Isolation of gangliosides by cloud-point extraction with a nonionic detergent.
A rapid, simple, and efficient chloroform/methanol-free method of isolating gangliosides is described. The nonionic polyoxyethylene detergent, hexaethyleneglycol mono-n-tetradecyl ether (C14EO6), forms clear micellar solutions in water, but two-phase separation can be achieved by centrifugation in the presence of ammonium sulfate at room temperature. A mixture of pure gangliosides, metabolically labeled gangliosides obtained from cultured hippocampal neurons, and gangliosides from rat cerebellar tissue were quantitatively recovered in the detergent-rich upper (coacervate) phase, with a partition coefficient K > 60. Gangliosides were subsequently separated from the detergent using an Iatrobead column prior to analysis by thin-layer chromatography. The procedure described here is as efficient as other methods of ganglioside extraction, such as that using chloroform/methanol/water/pyridine, but is less time-consuming inasmuch as extraction, purification, and TLC analysis can be completed within 1 day.
Schwarz
A
A
Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.
Terstappen
G C
GC
Futerman
A H
AH
D000818:Animals; D002531:Cerebellum / Q000737:chemistry; D002623:Chemistry Techniques, Analytical / Q000379:methods; D002855:Chromatography, Thin Layer; D003902:Detergents / Q000378:metabolism*; D005026:Ethylene Glycols / Q000378:metabolism*; D005677:G(M1) Ganglioside / Q000302:isolation & purification; D005678:G(M2) Ganglioside / Q000302:isolation & purification; D005679:G(M3) Ganglioside / Q000302:isolation & purification; D005732:Gangliosides / Q000302:isolation & purification*; D006624:Hippocampus / Q000737:chemistry; D009474:Neurons / Q000737:chemistry; D051381:Rats; D013109:Sphingomyelins / Q000378:metabolism; D014316:Tritium / Q000378:metabolism
C091181:4,5-dihydrosphingomyelin; D003902:Detergents; D005026:Ethylene Glycols; D005679:G(M3) Ganglioside; D005732:Gangliosides; D013109:Sphingomyelins; D014316:Tritium; C021298:ganglioside, GD1a; C033851:ganglioside, GD1b; D005678:G(M2) Ganglioside; D005677:G(M1) Ganglioside; C079073:hexaethyleneglycol mono-n-tetradecyl ether; C026226:ganglioside, GD3