Burkholderia Vietnamese and application thereof
By using Vietnamese Burkholderia CGMCC No. 33738 to produce chitinase with high enzyme activity, the problem of low extraction rate of enzymatic shrimp and crab shells was solved, and the effect of agricultural application was improved.
Patent Information
- Application Number
- CN202510636176.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-05-16
- Publication Date
- 2025-08-05
AI Technical Summary
The existing strains have low enzyme activity of chitinase, which leads to low extraction rate of enzymatic shrimp and crab shells, and has poor application in agriculture.
Using Burkholderella Vietnamese CGMCC No. 33738, chitinase containing high enzyme activity was produced by culturing in a specific culture medium, and used to enzymatically dissolve shrimp and crab shells.
The extraction rate of enzymatic shrimp and crab shells has been improved, and the effect of agricultural application has been significantly improved.
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Abstract
Description
Technical Field
[0001] The present invention relates to a Burkholderia vietnamiensis and its application. Background Art
[0002] In nature, chitin widely exists in the cells of lower plants such as fungi and algae, the shells of arthropods such as shrimps, crabs, maggots and insects, the shells and cartilages of shellfish and mollusks, and the cell walls of higher plants. It is the second largest biological resource on the earth after plant fiber, and the production amount of marine organisms is the largest, which can be said to be an inexhaustible biological resource.
[0003] Currently, chitin, chitosan and chitooligosaccharides are mainly extracted from shrimp and crab shells. Since chitin in shrimp and crab shells is covalently bound to proteins and exists in the form of proteoglycans, and calcium carbonate is also accompanied. Currently, chitosan and chitooligosaccharides prepared by enzymatic hydrolysis of shrimp and crab shells have been widely used in agriculture. However, the chitooligosaccharides prepared by degrading with chitinase have the problem of wasting nutrients beneficial to plants, and the application effect in agriculture is not very ideal.
[0004] Currently, there is no strain with high chitinase activity for enzymatic hydrolysis of shrimp and crab shells and high extraction rate, and it has a good effect when applied to agriculture. Summary of the Invention
[0005] The present invention provides a Burkholderia vietnamiensis and its application, and the technical problem to be solved is that the chitinase activity of the strain is not high, the extraction rate for enzymatic hydrolysis of chitin or chitosan is not high, and the application effect in agriculture is not satisfactory.
[0006] In order to solve the above technical problems, the present invention adopts the following technical solutions: A Burkholderia vietnamiensis, which is deposited in the General Microbiological Center of the China Committee for Culture Collection of Microorganisms, and the deposit number is CGMCC No. 33738.
[0007] The application of Burkholderia vietnamiensis is for producing an enzyme; the enzyme contains chitinase.
[0008] The produced enzyme is applied to enzymatic hydrolysis of chitin.
[0009] The invention has the following beneficial technical effects: The chitinase produced by the strain screened in this application has high activity, and it is a strain with high extraction rate for enzymatic hydrolysis of shrimp and crab shells, and it has a good effect when applied to agriculture.
[0010] RNA sequence of Burkholderia vietnamiensis Detailed implementation mode
[0011] In order to better understand the above technical solutions, the technical solutions of the present invention will be described in detail below in combination with preferred examples. Example 1
[0012] A Burkholderia vietnamiensis is preserved in the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, and the preservation number is CGMCC No. 33738.
[0013] The application of Burkholderia vietnamiensis is used for producing enzymes; the enzyme contains chitinase.
[0014] Inoculate Burkholderia vietnamiensis into the culture medium, and culture it at 45 °C for 60 h, then filter. The obtained clear liquid is the crude enzyme; the maximum chitinase enzyme activity detected for the crude enzyme is 4.57 U / mL; The inoculation amount is 4%; the culture medium is 2 g of peptone, 12 g of colloidal chitin, 21 g of sodium chloride, 0.02 g of ferrous sulfate, and the balance is water, pH 7.0, and the volume is fixed to 1000 ml. Example 2
[0015] A Burkholderia vietnamiensis is preserved in the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, and the preservation number is CGMCC No. 33738.
[0016] The application of Burkholderia vietnamiensis is used for producing enzymes; the enzyme contains chitinase.
[0017] Inoculate Burkholderia vietnamiensis into the culture medium, and culture it at 45 °C for 60 h, then filter. The obtained clear liquid is the crude enzyme; the maximum chitinase enzyme activity detected for the crude enzyme is 4.15 U / mL; The inoculation amount is 4%; the culture medium is 2 g of yeast powder, 15 g of colloidal chitin, 21 g of sodium chloride, 0.02 g of ferrous sulfate, 0.05 g of magnesium sulfate, and the balance is water, pH 7.0, and the volume is fixed to 1000 ml. Example 3
[0018] A Burkholderia vietnamiensis is preserved in the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, and the preservation number is CGMCC No. 33738.
[0019] The application of Burkholderia vietnamiensis is used for producing enzymes; the enzyme contains chitinase.
