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Methylotrophic bacillus and application thereof

A methylotrophic, bacillus-based technology applied to bacteria, microbe-based methods, microorganisms, etc.

Active Publication Date: 2014-12-17
青岛诺森生物技术有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The reported producing bacteria are mainly Bacillus subtilis, etc., and there is no relevant report on the preparation of acetoin by methylotrophic bacillus. The present invention provides a new resource for the preparation of acetoin by microbial fermentation

Method used

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  • Methylotrophic bacillus and application thereof
  • Methylotrophic bacillus and application thereof
  • Methylotrophic bacillus and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Embodiment 1: the screening of bacterial strain

[0020] Grind medium-high temperature Daqu (taken from Anhui Shuanglun Employment Co., Ltd.) into powder, accurately weigh 25 g and add it to a conical flask containing 200 ml of sterile normal saline. Add sterilized glass beads and put them on a shaker (180r / min) at room temperature for 30 minutes to shake and disperse, then take them out and let them stand for 30 minutes. Take 1 mL of supernatant and add 9 mL of sterile saline to prepare a bacterial suspension with a volume concentration of 10-1. By analogy, the volume concentration is prepared to be 10 -3 、10 -4 ...10 -7 bacterium suspension, evenly spread on the separation medium that has been cooled overnight (the preparation method of the separation medium is: beef extract 3g / L, peptone 10g / L, NaCl 5g / L, agar strip 20g / L, water 1000mL , mixed with nystatin 0.01g / L, adjusted the pH to 7.0, sterilized at 121°C for 20min.) on the plate, and placed the inoculated pl...

Embodiment 2

[0021] Example 2: Fermentation of bacterial strains

[0022] 1. Fermentation of strains

[0023] Pick the bacterial strain from the slope to the seed medium (the preparation method of the seed medium is: beef extract 20.0g, glucose 20.0g, K 2 HPO 4 1.0g, (NH 4 ) 2 SO 4 1.0g, MgSO 4 ·7H 2 O 1.0g, NaCl 0.5g, FeSO 4 ·7H 2 O0.01g, mix well, add distilled water to adjust to 1L, adjust pH to 7.0, sterilize at 121°C for 20min. ) to the logarithmic phase, and the bacterial strain that will be cultivated to the logarithmic phase with the seed medium is added to the fermentation medium by 5% inoculum (the preparation method of the fermentation medium is: beef extract 20.0g, glucose 20.0g, K 2 HPO 4 1.0g, (NH 4 ) 2 SO 4 1.0g, MgSO 4 ·7H 2 O1.0g, NaCl 0.5g, FeSO 4 ·7H 2 O 0.01g, mix well, add distilled water to adjust to 1L, adjust pH to 7.0, sterilize at 121°C for 20min. ), and then cultured with shaking at 37°C and 150r / min for 72h. The fermentation broth was collect...

Embodiment 3

[0026] Embodiment 3: the detection of acetoin

[0027] Gas chromatographic detection conditions: the fermentation broths of the different strains obtained in Example 2 were diluted 5 times respectively, and then 2 μL samples were taken for gas chromatographic analysis to detect the output of acetoin. After testing, the highest yield of acetoin was 15.71g / L.

[0028] Gas chromatographic parameters: CP-Wax 57C column for liquor, inner diameter: 250.00μm, length: 50.0m, liquid film thickness: 0.20μm, dead time: 2.983min, column temperature: 60°C for 5min, 5°C / min temperature program Keep at 140°C for 2min, inlet temperature: 150°C, detector temperature: 160°C, carrier gas: N2, column pressure 90kPa, makeup: 30mL / min, split ratio: 1:30, air: 50kPa, hydrogen : 50kPa.

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Abstract

The invention provides methylotrophic bacillus which can produce acetoin at high yield. The methylotrophic bacillus is collected in the China General Microbiological Culture Collection Center (CGMCC) and has the collection number of CGMCCC No.7485. When the methylotrophic bacillus is fermented under a certain conditions, acetoin with the concentration of 15.71g / L can be obtained. The production capacity of most of the reported acetoin producing bacteria is lower than or equivalent to that of the methylotrophic bacillus CGMCCC No.7485; however, according to the strain, except the glucose concentration is preliminarily optimized, other culture conditions and medium components are not subjected to breeding transformation. Thus, after the methylotrophic bacillus is subjected to further breeding transformation and condition optimization, the acetoin yield is further improved. Therefore, the methylotrophic bacillus has wide application prospects.

Description

technical field [0001] The invention belongs to the technical field of microorganisms and fermentation engineering, and in particular relates to a methylotrophic bacillus. Background technique [0002] Acetoin is also known as 3-hydroxybutanone, acetic acid, and methyl acetylmethanol. Existing in dairy products and some fruits, it is a widely used food spice, and it is allowed to be eaten according to GB 2760-86 in my country. As a spice, acetoin has a wide range of applications and a relatively large amount. Its main role is in the production of flavors such as cream, dairy products, liquor flavoring, yogurt and strawberry. In addition, 3-hydroxybutanone can also be used as a platform compound, which is widely used in many other industries. In 2004, the US Department of Energy listed it as one of the 30 priority platform compounds for development and utilization. [0003] At present, the production of acetoin at home and abroad is mainly through chemical synthesis, but th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P7/26C12R1/07C12N1/20
Inventor 薛正莲刘阳章宁娟王洲苏燕南
Owner 青岛诺森生物技术有限责任公司