Strain improvement of Bacillus subtilis for enhanced production of nattokinase and optimization of solid-state fermentation conditions

In this study, the nattokinase-producing capacity of Bacillus subtilis subsp. subtilis BS21076 was improved by sequential mutagenization with ultrasonic followed by UV light at the appropriate doses determined according to death and mutation rates. The optimal mutation conditions of strain BS21076 were established as 60 min ultrasonic treatment at 45 kHz and 280 W followed by 120 s UV irradiation at a distance of 30 cm. A high nattokinase-producing mutant strain named BSCZ-4, producing 1.96 times higher nattokinase than BS21076, was obtained after several rounds of primary and secondary screening. After 20 consecutive passages, the diameter of the zone of dissolution produced by BSCZ-4 remained stable in the range of 18-19 mm, suggesting good genetic stability. Using Box-Behnken response surface methodology, the optimal culture conditions for the enhanced production of nattokinase by BSCZ-4 in solid-state fermentation were determined as addition of 5% konjac power to the culture medium, an inoculum size of 8%, and 24 h culture at 34℃, resulting in a nattokinase activity of (4 087.83 ± 93.75) U/g. © 2016, Editorial Office of Food Science. All right reserved.