Fucoidans are sulfated polysaccharides derived from marine brown algae. In the current work the anti-HIV activity of three fucoidans, extracted from three brown seaweeds Sargassum mcclurei, Sargassum polycystum and Turbinara ornata and collected from Nha Trang bay, Vietnam was investigated. Fucoidans extracted from the three species displayed similar antiviral activities with a mean IC50 ranging from 0.33 to 0.7 μg/ml while displaying no cell toxicity. Our results showed that the anti-HIV activity of fucoidans is not primarily linked to the sulfate content and the appropriate position of sulfate groups in the fucoidan backbones was also not associated with the antiviral activity. Fucoidans inhibited HIV-1 infection when they were pre-incubated with the virus but not with the cells, and not after infection, blocking the early steps of HIV entry into target cells. These data contribute to a better understanding of the influence of fucoidans structural characteristics on their biological activity.

Two high-molecular water-soluble preparations with high anticomplementary, antioxidant, antilipoperoxidant and antiinflammatory activities were isolated from the roots of Symphytum asperum and S. caucasicum. Their main chemical constituent was found to be poly[oxy-1-carboxy-2-(3,4-dihydroxyphenyl)ethylene], according to IR and NMR spectroscopy. The Symphytum high-molecular preparations can modulate in vitro B- chronic lymphocytic leukaemia (B-CLL) cells apoptosis and cell cycle progression.

The conformational behavior of linear (1 --> 3)-linked propyl di-, tri-, tetrafucosides and 2,3-branched tetrafucosides with linear (1 --> 3)-linked trisaccharide backbone related to fragments of natural fucoidans were studied by theoretical molecular modeling and experimental determination of transglycosidic vicinal coupling constants (3)J(C,H). The application of NOE NMR-spectroscopy, which is traditionally used in conformational analysis of oligosaccharides, was accompanied by experimental difficulties in the case of tetrafucosides, due to the overlap of cross-peaks and their trend to be close to zero. It was shown that conformations of difucoside units in the studied compounds depend on their position within the oligosaccharide backbone, on the chain length, and on the presence or absence of 2,3-branch point. The comparison of experimental and calculated values of transglycosidic constants (3)J(C,H) showed good coincidence for the middle disaccharide units of tetrafucosides, indicating that these units are more rigid than terminal ones.

A fucoidan fraction consisting Of L-fucose, sulfate, and acetate in a molar proportion of 1:1.21:0.08 was isolated from the brown seaweed Fucus distichus collected from the Barents Sea. The C-13 NMR spectrum of the fraction was typical of regular polysaccharides containing disaccharide repeating units. According to 1D and 2D H-1 and 13 C NMR spectra, the fucoidan molecules are built up of alternating 3-linked alpha-L-fucopyranose 2,4-disulfate and 4-linked alpha-L-fucopyranose 2-sulfate residues: --> 3)-alpha-L-Fucp-(2,4-di-SO3-)-(1 --> 4)-alpha-L-Fucp-(2SO(3)(-))-(1 -->. The regular structure may be only slightly masked by random acetylation and undersulfation of several disaccharide repeating units. (C) 2003 Elsevier Ltd. All rights reserved.

The order-disorder conformational transition in the tetramethylammonium salt of gellan has been studied by high-sensitivity differential scanning calorimetry as a function of concentration of the background electrolyte, Me4NCl (0.0025-0.5 M). The transition temperature and enthalpy increase with increasing salt concentration following a logarithmic relation. At the same time, there is a linear correlation between the transition enthalpy and temperature with a positive slope of 0.18 J g(-1) K-1. According to Kirchhoff's law, the slope can be considered as a heat capacity increment of the transition. This value is much larger than that expected to arise from electrostatic effects. Thus, it has been suggested that the transition is accompanied by an increase in water accessibility to some less hydrophilic fragments of the primary structure of gellan buried in its ordered conformation. The transition has an lambda-like heat capacity profile and can be therefore defined as a second-order phase transition. This assignment is in agreement with predictions of a matching model of the double helix-coil transition. The model fits well to the integral transition curve at 0.5 M Me4NCl, assuming a cooperative length of gellan to be about eight repeating units. This value is close to the size of chain corresponding to the persistent length of gellan in the coil state. The cooperativity parameter in the matching model (sigma = 0.62 +/- 0.01) indicates a small contribution of the stacking effect into the cooperativity of the transition, while a larger contribution apparently originates from the loop factor. The free energy of transition at a reference temperature (273 K) is a logarithmic function of the concentration of the salt, Delta(t)G(C-s), as predicted by the counter condensation theory and the Poisson-Boltzmann (PB) model. The latter is preferable because of the low linear charge density parameter (xi < 1) for the double helix of gellan. Analysis of the Delta(t)G(C-s) function in terms of the PB model showed that the linear charge density of gellan in the coil conformation needs to be higher than that estimated for the fully stretched chain.

