Morgan Stefansson

Resolution of the branched forms of oligosaccharides by high-performance capillary electrophoresis

The heterogeneous nature of most polysaccharides found in nature includes distribution in molecular weight, primary sequence, and branching. The analytical methodology used in the characterization of these structural aspects must ensure high separation efficiency and selectivity. This paper reports on the high-performance capillary electrophoresis (HPCE) separation of branched forms of oligosaccharides as well as some variants in the primary structure. Oligosaccharide maps were obtained after selective debranching using isoamylase, laminarinase, and cellulase enzymes. The samples investigated were alpha-D-glucans (amylose, amylopectin, and pullulan) and beta-D-glucans (exemplified by lichenan). The solutes were separated as fluorescent derivatives with 8-aminonaphthalene-1,3,6-trisulfonate (ANTS) and detected by laser-induced fluorescence at 514 nm using a He/Cd laser (excitation at 325 nm). The number of theoretical plates was in excess of one million per meter. Baseline resolution of oligosaccharides with a degree of polymerization approximately 70 was obtained within 15 min using borate buffer as the electrolyte.

High-resolution studies of hyaluronic acid mixtures through capillary gel electrophoresis

Hyaluronic acid is a negatively charged polysaccharide with a high degree of polydispersity that makes the separation of its oligomers extremely difficult. Through the use of columns filled with a highly viscous polyacrylamide matrix, the unit resolution of hyaluronate oligomers could be achieved, up to at least 80 kDa of mass, through capillary electrophoresis. As analytical application examples, the fractions of enzymatically or ultrasonically degraded hyaluronates were monitored through this method. Because of the very high resolving power, peaks additional to the regular oligomers can be observed that are assumed to be conformers of this regular, unbranched biopolymer.