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The studies were carried out on 12 UV-mutants of B. subtilis B-3 which synthesise from 18 to 105% more hydrolytic enzymes (proteinases, amylases, cellulases, and lipases) than the wild strain. After five years' storage of the mutants, a 26-fold increase in amylase biosynthesis was observed for a G7 UV-mutant. This mutant showed also other phenotypic differences. The increased ability of mutant G7 to synthesise amylases was controlled by the maltose-substrate induction.
The activity of amylases obtained under chemostat conditions was found between 92 to 118 U/mL, but this highly effective biosynthesis was a short-lived process. Continuous amylase synthesis in a packed-bed reactor (carried out for 144 h) by the bacteria immobilized in calcium alginate resulted in slow reduction of cell productivity of amylases ranging from 69.5 to 44.7 U/g of biocatalyst. Amylase biosynthesis in this process was to a large extent dependent on the dilution rate (D). The highest effectiveness of bacteria production (Yp/x= 36.1 U/g beads) was obtained at D=0.8 h-1, while the highest volumetric productivity Qp= 98 kU/Lh was observed at D=6.4 h-1. The bacteria cells immobilized in chitosane gel beads exhibited high yield (70-73 U/g beads) for 96 h of incubation, which was followed by a rapid decrease.
The aim of the study was to determine optimal conditions for amylases biosynthesis by immobilized Bacillus polymyxa В-20 cells. Composition of the medium and immobilization conditions to a large extent affected the efficiency of enzyme production. Optimal production medium consisted of yeast extract (5-10 g/L), maltose, peptone and mineral salts. Stability of alginate biocatalyst beads was increased by 30 g/L gel concentration, increased bead size and chemical gel modification by glutaraldehyde. Increased quantities of synthesized enzymes were proportional to the increment of initial biomass present in beads, and to a lesser degree, depended on quantities of immobilized inoculum. In repeated batch culture, the bacteria immobilized in alginate were able to synthetize 32% more enzymes than those immobilized in chitosane. Operational stability of alginate and chitosane beads was maintained for 17 days.
Keratinolytic abilities of Bacillus polymyxa B20 and B. cereus B5esz were evaluated in liquid cultures in mineral media containing chicken feathers. Both tested strains were capable of effective liquefying and biodegradation of feather keratin, up to 56.5 – 72.1% in ten-day cultures, releasing considerable amounts of hydrolysis products. Tested bacteria were mesophilic species, producing highest activity of keratinases and proteases in the presence of keratin (1%) as a sole nutrient source or keratin supplemented with yeast extract, at 30℃. Keratinases of B. polymyxa were predominantly highly alkaline serine proteases, with optimum activity at 50℃, while B. cereus produced mainly a mixture of neutral proteases, optimally active at 45℃. Keratinolytic potential of tested bacterial strains could find a variety of applications, including utilization of keratinous waste from poultry industry and obtaining keratin hydrolysate-based soil fertilizers.
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