Hydrophobic interaction chromatography of mucopolysaccharides : Examination of fundamental conditions for fractionation of heparin on hydrophobic gels

Abstract

For hydrophobic interaction chromatography of mucopolysaccharides, some fundamental chromatographic conditions were examined mainly on a combination of Phenyl-Sepharose CL-4B gel and heparin. Every one of the conditions, such as column dimensions, amount of heparin applied, flow-rate, electrolyte and acidity of elution medium, and temperature, influenced the distribution of heparin among the fractions separated on the gel. The solutions of 4.0 M–1.0 M ammonium sulphate in water or in 0.01 M hydrochloric acid were excellent as elution media. Serious temperature effects were generally observed on the interaction between mucopolysaccharides and different types of hydrophobic gel. Commercially available hydrophobic gels of two types were examined: (1) hydrophobic gels without any ionizable function—Phenyl- and Octyl-Sepharose CL-4B gels and Benzyl- and Octyl-Agarose gels, and (2) hydrophobic gels with some ionizable groups, such as isoureide and primary amino groups—Alkyl-Agarose and ω-Aminoalkyl-Agarose gels.