Macrophage-stimulating activities of a novel low molecular weight saccharide fragment prepared from ascophyllan with alginate lyase

In this study, the polysaccharide ascophyllan, isolated from Ascophyllum nodosum, was degraded using alginate lyase (EC 4.2.2.3) to prepare low-molecular-weight fragments. LMWAs-L, an extremely low-molecular-weight saccharide (6.71 kDa) with low fucose- and sulphate-contents, was purified by gel filtration chromatography. LMWAs-L exhibited significant macrophage-stimulating activities, similar to ascophyllan, through intracellular signalling pathways. Methylation and GC/MS analyses indicated that the main glycosidic linkages of LMWAs-L were 1,2-linked-GlcpA, followed by T-linked-Glcp, 1,3,6-linked-Galp, 1,3-linked-Fucp, 1,4-linked-Xylp, and 1,4-linked-ManpA at a molar ratio of 3.37: 1.13: 1.11: 1.00: 0.96: 0.61. The proposal repeating structure of LMWAs-L was shown to be →2)-α-D-GlcpA-(1 → 2)-α-D-GlcpA-(1 → 6)-α-D-Galp-(1 → 2)-α-D-GlcpA-(1→ as main chain, branched by T-α-D-Glcp-(1 → 4)-β-D-Xylp-(1 → 3)-α-L-Fucp4S-(1→ at the O-3 of 3,6)-α-D-Galp-(1→ residues. 4-deoxy-L-erythro-hex-4-enuronosyluronate-4-β-D-ManpA-(1→ and →4)-β-D-ManpAred residues were attached to the ends of main chain as non-reducing end residue and reducing end residue, respectively. Our results suggest that the fragment LMWAs-L contain an essential structural element of ascophyllan that is responsible for the macrophage-stimulating activities.