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Clostridium butyricum and application of clostridium butyricum in immobilized fermentation production of 1, 3-propylene glycol

An immobilized fermentation, Clostridium butyricum technology, applied in fermentation, microorganism-based methods, bacteria, etc., can solve the problems of obvious inhibition, low substrate glycerol tolerance, low production intensity, etc.

Active Publication Date: 2021-02-12
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the biggest limitation of this bacterium is that the cell growth is slow under strict anaerobic conditions, resulting in low production intensity, and the cells have low tolerance to the substrate glycerol, especially the cheap substrate crude glycerol has obvious inhibitory effect on fermentation

Method used

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  • Clostridium butyricum and application of clostridium butyricum in immobilized fermentation production of 1, 3-propylene glycol
  • Clostridium butyricum and application of clostridium butyricum in immobilized fermentation production of 1, 3-propylene glycol
  • Clostridium butyricum and application of clostridium butyricum in immobilized fermentation production of 1, 3-propylene glycol

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1, Clostridium butyricum isolation and screening

[0036] Screening method: directly inject 5ml of the collected water sample into 45ml enrichment medium (enrichment medium composition: 41ml of liquid A, the composition is glucose 10g / L; 1ml of liquid B, the composition is tripotassium citrate 100g / L, Citric acid monohydrate 62.5g / L, Na 2 SO 4 50g / L, KH 2 PO 4 50g / L, NaHCO 3 125g / L; C solution 1ml, the composition is urea 250g / L, NH 4 Cl75g / L, yeast extract 50g / L; D liquid 1ml, the composition is cysteine ​​hydrochloride 50g / L, MgCl 2 ·6H 2 O50g / L, CaCl 2 2H 2 O10g / L, FeCl 2 4H 2 O5g / L; E liquid 1ml, the composition is pyridoxamine hydrochloride 1g / L, p-aminobenzoic acid 0.2g / L, Biotin0.1g / L, vitamin B12 0.2g / L, vitamin B1 0.2g / L), 37 ℃ After culturing for 2-4 days, dilute the culture medium to 10 -3 , spread on the screening plate (600ml in total, wherein A solution 450ml, composition is 3g glucose, 9g agar and 0.079g CaCl 2 2H 2 O. B liquid 6...

Embodiment 2

[0037] Embodiment 2, strain identification

[0038] The above strains were subjected to cell morphology identification, 16SrRNA identification, colony morphology identification and fermentation product identification using glycerol as substrate

[0039] 1. Morphological identification of bacterial strains

[0040] A single colony isolated in Example 1 was picked and inoculated into a centrifuge tube containing enriched medium, and after anaerobic culture at 37°C for 24 hours, a small amount of bacterial liquid was taken for Gram staining microscope observation. Gram stain test:

[0041] (1) Staining: Take an appropriate amount of bacteria and drop them on the glass slide, fix them by flame 4-6 times, add ammonium oxalate crystal violet solution dropwise, and stain the fixed smears for 1 min.

[0042] (2) Water washing: Slowly rinse the staining solution on the smear with water, and dry it with absorbent paper. Cell morphology can be observed after simple staining.

[0043]...

Embodiment 3

[0057] Embodiment 3, Clostridium butyricum tolerance test to substrate glycerol

[0058] Seed solution preparation: Prepare the fermentation solution as described in Example 2, and after standing for 10 hours, inoculate it into a 100ml shake flask when the OD is 1.5.

[0059] Fermentation medium ( / L): add about 40g, 80g, 120g, 160g, 200g, 240g of pure glycerin; yeast powder 5g; K 2 PO 4 ·3H 2 O1g; KH 2 PO 4 0.5g; (NH 4 ) 2 SO 4 2g; MgSO 4 ·7H 2 O 0.2g; CaCl 2 0.02g; Clostridium butyricum trace element solution 1ml / L; Clostridium butyricum Fe solution 1ml / L.

[0060] Shake flask culture: at 37 ° C, the above seed liquid was inserted into the shake flask by 10% of the fermentation volume, the fermentation volume was 50 ml, and the culture was static, and a sample was taken every about 1 h to test the substrate tolerance ( Figure 5 ).

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Abstract

The invention discloses clostridium butyricum and an application thereof in immobilized fermentation production of 1, 3-propylene glycol, the clostridium butyricum is clostridium butyricum SCUT343-4 and is preserved in Guangdong province microbial strain preservation center on October 12, 2019, and the preservation number is GDMCC NO: 60805. The strain can be used for directly converting pure glycerol or crude glycerol into 1, 3-propylene glycol, has higher substrate conversion rate and substrate tolerance, has few types and contents of byproducts, and is beneficial to separation and purification of subsequent products; the strain has the most outstanding tolerance to a substrate crude glycerol, and the conversion rate and the production efficiency are not reduced during crude glycerol fermentation, so that the processing and utilization of the crude glycerol are facilitated, and the production cost of the 1, 3-propylene glycol is remarkably reduced. The strain is applied to immobilized fermentation production of 1, 3-propylene glycol, the production efficiency can be remarkably improved and reaches 4.20 g / L.h.

Description

technical field [0001] The invention belongs to the technical field of bioengineering fermentation, and in particular relates to a high-yield 1,3-propanediol Clostridium butyricum and an immobilized fermentation method. Background technique [0002] 1,3-propanediol (1,3-propanediol), the molecular formula is C 3 h 8 o 2 As an important chemical raw material, it is widely used in many industries such as pharmaceuticals, food, cosmetics and textiles. At present, chemical synthesis is mostly used in industrial production, and the mature routes are (1) DuPont: acrolein route, that is, acrolein is converted into 3-hydroxypropionaldehyde (3-HPA) through hydration, and then hydrogenated to generate 1,3 -Propylene glycol; (2) Royal Shell Petroleum Company: Ethylene oxide route, that is, ethylene oxide hydroformylation method to prepare 3-hydroxypropionaldehyde, and then hydrogenation to generate 1,3-propanediol. However, the above two chemical pathways are not only technically d...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P7/18C12R1/145
CPCC12N1/20C12P7/18C12R2001/145C12N1/205
Inventor 王菊芳兰洋冯骏傅宏鑫张凌蔚
Owner SOUTH CHINA UNIV OF TECH
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