Fermentation Conditions Of Chitosanase-producing Strain And The Characteristics Of Chitosanase

A strain with high chitosanase activity was isolated from soil samples and identified as Aeromonas sp. The optimal compositions of the medium for chitosanase production by strain YJ02 were as follows(%, w/v): powder chitosan 2.0, glucose 0.1, NH4NO3 1.0, yeast extract 0.5, K2HPO40.07, KH2PO40.03, NaCl 0.5, MgSO4 -TH2O 0.05, the initial pH6.0. The optimal cultivation conditions for chitosanase production by YJ02 were 500mL flask containing 150mL medium, 2%(v/v) amount of inoculation , 30 and shaking cultivation at 150rpm for 72 hours. The chitosanase activity of fermented broth could reach 17.04U/mL when YJ02 was incubated in flask under the optimal cultivition conditions. The chitosanase from YJ02 strain was an inducible and secreted extracellar enzyme. The chitosanase of the strain was extracted by ammonium sulfate precipitation(20-70% saturation) , and purified by gel filtration (Sephadex G-75 colum) and ion-exchange chromatography (Q-Sepharose Fast Flow colum), the results in SDS-PAGE show a single band for the purified chitosanase and the molecular weight was estimated to be 66kD. The optimal temperature and pH for hydrolysis of chitosan by the chitosanase were 50 and 5.6, respectively; The chitosanase activity was stable in the pH range of 4.0-9.0 and the temperature under 50; Mn2+ enhanced the enzyme activity significantly, whereas Cu2+ Fe3+and Hg2+ inhibited the enzyme activity; the enzyme show activity for hydrolysis of chitosan and colloidal chitin, and 60-93% deacetylated chitosan could be degraded most effectively. Chitooligosaccharides were prepared by enzymatic hydrolysis with the chitosanase, the components of the hydrolysis product were analyzed by HPLC, the average molecular weight of hydrolysis product was estimated to be about 3000Da.