The cell wall of Brevibacterium sp. VKM Ac-2118 isolated from a frozen (mean annual temperature -12 degrees C) late Pliocene layer, 1.8-3 Myr, Kolyma lowland, Russia, contains mannitol teichoic acid with a previously unknown structure. This is 1,6-poly(mannitol phosphate) with the majority of the mannitol residues bearing side phosphate groups at O-4(3). The structure of the polymer was established by chemical methods, NMR spectroscopy, and MALDI-TOF mass spectrometry.

Propyl 4,4',4"-tri-O-sulfated and non-sulfated (1 --> 3)-alpha-L-fucotriosides which are related to fragments of natural fucoidans have been synthesized. Their spectral and conformational properties have been investigated by H-1 and C-13 NMR, NOE and molecular modeling. Molecular mechanics calculations of the tri-O-sulfated compound as a trianion did not give agreement with the experimental NOE values, while the model with the non-dissociated sulfo group on the non-reducing end worked successfully. (1 --> 3)-Fucobioside fragments in both trisaccharides investigated were shown to have the same range of conformations as in previously described propyl (1 --> 3)-alpha-L-fucobiosides, but with the increase of the relative population of the conformer with the spatial proximity of H-1' and H-4 in the case of non-sulfated fucotrioside.

Carrageenans represent one of the major texturising ingredients in the food industry. They are natural ingredients, which are used for decades in food applications. Carrageenan is a generic name for a family of linear, sulfated galactans, obtained by extraction from certain species of red seaweeds (Rhodophyta). Since natural carrageenans are mixtures of different sulfated polysaccharides, their composition differs from batch to batch. Therefore, the quantitative analysis of carrageenan batches is of greatest importance for both ingredient suppliers and food industries to deliver a constant consumer product and to develop new applications based on their unique intrinsic properties. Nowadays NMR spectroscopy is one of the standard tools for the determination of the chemical structure of carrageenan samples. This review gives an overview of NMR-spectroscopy (both 1H- and 13C-NMR) as a powerful tool for the qualitative and quantitative analysis of carrageenan samples. In addition to tables containing chemical shift data for both 1H- and 13C-spectra, details about sample preparation, selective degradation and fractionation techniques are included.

The composition and structure of polysaccharides from several groups of calcareous algae (including calcareous cyanobacteria), which differ in the calcification mode (extracellular, cell wall, or intracellular), are reviewed, Two families of marine algae, Corallinaceae (Rhodophyta) and Coccolithophoraceae (Prymnesiophyta = Haptophyta), are considered in detail; they exhibit the cell wall and intracellular calcification modes, respectively, and synthesize unusual polysaccharides that seem to directly participate in the calcification process.

A water-soluble high-molecular preparation with strong anticomplementary and antioxidative activity isolated from the roots of Symphytum asperum is principally polyoxy-1-carboxy-2-(3,4-dihydroxyphenyl)ethylene.

Two fucotriosides with vicinal disubstitution alpha -L-Fuc-(1-->2)[alpha -L-Fuc-(1-->3)[alpha -L-Fuc-OPr (1) and alpha -L-Fuc-(1-->3)[alpha -L-Fuc-(1-->4)]alpha -L-Fuc-OPr (2), which are related to fragments of natural polysaccharides fucoidans, have been synthesized together with constituent disaccharides 3-5, Spectral and conformational properties of tri- and disaccharides have been investigated by (1)H, (13)C and NOE NMR spectroscopy.

Fucoidan fractions from the brown seaweed Chorda filum were studied using solvolytic desulfation. Methylation analysis and NMR spectroscopy were applied for native and desulfated polysaccharides. Homofucan sulfate from C. filum was shown to contain poly-α-(1→3)-fucopyranoside backbone with a high degree of branching, mainly of α-(1→2)-linked single units. Some fucopyranose residues are sulfated at O-4 (mainly) and O-2 positions. Some α-(1→3)-linked fucose residues were shown by NMR to be 2-O-acetylated. The 1H and 13C NMR spectra of desulfated, deacetylated fucan were completely assigned. The spectral data obtained correspond to a quasiregular polysaccharide structure with a branched hexasaccharide repeating unit. Other fucoidan fractions from C. filum have more complex carbohydrate composition and give rather complex methylation patterns.

