, laying the groundwork for developing clinically applicable triggered release formulations.
Assuntos
Histidina , Lipossomos , Lipossomos/química , Concentração de Íons de Hidrogênio , Polímeros/química , Endossomos , Sistemas de Liberação de Medicamentos/métodosRESUMO
Removal of linear and branched alkylphenols with different alkyl chain lengths or different branchings (normal, secondary, and tertiary), some of which are suspected as endocrine disrupting chemicals, from an aqueous medium were investigated through quinone oxidation by polyphenol oxidase (PPO) and subsequent quinone adsorption on chitosan beads or powders at pH 7.0 and 40 °C. PPO-catalyzed quinone oxidation increased with an increase in alkyl chain length of the alkylphenols used. Although a higher PPO dose was required for quinone oxidation of branched alkylphenols, they were completely or mostly removed by quinone adsorption on chitosan beads or powders. The apparent activity of PPO increased by a decrease in quinone concentration. On the other hand, in the homogeneous systems with solutions of chitosan and PPO at pH 6.0, longer reaction times were required to generate insoluble aggregates, and a small amount of quinone derivatives were left in the solution even under optimum conditions. These results support that the two-step reaction, that is, PPO-catalyzed quinone oxidation and subsequent quinone adsorption on chitosan beads or powders, in the heterogeneous system is a good procedure for removing linear and branched alkylphenols from aqueous medium.
RESUMO
Scanning tunneling microscopy (STM) investigation was performed for the blends of isobutenyl compounds, in which the long alkyl chains were connected with ester or carbamoyl linkages. Each component by itself did not show the odd-even effect of alkyl chain length, whereas after blending them, the 2D structures drastically changed and modulated to exhibit odd-even effect. Star, lozenge, twist-like, and linear structures were found, dependent on the blend ratio and alkyl chain length. The blend ratio dependence of 2D structures was explained in terms of homogeneous and heterogeneous dimerization due to the interdigitation of alkyl chains.
RESUMO
In this study, enzymatic removal of bisphenol A (BPA) from the aqueous medium was investigated through the generation of water-insoluble oligomers, and this procedure was applied to removal of bisphenol derivatives. The experimental parameters, such as the temperature, pH value, enzyme concentration, and concentration and molecular weight of polyethylene glycol (PEG), were determined for the laccase-catalyzed treatment of BPA. The optimum conditions were determined to be pH 7.0 and 40°C in the absence of PEG. Water-insoluble oligomers generated under these conditions were readily removed by filtration or centrifugation. The optimum pH value was decreased to 5.0 in the presence of PEG and the laccase dose was reduced to one-fiftieth of that in the absence of PEG. This indicates that the addition of PEG protects the enzymatic activity and prevents capture of laccase molecules in the oligomers. The oligomers generated in the presence of PEG were removed from the aqueous medium by filtration with a membrane filter or by centrifugation. The oligomers were completely filtrated out with a filter paper by decreasing the pH value to 3.0. In addition, several bisphenol derivatives were also treated and subsequently removed by adjusting the laccase dose in the presence of PEG using the above procedure.
Assuntos
Compostos Benzidrílicos/química , Lacase/química , Fenóis/química , Polietilenoglicóis/química , Poluentes Químicos da Água/química , Purificação da Água/métodos , Compostos Benzidrílicos/análise , Concentração de Íons de Hidrogênio , Peso Molecular , Fenóis/análise , Temperatura , Poluentes Químicos da Água/análiseRESUMO
In this study, the combined use of a biopolymer chitosan and an oxidoreductase polyphenol oxidase (PPO) was systematically investigated for the removal of bisphenol derivatives from aqueous medium. The process parameters, such as the pH value, temperature, and PPO concentration, were estimated to conduct the enzymatic quinone oxidation of bisphenol derivatives by as little enzyme as possible. Bisphenol derivatives effectively underwent PPO-catalysed quinone oxidation without H2O2 unlike other oxidoreductases, such as peroxidase and tyrosinase, and the optimum conditions were determined to be pH 7.0 and 40°C for bisphenol B, bisphenol E, bisphenol O, and bisphenol Z; pH 7.0 and 30°C for bisphenol C and bisphenol F; and pH 8.0 and 40°C for bisphenol T. They were completely removed through adsorption of enzymatically generated quinone derivatives on chitosan beads or chitosan powders. Quinone adsorption on chitosan beads or chitosan powders in the heterogeneous system was found to be a more effective procedure than generation of aggregates in the homogeneous system with chitosan solution. The removal time was shortened by increasing the amount of chitosan beads or decreasing the size of the chitosan powders.
