Pseudomonas oryzihabitans capable of secreting ferulic acid esterase and application of pseudomonas oryzihabitans

A technology of ferulic acid esterase and pseudomonas, which is applied in the direction of enzymes, bacteria, hydrolytic enzymes, etc., can solve the problems of rice wine quality change, specific role is not very clear, microbial composition and quantity complexity, etc., to increase the content Effect

Active Publication Date: 2019-07-30
JIANGNAN UNIV
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  • Claims
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AI Technical Summary

Problems solved by technology

However, at present, during the fermentation process of wheat koji and rice wine, microorganisms can only release a small amount of bound ferulic acid in wheat koji, which cannot achieve the goal of increasing the content of ferulic acid in rice wine. Ferulic acid content
Some studies have used a fungus that can produce ferulic acid esterase - Cladosporium cladoides to increase the content of ferulic acid in rice wine, but the fungal culture is easy to form hyphae or balls, which brings great difficulties to the inoculation operation. difficulty
Moreover, since rice wine fermentation is a complex process involving multiple strains, the composition and quantity of microorganisms in wheat koji are very complex, and the specific role of different strains in the fermentation process of rice wine is not very clear. The introduction of bacteria into wheat koji may cause the migration of the original microbial flora structure of wheat koji, which will change the quality of rice wine obtained from fermentation

Method used

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  • Pseudomonas oryzihabitans capable of secreting ferulic acid esterase and application of pseudomonas oryzihabitans
  • Pseudomonas oryzihabitans capable of secreting ferulic acid esterase and application of pseudomonas oryzihabitans
  • Pseudomonas oryzihabitans capable of secreting ferulic acid esterase and application of pseudomonas oryzihabitans

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1: Screening and identification of high-yield ferulic acid esterase strains in wheat koji

[0029] (1) Sample collection and processing

[0030] The wheat koji was collected from a rice wine factory in Shaoxing City, Zhejiang Province, and the collected wheat koji was stored in a sealed sterile plastic bag at 4 °C. Weigh 5.0 g of wheat koji sample, add it to 45 mL of sterilized enrichment medium, place it on a shaker at 150-200 rpm for enrichment culture for 2-4 hours, and the temperature is 28-30°C.

[0031] Enrichment medium: Ammonium sulfate 1.3g, KH 2 PO 4 0.37g, MgSO 4 ·7H 2 O 0.25g, CaCl 2 2H 2 O 0.07g, FeCl 3 0.02g, 1.0g of yeast extract, 5mL of ethyl ferulate (10% W / V dissolved in dimethylformamide), 1000mL of distilled water, natural pH, sterilized at 115°C for 20min.

[0032] (2) Screening of strains

[0033] Screening medium: NaCl 0.3g, ammonium sulfate 1.3g, K 2 HPO 4 0.3g, MgSO 4 ·7H 2 O 0.3g, agar powder 18.0g, 15mL ethyl ferulate (1...

Embodiment 2

[0036] Example 2: Physiological and biochemical properties of Pseudomonas oryzihabitans Y-5

[0037] The above-mentioned Pseudomonas oryzihabitansY-5 was inoculated on LB solid medium, and the colonies formed were dot-shaped, waxy, opaque, light yellow, raised on the surface, with neat edges, and Gram staining was negative. The body is rod-shaped, appearing singly or in short chains; its physiological and biochemical characteristics are: catalase negative, gelatin liquefaction negative, oxidase negative, can utilize glucose, xylose, fructose, maltose, mannitol, sucrose, galactose , Mannose produces acid, cannot use starch and lactose, and can grow at 42°C. The physiological and biochemical characteristics of the strains are shown in Table 1.

[0038] Table 1 Physiological and biochemical characteristics of Pseudomonas oryzihabitans Y-5

[0039]

[0040] Note: +: Indicates that the strain is positive in reaction, i.e. producing gas / fermentation acid; -: Indicates that the ...

Embodiment 3

[0041] Embodiment 3: Determination of ferulic esterase enzyme activity in the fermented liquid of Pseudomonas oryzihabitans Y-5

[0042] Pseudomonas (Pseudomonas oryzihabitans) Y-5 was inoculated into LB liquid medium and cultured at 28±2°C for 24-32 hours to obtain Pseudomonas seed liquid.

[0043] Preparation method of crude enzyme solution: inoculate 100mL induction medium with an inoculum volume ratio of 4% of the strain seed solution cultivated for 24 hours, culture at 30°C and 200r / min for 3 days, centrifuge at 10000g for 15 minutes, and the supernatant is crude enzyme solution.

[0044] Induction medium: 1.0g NaCl, 1.0g tryptone, 0.5g yeast extract, 10g wheat bran, 100mL distilled water, natural pH, sterilized at 115°C for 20min.

[0045] Determination of ferulic acid esterase enzyme activity: Take 1.0mL of crude enzyme solution in a 5mL centrifuge tube, preheat at 30°C for 5min, then add 1.0mL methyl ferulic acid solution (pH 6.0), mix and incubate for 10min, then qui...

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Abstract

The invention discloses a strain of pseudomonas oryzihabitans capable of secreting ferulic acid esterase and application of the pseudomonas oryzihabitans and belongs to the technical field of fermentation engineering and biology. The pseudomonas oryzihabitans Y-5 is preserved in the China Center for Type Culture Collection in July, 13th, 2018, and the preservation number is CCTCC NO:M2018480. Thestrain can secrete ferulic acid esterase, and under the induction of bran, the enzyme activity of the ferulic acid esterase of a fermented crude enzyme liquid of the strain is 116 U/L, and by adoptingthe bran for substrate fermentation to produce ferulic acid, the yield of ferulic acid can reach 59.51+/-0.6 mg/L. According to the pseudomonas oryzihabitans, the content of ferulic acid in yellow rice liquor can be increased when the alcoholic strength, pH value and reducing sugar content of the yellow rice liquor are not influenced, and after fermentation of strengthened wheat koji to which thepseudomonas oryzihabitans is inoculated, the content of ferulic acid in the yellow rice liquor is about 2.6 times higher than that of common yellow rice liquor fermented by traditional wheat koji.

Description

technical field [0001] The invention relates to a strain of pseudomonas producing ferulic acid esterase and application thereof, belonging to the fields of fermentation engineering and biotechnology. Background technique [0002] The chemical name of ferulic acid (Ferulic Acid, FA) is 4-hydroxy-3-methoxycinnamic acid, which has the functions of scavenging free radicals, antithrombotic, antibacterial and anti-inflammatory, inhibiting tumors, anti-aging, and inhibiting low-density lipoproteins. The physiological functions of oxidation and anti-platelet aggregation are widely used in the fields of health care products, food additives, cosmetics and medicine; at the same time, ferulic acid is not only a flavor substance in rice wine, but also an important material basis for the health function of rice wine. [0003] Ferulic acid esterase (EC.3.1.1.73, ferulic acid esterase, FAE) belongs to the extracellular enzyme, which can cut the ester bond between ferulic acid and polysaccha...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12P7/42C12R1/38
CPCC12N9/18C12P7/42C12Y301/01073
Inventor 毛健王宗敏张晶鲁思苑
Owner JIANGNAN UNIV
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