T. F. Solov'eva

Thin-layer chromatography of carbohydrates

The separation of carbohydrates on thin layers of silica gel is due first of all to the type and quantity of impregnating salt. The results of chromatography of carbohydrates on thin layers of silica gel in the presence of sodium mono- and dihydrogen phosphate are often better than those of paper chromatography (e.g. a good separation of complex mixtures, less sensitivity to inorganic impurities, and faster separation). The present method can be recommended for the study of the monosaccharide composition of oligo- and polysaccharides.

Comparative study of carrageenans from reproductive and sterile forms of Tichocarpus crinitus (Gmel.) Rupr (Rhodophyta, Tichocarpaceae)

A comparative study of the structure and properties of the sulfated polysaccharides (carrageenans) isolated from the vegetative and reproductive forms of the red alga Tichocarpus crinitus was performed. The polysaccharides were separated into the gelling (KCl-insoluble) and non-gelling (KCl-soluble) fractions by precipitation with 4% KCl. The total content of polysaccharides extracted from the reproductive form of the alga was 1.8-fold more than that extracted from the vegetative form, and in the first case, the gelling polysaccharides mostly accumulated. The gelling polysaccharides from the vegetative form have the highest molecular weight (354 kD). According to the results of FT-IR and 13C-NMR spectroscopy, the gelling polysaccharide fractions from both forms are κ/β carrageenans. The differences concern the content of the κ- and β-disaccharide units and the presence of a small content of the sulfated disaccharide segments (precursors of the κ-carrageenans) in the polysaccharide from the reproductive form of the alga. The non-gelling polysaccharide fractions from both forms of the plant are mixtures of sulfated galactans with a low content of 3,6-anhydrogalactose.

New glycolipids (chitooligosaccharide derivatives) possessing immunostimulating and antitumor activities

New glycolipids, derived from chitooligosaccharides of dp 2-4 and containing both free and acylated amino groups, were synthesized. The structure of the key compounds (di-, tri-, and tetra-saccharides acylated with different fatty acids) were elucidated by 13C NMR spectroscopy. Only the amino group of the reducing end of the chitooligosaccharides was found to be acylated when equimolecular amounts of reagents were used. The compounds obtained were shown to possess a low toxicity and certain immunostimulatory and antitumor activities. An induction of interleukin-1 and tumor necrosis factor by the immunocompetent cells and an augmentation by 140-180% of the mean life of mice with the Erlich carcinoma were observed.

Some structural features of Panax ginseng C. A. Mey pectin

Abstract The pectin isolated from roots of Panax ginseng C. A. Mey, contains residues of d -galacturonic acid, d -galactose, and l -arabinose, as main components, and those of d -xylose, l -rhamnose, and an unidentified sugar, as minor components of its carbohydrate chain. On partial hydrolysis with acid, the pectin afforded a galacturonan having a linear chain of (1 → 4)- and, possibly, (1 → 5)-linked α- d -galacturonic acid residues. Pectinase digestion of the pectin furnished a neutral, pectic polysaccharide in which l -arabinofuranose residues apparently occur in the exterior chains. On partial hydrolysis with acid, the polysaccharide afforded a branched galactan and an acidic heteropolysaccharide. The galactan contains chains of (1 → 3)- and (1 → 6)-linked d -galactopyranose residues. The heteropolysaccharide appears to contain sequences involving d -galactose residues, and others involving residues of d -galacturonic acid, d -xylose, and l -rhamnose.

Carrageenans from cystocarpic and sterile plants of Chondrus pinnulatus (Gigartinaceae, Rhodophyta) collected from the Russian Pacific coast

The chemical structure, gel properties and biological activity of the carrageenans isolated from cystocarpic and sterile plants of Chondrus pinnulatus were investigated. The total carrageenan content of the sterile plant was observed to be twice that of the cystocarpic plants. According to data obtained by 13C-NMR and FT IR, the gelling polysaccharides from cystocarpic and sterile plants of C. pinulatus have similar structures and were identified as κ/ι-carrageenans. The difference between these polysaccharides was in the ratio of the κ- and ι-segments, with a predominant content of κ-segments in cystocarpic plants (80%). Moreover, KCl-insoluble fractions possibly contain hetero-disperse μ/ν precursor: amounts of this in the polysaccharide from sterile plants were more than that extracted from the cystocarpic plants. The KCl-soluble fractions (non gelling) were λ-carrageenans with another carrageenan type that had a low amount of 3,6-anhydrogalactose. Carrageenans from cystocarpic stages showed good gelling properties, whereas those from sterile plants formed a very weak gel. Structural differences and molecular weight of carrageenans obviously determine the biological activity of the polysaccharides. Non gelling-carrageenans from both types of ιt C. pinnulatus plants showed high macrophage-phosphatase activity and κ/ι-carrageenan from cystocarpic plant possessed a potent anti-coagulant activity, which was extremely strong in a low concentration of 100 μg ml−1.

