J. Jozefonvicz

Anticoagulant properties of a fucoidan fraction

Fucoïdans are a family of high molecular weight sulphated polysaccharides in the Mr range 8 x 10(5) -10(6), widely dispersed in brown seaweed cell wall. When extracted from several brown algae, they exhibit anticoagulant properties. The chemical degradation of a crude extract, from Pelvetia canaliculata, was undertaken to obtain a low molecular weight polysaccharide (Mr 20,000 +/- 5,000) with the purpose of a possible clinical use. Its anticoagulant potency was investigated through the inhibition of factor IIa and factor Xa in the presence of antithrombin III or heparin cofactor II. The degraded fucoïdan revealed a potent antithrombin activity: studied in an antithrombin III depleted plasma or in the presence of purified heparin cofactor II, the fucoïdan was as efficient as heparin and dermatan sulphate on heparin cofactor II potentiation, at the same concentration by weight. In whole plasma or in the presence of the purified inhibitor, an anti-factor IIa activity mediated by antithrombin III was detected (30 times less potent than for heparin, on a weight to weight basis). In contrast, no anti-factor Xa activity was detected in the presence of the degraded fucoïdan, under the same experimental conditions. These fucoïdans, by-products of alginates preparation in the food and cosmetologic industries, are obtained easily. Thus, they may represent a cheap and easy source of a new type of anticoagulants.

Anticoagulant activity of dextran derivatives. Part I: Synthesis and characterization

Substituted dextrans bearing carboxymethyl and benzylsulphonate groups have been prepared. These materials exhibit an antithrombic activity correlated with the ratio of each substituent. The highest activity is obtained when the dextran derivative contains more than 50% of carboxylic acid groups and simultaneously about 15% of benzylsulphonate functions.

New natural polysaccharides with potent antithrombic activity fucans from brown algae

Fucans extracted from brown algae exhibit anticoagulant properties, as demonstrated by coagulation assays and kinetic study of the inactivation of thrombin in the presence of antithrombin III. These derivatives exert a direct antithrombic activity by an antithrombin III-independent pathway. Besides, their anticoagulant activity is not mediated by a potency to inhibit factor Xa. Relatively low-molecular-mass fractions (50,000 Daltons) prepared by chromatographic fractionation of the crude fucans also exhibit anticoagulant activity. They may prove useful as anticoagulant drugs.

A low molecular weight fucoidan fraction from the brown seaweed Pelvetia canaliculata

Abstract A homogeneous highly purified fucoidan fraction (F2) with a low molecular weight was obtained by hydrolysis of crude fucoidan extracted from the brown seaweed Pelvetia canaliculata. Moreover, this fraction exhibited anticoagulant properties, and with both low viscosity and low molecular weight compatible with further in vivo experiments and clinical use, could be a promising anticoagulant agent.

Inhibition of complement activation by natural sulfated polysaccharides (fucans) from brown seaweed

In the present study, we demonstrate that natural sulfated polysaccharides (fucans) isolated from brown seaweed are potent inhibitors of human complement activation. A fucan fraction of chromatographic molecular weight 22,600, termed BS8, was found to inhibit classical and alternative pathway activation in whole serum in a dose-dependent fashion. Fucan BS8 inhibited formation of the classical pathway C3 convertase by interfering with C1 activation or by inhibiting C4 cleavage and the interaction between C4b and C2. The fucan also inhibited formation/function of the alternative pathway C3 convertase by suppressing the binding of B to C3b and by interfering with the stabilizing function of Properdin. The inhibitory effect of fucans on formation of the C3 convertases was dependent on the molecular weight of the polysaccharide for compounds of chromatographic molecular weight below 16,600. Fucan had no effect on the function of the terminal complex. Since fucans were more efficient than heparin in inhibiting activation of the classical pathway in whole serum and exhibited a lesser specific anticoagulant activity on a molar basis, our results suggest that these natural sulfated polysaccharides have a potential for use as anti-complementary and anti-inflammatory agents.

