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Menaquinone-7 production from maize meal hydrolysate by Bacillus isolates with diphenylamine and analogue resistance
Jian-Zhong Xu1, Wei-Guo Zhang1
1The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China.
Abstract:
A menaquinone-7 (MK-7) high-producing strain needs to be isolated to increase MK-7 production, in order to meet a requirement of MK-7 given the low MK-7 content in food products. This article focuses on developing MK-7 high-producing strains via screening and mutagenesis by an atmospheric and room temperature plasma (ARTP) mutation breeding system. We isolated an MK-7-producing strain Y-2 and identified it as Bacillus amyloliquefaciens, which produced (7.1±0.5) mg/L of MK-7 with maize meal hydrolysate as carbon source. Then, an MK-7 high-producing strain B. amyloliquefaciens H.β.D.R.-5 with resistance to 1-hydroxy-2-naphthoic acid, β-2-thienylalanine, and diphenylamine was obtained from the mutation of the strain Y-2 using an ARTP mutation breeding system. Using strain H.β.D.R.-5, efficient production of MK-7 was achieved ((30.2±2.7) mg/L). In addition, the effects of nitrogen sources, prenyl alcohols, and MgSO4 on MK-7 production were investigated, suggesting that soymeal extract combined with yeast extract, isopentenol, and MgSO4 was beneficial. Under the optimized condition, the MK-7 production and biomass-specific yield reached (61.3±5.2) mg/L and 2.59 mg/L per OD600 unit respectively in a 7-L fermenter. These results demonstrated that strain H.β.D.R.-5 has the capacity to produce MK-7 from maize meal hydrolysate, which could reduce the substrate cost.
Insights
Researchers developed a high-yield menaquinone-7 (MK-7) producing strain, Bacillus amyloliquefaciens H.β.D.R.-5, using atmospheric and room temperature plasma (ARTP) mutagenesis. This strain significantly increased MK-7 production for potential food fortification applications.
Area of Science:
- Biotechnology
- Microbiology
- Biochemical Engineering
Background:
- Menaquinone-7 (MK-7) is essential but deficient in many foods.
- Current MK-7 production methods require enhancement to meet demand.
- Developing high-yield MK-7 producing strains is crucial for industrial applications.
Purpose of the Study:
- To isolate and develop a high-yield menaquinone-7 (MK-7) producing strain.
- To optimize fermentation conditions for enhanced MK-7 production.
- To evaluate the feasibility of using low-cost substrates for MK-7 biosynthesis.
Main Methods:
- Screening and isolation of MK-7 producing Bacillus strains.
- Utilizing atmospheric and room temperature plasma (ARTP) for mutagenesis and strain improvement.
- Optimization of fermentation parameters including carbon and nitrogen sources, and additives.
- Fermentation in a 7-L bioreactor to assess large-scale production potential.
Main Results:
- Isolated Bacillus amyloliquefaciens Y-2 producing 7.1±0.5 mg/L MK-7.
- Developed a mutant strain, H.β.D.R.-5, exhibiting enhanced MK-7 production (30.2±2.7 mg/L).
- Optimized conditions using soymeal extract, yeast extract, isopentenol, and MgSO4 increased MK-7 yield to 61.3±5.2 mg/L in a 7-L fermenter.
Conclusions:
- The engineered Bacillus amyloliquefaciens H.β.D.R.-5 strain demonstrates significant potential for high-level MK-7 production.
- Optimized fermentation using cost-effective substrates like maize meal hydrolysate is viable.
- This study provides a foundation for industrial-scale MK-7 production to address dietary deficiencies.
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