Solubility-based Separation and Purification of Long-Chain Chitin Oligosaccharides with an Organic-Water Mixed Solvent.

A simple and rapid method for separation and purification of chitin oligosaccharides, (GlcNAc)n, with n ≥ 5 is presented. A commercially available chitin oligosaccharides sample, consisting of (GlcNAc)n with n = 1 - 7, was used as the starting material. Ten milligrams of the material was mixed with 100 μL of the 1 mol/L HCl. All the (GlcNAc)n species were dissolved in the aqueous medium. The aqueous solution was mixed with 900 μL of EtOH; the mixture was centrifuged, and the supernatant was removed to obtain a precipitate. The precipitate was found to consist mainly of (GlcNAc)n with n ≥ 5, indicating the significant difference in solubility between the short-chain (GlcNAc)n species with n ≤ 3 and the longer ones. By the repetition of the operations, a high purity long-chain (GlcNAc)n sample with n ≥ 5 could be prepared successfully. Since the long-chain (GlcNAc)n species are known to have excellent elicitor activity, this sample would be useful in the study of plant pathology, as well as chitin and chitosan chemistry.