Marine Compounds with Therapeutic Potential in Gram-Negative Sepsis

This paper concerns the potential use of compounds, including lipid A, chitosan, and carrageenan, from marine sources as agents for treating endotoxemic complications from Gram-negative infections, such as sepsis and endotoxic shock. Lipid A, which can be isolated from various species of marine bacteria, is a potential antagonist of bacterial endotoxins (lipopolysaccharide (LPSs)). Chitosan is a widespread marine polysaccharide that is derived from chitin, the major component of crustacean shells. The potential of chitosan as an LPS-binding and endotoxin-neutralizing agent is also examined in this paper, including a discussion on the generation of hydrophobic chitosan derivatives to increase the binding affinity of chitosan to LPS. In addition, the ability of carrageenan, which is the polysaccharide of red alga, to decrease the toxicity of LPS is discussed. We also review data obtained using animal models that demonstrate the potency of carrageenan and chitosan as antiendotoxin agents.

Molecular characteristics of OmpF-like porins from pathogenic yersinia

Nonspecific pore-forming proteins (porins) are the major proteins of the outer membrane of Gram-negative bacteria responsible for diffusion of low-molecular-weight compounds. Nucleotide sequences of the OmpF-like porins from the pathogenic bacteria Yersinia pseudotuberculosis (YPS) and Yersinia enterocolitica (YE) were cloned and determined. Values of molecular weights (MW) and isoelectric points (IEP) calculated for these proteins (for OmpF-YPS: MW 37.7 kD, IEP 4.45; for OmpF-YE: MW 39.5 kD, IEP 4.34) are in good agreement with experimental data. The OmpF-like Yersinia porins are highly homologous to each other (83–92%) and also to the OmpF protein from Serratia marcescens (70%); the homology to the OmpF porin from E. coli is significantly lower (52–58%). Multiple alignment of the amino acid sequences of mature OmpF proteins provided the distribution of conservative amino acid residues typical for porins. Moreover, the OmpF-like porins from Yersinia are characterized by the presence of extended regions with high and low homologies, which coincide with the transmembrane domains and “external” loops, respectively, of the topological model of the OmpF porin from E. coli. By predictive methods, the secondary structure of the OmpF-like porins from Yersinia was obtained. This structure is represented by 16 β-strands connected by short “ periplasmic” and longer “external” loops with unordered structure.

Determination of binding constants of lipopolysaccharides of different structure with chitosan

The interaction of endotoxins—lipopolysaccharides (LPS) different in degree of the O-specific chain polymerization—with 20-and 130-kD chitosan was studied using the competitive binding of LPS with the complex of chitosananionic dye (tropaeolin 000-2) and the direct binding of 125I-labeled LPS with chitosan immobilized on Sepharose 4B. The interaction of 20-kD chitosan with LPS was non-cooperative, and immobilization of the polycation on Sepharose resulted in its binding to 125I-labeled LPS with a positive cooperativity. The interaction of LPS possessing a long O-specific chain with 130-kD chitosan was characterized by negative cooperativity. Binding constants of LPS with the polycation and the number of binding sites per amino group of chitosan were determined. The interaction affinity and stoichiometry of the LPS-chitosan complexes significantly depend on the LPS structure and concentration in the reaction mixture. The increase in the length of carbohydrate chains of LPS results in increase in the binding constants and decrease in the bound endotoxin amount.

[Elucidation of structure of lipid A from the marine Gram-negative bacterium Pseudoalteromonas haloplanktis ATCC 14393T].

The chemical structure of lipid A, from the marine γ-proteobacterium Pseudoalteromonashaloplanktis АТСС 14393Т, a main product of lipopolysaccharide hydrolysis (1% AcOH), was determined using chemical methods and NMR spectroscopy. The lipid A was shown to be β-1,6-glucosaminobiose 1,4′-diphosphate acylated with two (R)-3-hydroxyalkanoic acid residues at C3 and C3′ and amidated with one (R)-3-hydroxydodecanoyl and one (R)-3-dodecanoyloxydodecanoyl residue at N2 and N2′, respectively.

Chemical characterization of lipid A from some marine proteobacteria.

Lipids A from type and wild strains of marine Proteobacteria belonging to Alteromonadaceae (Alteromonas (1 species), Idiomarina (1 species), and Pseudoalteromonas (8 species) genera) and Vibrionaceae (Shewanella (1 species) and Vibrio (1 species) genera) families and Marinomonas genus (1 species) were isolated by hydrolysis of their respective lipopolysaccharides with 1% acetic acid. Based on thin-layer chromatography data, the lipids A studied had low heterogeneity and generated family-specific patterns varying in numbers of bands and their chromatographic mobility. Total chemical analysis of the compounds showed that they contained glucosamine, phosphate, and fatty acids with decanoate (I. zobellii KMM 231T lipid A) or dodecanoate (lipids A of the other bacteria) and 3-hydroxy alkanoates as the major fatty acid components. Unlike terrestrial bacterial lipids A, lipids A of marine Proteobacteria had basically monophosphoryl (except V. fluvialis AQ 0002B lipid A with its two phosphate groups) and pentaacyl (except S. alga 48055 and V. fluvialis AQ 0002B lipids A which were found to have six residues of fatty acids per molecule of glucosamine disaccharide) structural types, low toxicity, and may be useful as potential endotoxin antagonists.