Assuntos
Compostos Benzidrílicos/isolamento & purificação , Catecol Oxidase/metabolismo , Quitosana/química , Fenóis/isolamento & purificação , Quinonas/metabolismo , Poluentes Químicos da Água/isolamento & purificação , Purificação da Água/métodos , Adsorção , Compostos Benzidrílicos/metabolismo , Oxirredução , Fenóis/metabolismo , Quinonas/isolamento & purificação , Poluentes Químicos da Água/metabolismoRESUMO
In this study, the combined use of an amino group-containing polymer chitosan and an oxidoreductase polyphenol oxidase (PPO) was applied to the removal of naphthols and dihydroxynaphthalenes (DHNs) from aqueous solutions. The process parameters, such as the pH value, temperature and enzyme dose, were discussed for PPO-catalysed oxidation of 1-naphthol. The optimum conditions of enzymatic oxidation of 1-naphthol were determined to be pH 8.0 and 40 °C. Under the optimum conditions, PPO-catalysed oxidation of 1-naphthol increased with an increase in the enzyme dose. Quinone derivatives enzymatically generated were chemisorbed on chitosan beads and the initial velocity of PPO-catalysed oxidation increased with an increase in the amount of added chitosan beads. A specific initial velocity of 0.0675 µmol/U·min was obtained in the PPO concentration range below 200 U/cm³ and 1-naphthol was completely removed within 24 h by quinone adsorption on chitosan beads (0.20 cm³/cm³) at a PPO concentration of 100 U/cm³. The removal time was shortened by increasing the enzyme dose or the amount of added chitosan beads. 2-Naphthol was also completely removed at an initial concentration of 0.05 mM or less by prolonging the reaction time, since PPO-catalysed oxidation of 2-naphthol was much slower than that of 1-naphthol. In addition, this procedure was also applied to the removal of DHNs. These results revealed that the procedure constructed in this study was an effective technique to remove naphthols and DHNs from the aqueous medium.
Assuntos
Catecol Oxidase/química , Quitosana/química , Naftóis/química , Poluentes Químicos da Água/química , Concentração de Íons de Hidrogênio , Oxirredução , Soluções , Temperatura , Purificação da ÁguaRESUMO
Only one methylene unit difference in the long alkyl group of isobutenyl ether compounds is amplified by blending, resulting in the diversification of 2D structures, although each component formed individual 2D patterns, regardless of alkyl chain lengths. The 2D structures were revealed by using scanning tunneling microscopy at the solid/liquid interface.
RESUMO
The two-dimensional molecular assembly was accomplished of bacteriochlorophyll a (BChl a) and zinc-substituted BChl a (Zn-BChl a) together with synthetic poly(ethylene glycol)(PEG)-linked light-harvesting (LH) model polypeptides on a gold Au(111) electrode modified with supported lipid bilayers. Model polypeptides for LH1-α from Rhodospirillum (Rs.) rubrum were successfully synthesized and stably assembled with Zn-BChl a in 1,2-dioleoyl-sn-glycero-3-[phospho-rac-(1'-glycerol)] (DOPG) lipid bilayers on an electrode at room temperature, as well as in liposomal solution, in which the Zn-BChl a complex unlike BChl a, was stably assembled. The PEG moiety of the model polypeptide assisted the stable assembly with an α-helical conformation of the LH1-α model peptides together with these pigments onto the gold electrode with defined orientation. The photocurrent response depended on the combinations of the pigments and synthetic LH model polypeptides. The results presented herein will be useful for the self-assembly of these complexes on electrodes to construct efficient energy-transfer and electron-transfer reactions between individual pigments in lipid bilayers.
RESUMO
Membrane fusion proteins such as the hemagglutinin glycoprotein have target recognition and fusion accelerative domains, where some synergistically working elements are essential for target-selective and highly effective native membrane fusion systems. In this work, novel membrane fusion devices bearing such domains were designed and constructed. We selected a phenylboronic acid derivative as a recognition domain for a sugar-like target and a transmembrane-peptide (Leu-Ala sequence) domain interacting with the target membrane, forming a stable hydrophobic α-helix and accelerating the fusion process. Artificial membrane fusion behavior between the synthetic devices in which pilot and target liposomes were incorporated was characterized by lipid-mixing and inner-leaflet lipid-mixing assays. Consequently, the devices bearing both the recognition and transmembrane domains brought about a remarkable increase in the initial rate for the membrane fusion compared with the devices containing the recognition domain alone. In addition, a weakly acidic pH-responsive device was also constructed by replacing three Leu residues in the transmembrane-peptide domain by Glu residues. The presence of Glu residues made the acidic pH-dependent hydrophobic α-helix formation possible as expected. The target-selective liposome-liposome fusion was accelerated in a weakly acidic pH range when the Glu-substituted device was incorporated in pilot liposomes. The use of this pH-responsive device seems to be a potential strategy for novel applications in a liposome-based delivery system.
