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A Michael reaction of chitosan was conducted in water containing acetic acid with various acryl reagents. The degree of substitution could be controlled by temperature, reaction time, and the amount of acryl reagents. Although the modified chitosan derivatives with acrylic acid esters showed water-solubility, that with poly(ethylene glycol) acrylate, however, turned to water-insoluble material by lyophilization. Good biodegradation was observed in modified chitosan derivatives by standard activated sludge.

The hierarchical self-assemblies of hydrophobically modified pullulan, such as cholesteryl-bearing pullulan (CHP) and hexadecyl group-bearing pullulan (C16P), were investigated. In a dilute aqueous solution, they formed hydrogel nanoparticles. In semidilute solution above approximately 2% (w/w), the viscosity of the CHP or C16P solution drastically increased. At higher concentration, they formed macroscopic gels:  CHP gave a structure in which the nanoparticles are linked, while the C16P macrogel has mainly a fibrous network structure. In rheological measurements of CHP gel, G‘ ‘ exhibited a maximum at ω = 0.1 rad s^-1, while G‘ reached a plateau in the high-frequency regime. On the other hand, C16P gel showed G‘ ‘ with a peak at ω > 10 rad s^-1. The G‘ and G‘ ‘ values for CHP and C16P gels obeyed an Arrhenius plot. The activation energy of the shift factor for CHP was higher than that for C16P. Thus, we can successfully control the association of hydrophobically modified pullulans and organize them from the molecular level to the nanoscale and macroscopic region.

Thermoresponsive hydrogel nanoparticles were prepared by self-assembly of two different hydrophobically modified polymers, namely a cholesterol-bearing pullulan (CHP) and a copolymer of N-isopropylacrylamide (NIPAM) and N-[4-(1-pyrenyl)butyl]-N-n-octadecylacrylamide] (PNIPAM-C18Py). The interactions between CHP and PNIPAM-C18Py were investigated by fluorescence spectroscopy, dynamic light scattering, and size exclusion chromatography. After ultrasonication of a mixture of CHP and PNIPAM-C18Py (5:1 by weight) at 25 °C, monodisperse nanoparticles (Dh = 45 nm) were obtained, consisting of self-assembly of the two polymers associated via their hydrophobic moieties. Evidence from fluorescence and dynamic light scattering demonstrated that, above 32 °C, the lower critical solution temperature (LCST) of PNIPAM-C18Py, the colloidal mixed nanoparticles increase in diameter (from 47 to 160 nm), but no macroscopic aggregation could be detected. This phenomenon was thermoreversible:  upon cooling the particles recovered their original diameter.

We have been studying the formation of hydrogel nanoparticles by the self-aggregation of hydrophobized polysaccharide and the effective complexation between these nanoparticles as a host and various globular soluble proteins as a guest. This paper describes a new finding that refolding of the heat-denatured enzyme effectively occurs with the nanoparticles and β-cyclodextrin according to a mechanism similar to that of a molecular chaperone. In particular, the irreversible aggregation of carbonic anhydrase B (CAB) upon heating was completely prevented by complexation between the heat-denatured enzyme and hydrogel nanoparticles formed by the self-aggregation of cholesteryl group-bearing pullulan (CHP). The complexed CAB was released by dissociation of the self-aggregate upon the addition of β-cyclodextrin. The released CAB refolded to the native form, and almost 100% recovery of the activity was achieved. The thermal stability of CAB was drastically improved by capture of the unfolded form which was then released to undergo refolding.

Various cholesterol-bearing pullulans (CHPs) with different molecular weights of the parent pullulan and degrees of substitution (DS) of the cholesteryl moiety were synthesized. The structural characteristics of CHPs in water were studied by static (SLS) and dynamic light scattering (DLS) and the fluorescence probe method. Irrespective of the molecular weight of the parent pullulan and the DS, all of CHPs provided unimodal and monodisperse self-aggregates in water. The size of the self-aggregate decreased with an increase in the DS of the cholesteryl moiety (hydrodynamic radius, 8.4−13.7 nm). However, the aggregation number of CHP in one nanoparticle was almost independent of the DS. The polysaccharide density within the self-aggregate (0.13−0.50 g/mL) was affected by both the molecular weight and the DS of CHPs. The mean aggregation number of the cholesteryl moiety (3.5−5.7), which was estimated by the fluorescence quenching method using pyrene and cetylpyridinium chloride, was almost same for all the CHP self-aggregates. The CHP self-aggregate is regarded as a hydrogel nanoparticle, in which pullulan chains are cross-linked noncovalently by associating cholesteryl moieties. The microenvironment inside or the structural characteristic of the self-aggregate was spectrometrically studied using a fluorescence probe, ANS. The characteristic temperature to cause a structural change of the nanoparticle (T*) decreased with an increase in the DS of CHP and the ionic strength of the medium. The thermoresponsiveness of the nanoparticle hydrogel is related to the partial dehydration of the hydrophobized pullulan upon heating.

The miscibility of dimyristoylphosphatidylcholine (DMPC) and poly(ethylene oxide) lipids (PEO lipids) in monolayers has been assessed through surface pressure and surface potential measurements. These experiments performed under equilibrium conditions at constant area revealed a strong dependence of the interfacial behavior of these mixtures on the polymer chain length. While for the lipid with the short PEO chain composed of 5 monomer units the mixtures with DMPC displayed almost an ideal behavior, for longer PEO chains (13 and 31 monomer units) π−δ isotherms considerably deviated from ideality. The effect of hydrocarbon and polymer chain orientation and of the structuring of interfacial water induced by PEO lipids on their mixing behavior with DMPC and on calculated effective dipole moments is discussed and analyzed.