Combined strong anion-exchange HPLC and PAGE approach for the purificationof heparan sulphate oligosaccharides

Heparan sulphates are highly sulphated linear polysaccharides involved in m any cellular functions. Their biological properties stem from their ability to interact with a wide range of proteins. An increasing number of studies , using heparan sulphate-derived oligosaccharides, suggest that specific st ructural features within the polysaccharide are responsible for ligand reco gnition and regulation. In the present study, we show that strong anion-exc hange HPLC alone, a commonly used technique for purification of heparan sul phate-derived oligosaccharides, may not permit the isolation of highly pure heparan sulphate oligosaccharide species. This was determined by PAGE anal ysis of hexa-, octa- and decasaccharide samples deemed to be pure by strong anion-exchange HPLC. In addition, subtle differences in the positioning of sulphate groups within heparan sulphate hexasaccharides were impossible to detect by strong anion-exchange HPLC. PAGE analysis on the other hand affo rded excellent resolution of these structural isomers. The precise position ing of specific sulphate groups has been implicated in determining the spec ificity of heparan sulphate interactions and biological activities; hence, the purification of oligosaccharide species that differ in this way becomes an important issue. In this study, we have used strong anion-exchange HPLC and PAGE techniques to allow production of the homogeneous heparan sulphat e oligosaccharide species that will be required for the detailed study of s tructure/activity relationships.