Application of saccharomyces cerevisiae in biosynthetic L-phenylacetylcarbinol

A technology of phenylacetyl carbinol and Saccharomyces cerevisiae is applied in the field of bioengineering, which can solve the problems of complex process and high cost, and achieve the effects of simplified separation procedure, stable growth characteristics and high tolerance

Inactive Publication Date: 2016-02-17
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the process is complicated and the cost is high

Method used

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  • Application of saccharomyces cerevisiae in biosynthetic L-phenylacetylcarbinol
  • Application of saccharomyces cerevisiae in biosynthetic L-phenylacetylcarbinol
  • Application of saccharomyces cerevisiae in biosynthetic L-phenylacetylcarbinol

Examples

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Embodiment 1

[0021] The acquisition of embodiment 1 benzaldehyde tolerant yeast strain

[0022] For the acquisition of benzaldehyde-tolerant yeast strains, see figure 1 ,Proceed as follows:

[0023] (1) Screening of benzaldehyde-resistant strains: Activate and pick a single colony for fermentation from commercially available yeast strains to obtain yeast cells; take 1 g of yeast cells and suspend them in 10 ml of normal saline, and take 50 μl of the cell fluid and inoculate them in 0.05 %, 0.1%, 0.2%, 0.4%, 0.6%, 0.8% on solid culture plates, cultured at 30°C, and observed the growth of plate colonies every 12 hours;

[0024] (2) Tolerance analysis: pick the tolerant single colony on the solid culture plate containing benzaldehyde 0.2%, 0.4%, 0.6%, inoculate to 150ml containing 0.2%, 0.4%, 0.6%, 0.8% benzaldehyde respectively Fermentation 24h' in the seed culture medium

[0025] (3) by OD 600 =0.1 initial cell concentration inoculated into the same high-concentration seed culture mediu...

Embodiment 2

[0027] Example 2 Biosynthesis of L-PAC

[0028] (1) Construction of biocatalyst

[0029] Utilize composite carrier to immobilize benzaldehyde tolerant yeast strain, be biocatalyst; Wherein, described composite carrier is the immobilization buffer containing polyvinyl alcohol and sodium alginate, and described immobilization buffer is containing 20mmol / LMgSO 4 and 0.5MKH of 1mmol / L thiamine pyrophosphate (TPP) 2 PO 4 / K 2 HPO 4 Buffer, the mass percentage concentration of the polyvinyl alcohol is 5-15%; the mass percentage concentration of the sodium alginate is 1.5-2.5%;

[0030] The specific operation is: add polyvinyl alcohol and sodium alginate and 1g of benzaldehyde-tolerant yeast strain to 4ml of 20mmol / LMgSO 4 and 0.5MKH of 1mmol / L thiamine pyrophosphate (TPP) 2 PO 4 / K 2 HPO 4 In the buffer solution, after mixing, use a 20ml syringe to draw the mixture and drop it into 4% CaCl containing 0.2% glutaraldehyde at 4°C 2 Granulate and solidify in saturated boric ac...

Embodiment 3

[0041] This example is basically the same as Example 2, except that in step (3), 1.2% additive β-cyclodextrin is added to the reaction system.

[0042] HPLC detection was performed on the finally obtained reaction liquid, and the concentration of the prepared L-PAC was 34 g / L.

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Abstract

The invention provides an application of saccharomyces cerevisiae in biosynthetic L-phenylacetylcarbinol. The application comprises the steps of obtaining formaldehyde tolerant yeast strains, constructing a biocatalyst, converting microorganisms and the like. According to a method, one the one hand, the yeast strains are screened to obtain the yeast strain which is high in tolerance to benzaldehyde and stable in growth trait; on the other hand, a composite carrier stabilized benzaldehyde tolerant yeast strain is used for improving the metabolism condition of thallus, the catalyzing time of enzyme is stabilized and prolonged, the reaction condition is improved, the concentration of key substrate benzaldehyde is improved, the separation procedure of the thallus and conversion liquid is simplified, and therefore the cumulant of L-PAC is improved, wherein the concentration of L-PAC in the conversion liquid is close to 45 g / L. According to the method, the utilization of the raw materials is greatly improved, production cost is reduced, the yield of L-PAC is remarkably increased, and the foundation can be laid for industrialization of biologically synthesized ephedrine.

Description

technical field [0001] The invention relates to the application of Saccharomyces cerevisiae in the biosynthesis of L-phenylacetylcarbinol, belonging to the technical field of bioengineering. Background technique [0002] Benzaldehyde, also known as benzoin aldehyde and bitter almond oil, is an important pharmaceutical intermediate widely used in pharmaceuticals, spices, pesticides, dyes, plastic additives and other industries. Using starch and sugar as fermentation materials and benzaldehyde as substrate to biosynthesize drug intermediates by microbial method is a technology to alleviate the energy crisis, but the cytotoxicity of substrate benzaldehyde limits its industrial application. [0003] L-phenylacetylcarbinol (L-PAC) is a key chiral precursor for the synthesis of drugs L-ephedrine (L-ephedrine) and D-pseudoephedrine (D-pseudoephedrine). The research of substrate enzyme-catalyzed synthesis of L-PAC focuses on finding biocatalysts with low cost, high yield and high y...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/26C12R1/865
Inventor 许激扬卞筱泓张修芹朱兴宇孙天娇
Owner CHINA PHARM UNIV
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