Screening and application of high-yield 2-phenylethanol microzyme strain

A phenethyl alcohol and yeast technology, applied in the field of fermentation engineering, can solve the problems of inability to meet industrial production, low 2-phenylethyl alcohol yield and the like, and achieve the effect of good application prospects

Active Publication Date: 2020-12-18
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, using phenylalanine as a substrate, the yield of 2-phenylethanol produced by the whole cell conversion method is still relatively low, which cannot meet the needs of industrial production.

Method used

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  • Screening and application of high-yield 2-phenylethanol microzyme strain
  • Screening and application of high-yield 2-phenylethanol microzyme strain
  • Screening and application of high-yield 2-phenylethanol microzyme strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059]Example 1: Screening and isolation of strains

[0060](1) Weigh 1g of medicinal koji, add 9mL of sterile water, soak for 15 minutes and then shake well to make a bacterial suspension; under aseptic operating conditions, dilute to 10 with sterile water.-6, 10-5, 10-4, 10-3, 10-2, 10-1Gradient bacterial suspension; take 100μL of each gradient bacterial suspension and spread it on YPD plates, and make three parallel dilutions of each gradient; invert the medium plate in a 30℃ constant temperature incubator for 2 days, and select the protrusions on the surface of the plate. Single colony with milky white, irregular edges and opaque, streak on the YPD plate until a single colony grows, repeat the streak on the plate until each plate grows a single colony (that is, the colony form on each plate Consistent); Pick a single colony from each plate and store it on YPD slant medium.

[0061](2) Preliminary screening of strains: Inoculate the strains obtained in step (1) into YPD medium, culture...

Embodiment 2

[0070]Example 2: Production of 2-Phenylethanol by Yeast Shaking Flask Culture

[0071](1) Fermentation culture of strains in SC medium

[0072]The screened abnormal Wickham's yeast 1D6 was activated in YPD liquid medium to the initial OD600The inoculum values ​​of 0.1 and 30mL / 250mL were inoculated into SC medium, and the fermentation broth was taken every 8h and cultured for 56h at 30°C and rotating speed of 220r / min. The fermentation broth was centrifuged at 12000r / min for 10min at 4°C, filtered through a 0.22μm organic filter membrane, and analyzed by HPLC.

[0073]The results showed that after 24 hours of culture, the yield basically stabilized, and the yield of 2-PE was as high as 2274.73mg / L (Figure 5 ).

[0074](2) The effect of glucose concentration in SC medium on the production of 2-phenylethanol

[0075]The specific method is shown in step (1), the difference is that the glucose concentration in the SC medium is replaced, the fermentation is carried out for 48 hours, and the production ...

Embodiment 3

[0084]Example 3: Production of 2-Phenylethanol by Yeast Fermentation Tank Culture

[0085]The abnormal Wickham's yeast 1D6 was cultured at scale and cultured on YPD plates for 2-3 days. Then pick a single colony and inoculate it into a medium containing 100mL / 500ml YPD, culture it at 30°C, 220rpm / min shaker for 24h as seed solution. Both batch and fed-batch fermentation were carried out in a 5L fermentor equipped with 2.5L of fermentation medium (ie, optimized SC medium).

[0086]OD6004.0 is the seed liquid, inoculated into the fermentation medium at an inoculum of 10mL / 100mL, the stirring speed is 400rpm, and the aeration rate is 2.0vvm. Use sterile 4mol / L NaOH solution and 4mol / L HCl solution to control the pH of the fermentation medium to 4.5. All cultures were performed at 30°C. The content of 2-phenylethanol in the fermentation broth was measured every 12 hours, and the results are shown in Table 3. At 24h of fermentation, L-phenylalanine with a final concentration of 5g / L was added ...

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Abstract

The invention discloses screening and application of a high-yield 2-phenylethanol microzyme strain, and belongs to the technical field of fermentation engineering. According to the invention, a high-yield 2-phenylethanol microzyme strain, wickerhamomyces anomalus 1D6, is obtained by screening; and the obtained wickerhamomyces anomalus 1D6 has a relatively high L-phenylalanine conversion rate, as well as relatively low glucose content requirement in a reaction system so that relatively high yield can be reached when the glucose concentration is only 20-40 g / L. Moreover, the wickerhamomyces anomalus 1D6 has low L-phenylalanine substrate concentration requirement so that yield of 2500 mg / L or above can be reached when the L-phenylalanine substrate concentration is only 5-7 g / L under shake-flask fermentation conditions. In addition, through enlarged culture in a fermentation tank, the yield can be up to 4727.29 mg / L; and thus, the wickerhamomyces anomalus 1D6 has promising application prospects.

Description

Technical field[0001]The invention relates to the screening of a high-yield 2-phenylethanol yeast strain and its application, and belongs to the technical field of fermentation engineering.Background technique[0002]2-Phenylethanol (2-PE) is a high-grade aromatic alcohol with rose fragrance. Due to this feature, 2-Phenylethanol has become one of the most commonly used aromatic compounds in the perfume, cosmetics and food industries, as the sensory final product Enhancer. However, 2-PE also shows other interesting properties, such as its antifungal and antibacterial properties, making this compound a popular additive in disinfectants, pest control, cleaning and personal care products. In addition, 2-PE can be easily esterified to produce an important flavor substance, ethyl 2-phenylacetate. It also has a floral and rose-like smell, and its use is similar to 2-PE. At present, the production methods of 2-PE mainly include chemical synthesis and plant extraction.[0003]Most of 2-PE is syn...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/16C12P7/22C12R1/645
CPCC12N1/16C12P7/22C12R2001/645C12N1/145Y02E50/10
Inventor 周景文田淑芳陈宇陈坚曾伟主堵国成
Owner JIANGNAN UNIV
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