Assuntos
Materiais Biomiméticos/química , Materiais Biomiméticos/metabolismo , Ácidos Borônicos/química , Desenho de Fármacos , Lipossomos/metabolismo , Fusão de Membrana , Peptídeos/química , Sequência de Aminoácidos , Membrana Celular/metabolismo , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Peptídeos/metabolismo , Especificidade por SubstratoRESUMO
A weakly acidic pH-responsive polypeptide is believed to have the potential for an endosome escape function in a polypeptide-triggered delivery system. For constructing a membrane fusion device with pH-responsiveness, we have designed novel polypeptides that are capable of forming an α2 coiled coil structure. Circular dichroism spectroscopy reveals that a polypeptide, AP-LZ(EH5), with a Glu and His salt-bridge pair at a staggered position in the hydrophobic core forms a stable coiled coil structure only at endosomal pH values (pH 5.0 to 5.5). On the basis of their endosomal-pH responsiveness, a boronic acid/polypeptide conjugate (BA-H5-St) was also designed as a pilot molecule to construct a pH-responsive, one-way membrane fusion system with a sugarlike compound (phosphatidylinositol: PI)-containing liposome as a target. Membrane fusion behavior was characterized by lipid-mixing, inner-leaflet lipid-mixing, and contents-mixing assays. These studies reveal that membrane fusion is clearly observed when the pH of the experimental system is changed from 7.4 (physiological condition) to 5.0 (endosomal condition).
Assuntos
Ácidos Borônicos/química , Endossomos/química , Ácido Glutâmico/química , Lipossomos/química , Peptídeos/química , Dicroísmo Circular , Concentração de Íons de Hidrogênio , Fusão de Membrana , Membranas Artificiais , Dados de Sequência Molecular , Estrutura Molecular , TermodinâmicaRESUMO
The coiled coil trimer structure is a common motif observed in membrane fusion processes of specific fusion proteins such as the hemagglutinin glycoprotein. The HA2 subunit in the hemagglutinin changes its conformation or geometry to be favorable to membrane fusion in response to endosomal weakly acidic pH. This pH responsiveness is indispensable to an artificial polypeptide-triggered delivery system as well as the membrane fusion reaction in biology. In this study, we have constructed an AAB-type coiled coil heteroassembled system that is sensitive to weakly acidic pH. The heterotrimer is formed from two kinds of polypeptides containing an Ala or a Trp residue at a hydrophobic a position, and it was observed that the Glu residue at the other a position induced an acidic pH-dependent conformational change. On the basis of this pH-responsive coiled coil heteroassembled system, a boronic acid coupled working polypeptide for the combination of an intervesicular complex with a sugarlike compound on the surface of the target liposome, and a supporting polypeptide for the construction of a pH-responsive heterotrimer with the working polypeptide were designed and synthesized. The process of membrane fusion was characterized by lipid-mixing, inner-leaflet lipid-mixing, and content-mixing assays. The target selective vesicle fusion is clearly observed at a weakly acidic pH, where the working polypeptides form a heterotrimeric coiled coil with the supporting polypeptides in a 1:2 binding stoichiometry and the surfaces between pilot and target vesicles come into close proximity to each other.
Assuntos
Lipossomos/química , Dicroísmo Circular , Hemaglutininas/química , Concentração de Íons de Hidrogênio , UltracentrifugaçãoRESUMO
Systematic investigations were carried out to determine the optimum process parameters such as the hydrogen peroxide (H2O2) concentration, concentration and molar mass of poly(ethylene glycol) (PEG) as an additive, pH value, temperature and enzyme dose for treatment of bisphenol A (BPA) with horseradish peroxidase (HRP). The HRP-catalysed treatment of BPA was effectively enhanced by adding PEG, and BPA was completely converted into phenoxy radicals by HRP dose of 0.10 U/cm3. The optimum conditions for HRP-catalysed treatment of BPA at 0.3 mM was determined to be 0.3 mM for H2O2 and 0.10 mg/cm3 for PEG with a molar mass of 1.0 x 10(4) in a pH 6.0 buffer at 30 degrees C. Different kinds of bisphenol derivatives were completely or effectively treated by HRP under the optimum conditions determined for treatment of BPA, although the HRP dose was further increased as necessary for some of them. The aggregation of water-insoluble oligomers generated by the enzymatic radicalization and radical coupling reaction was enhanced by decreasing the pH values to 4.0 with HCl after the enzymatic treatment, and BPA and bisphenol derivatives were removed from aqueous solutions by filtering out the oligomer precipitates.
