Elucidation of substituent distribution states for carboxymethyl chitosan by detailed NMR analysis

Abstract

A series of carboxymethyl chitosan (CMCh) samples with a wide degree of substitution (0.65–2.03) was prepared by multi-step carboxymethylation of chitosan using a 2-propanol/aqueous sodium hydroxide solution mixture as the solvent and monochloroacetic acid as the etherification reagent. Two-dimensional NMR correlation maps of the direct and long-range coupled 1H and 13C spin pairs of the CMCh samples allowed complete assignment of the 1H and 13C chemical shifts of eight anhydroglucosamine units (AGUs) comprising CMCh chains, namely un-, 2-mono-, 3-mono-, 6-mono-, 2,3-di-, 2,6-di-, 3,6-di-, and 2,3,6-tri-substituted AGUs. Line-shape analysis of the anomeric carbon resonance in the quantitative 13C NMR spectra determined the mole fractions of these AGUs. Plots of the mole fractions against the change in the degree of substitution clarified the detailed substitution state at the 2-amino and 3-/6‑hydroxyl groups of chitosan and the transition of the monomer fractions during the change from the unsubstituted to the fully substituted state. Therefore, the method estimating the mole fractions of the eight AGUs in CMCh is expected to be an effective and powerful tool for elucidating functionalities from a structural perspective.

Keywords

• Carboxymethyl chitosan
• Substituent distribution
• Monomer mole fraction
• NMR