SCREENING OF A GROWING CELL IMMOBILIZATION PROCEDURE FOR THE BIOSYNTHESIS OF THERMOSTABLE ALPHA-AMYLASES

Studies were carried out on alpha-amylase production with immobilized cells of two Bacillus strains. High yields of thermostable alpha-amylases were obtained by Bacillus licheniformis 44MB82-G, resistant to glucose catabolite repression and a thermophile Bacillus brevis 174, after repeated batch cultivation (270-600 h) of the immobilized biocatalysts. Various cell immobilization techniques were compared, including entrapment in gel matrices (Ca-alginate, kappa-carrageenan, agar, and their combinations with polyethylene oxide), adsorption on cut disks of polymerized polyethylene oxide, and fixation on formaldehyde activated acrylonitrile-acrylamide membranes. The optimal immobilization parameters (gel and biocatalyst concentration, initial cell quantity) were determined. Among the gels and supports tested, agar, kappa-carrageenan, agar/polyethylene oxide gels, and the membranes were found to be suitable for immobilization and biocatalysts with high operational stabilities were obtained. An enzyme yield of 2750 U/mL culture medium was reached in the fifth repeated batch run with membrane-immobilized Bacillus licheniformis cells. This activity represented 176% of the corresponding yield obtained in batch fermentation with free cells. Higher amylase yields than the activity of the control were reached in all experiments and repeated batch runs with immobilized Bacillus brevis cells.