ABSTRACT
Chitosan, (1-4)-2-amino-2-deoxy-beta-d-glucan, is a deacetylated form of chitin, an abundant biodegradable, positively charged natural polysaccharide. Chitosan is used for antigen delivery through mucosal barrier due to its ability to disrupt tight junctions. Here we produced new water-soluble low-molecular weight chitosan (LMW-Chi) lipid derivatives and compared their ability to stimulate humoral response with the effect of unmodified LMW-Chi or its oligosaccharide derivatives. LMW-Chi effectively penetrated into macrophage-like, lymphoid and epithelial cells. It also stimulated in mice IgG production to model proteins delivered either by subcutaneous or intranasal routes. Adjuvant effect of chitosan derivatives was comparable to or lower than that of unmodified LMW-Chi. Thus, it is possible that adjuvant effect is induced by unmodified glucosamine units of chitosan.
Subject(s)
Adjuvants, Immunologic/chemistry , Antibody Formation/drug effects , Chitin/analogs & derivatives , Animals , Chitin/chemistry , Chitin/immunology , Chitosan , Glycoconjugates , Immunoglobulin G/biosynthesis , Lipids , Mice , Molecular Weight , Oligosaccharides , Solubility , Structure-Activity RelationshipABSTRACT
The use of the traditional scheme for the isolation of bovine dopamine-beta-hydroxylase (bDBH) from bovine adrenal medulla resulted in active but not pure bDBH, containing about 50% of admixtures. Immobilized metal chelate affinity chromatography on agarose modified with iminodiacetic acid residues and charged with cobalt ions was applied in the final stage to obtain more than 90% pure and active bDBH. Final purification of bDBH using step elution with 0-0.5 M methyl-D-mannoside in buffer solution from concanavalin A-Sepharose was studied. The determination of bDBH in various samples was performed using size-exclusion chromatography.
Subject(s)
Chromatography/methods , Dopamine beta-Hydroxylase/isolation & purification , Adrenal Medulla/enzymology , Animals , Cattle , Chromatography, Affinity/methods , Chromatography, Gel/methods , Chromatography, High Pressure Liquid/methods , Electrophoresis, Polyacrylamide Gel , Spectrophotometry, UltravioletABSTRACT
The preparation of two affinity-chromatography sorbents based on cross-linked chitin are described. Both sorbents retained selectively one of the four extracellular chitinases in the culture supernatant produced by Streptomyces kurssanovii. Chitinases with molecular masses of 42 kDa and 26 kDa were isolated in homogeneous form using one-step affinity-chromatography procedures involving either a fully N-acetylated or a partially N-acetylated cross-linked chitin-type sorbent. Two other chitinases were not selectively bound by the sorbents and, therefore, were not isolated in a homogeneous form. The affinity sorbents were shown to be stable over the period of separation and could be used repeatedly.
Subject(s)
Chitinases/isolation & purification , Chromatography, Affinity/methods , Streptomyces/enzymology , Anticholesteremic Agents , Cells, Cultured , Chitin/analogs & derivatives , Chitin/chemistry , Chitosan , Cross-Linking Reagents/chemistry , Culture Media/chemistry , HemostaticsABSTRACT
One of the four chitinases with a molecular mass of 42 kDa existing in the chitinolytic enzyme complex produced by Streptomyces kurssanovii was separated in homogeneous form using one-step affinity chromatography on cross-linked and phosphorylated chitin-type sorbents. Three other chitinases were not selectively bound by the sorbents and were not obtained in homogeneous form. The affinity sorbents were shown to be stable over the time of separation and could be used repeatedly.
Subject(s)
Chitinases/isolation & purification , Streptomyces/enzymology , Adsorption , Chromatography, Affinity , Cross-Linking Reagents , Culture Media , Electrophoresis, Polyacrylamide Gel , PhosphorylationABSTRACT
The action of a microbial chitinolytic enzyme complex from Streptomyces kurssanovii on chitosan gels with various degrees of cross-linking and on the O-derivatives of the gels was examined. The rate of digestion of the gels was shown to depend on the degree of cross-linking as well as on the size of the O-substituent, decreasing greatly with a rise in substitution.
