Depolymerization and De-N-acetylation of Chitin Oligomers in Hydrochloric Acid

The monosaccharide 2-amino-2-deoxy-d-glucose (glucosamine, GlcN) has recently drawn much attention in relation to its use to treat or prevent osteoarthritis in humans. Glucosamine is prepared from chitin, a process that is performed in concentrated acid, such as hydrochloric acid. This process involves two acid-catalyzed processes, that is, the hydrolysis of the glycosidic linkages (depolymerization) and of the N-acetyl linkages (de-N-acetylation). The depolymerization reaction has previously been found to be much faster compared to the deacetylation, with the consequence that the chitin chain will first be hydrolyzed to the monomer 2-acetamido-2-deoxy-d-glucose (N-acetylglucosamine, GlcNAc) which is subsequently deacetylated. We have found that the chitin disaccharide GlcNAc(1→4)GlcNAc could be completely hydrolyzed to the monosaccharide GlcNAc with negligible concomitant de-N-acetylation, and the chitin disaccharide and monosaccharide were further used to study the depolymerization reaction and the de-N-acetylation reaction, respectively. The reactions were performed in hydrochloric acid as a function of acid concentration (3−12 M) and temperature (20−35 °C), and 1H− NMR spectroscopy was used to monitor the reaction rates. The 1H NMR spectrum of GlcNAc in concentrated (12 M) and deuterated hydrochloric acid at 25 °C was assigned. The glucofuranosyl oxazolinium (3) ion was found to exist in equilibrium with the α- and β-anomers of the pyranose form of GlcNAc, where 3 was present in half the total molar concentrations of the two anomeric forms of GlcNAc. At lower acid concentration (3−6 M), only trace concentrations of 3 could be detected. The rate of de-N-acetylation of GlcNAc was determined as a function of hydrochloric acid concentration, showing a maximum at 6 M and decreasing by a factor of 2 upon decreasing or increasing the acid concentration to 3 or 12 M. The activation energy for hydrolysis of the N-acetyl linkage of GlcNAc was determined to be 102 ± 7, 116 ± 8, and 110 ± 8 kJ mol-1 at 3, 6, and 12 M hydrochloric acid concentration, respectively. The results are in accordance with the proposed SN2 reaction mechanism of the acid-catalyzed hydrolysis of the N-acetyl linkage where the rate-limiting step is the addition of water to the carbonium ion. The 1H NMR spectrum of the dimer GlcNAc−GlcNAc in concentrated (12 M) and deuterated hydrochloric acid at 25 °C was assigned. The rate of the acid-catalyzed cleavage of the glycosidic linkage of the dimer