The fermentation optimization for alkaline protease production by Bacillus subtilis BS-QR-052

Biao Sun; Kai Zou; Yingqing Zhao; Yinfang Tang; Fuming Zhang; Weijing Chen; Xiaoting Tang; Chenran Chang; Yan Zheng

doi:10.3389/fmicb.2023.1301065

The fermentation optimization for alkaline protease production by Bacillus subtilis BS-QR-052

Front Microbiol. 2023 Dec 19:14:1301065. doi: 10.3389/fmicb.2023.1301065. eCollection 2023.

Authors

Biao Sun^{1

2

3}, Kai Zou², Yingqing Zhao⁴, Yinfang Tang¹, Fuming Zhang¹, Weijing Chen³, Xiaoting Tang², Chenran Chang², Yan Zheng²

Affiliations

¹ Zhejiang Cheery Bio-Technology Co., Ltd., Jiaxing, China.
² College of Advanced Materials Engineering, Jiaxing Nanhu University, Jiaxing, China.
³ Zhejiang Institute of Advanced Technology Chinese Academy of Sciences, Jiaxing, China.
⁴ Sinochem Linyi Crop Nutrition Co., Ltd., Linyi, China.

Abstract

Introduction: Proteases exhibit a wide range of applications, and among them, alkaline proteases have become a prominent area of research due to their stability in highly alkaline environments. To optimize the production yield and activity of alkaline proteases, researchers are continuously exploring different fermentation conditions and culture medium components.

Methods: In this paper, the fermentation conditions of the alkaline protease (EC 3.4.21.14) production by Bacillus subtilis BS-QR-052 were optimized, and the effect of different nutrition and fermentation conditions was investigated. Based on the single-variable experiments, the Plackett-Burman design was used to explore the significant factors, and then the optimized fermentation conditions, as well as the interaction between these factors, were evaluated by response surface methodology through the Box-Behnken design.

Results and discussion: The results showed that 1.03% corn syrup powder, 0.05% MgSO₄, 8.02% inoculation volume, 1:1.22 vvm airflow rate, as well as 0.5% corn starch, 0.05% MnSO₄, 180 rpm agitation speed, 36°C fermentation temperature, 8.0 initial pH and 96 h incubation time were predicted to be the optimal fermentation conditions. The alkaline protease enzyme activity was estimated to be approximately 1787.91 U/mL, whereas subsequent experimental validation confirmed it reached 1780.03 U/mL, while that of 500 L scale-up fermentation reached 1798.33 U/mL. This study optimized the fermentation conditions for alkaline protease production by B. subtilis through systematic experimental design and data analysis, and the activity of the alkaline protease increased to 300.72% of its original level. The established model for predicting alkaline protease activity was validated, achieving significantly higher levels of enzymatic activity. The findings provide valuable references for further enhancing the yield and activity of alkaline protease, thereby holding substantial practical significance and economic benefits for industrial applications.

Keywords: Bacillus subtilis; alkaline protease; fermentation optimization; response surface methodology; single-variable experiments.

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was supported by the following grants: Nanhu District Science and Technology Plan Project of Jiaxing City (SQ2020000016), The “14th Five-Year Plan” Teaching Reform Project of Ordinary Undergraduate Universities in Zhejiang Province (jg20220697), Scientific Research Project of Jiaxing Nanhu University (62203ZL), and Research Start-up Funding Project of Jiaxing Nanhu University (QD61220020).