Orsolya Geda1, Tamás Tábi1, Éva Szökő1
1Department of Pharmacodynamics, Semmelweis University, Budapest
Introduction: Gangliosides are glycolipids that are highly abundant in the outer leaflet of specialized membrane microdomains; the lipid rafts. Recent studies indicate that altered concentration may have a role in the development of insulin resistance by altering insulin receptor signaling in the membrane. Gangliosides have heterogeneous molecular structures both in their hydrophobic lipid moiety and their hydrophilic carbohydrate residue which contains one or more sialic acid thus negative charges. They form micelles in aqueous solutions and it was reported previously that cyclodextrins (CDs) are able to break-up the micellar aggregates allowing their separation as monomers by capillary electrophoresis (CE).
Aims: We aimed at developing a CE method capable of the separation of major gangliosides with biological relevance.
Method: Several parameters of CE were investigated to optimize the separation of gangliosides GM3, GM1, GD1a, GD1b and GT1b, including buffer concentration, pH and type and concentration of CDs. The applicability of the methods was also tested on biological samples.
Results: Previously alpha-cyclodextrin (ACD) was used for CE separation of various gangliosides. In our current study ACD was found appropriate for separation of gangliosides with different number of sialic acid but the resolution of GM3 and GM1 and the isomeric GD1a and GD1b was only partial. Using more alkaline pH (10.0) slightly improved their separation as did the increased buffer concentration. However complete resolution of GM3 and GM1 was not achieved. The effect of modified ACD derivatives was also studied. The use of hydroxypropyl-alpha-cyclodextrin (HPACD) resulted in similar resolution as seen with ACD, however methylated-alpha-cyclodextrin (RAMEA) showed improved selectivity for compounds having differences only in the carbohydrate residues.
Conclusion: Using the optimized method with RAMEA separation of all five studied gangliosides was achieved. Furthermore this method was found appropriate for analysis of real biological samples.
Doctoral School: Pharmaceutical Doctoral School
Program: Modern Trends in Pharmaceutical Scientific Research
Supervisor: Prof. Éva Szökő
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