Preparation of N-acetyl-D-glucosamine using enzyme from Aspergillus fumigatus

Abstract

N-acetyl-D-glucosamine (GlcNAc) is one of the basic constituents of mammal cartilage, which is known to help repair deteriorating cartilage and relieve pain and inflammation in osteoarthritis patients. The preparation of GlcNAc in multigram scale was achieved by hydrolyzing squid pen chitin with crude enzyme from Aspergillus fumigatus. The enzyme was obtained by cultivation of the Aspergillus fumigatus in colloidal chitin minimum media at pH 3.5 and 40 C. The highest chitinolytic activity of 8.0  0.4 U/mL was obtained after 3 days of cultivation. The hydrolysis of milled squid pen chitin (5 g) with the crude enzymes gave N-acetyl-D-glucosamine (GlcNAc) over 80% isolated yield within 2 days. The optimum ratio of enzyme to chitin was found to be 22 U/g chitin and the optimum chitin concentration was 2.5% w/w of the reaction volume. A 100 g scale hydrolysis of squid pen chitin at 45 C and pH 4, adjusted with 2 M acetic acid, with 2.2 kU of enzyme gave GlcNAc in 65% isolated yield after 2 days of hydrolysis. The high purity of GlcNAc (100%) was obtained by precipitation of the crude GlcNAc in absolute ethanol from its concentrated charcoal decolorized aqueous solution.