Isolation of unreduced oligosaccharides
AssigneeMonsanto Company
InventorsThomas W. Rademacher, Raymond A. Dwek
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Abstract
A preparative scale method for the isolation of unreduced oligosaccharides of glycoproteins and glycohormone having a N-linked oligosaccharide structure is disclosed. The method employs hydrazinolysis of the glycoprotein or glycohormone under reaction conditions to cause cleavage at the N-linked sites, producing a mixture having as a major component a de-N-acetylated hydrazone derivative of the oligosaccharides, followed by N-acylation of the hydrazone derivative, acid-catalysis of the hydrazone derivative to produce unreduced oligosaccharides, and subjecting the resulting unreduced oligosaccharides to cellulose column chromatography to remove contaminants and to recover the unreduced oligosaccharides.
Claims1
A method for isolating unreduced oligosaccharides from a glycoprotein or glycohormone having and N-linked oligosaccharide structure comprising the steps of:
- (a) hydrazinolyzing the glycoprotein or glycohormone by heating an essentially water-free and essentially salt-free glycoprotein or glycohormone in the presence of essentially water-free hydrazine under reaction conditions sufficient to cause cleavage at the N-linked sites to produce a mixture having as a major component a de-N-acetylated hydrazone derivative of an oligosaccharide;
- (b) N-acylating the hydrazone derivative of (a) with acylating agent to form a N-acylated hydrazone derivative of said oligosaccharide;
- (c) subjecting the hydrazone derivative of (b) to acid catalysis with a strongly acidic cationic ion exchange resin followed by treatment with a stoichiometric amount of Cu(II) ions to produce said unreduced oligosaccharides;
- and
- (d) subjecting the unreduced oligosaccharides of (c) to cellulose column chromatography to remove contaminants and recover therefrom said unreduced oligosaccharides.
Dependencies are inferred from the claim text.