Assuntos
Peroxidase/química , Fenóis/química , Polietilenoglicóis/química , Purificação da Água/métodos , Compostos Benzidrílicos , Catálise , Relação Dose-Resposta a Droga , Peroxidase do Rábano Silvestre/química , Peróxido de Hidrogênio/química , Concentração de Íons de Hidrogênio , Modelos Químicos , Fenol/química , Fenóis/isolamento & purificação , Polímeros/química , Temperatura , Água/química , Poluentes Químicos da Água/isolamento & purificaçãoRESUMO
A novel programmable membrane fusion system driven by selective molecular recognition between diols and boronic acids on the different vesicles was constructed in this study.
Assuntos
Bicamadas Lipídicas/metabolismo , Fusão de Membrana , Ácidos Borônicos/química , Ácidos Borônicos/metabolismo , Fluorescência , Inositol/química , Inositol/metabolismo , Fosfatidilinositóis/metabolismo , Especificidade por SubstratoRESUMO
In many viruses, pH-responsive coiled-coil domains in the specific fusion proteins play important roles in membrane fusion and the infection of viruses into host cells. To investigate the relationship between the conformational change of the coiled coil and the fusion process, we have introduced a de novo designed polypeptide as a model system of the coiled-coil domain. This system enables the systematic study of the dynamics of pH-responsive coiled-coil polypeptide-membrane interactions. First, we designed and synthesized pH-responsive isoleucine-zipper triple-stranded coiled-coil polypeptides. Then the relationship between the pH-induced conformational change of the polypeptide and the membrane's interactive properties was studied by physicochemical methods. Structural changes in the designed polypeptides were examined by means of circular dichroism measurements. And finally, the behavior of the membrane fusion was investigated by leakage of liposomal contents, turbidity analysis, dynamic light scattering, and lipid mixing experiments. Our data show that coiled-coil formation under acidic pH conditions enhances polypeptide-induced membrane fusion. The results in this study demonstrate that an artificial membrane fusion system can be constructed on a molecular level by the use of a pH-responsive isoleucine-zipper triple-stranded coiled-coil polypeptide.
Assuntos
Concentração de Íons de Hidrogênio , Fusão de Membrana , Membranas Artificiais , Peptídeos/química , Dicroísmo Circular , Transferência Ressonante de Energia de Fluorescência , Lipossomos , Conformação ProteicaRESUMO
Alkylphenols were effectively treated with horseradish peroxidase at pH 7.0 and 30 degrees C in the presence of H(2)O(2) and poly(ethylene glycol) irrespective of the relative position or isomeric form of the alkyl chains. Water-insoluble oligomer precipitates were readily filtered out after enzymatic treatment, and transparent and colorless solutions were obtained for all p- and m-alkylphenols used.
Assuntos
Peroxidase do Rábano Silvestre/metabolismo , Fenóis/química , Fenóis/isolamento & purificação , Água/química , Peróxido de Hidrogênio/metabolismo , Concentração de Íons de Hidrogênio , Isomerismo , Polietilenoglicóis/metabolismo , Soluções , TemperaturaRESUMO
Enzymatic treatment of o-, m-, and p-chlorophenols and o-, m-, and p-cresols from artificial wastewater was undertaken through the enzymatic conversion into the corresponding phenoxy radicals with horseradish peroxidase (HRP) and nonenzymatic radical coupling reaction. The concentration of chlorophenols and cresols decreased sharply over the reaction time and water-insoluble oligomer precipitates were generated. The optimum conditions were determined to be the H2O2 concentration of 2.5 mM and poly(ethylene glycol) with molecular mass of 1.0 x 10(4) (10K-PEG) of 0.10 mg/cm3 at 30 degrees C for treatment of p-chlorophenol at 2.5 mM. The optimum pH values depended on the relative position of a chlorine atom for chlorophenols and on a methyl group for cresols. Concentrations of HRP and 10K-PEG were increased to 1.0 U/cm3 and 1.0 mg/cm3 respectively to treat m-chlorophenol highly. For o-chlorophenol, a decrease in the pH value to 3.0 after the enzymatic treatment led to the enhancement of the aggregation of oligomer precipitates. The % residual value for o-cresol effectively decreased by absorbing water-soluble intermediates on the chitosan films. These results indicate that chlorophenols and cresols were removed to a great degree by this technique, although the detailed procedure depended on the position of substituent groups of chlorophenols and cresols.
