Preparation of pentamer to heptamer chitooligosaccharides by hydrochloric acidic degradation of chitosan

Abstract:

This study outlines a method to prepare chitooligosaccharides with a specific degree of polymerization (5-6). It was carried out by hydrochloric acid hydrolysis in a chitosan degraded system and a following procedure by gel filtration chromatography to fractionate chitosan oligomers. The degradation of chitosan over a range of temperatures (40 ℃, 60 ℃ and 80 ℃) and hydrochloric acid concentrations (6 mol/L, 9 mol/L and 11 mol/L) was investigated. The results showed that chitosan was degraded effectively and various chitooligosaccharide mixtures with different oligomer constitutions were obtained. Among them, the optimum conditions were 9 mol/L HCl at 60 ℃ and the yield of chitopentaose and chitohexose reached 16.2%. A procedure using Sephadex G 15 onto size exclusion column was further developed to narrow the range of oligomers and obtain expected fraction containing mainly chitosan pentamer and hexamer detected by thin layer chromatography. Furthermore, it was identified by high performance liquid chromatography. Alongside peaks for chitopentaose and chitohexaose, a new peak designating an unknown compound occurred. It was identified as chitooligosaccharide by 13C and 1H nuclear magnetic resonance. Enzymatic depolymeriza tion by chitosanase produced mainly trimer and tetramer, further confirming the presence of a heptamer.