Nobuo Sakairi

Facile preparation and inclusion ability of a chitosan derivative bearing carboxymethyl-β-cyclodextrin

Abstract Coupling carboxymethylated β-cyclodextrin and partially deacetylated chitin (Mw = 7300) afforded a new type of functional chitosan derivative having an ability to form an inclusion complex. The property to form an inclusion complex was studied using a fluorescent dye, 6-(p-toluidino)-2-naphthalene-6-sulfonate (TNS), as the guest molecule. It was found that the presence of the cyclodextrin bonded to chitosan enhanced the relative intensity of TNS fluorescence significantly (3.6 times as much as that of β-cyclodextrin). Fluorometric titration revealed that the 1:1 stoichiometrical complex was formed and that the equilibrium constant was 1.13-1.68 × 103 M−1. Ionic interaction between the sulfonic acid moiety of TNS and the amino group of chitosan was suggested from experiments which involved changing the pH and ionic strength of the buffer solution as well as adding a surface-active agent.

Laminin-1 peptide-conjugated chitosan membranes as a novel approach for cell engineering

Laminin, a major component of the basement membrane, has diverse biological activities. Recently, we identified various biologically active sequences on laminin‐1 by using a large set of synthetic peptides. Chitosan, a polysaccharide, is biodegradable and has been used as a biomaterial. Here, we conjugated several biologically active laminin peptides onto chitosan membranes and measured the cell attachment activity of peptide‐conjugated chitosan membranes with various cell types. The active laminin peptide‐conjugated chitosan membranes promoted cell attachment with cell type specificity. A99 (AGTFALRGDNPQG)‐chitosan membrane promoted cell attachment with well‐organized actin stress fibers. This adhesion was inhibited by EDTA but not by heparin. AG73 (RKRLQVQLSIRT)‐chitosan membrane promoted cell attachment with filopodia formation, and this adhesion was inhibited by heparin but not by EDTA. These data suggest that the A99‐chitosan membrane interacted with an integrin cellular receptor and that the AG73‐chitosan membrane promoted proteoglycan‐mediated cell attachment, as previously reported. Furthermore, both AG73‐chitosan and A99‐chitosan membranes effectively promoted neurite outgrowth with PC12 rat pheochromocytoma cells. We conclude that conjugation on a chitosan membrane is applicable for testing quantitatively the biological activity of synthetic peptides and that these constructs have a potential ability to serve as bioadhesive materials for tissue regeneration and engineering.

Surface modification of nonporous glass beads with chitosan and their adsorption property for transition metal ions

Abstract A new hybrid material that adsorbs transition metal ions was prepared by immobilizing chitosan on the surface of nonporous glass beads. The glass beads, prepared by etching in aqueous NaOH at 100°C, were first reacted with γ-aminopropyltriethoxysilane (APES) to introduce amino groups on the surface. Subsequently, the resulting aminated beads were treated with glutaraldehyde at 25°C to change the amino groups into aldehyde groups. Finally, chitosan of average molecular weight 40,000 was introduced via the aldehyde groups through a Schiffs reaction. After complete acid-hydrolysis of the immobilized chitosan, the Svennerholm method for glucosamine analysis showed that 0.3% (w/w) chitosan had been successfully introduced on the glass beads. Atomic absorption spectroscopic analysis of eluants of a column of the chitosan-modified glass beads showed that metal ions such as Cu 2+ , Ag + , Pb 2+ , Fe 3+ , and Cd 2+ were more than 90% entrapped on a column of beads prepared in this manner.

Preparation and characterization of DNA films induced by UV irradiation.

Large amounts of DNA-enriched materials, such as salmon milts and shellfish gonads, are discarded as industrial waste. We have been able to convert the discarded DNA to a useful material by preparing novel DNA films by UV irradiation. When DNA films were irradiated with UV light, the molecular weight of DNA was greatly increased. The reaction was inhibited by addition of the radical scavenger galvinoxyl suggesting that the DNA polymerization with UV irradiation proceeded by a radical reaction. Although this UV-irradiated DNA film was water-insoluble and resistant to hydrolysis by nuclease, the structure of the DNA film in water was similar to non-irradiated DNA and maintained B-form structure. In addition, the UV-irradiated DNA film could effectively accumulate and condense harmful DNA-intercalating compounds, such as ethidium bromide and acridine orange, from diluted aqueous solutions. The binding constant and exclusion number of ethidium bromide for UV-irradiated DNA were determined to be 6.8 +/- 0.3 x 10(4) M(-1) and 1.6 +/- 0.2, respectively; these values are consisted with reported results for non-irradiated DNA. The UV-irradiated DNA films have potential uses as a biomaterial filter for the removal of harmful DNA intercalating compounds.

