Isolation and Optimal Fermentation Condition of the Bacillus subtilis Subsp. natto Strain WTC016 for Nattokinase Production
by
Shouyong Ju 1,2,†, Zhilin Cao 1,3,†, Christina Wong 4, Yangyang Liu 1, Mohamed F. Foda 1,5,6
, Zhenyu Zhang 1,* and Jinshan Li 1,*
Shouyong Ju 1,2,†, Zhilin Cao 1,3,†, Christina Wong 4, Yangyang Liu 1, Mohamed F. Foda 1,5,6, Zhenyu Zhang 1,* and Jinshan Li 1,*
1
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
2
College of Bioengineering, Wuhan Polytechnic, Wuhan 430074, China
3
Department of Resources and Environment, Henan University of Engineering, Zhengzhou 451191, China
4
College of Arts and Sciences, Boston University, Boston, MA 02215, USA
5
Department of Biochemistry, Faculty of Agriculture, Benha University, Moshtohor, Toukh 13736, Egypt
6
College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China
*
Authors to whom correspondence should be addressed.
†
These authors have equally contributed to this work.
Fermentation 2019, 5(4), 92; https://doi.org/10.3390/fermentation5040092
Received: 5 August 2019 / Revised: 15 October 2019 / Accepted: 25 October 2019 / Published: 29 October 2019
Nattokinase is a serine protease in the subtilisin family which is produced by Bacillus subtilis subsp. natto and exhibits vigorous fibrinolytic activity that has been suggested to be able to prevent and treat thromboembolic diseases. In this study, WTC016, a spore-forming and rod-shaped bacterium with fibrinolytic activity was successfully isolated from soil, which was identified as Bacillus subtilis subsp. natto based on morphological and physiological tests, and phylogenetic analysis of 16S rRNA and gyrA. According to the growth curve of WTC016, the nattokinase production reached the highest amount in the stationary phase. To optimize the liquid fermentation condition for nattokinase yield of WTC016, further optimal tests of four factors, including the temperature, pH, inoculum size, and loading volume, followed by orthogonal test of all these factors, was performed. The optimal fermentation conditions were determined as 30 °C, 7.0 pH, 2% inoculum size, and 60 mL of loading volume in 250 mL conical flask, which indicates the highest nattokinase production of 3284 ± 58 IU/mL while fermented for 26 h. This work laid the foundation for producing nattokinase using Bacillus subtilis subsp. natto WTC016.
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Keywords:
nattokinase; Bacillus subtilis subsp. subtilis natto; isolation; optimization of fermentation conditions
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MDPI and ACS Style
Ju, S.; Cao, Z.; Wong, C.; Liu, Y.; Foda, M.F.; Zhang, Z.; Li, J. Isolation and Optimal Fermentation Condition of the Bacillus subtilis Subsp. natto Strain WTC016 for Nattokinase Production. Fermentation 2019, 5, 92.
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