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浙江大学学报（农业与生命科学版）
浙江大学学报(农业与生命科学版)  2016, Vol. 42 Issue (04): 485-494    DOI: 10.3785/j.issn.1008-9209.2015.08.291
资源与环境科学     
利用味精废液发酵枯草芽孢杆菌的培养基配方优化
刘丽1,2， 曾真1， 方萍1.3*
1.浙江大学环境与资源学院，污染环境修复与生态健康教育部重点实验室，杭州 310058；2.贵州大学农学院农业资源与环境系，贵阳 550025；3.浙江大学环境与资源学院，浙江省亚热带土壤与植物营养重点研究实验室，杭州 310058
Optimization of a culture medium for Bacillus subtilis based on monosodium glutamate wastewater
LIU Li1,2, ZENG Zhen1, FANG Ping1,3*
(1. Key Laboratory of Environment Remediation and Ecological Health of the Ministry of Education, College of Environmental and Resource Sciences, Zhejiang University, Hangzhou 310058, China; 2. Department of Agricultural Resource and Environment, College of Agriculture, Guizhou University, Guiyang 550025, China; 3. Zhejiang Provincial Key Laboratory of Subtropical Soil and Plant Nutrition, College of Environmental and Resource Sciences, Zhejiang University, Hangzhou 310058, China)
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摘要： 以营养肉汤（nutrient broth，NB）培养基为对照，通过对比试验、正交试验和单因素试验，对以味精废液为主要营养源的摇瓶培养的枯草芽孢杆菌F-2培养基配方及培养条件进行优化，以提高F-2发酵液的活菌密度并实现味精废液的资源化利用。对比试验表明，用12.5 g/L浓缩味精废液（concentrated monosodium glutamate wastewater，CMGW）培养的F-2菌悬液的D(600 nm)值及活菌密度显著高于NB培养基，其培养F-2后的氨基酸含量显著降低。通过L16（43×26）正交试验筛选出F-2的优化配方为CMGW 12.5 g/L，牛肉膏1.0 g/L，蛋白胨4.0 g/L，MnSO4·2H2O 0.5 g/L，H3BO3 0.02 g/L，FeSO4·7H2O 0.1 g/L，MgSO4·7H2O 0.5 g/L。按此优化配方接种培养F-2菌株，其菌液的活菌密度分别是未经优化的CMGW培养基和NB培养基的2.9倍和6.3倍。通过单因素试验，筛选出基于该优化配方的F-2菌株适宜的初始pH范围为6.5~7.5，适宜的培养温度为30~35 ℃。以上结果显示，培养基CMGW对菌株F-2的发酵效果优于NB培养基，其优化配方的效果更佳。
Abstract: Concentrated monosodium glutamate wastewater (CMGW) generated from the production of monosodium glutamate is an organic wastewater with high concentration of ammonia, chemical oxygen demand, biochemical oxygen demand and SO42-and low pH. Discharge of CMGW has raised serious environmental problems, and potential secondary pollution existed even treated with traditional physical and chemical processes. It has already been reported that the richness of nitrogen and carbon makes the recycling of this wastewater possible in the way of microbial fermentation as medium. However, the differences of strain and fermentation purpose require that the medium contains different nutritional compositions with a certain dosage. Bacillus subtilis F-2, isolated from a commercial organic fertilizer, can inhibit the growth of 18 plant pathogenic fungi with varying degrees, especially in F. oxysoporum f.sp niweum with inhibition rate of 93%, which reveal the potential to prepare bio-organic fertilizer under biocontrol. In addition, F-2 also plays an important role in decomposition of organic waste. The study on degradation of food waste has pointed to its dominance in microbial community. Therefore, F-2 as a prospective functional bacteria in agriculture and environment, it is necessary to improve F-2 viable cell production as much as possible. Massive studies were focused on medium optimization of B. subtilis to enhance its cell density in fermentation liquor. However, the main ingredients of medium used in most of these studies were peptone, yeast extracts, beef extracts, soy or corn flour, associated with great cost and excessive consumption of resources. The purpose of this study was to investigate a cost-effective resource utilization of CMGW by increasing the density of B. subtilis F-2 viable cells in cultures. The CMGW medium which was 80-fold diluent of the wastewater, i.e., 12.5 g/L CMGW, was served as the main carbon and nitrogen sources of strain F-2. Contrast experiments of shaking cultures, orthogonal and single factor experiments were applied to find the optimal CMGW medium formula and its culturing condition.  The contrast experiments showed that the D(600 nm) value of F-2 suspension and viable cell density of the CMGW medium was significantly higher than that of nutrient broth (NB) medium, and the amino acid content in the CMGW medium decreased significantly after culturing. The optimal formula identifying F-2 by the orthogonal experiment of L16(43×26) was as follows: 12.5 g/L CMGW, 1.0 g/L beef extracts, 4.0 g/L peptone, 0.5 g/L MnSO4·2H2O, 0.02 g/L H3BO3, 0.1 g/L FeSO4·7H2O and 0.5 g/L MnSO4·7H2O. The viable cell density in the optimized CMGW medium was 2.9 and 6.3 times of the CMGW medium and NB medium, respectively. Furthermore, according to the results of single factor experiment, the proper initial pH and culturing temperature of the optimized CMGW medium were screened as pH 6.5-7.5 and 30-35 ℃. In conclusion, the CMGW medium is an ideal substitute for NB medium in the fermentation of B. subtilis F-2, providing a practicable and promising approach to reuse and recover CMGW. Moreover, the CMGW medium is superior to NB medium, and the fermentation effect is even better after the medium and culturing condition are optimized.
出版日期: 2016-07-20
CLC:  TQ 920.6  
通讯作者: 方萍（http://orcid.org/0000-0001-7784-7873），Tel:+86-571-88982480,E-mail:pfang@zju.edu.cn   
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引用本文:

刘丽，曾真，方萍. 利用味精废液发酵枯草芽孢杆菌的培养基配方优化[J]. 浙江大学学报(农业与生命科学版), 2016, 42(04): 485-494.

LIU Li, ZENG Zhen, FANG Ping. Optimization of a culture medium for Bacillus subtilis based on monosodium glutamate wastewater. Journal of Zhejiang University (Agriculture and Life Sciences), 2016, 42(04): 485-494.

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http://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2015.08.291    或    http://www.zjujournals.com/agr/CN/Y2016/V42/I04/485

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