Shake-flask and bench-scale stirred tank bioreactor production optimization of a thermoalkaline protease from Bacillus cereus SIU1 using one-factor-at-a-time and response surface (statistical) methodologies

Abstract

We report the optimization of production of a halotolerant, thermoalkaline protease by Bacillus cereus SIU1, at shake-flask and bench-scale bioreactor level, using conventional and response surface methods. The basal medium supplemented with optimized (w/v) 0.8% glucose, 1.5% peptone, and 0.4% yeast extract produced 224 Uml− 1 alkaline protease after 20 h incubation. Enzyme yield was further increased to 491 Uml− 1 when the fermentation broth was supplemented with 0.02% (w/v) Ca2+. Optimization of physical factors resulted in still higher protease level of 651 Uml− 1 within 18 h fermentation at initial pH 9.0, 50°C, and 150 rpm agitation. Statistically designed experiments revealed significant effects of peptone and CaCl2 on protease production. A maximum of 749 protease Uml− 1 was produced at optimum factor levels (w/v) of peptone 1.75%, yeast extract 0.4%, CaCl2 0.025%, and pH 9.0 after 18 h incubation. Optimization of agitation and aeration rates in bench-scale bioreactors further enhanced the enzyme yield to 941 protease Uml− 1 at 125 rpm and 2.0 vvm aeration. Optimization of protease production by conventional and statistical approaches resulted in a ∼10.7-fold increase (941 Uml− 1) compared to un-optimized conditions (88 Uml− 1).