Enhanced Production of Thermostable Pullulanase Type 1 Using Bacillus cereus FDTA 13 and Its Mutant

This paper describes optimization of fermentation conditions for the production of thermostable pullulanase from a new strain Bacillus cereus FDTA 13, isolated from tapioca soil. By using one-factor-at-a-time and orthogonal array method, a simple medium with effective and minimal components was optimized for maximal pullulanase production. The optimized medium containing (in g/L): starch 20, yeast extract 5, NaCl 2, MgSO4 0.1 and K2HPO4 0.17 gave nearly 3.6-fold increase in pullulanase activity. Addition of Mn ions in the optimized medium further enhanced the production by 1.4-fold. Effects of proteolytic inhibitors on stability of pullulanase in the medium were studied. A mutant Bacillus cereus FDTA 13/NTG04-B4, obtained by chemical mutation of the parent strain, further increased the pullulanase production by 2-fold in optimized medium, and gave similar yield as the parent strain in a medium with lower levels of carbon source.