High performance liquid chromatography preparation of the molecular species of GM1 and GD1a gangliosides with homogeneous long chain base composition.
A semi-preparative, analytical high performance liquid chromatographic (HPLC) procedure is described for the isolation of molecular species of GM1 and GD1a gangliosides containing a single long chain base, C18 or C20 sphingosine, C18 or C20 sphinganine, each in its natural erythro or unnatural threo form. The threo forms were obtained from 2,3-dichloro-5,6-dicyanobenzoquinone/NaBH4 -treated gangliosides. The ganglioside molecular species separated by HPLC were analyzed for carbohydrate, fatty acid, and long chain base composition. In particular, long chain bases were submitted to gas-liquid chromatographic-mass spectrometric analyses as their trimethylsilyl (TMS) or N-acetyl-TMS derivatives, and chain length, presence or absence of C4-C5 double bond, and C-3 steric configuration were ascertained. The final preparations of individual molecular species of GM1 and GD1a gangliosides were more than 99% homogeneous in their saccharide moiety, contained a single long chain base (homogeneity higher than 99%), and had a fatty acid composition primarily of stearic acid (92 to 97%). All the individual molecular species of GM1 and GD1a gangliosides were also prepared in radioactive form by selective tritiation at C-3 of the long chain base. Their specific radioactivity ranged from 1.3 to 1.45 Ci/mmol. The availability of these molecular species of gangliosides is expected to facilitate studies aimed at ascertaining the role played by the hydrophobic portion in the functional behavior of gangliosides.
D002851:Chromatography, High Pressure Liquid; D005227:Fatty Acids / Q000032:analysis; D005677:G(M1) Ganglioside / Q000032:analysis*; D005732:Gangliosides / Q000032:analysis*; D013058:Mass Spectrometry; D013110:Sphingosine / Q000031:analogs & derivatives / Q000032:analysis
D005227:Fatty Acids; D005732:Gangliosides; C021298:ganglioside, GD1a; D005677:G(M1) Ganglioside; D013110:Sphingosine; C005682:safingol