UV-irradiation-induced DNA immobilization and functional utilization of DNA on nonwoven cellulose fabric

Immobilization of double-stranded DNA onto nonwoven cellulose fabric by UV irradiation and utilization of DNA-immobilized cloth were examined. The immobilized DNA was found to be stable in water, with the maximum amount of fabric-immobilized DNA being approximately 20 mg/g of nonwoven fabric. The DNA-immobilized cloth could effectively accumulate endocrine disruptors and harmful DNA intercalating pollutants, such as dibenzo-p-dioxin, dibenzofuran, biphenyl, benzo[a]pyrene and ethidium bromide. Additionally, DNA-immobilized cloth was found to bind metal ions, such as Ag+, Cu2+, and Zn2+. The maximum amounts of bound Ag+, Cu2+, and Zn2+ onto DNA-immobilized cloth (1 g) were approximately 5, 2, and 1 mg, respectively. DNA-immobilized cloth containing Ag+ showed antibacterial activity against Escherichia coli and Staphylococcus aureus. DNA-immobilized cloth without metal ion and with Cu2+ or Zn2+ did not show antibacterial activity. These results suggest that immobilized DNA imparts useful functionality to cloth. DNA-immobilized cloth prepared by UV irradiation has potential to serve as a useful biomaterial for medical, engineering, and environmental application.

Synthesis of amylostatin (XG), α-glucosidase inhibitor with basic pseudotrisaccharide structure

Abstract The basic pseudotrisaccharide ( 1 constituting the common and essential building block of several α-glucosidase inhibitors of microbial origin was synthesized by coupling two synthons: the chiral cyclohexyl halide ( 11 ) and the 4′-amino-4′-deoxy-disaccharide ( 14 ).

A method for direct harvest of bacterial cellulose filaments during continuous cultivation of Acetobacter xylinum

Abstract Continuous filamentation of bacterial cellulose (BC) was successfully achieved by using shallow pan for the incubation to regulate thickness of the BC gel produced by Acetobacter xylinum . The BC filament was harvested and prepared directly by picking up BC pellicles, the thin BC gel, and winding slowly from the surface of the culture medium passed through a preliminary bactericidal washing bath. The X-ray diffraction analysis and scanning electron microscopic observation of the BC filament thus obtained showed that the filament was smooth and the fairly good orientation of BC molecules. The average tensile strength was 4.4 g denier −1 for the filament prepared by hot alkaline treatment and subsequent washing with distilled water and dried under tension (Filament W): 3.4 g denier −1 for washing with 10% aqueous ethylene glycol after alkaline treatment followed by drying under tension (Filament E) and 2.4 g denier −1 for the treatment with 10% ethylene glycol after normal water-washing followed by drying under tension.

Novel synthesis of a water-soluble cyclodextrin-polymer having a chitosan skeleton

Formation of Schiffs base between 2-O-(formylmethyl)-β-cyclodextrin and chitosan with an average molecular weight of 40 000 in acetate buffer at pH 4.4, followed by reduction with sodium cyanoborohydride produced a β-cyclodextrin-linked chitosan in a one-pot reaction. The product, which had a degree of substitution of 37%, was soluble in water at neutral and alkaline conditions. UV-visible and circular dichroism spectroscopic examinations revealed that the product had the ability to form a host-guest complex with p-nitrophenolate.

Chitosan beads with pendant α-cyclodextrin: preparation and inclusion property to nitrophenolates

Highly porous beads having an ability to form inclusion complexes with specific substrates have been synthesized and preliminary experiments for the application to adsorbent for affinity column chromatography and controlled release were carried out. Water-insoluble chitosan beads were synthesized by adding an aqueous acetic acid solution of chitosan into ethanolic aqueous sodium hydroxide and subsequent crosslinking with hexamethylene diisocyanate in N, N-dimethylformamide. The resulting beads were further treated with 2-O-formylmethyl-α-cyclodextrin in the presence of sodium cyanoborohydride in acetate buffer at pH 4.4, giving the cyclodextrin-linked chitosan beads. Their inclusion ability was examined by the use of p-nitrophenol and its analogue as model compounds.

Biosynthesis of hetero-polysaccharides by Acetobacter xylinum - Synthesis and characterization of metal-ion adsorptive properties of partially carboxymethylated cellulose

Abstract Biological reconstruction of water-soluble carboxymethylated cellulose (CMC; D.S. =0.47) has been achieved by culturing Acetobacter xylinum in medium containing CMC and d -glucose to give a novel hetero-polysaccharide having a carboxymethyl function. The novel extracellular polysaccharide, carboxymethylated-bacterial cellulose (CM-BC), had an ion exchange ability with enhanced specific adsorption for lead and uranyl ions compared to the original CMC and bacterial cellulose. The contribution of the hydroxy group at C-2 was confirmed by applying carboxymethylated chitin, which possesses acetamido group at C-2 of the glucose residue, as the carbon source of the incubation.