Dextran-coated silica packings for high-performance size-exclusion chromatography of proteins

Porous silica beads have excellent mechanical properties for the high-performance liquid chromatography of proteins. However, the importance of non-specific interactions between silanols on the silica surface and the proteins requires a modification of these supports before they can be used as stationary phases for size-exclusion chromatography (SEC). Silica beads were coated with dextran bearing a small number of positive charges in order to neutralize the negatively charged silanol groups. Diethylaminoethyl-dextrans (DEAE-dextrans) with a relatively low percentage of dextran units bearing DEAE functions were layered on the silica beads. The influence of several characteristics of these packings on chromatographic performance were studied in order to determine the optimal conditions for the SEC of proteins.

Relationships between chemical characteristics and anticomplementary activity of fucans.

We have shown previously that a low-molecular-weight fucan extracted from the brown seaweed Ascophylum nodosum strongly inhibited human complement activation in vitro and its mechanism of action was largely elucidated. We further investigated the influence of molecular weight and chemical composition of fucan on its anticomplementary activity. The capacity of 12 fragments of fucan (ranging from a molecular weight of 4100 to 214,000) to prevent complement-mediated haemolysis of sheep erythrocytes (classical pathway) and of rabbit erythrocytes (alternative pathway) increased with increasing molecular weight, and reached a plateau for 40,000 and 13,500, respectively. The most potent fucan fractions were 40-fold more active than heparin in inhibiting the classical pathway. They were, however, as active as heparin in inhibiting the alternative pathway. In addition, we have developed a haemolytic test based on the CH50 protocol, which allows discrimination between activators and inhibitors of complement proteins. Although the mannose content within the different fucan fragments did not vary, the galactose and glucuronic acid contents increased with increasing activity, suggesting that these residues should be essential for full anticomplementary activity. Meanwhile, sulphate groups appeared to be necessary, but were clearly not a sufficient requirement for anticomplementary activity of fucans. Taken together, these data illustrate the prospects for the use of fucans as potential anti-inflammatory agents.

Degradation of algal (Ascophyllum nodosum) fucoidan by an enzymatic activity contained in digestive glands of the marine mollusc Pecten maximus

Abstract A protein extract from digestive glands of the marine mollusc Pecten maximus was shown to possess fucoidan-degrading activity. This activity was able to release l -fucose from fucoidan derived from the brown algae Ascophyllum nodosum, and markedly reduce the molecular size of the polysaccharide. An enzymatically degraded fucoidan was produced and analysed by NMR spectroscopy. The 2D 1H NMR data obtained for the first time on low-molecular-weight fractions of algal fucoidan provided new insight into the structure of the polysaccharide. The latter has a randomly organised structure, involving (1→3)- and (1→4)-linked unsulfated and 2-sulfated-α- l -fucose residues.

Molecular weight dependency of the acquired anticomplementary and anticoagulant activities of specifically substituted dextrans

Dextran that had been substituted with carboxylic and benzylamine sulphonated groups was fractionated by gel chromatography into fractions, of narrow molecular weight distribution from 6000 to 190,000 daltons and of similar chemical composition. The fractions exhibited anticomplementary and anticoagulant activities that rapidly increased with molecular weight and tended to plateau above approximately 20,000 and 40,000 daltons respectively. Anticoagulant activity was lower than that of heparin, whereas the capacity of the fractions to inhibit formation of the classical and alternative C3 convertases in a purified system was similar to that of heparin and their ability to inhibit CH50 in whole serum was higher than that of heparin. The data argue for a random distribution of structurally independent anticoagulant and anticomplementary sites along the macromolecular chains of substituted dextrans.

Interactions of biospecific functional polymers with blood proteins and cells.

Biospecific functional polymers, i.e. synthetic or artificial polymers substituted with specific chemical functional groups carried by the macromolecular chain are designed to interact with living systems. These polymers are either insoluble or soluble, derived from polystyrene and dextran. Polymers substituted with aryl sulfonate and carboxyl groups specifically interact with antithrombin III and serine-proteases involved in the coagulation of blood. As a consequence, these polymers possess heparin-like activity and are therefore of low thrombogenicity when exposed to flowing blood. Other functional polymers have been prepared in order to interact with various components of the immune system. Soluble and insoluble functional polymers in contact with cells can affect both cell proliferation and metabolism. Some functional polymers have the ability to inhibit or to stimulate cell growth while others can alter cell function without a change in growth characteristics. The functional polymers described have possible applications as plasma expanders, non-thrombogenic catheters, non-complement activating surfaces and other applications in oncology, biotechnology and immunochemistry.