Separation of underivatized gangliosides by ion exchange high performance liquid chromatography

A procedure is described which separates neutral glycolipids from gangliosides, and which separates the gangliosides into classes, based on their number of sialic acid residues. In addition to separation into mono-, di- and trisialoganglioside classes, there is purification of individual disialoganglioside species. The procedure uses a commercially available--NH2 high performance liquid chromatography (HPLC) column, which normally is used as an adsorption column. In this method the column is modified by protonation, to pH 5.4, so that it exhibits ion exchange as well as adsorption properties. Sensitive, nondestructive detection of eluent peaks is accomplished by monitoring continuously at 210 nm, a wavelength near which underivatized glycosphingolipids have absorbance maxima and which also will clearly detect most possible contaminants. An entire run, including re-equilibration of the column, takes two hours.