[0020] Inoculate Burkholderia vietnamiensis into a culture medium, incubate at 50 °C for 60 h, filter, and the obtained supernatant is the crude enzyme; the maximum chitinase activity detected for the crude enzyme is 4.69 U / mL. The inoculation amount is 5%; the culture medium consists of 1 g of yeast powder, 1 g of peptone, 12 g of colloidal chitin, 20 g of sodium chloride, 0.02 g of ferrous sulfate, with the balance being water, pH 7.0, and the volume is made up to 1000 ml.
[0021] The beneficial effects of the present invention will be further illustrated below in conjunction with experimental data: Experiment 1 1.1 Test site: Shandong Aiguozhe Biotechnology Co., Ltd.
[0022] 1.2 Experimental detection: Solid content (%).
[0023] 1.3 Test materials: Comparison 1 (except that the crude enzyme is replaced with diluted chitinase, and the others are the same as in Example 1. The chitinase is purchased from Guangdong Wengjiang Chemical Reagent Co., Ltd. with an enzyme activity of 100,000) and Examples 1 to 3. Dilute the chitinase with water to 4.70 U / mL to obtain diluted chitinase.
[0024] Dilute the chitinase with water to 4.70 U / mL to obtain diluted chitinase.
[0025] 1.4 Test method: Take the cleaned crab shells, remove the crab meat, crush them into fine powder with a high-speed universal grinder, then add water and yeast powder, add the diluted chitinase or the crude enzyme prepared in Examples 1 to 3, and carry out enzymatic hydrolysis at 45 °C for 72 h, filter, and the obtained filtrate is the enzymatic hydrolysate, and detect the solid content of the enzymatic hydrolysate; The mass ratio of crab shells, water, yeast powder and the diluted chitinase or the crude enzyme prepared in Examples 1 to 3 is 20:74:5:1; The yeast powder is purchased from Xi'an Tianbao Biotechnology Co., Ltd.
[0026] Solid content detection: Use a dropper to suck about 1 g of the aqueous chitin mixture, drop it on an evaporating dish, weigh it as m0, put the evaporating dish into a 60 °C vacuum oven, dry it, weigh it after 4 hours, put it back into the oven again, weigh it after an interval of 1 hour, until the difference between the two weighings does not exceed 0.04 g, record it as m1, and the solid content = ((m0 - m1) / m0) * 100%.
[0027] Except for the different treatments in this experiment, the other treatments are the same.
[0028] 2 Results and analysis The solid content (%) is shown in Table 1 Table 1
[0029] It can be seen that the enzyme prepared by Burkholderia vietnam in the present application has a good effect of degrading chitin. Of course, its high solid content may also be related to the fact that the crude enzyme contains not only chitinase but also other enzymes. The solid content of the crude enzyme hydrolysis product contains not only chitosan oligosaccharides but also other substances. Therefore, the solid content is high.
[0030] Experiment 2 1.1 Experimental location: Luwa Village, Jinzhongzi Town, Anqiu City, 1 mu of strawberry greenhouse, the variety is Zhangji.
[0031] 1.2 Experimental tests: soluble sugar (%), vitamin C content (mg / 100g) and organic acid content (%).
[0032] 1.3 Test materials: Experiment 1, Comparison 1 (except that the crude enzyme was replaced by diluted chitinase, all other materials were the same as in Example 1) Chitinase was purchased from Guangdong Hongjiang Chemical Reagent Co., Ltd. (enzyme activity 100,000) and the enzymatic hydrolysates prepared in Examples 1 to 3.
[0033] 1.4 Experimental method: The greenhouse was randomly divided into 4 small areas, each area was 150 square meters, and isolation zones were set between the small areas. The plots were randomly matched to the experimental treatments. The enzymatic hydrolysates prepared in Comparative 1 (except that the crude enzyme was replaced by diluted chitinase, all other conditions were the same as in Example 1; chitinase was purchased from Guangdong Hongjiang Chemical Reagent Co., Ltd., with an enzyme activity of 100,000) and Examples 1 to 3 were diluted 500-fold with water to obtain dilutions. Fertilization was performed by foliar spraying at intervals of 15 days, with an application rate of 10 kg of the dilution per plot.
[0034] 1.5 Detection method: 20 mature fruits of similar size were randomly picked from each plot, and the soluble sugar content was determined by the anthrone method; the organic acid content was determined by the NaOH titration method; and the Vc content was determined by the 2,6-dichlorophenol indophenol reduction titration method.
[0035] Except for the different treatments in this experiment, the other treatments were the same.
[0036] 2 Results and Analysis Soluble sugar (%), Vc (mg / 100g) and organic acid (%), see Table 2 Table 2
[0037] As can be seen from Table 2, the chitin mixture obtained by enzymatic hydrolysis using the chitinase prepared by the Burkholderia vietnamese of the present application can significantly improve the quality of fruits when applied to agriculture.
Claims
1. A Burkholderia vietnamese strain, deposited in the General Microbiology Center of China Culture Collection Administration of Microorganisms, with the accession number being CGMCC No. 33738.
2. The use of Burkholderia vietnamese according to claim 1, characterized in that Used for producing enzymes; the enzymes contain chitinase.
3. The use of Burkholderia vietnamese according to claim 2, characterized in that The produced enzyme is used to enzymatically hydrolyze chitin.