Capillary electrophoresis separation of a mixture of chitin and chitosan oligosaccharides derivatized using a modified fluorophore conjugation procedure

A capillary electrophoresis (CE) method was developed for the simultaneous analysis of small chitin and chitosan oligosaccharides. For detection purposes, the oligomers were derivatized with 8-aminopyrene-1,3,6-trisulfonic acid (APTS), a well known fluorophore for oligosaccharides analysis. The detection was performed by laser-induced fluorescence (LIF) with an argon ion laser having an excitation wavelength of 488 nm and with emission monitored at 520 nm. Derivatization parameters such as reaction time and conditions were examined. Separation conditions were also varied by testing a range of buffer pHs and concentrations. The best conditions were found using an 80 mM borate buffer at pH 8.4. This CE-LIF optimized method was used for the analysis of an enzymatically produced oligo-chitosan sample composed of a complex mixture and having an average degree of polymerization of 3.7 monomer units and 80% deacetylation. The oligo-chitosan sample was treated with a chitin deacetylase-like enzyme, the products were derivatized with APTS, and then analyzed without purification. The goal was to determine whether the deacetylase-like enzyme could increase the extent of deacetylation of the oligo-chitosan sample.