Quantitative analysis of brain gangliosides by high performance

  • H. McCluer
  • Published 2002


This report describes a convenient, highly sensitive, and reproducible HPLC procedure for the quantitative analysis of gangliosides from brain tissues. The procedure involves the conversion of gangliosides to their perbenzoyl derivatives, isolation of the derivatives on a C18-reversed-phase cartridge, separation of the derivatives on a column (3-micron silica) maintained at an elevated temperature, and UV detection of the derivatives at 230 nm. The convenience of the procedure, its sensitivity, reproducibility, and application to the analysis of gangliosides from tissue sources make it the method of choice for ganglioside quantification in our laboratories. Three aspects of the procedure contribute to its convenience: reaction conditions that lead to single products, a convenient isolation procedure for the derivatives, and chromatographic conditions that provide resolution of the derivatives. -Ullman, M. D., and R. H. McCluer. Quantitative analysis of brain gangliosides by high performance liquid chromatography of their perbenzoyl derivatives. J. Lipid Res. 1985. 26: 501-506. Supplementary key words polysialogangliosides HPLC A rapid, sensitive, quantitative HPLC technique would aid in studies of the function, distribution, and metabolism of gangliosides. This study reports such a technique that involves high performance liquid chromatography (HPLC) of perbenzoylated gangliosides. Gangliosides are acidic glycosphingolipids that contain sialic acid. The high concentration of gangliosides in the CNS, as opposed to non-neural tissue, implies that gangliosides play an important role in CNS function. The major gangliosides of mammalian brain are G,, , GD1,, GDlb, and GTlb. These, and other gangliosides, may function in the central nervous system as receptor site determinants (1) or modifiers (2), and in neural transmission (3). Although significant progress has been made in the delineation of ganglioside structure, distribution, and function, that progress has been limited by the available quantitative methods. Current methods for ganglioside analysis include thinlayer chromatography (TLC) (4) or high performance thin-layer chromatography (HF'TLC) (5, 6) followed by destructive densitometry, colorimetry, or gas-liquid chromatography (GLC) (7). The use of ozonolysis to form the p-nitrobenzyloxyamine derivatives, which are separated and quantitated by reversed-phase HPLC, has also been reported (8). The analysis times, limited sensitivity, and variability of these techniques (9), however, restrict the number, sample size, and reproducibility of the ganglioside determinations. Further, even though gangliosides have been quantitated in the picomolar range by HPTLC (6 ) , the destructive detection does not allow further characterization of the separated gangliosides. Convenient and sensitive HPLC methods for the quantification of monosialogangliosides (10) or monosialogangliosides and GD1, (11) as their perbenzoyl derivatives have been reported. However, those techniques do not simultaneously quantify the monoand polysialogangliosides. The method reported here is reasonably simple to perform, highly sensitive, reproducible, and measures monoand polysialogangliosides in a single chromatographic run. It also permits collection of the derivatives for further characterization and analysis. MATERIALS AND METHODS

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@inproceedings{McCluer2002QuantitativeAO, title={Quantitative analysis of brain gangliosides by high performance}, author={H. McCluer}, year={2002} }