The red alga Ahnfeltia tobuchiensis was successively extracted six times, and the resulting crude agar extracts were characterized by high-performance size exclusion chromatography. An agar fraction (13.1-42.7%), protein-polysaccharide contaminants (53.4%-77.2%), and low-molecular mass impurities (3.9-9.7%) were found in the extracts. In successive extracts, the degree of methylation of agar molecules was shown to increase, the methyl groups being located only in position 2 of 3,6-anhydro-alpha-L-galactose residues. The differences in molecular mass and sulfate content between the different agar extracts were insignificant. An increase in the methylation degree of agars does not affect their gelation temperature and only decreases the melting point of the agar gels.

Several species of freshwater green algae belonging to the order Zygnematales (Spirogyra crassa (Ktz.) Czurda, S. condensata (Vauch.) Czurda, S. longata (Vauch.) Ktz., S. juergensii Ktz., S. olivascens Rabenh. and Mougeotia viridis (Ktz.) Wittr.) were shown to have specific sterol content and characteristic monosaccharide composition of the biomass. Polysaccharide fractions were obtained by stepwise extraction of S, condensata and characterised by a determination of monosaccharides liberated after acid hydrolysis. Action of amyloglucosidase was used to prove the presence of starch in algal biomass. The main polysaccharide seems to be a complex mucilage composed of rhamnose, arabinose, xylose, galactose, and uronic acid, and the structure of this mucilage needs further investigation.

The structure of sulfated polysaccharide isolated from the calcareous red alga Bossiella cretacea was studied by C-13 NMR and by the methylation analysis, which was carried out both before and after desulfation and the Smith degradation. It was shown that the polysaccharide contains D-galactose, L-galactose, 2-O-methyl-L-galactose, 3-O-methyl-L-galactose, 6-O-methyl-D-galactose, D-xylose, and sulfate. The molar ratio of these components in the acid hydrolyzate was 27 : 20 : 5 : 4 : 1 : 22 : 21, respectively. The molecule of this polysaccharide was found to contain a backbone characteristic of agar galactans composed of alternating 3-linked residues of beta-D-galactopyranose (or 6-O-methyl-beta-D-galactopyranose) and 4-linked residues of alpha-L-galactopyranose (partially 2-O- or 3-O-methylated). All D-xylose residues of the polysaccharide are in pyranose form with the beta-configuration; they represent terminal nonreducing groups attached at position 6 of the beta-D-galactose residues of the backbone. Sulfate groups occupy position 2 of the alpha-L-galactose residues and positions 4 and 6 of the beta-D-galactose residues.

MALDI and FAB mass spectrometry were applied to eight samples of laminarans from different algal species. The existence of both M-chains (mannitol-containing) and G-chains (mannitol-free) was confirmed for six of them, as well as the absence of M-chains for laminarans from two Cystoseira sp. It was found that Cystoseira barbata and C. crinita glucans contain a small percentage of N-acetylhexosamine-terminated chains. This is the first observation of nitrogen-containing sugars in laminarans. The presence of cyclic structures in some laminaran samples is suggested by the MALDI and FAB data. This study demonstrates the power of modern mass spectrometry for defining d.p. profiles in complex mixtures of polysaccharides and for rapidly and sensitively revealing the presence of novel components.

Brown alga Laminaria saccharina was investigated as a potential source of fucoidan. Two samples of alga collected in the White Sea and the Barents Sea showed a qualitatively similar polysaccharide composition and considerably differed only in the content of stock carbohydrates mannitol and laminaran. The polysaccharides were isolated by fractional extraction, and the laminaran and sodium alginate preparations were characterized by C-13 NMR spectra. Fucoidan was found to contain L-fucose and sulfate as major components and galactose, xylose, and glucuronic acid as minor components. It was efficiently desulfated by solvolysis in a DMSO-methanol mixture. The structures of the starting and desulfated polysaccharides were studied by the methylation procedure, periodate oxidation, and NMR spectroscopy. The fucoidan backbone was found to be composed of 1 ---> 3-linked alpha-L-fucopyranose residues sulfated in position 4. It contains single branchings in position 2. Fucoidan possesses anticoagulant properties and efficiently inhibits interaction of P-selectin with its carbohydrate ligands.

This review is concerned with the progress in the methods of structural analysis of sulfated galactans isolated from the red seaweeds, including determination of monosaccharide constituents and partial depolymerization by reductive hydrolysis, identification of disaccharide repeating units by NMR spectroscopy, and sequence analysis by enzymatic degradation. Examples of elucidation of primary structures for several complex sulfated galactans and xylogalactans are given.