Assuntos
Clorofenóis/química , Cresóis/química , Peroxidase do Rábano Silvestre/farmacologia , Eliminação de Resíduos Líquidos/métodos , Poluentes Químicos da Água , Purificação da Água/métodos , Relação Dose-Resposta a Droga , Concentração de Íons de Hidrogênio , Peso Molecular , Polietilenoglicóis/química , TemperaturaRESUMO
Enzymatic removal of p-alkylphenols from aqueous solutions was investigated through the two-step approach, the quinone conversion of p-alkylphenols with mushroom tyrosinase (EC 1.14.18.1) and the subsequent adsorption of quinone derivatives enzymatically generated on chitosan beads at pH 7.0 and 45 degrees C as the optimum conditions. This technique is quite effective for removal of various p-alkylphenols from an aqueous solution. The % removal values of 97-100% were obtained for p-n-alkylphenols with carbon chain lengths of 5 to 9. In addition, removal of other p-alkylphenols was enhanced by increasing either the tyrosinase concentration or the amount of added chitosan beads, and their % removal values reached >93 except for 4-tert-pentylphenol. This technique was also applicable to remove 4-n-octylphenol (4NOP) and 4-n-nonylphenol (4NNP) as suspected endocrine disrupting chemicals. The reaction of quinone derivatives enzymatically generated with the chitosan's amino groups was confirmed by the appearance of peaks for UV-visible spectrum measurements of the chitosan films incubated in the p-alkylphenol and tyrosinase mixture solutions. In addition, 4-tert-pentylphenol underwent tyrosinase-catalyzed oxidation in the presence of hydrogen peroxide.
Assuntos
Agaricales/enzimologia , Quitosana , Monofenol Mono-Oxigenase/metabolismo , Fenóis/metabolismo , Purificação da Água/métodos , Disruptores Endócrinos/metabolismo , Poluentes Ambientais/metabolismo , Soluções , ÁguaRESUMO
Enzymatic removal of various phenol compounds from artificial wastewater was undertaken by the combined use of mushroom tyrosinase (EC 1.14.18.1) and chitosan beads as function of pH value, temperature, tyrosinase dose, and hydrogen peroxide-to-substrate ratio. Chitosan film incubated in a p-crersol+tyrosinase mixture had the main peaks at 400-470 nm assigned to chemically adsorbed quinone derivatives, which increased over the immersion time. These results indicate that removal of phenol compounds is caused by their tyrosinase-catalyzed oxidation to the corresponding quinone derivatives and the subsequent chemical adsorption on the chitosan film. The optimum conditions for quinone adsorption were determined to be pH 7 and 45 degrees C for p-cresol. Some alkyl-substituted phenol compounds were removed by adsorption of quinone derivatives enzymatically generated on the chitosan beads, and the % removal for p-cresol, 4-ethylphenol, 4-n-propylphenol, 4-n-butylphenol, and p-chlorophenol went up to 93%. In addition, 4-tert-butylphenol underwent tyrosinase-catalyzed oxidation in the presence of hydrogen peroxide. This procedure was applicable to removal of chlorophenols and alkyl-substituted phenols.
Assuntos
Quitosana/química , Monofenol Mono-Oxigenase/química , Fenóis/química , Fenóis/isolamento & purificação , Purificação da Água/métodos , Adsorção , Agaricales/enzimologia , Biodegradação Ambiental , Materiais Revestidos Biocompatíveis/química , Enzimas Imobilizadas/química , Concentração de Íons de Hidrogênio , Microesferas , Ligação Proteica , Poluentes Químicos da Água/isolamento & purificaçãoRESUMO
We prepared amide compounds which were derived from ferulic acid using various amines, and investigated their stimulatory effects on insulin secretion using rat pancreatic RIN-5F cells. Most of these compounds exhibited significant promotion of the insulin-release at a concentration of 10 microM and in particular, the amides having n-butyl, n-pentyl, pyrrolidine, and piperidine groups showed high activity.