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Glucose oxidase from Aspergillus niger: Production, characterization and immobilization for glucose oxidation

Jagdish Singh and Neelam Verma

Glucose oxidase (GOx), a flavoenzyme, from Aspergillus niger was produced, purified and immobilized for glucose oxidation. Maximum activities of 0.6 and 0.31U mg-1 fungus dry weight were obtained as intracellular and extracellular respectively. Optimal production (4000 IU/L) was achieved when basal salt medium was supplemented with sucrose (7.5%), peptone (1.5%) phosphorus (0.2%) and MgSO4 (0.2%) at pH 5.7 after 48 h at 250 rev. min–1. Enzyme purified by ammonium sulphate precipitation (75%), gel filtration, Q-Sepharose and DEAE Sepharoses has 30.08, 63.3% and 22.3 fold purification, % recovery and specific activity respectively. Enzyme was dimeric consisting of two equal subunits with molecular weight of 80 kDa. and displayed temperature and pH optima 25- 30°C and 5.5-6.0 respectively for the oxidation of-D-glucose. The enzyme was stable at 50 °C for 1 h without any prior stabilisation. Vmax, and Km value 17 Umg-1 and Km, 7.1mM respectively. GOx was inhibited by Cu2+ (56.5%), and Ag2+ (48%) appreciably and to a lower extent by NaF (25%). Covalent immobilization of enzyme with oxirane beads was less effective and enzyme lost activity, but entrapment methods of sodium alginate and poly acrylamide was effective for the oxidation of glucose