SEC-MALS study on aggregates of chitosan molecules in aqueous solvents: Influence of residual N-acetyl groups

The size-exclusion chromatography equipped with a multi-angle light scattering detector (SEC-MALS) method was applied to chitosan samples with different average molecular mass (MM) and degree of N-acetylation (DNAc) values. The influence of these parameters on dispersion state of chitosan molecules in aqueous media were preliminarily examined for conformation analysis. It was shown that even a slight amount of residual N-acetyl groups (as low as 2% DNAc) caused the presence of aggregates or insufficiently dispersed residues of chitosan molecules in aqueous solutions, which gave critical effects on the data for the conformation analysis. Such aggregates could not be removed or prevented by common techniques such as preparative centrifugation (7740g, 10 min), microfiltration (0.1 μm) or lowering ionic strength of the solution. On the other hand, completely deacetylated samples had no such structures, and gave consistent data regardless of their MM values. Therefore, complete deacetylation is highly recommended in advance of the conformation analysis of chitosan, although some depolymerization is unavoidable during the repeated deacetylation treatments.