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Method for fermentation production of succinic acid by circulating utilization of recombinant Bacillus coli cells

A technology of recombinant Escherichia coli and cell fermentation, which is applied in the field of bioengineering, can solve problems such as the decrease of yield and production intensity, the decrease of cell density and specific production intensity of cells, and prolong the production time of succinic acid, so as to improve the specific production intensity , the effect of prolonging the production time

Inactive Publication Date: 2013-06-19
NANJING TECH UNIV
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  • Description
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Problems solved by technology

But if you take into account the sugar consumed and the time spent in the aerobic stage, the yield and production intensity will drop
Andersson et al. recovered anaerobic fermentation cells in time and transformed fresh medium to continue anaerobic succinic acid production, which relieved product inhibition and extended the production time of succinic acid. Compared with two-stage fermentation in the same time period, the product concentration increased 60%; but during the process, the density of bacteria and the specific production strength of cells are constantly decreasing (Bioprocess and Biosystems Engineering.2009, DOI10.1007 / s00449-009-0393-y)

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  • Method for fermentation production of succinic acid by circulating utilization of recombinant Bacillus coli cells
  • Method for fermentation production of succinic acid by circulating utilization of recombinant Bacillus coli cells
  • Method for fermentation production of succinic acid by circulating utilization of recombinant Bacillus coli cells

Examples

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Embodiment 1

[0028] This example illustrates that Escherichia coli AFP111 can use metabolites (succinic acid, acetic acid, etc.) in the culture medium to maintain cell growth and function during the aerobic induction process after the two-stage fermentation. Its specific culture conditions and steps are as follows:

[0029] 1. Production of succinic acid by two-stage fermentation: first, LB culture medium activates the thallus, inserts the inoculum into the Erlenmeyer flask from the cryopreservation tube according to 1% (v / v), and cultivates the thallus for about 12 hours as the seed liquid; Inoculum size 3% was transferred to JSM fermentation medium, and glucose was added to an initial concentration of 25g L -1 , cultured aerobically until the dry weight of the cells was 10-12g·L -1 , the dissolved oxygen in the cultivation process was maintained above 40%; 2 Gas, for anaerobic fermentation to produce acid, adopt the method of adding glucose in batches, and the maximum concentration sha...

Embodiment 2

[0041] In this example, Escherichia coli AFP111 cells that have been induced to increase the activity of key enzymes in the pathway of succinic acid synthesis were used to ferment and produce succinic acid in fresh medium. The specific steps are as follows:

[0042] 1. Production of succinic acid by two-stage fermentation: first, LB culture medium activates the thallus, inserts the inoculum into the Erlenmeyer flask from the cryopreservation tube according to 1% (v / v), and cultivates the thallus for about 12 hours as the seed liquid; Inoculum size 3% was transferred to JSM fermentation medium, and glucose was added to an initial concentration of 25g L -1 , cultured aerobically until the dry weight of the cells was 10-12g·L -1 , the dissolved oxygen in the cultivation process was maintained above 40%; 2 Gas, for anaerobic fermentation to produce acid, adopt the method of adding glucose in batches, and the maximum concentration shall not exceed 50g L -1 , while basic magnesiu...

Embodiment 3

[0046] In this example, Escherichia coli AFP111 cells that have been induced to increase the activity of key enzymes in the pathway of succinic acid synthesis were used to recycle the cells to ferment and produce succinic acid in fresh medium. The specific steps are as follows:

[0047] 1. Production of succinic acid by two-stage fermentation: first, LB culture medium activates the thallus, inserts the inoculum into the Erlenmeyer flask from the cryopreservation tube according to 1% (v / v), and cultivates the thallus for about 12 hours as the seed liquid; Inoculum size 3% was transferred to JSM fermentation medium, and glucose was added to an initial concentration of 25g L -1 , cultured aerobically until the dry weight of the cells was 10-12g·L -1 , the dissolved oxygen in the cultivation process was maintained above 40%; 2 Gas, for anaerobic fermentation to produce acid, adopt the method of adding glucose in batches, and the maximum concentration shall not exceed 50g L -1 ,...

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Abstract

The invention relates to a method for the fermentation production of succinic acid by the circulating utilization of recombinant Bacillus coli cells, which is characterized by comprising the following steps: adopting the two-stage fermentation mode of first aerobic culture of Bacillus coli and later anaerobic acid production to produce succinic acid by utilizing Bacillus coli for producing succinic acid; when the two-stage fermentation is finished, utilizing metabolites (succinic acid, acetic acid, pyruvic acid and the like) produced in the fermentation process to aerobically induce cells; aseptically collecting Bacillus coli cells, and anaerobically biotransforming and synthesizing the succinic acid in a fresh culture medium; after finishing the anaerobic biotransformation and synthesis,aerobically inducing again, and recovering the cells to continue to biotransform and synthesize the succinic acid in the fresh culture medium, and achieving the purpose of circulatingly utilizing thecells to biotransforme and synthesize the succinic acid. The technical scheme of the invention solves the problems that the yield and production intensity of the succinic acid is on the low side in the whole process of two-stage fermentation due to the consumption of a great deal of carbon sources and time in the process of aerobic Bacillus coli culture.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and relates to a method for recycling recombinant Escherichia coli cells to ferment and produce succinic acid, in particular to using metabolites produced during two-stage fermentation to induce and improve intracellular enzyme activity of recombinant Escherichia coli cells, and A method for recycling cells that have increased intracellular enzyme activity to produce succinic acid. Background technique [0002] Succinic acid (succinic acid) is widely used in industries such as medicine, pesticides, dyes, spices, paints, food and plastics, and as an excellent C4 platform compound, it can be used to synthesize 1,4-butanediol, tetrahydrofuran, γ- Organic chemicals such as butyrolactone and biodegradable materials such as polybutylene succinate (PBS) are considered by the US Department of Energy to be among the 12 most valuable biorefinery products in the future. [0003] The production metho...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P7/46C12R1/19
CPCY02P20/142Y02P20/141
Inventor 姜岷马江锋陈可泉应汉杰韦萍欧阳平凯
Owner NANJING TECH UNIV
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