Abstract
Treatment of heparin with HONO at pH 1.5 cleaves the polymer at N-sulfated, but not at N-acetylated GlcN residues, and yields di- and tetrasaccharides. The GlcNSO3 residues at the sites of cleavage are converted into anhydromannose (AMan) residues. Reduction of heparin cleavage products with NaB3H4 yields mixtures of di- and tetrasaccharides with reducing terminal [3H]anhydromannitol residues. The identification and quantification of these oligosaccharides by HPLC procedures have been described1,2 These procedures have been used to determine the rates of periodate oxidation of the susceptible unsulfated GlcA and IdoA residues in heparin by measuring the disappearance of the di- and tetrasaccharides that contain GlcA and IdoA. Complete oxidation with IO4 − results in the total loss of the unsulfated uronic acid-containing oligosaccharides, but kinetic studies reported here show that IdoA is oxidized much more rapidly than the major fraction of the GlcA under all reaction conditions. As the pH is lowered from 7 down to 3, the overall rate of the oxidation slows markedly, but the relative rates of GlcA and IdoA oxidation do not change. The slow rate of oxidation of GlcA residues at all pH’s yields oxidation products early in the reaction progress in which all of the unsulfated IdoA residues are oxidized while 70–80% of the pH 1.5 nitrous acid-releasable GlcA→AMan(3,6-(SO4)2) are retained. The anticoagulant activity (APTT) of the partially oxidized product is reduced from 170 IU/mg to 38 IU/mg. Further studies show that the GlcA residue in the antithrombin III binding pentasaccharide is oxidized much more rapidly than the bulk of the GlcA residues in heparin. The results suggest that heparin contains GlcA→AMan(3,6-(SO4)2) sequences that lie outside of the antithrombin-binding pentasaccharide.
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© 1992 Springer Science+Business Media New York
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Conrad, H.E., Guo, Y. (1992). Structural Analysis of Periodate-Oxidized Heparin. In: Lane, D.A., Björk, I., Lindahl, U. (eds) Heparin and Related Polysaccharides. Advances in Experimental Medicine and Biology, vol 313. Springer, Boston, MA. https://doi.org/10.1007/978-1-4899-2444-5_3
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DOI: https://doi.org/10.1007/978-1-4899-2444-5_3
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