José Eduardo Olivo

Evaluation of Supports and Methods for Immobilization of Enzyme Cyclodextringlycosyltransferase

An experimental design with factorial planning was used for the immobilization of the enzyme cyclodextringlycosyltransferase (CGTase) from Bacillus firmus (strain no. 37) to select the best combination of support, method of immobilization, and conditions that gives primarily higher average values for the specific immobilized enzyme activity, and secondarily, higher average values for the percentage of protein fixation. The experimental design factors were as follows: supports-controlled-pore silica, chitosan, and alumina; immobilization methods-adsorption, and two covalent bonding methods, either with gamma-aminopropyltriethoxysilane or hexamethylenediamine (HEMDA); conditions-7 degrees C without agitation and 26 degrees C with stirring. The best combination of factors that lead to higher average values of the response variables was obtained with immobilization of CGTase in silica with HEMDA at 7 degrees C. However, immobilization in chitosan at 7 degrees C gave the highest immobilized CGTase specific activity, 0.25 micromole of beta-CD/ (min mg protein). Physical adsorption gave low specific enzyme activities, and, in general, a high load of enzyme leads to lower specific enzyme activity.

Production of the Enzyme Cyclodextringlycosyltransferase from Bacillus firmus Alkalophilic

Different culture media have been testedfor the production of the enzyme CGTase (cyclodextringlycosyltransferase) from Bacillus firmus (strain #37). The concentration of different carbon and nitrogen sources have been varied and the enzyme activity, cell concentration, reducing sugars, total reducing sugars, soluble protein and pH have been followed during cultivation. Results indicate that higher concentrations of yeast extract and polypeptone lead to increased synthesis of CGTase, whereas when starch is substituted by glucose there is a drastic inhibition of CGTase production.