Enhanced thin-layer chromatographic separation of GM1b-type gangliosides by automated multiple development

Enhancement in separation of gangliosides on silica gel precoated high-performance TLC plates has been obtained by automated multiple development chromatography. A less polar mixture of the standard solvent chloroform-methanol-20 mM aqueous CaCl2 (120:85:20, v/v) was used. Lowering the water content achieved separation of two complex monosialoganglioside fractions, isolated from murine YAC 1 T lymphoma and MDAY-D2 lymphoreticular cells. Three-fold chromatography in the solvent chloroform-methanol-20 mM aqueous CaCl2 (120:85:14, v/v) resulted in TLC separation of GM1b-type gangliosides, substituted with C24 and C16 fatty acids and with Neu5Ac and Neu5Gc as well, which could not be achieved by unidirectional standard chromatography. Compared to conventional single chromatography, the technique described allows high-resolution separation of extremely heterogenous ganglioside mixtures and offers a convenient tool for both analytical and preparative TLC.