Method for increasing content of total flavonoids in red orange peel through microbial fermentation
By fermenting the red orange peel with specific microbial agents, the problem of increasing the total flavonoid content and long aging cycle of tangerine peel was solved, and the effect of rapidly improving the total flavonoid content and quality was achieved.
Patent Information
- Application Number
- CN202510468822.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-04-15
- Publication Date
- 2025-08-15
AI Technical Summary
The prior art is difficult to efficiently and safely increase the total flavonoid content in tangerine peel, especially during the aging process, which has the problem of long cycles.
The mixed microbial agent of Aspergillus niger strain Aspergillus niger SFCC-M19, Coprinelus radians SFCC-M18 and Polycysticus Stagonosporopsis sp. SFCC-M17 was used to ferment the red orange peel. The spore suspension was sprayed and fermented with fermentation temperature and humidity, and the aging time was shortened.
In a very short time, the total flavonoid content of red orange peel residue was significantly increased, an increase of 94.16%-115.94% compared to natural fermentation, accelerating the tangerine peel tangerine peel process and improving quality and nutritional value.
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Figure CN120484967A_ABST
Abstract
Description
Technical Field
[0001] The invention belongs to the technical field of microorganisms, and in particular relates to a method for increasing the total flavonoid content in tangerine peel by microbial fermentation. Background Art
[0002] Tangerine peel (Dried Tangerine Peel) is a substance with both edible and medicinal properties. It is obtained by drying and aging the mature peel of the Rutaceae plant (Citrus reticulata Blanco) and its cultivated varieties. Cultivated varieties include the tea-branched tangerine peel (Guangcheng tangerine peel), Dahongpao (Sichuan tangerine peel), Wenzhou mandarin orange, and Fuju tangerine. The peel of the red tangerine (Dahongpao) is a key raw material for the production of Sichuan tangerine peel. During the aging process, the content of flavonoids—the main active ingredients—such as hesperidin, nobiletin, tangeretin, and polymethoxyflavonoids, gradually increases. Therefore, total flavonoid content is a key indicator of tangerine peel quality.
[0003] At present, how to efficiently and safely increase the content of total flavonoids in dried tangerine peel is an urgent problem to be solved. Summary of the Invention
[0004] In response to the problems existing in the prior art, the purpose of the present invention is to provide a method for increasing the total flavonoids content in red tangerine peel through microbial fermentation. This method increases the total flavonoids content in aged red tangerine peel through microbial fermentation, which not only greatly shortens the aging cycle, but also helps to improve and maintain the quality of Sichuan tangerine peel, and has great market application prospects.
[0005] The purpose of the present invention is achieved through the following technical solutions:
[0006] A first aspect of the present invention provides a microbial agent, wherein the microbial agent is a mixture of one or more strains of Aspergillus niger SFCC-M19, Coprinellus radians SFCC-M18, and Stagonosporopsis sp. SFCC-M17;
[0007] Aspergillus niger SFCC-M19 was deposited in China Center for Type Culture Collection, Wuhan University, Wuhan, China, 430072, on January 10, 2025; the deposit number is CCTCCNO: M 2025088;
[0008] Coprinus radiatus SFCC-M18 was deposited in China Center for Type Culture Collection, Wuhan University, Wuhan, China, 430072, on January 10, 2025; the deposit number is CCTCCNO: M 2025089.
[0009] Sporocystis SFCC-M17 was deposited in the China Center for Type Culture Collection on January 10, 2025, address: Wuhan University, Wuhan, China, Postal Code: 430072; the deposit number is CCTCC NO: M 2025090.
[0010] The second aspect of the present invention provides the use of the microbial agent described in the first aspect in accelerating the aging and fermentation of red tangerine peel.
[0011] A third aspect of the present invention provides a method for increasing the total flavonoid content in tangerine peel by microbial fermentation, the method using the microbial agent described in the first aspect, comprising the following steps:
[0012] (1) Cultivating the microbial agent described in the first aspect in a culture medium, then taking spores and adding them to sterile water to prepare a spore suspension;
[0013] (2) selecting fresh tangerines, squeezing the juice to obtain the tangerine peel residue, and then drying and sterilizing the peel residue;
[0014] (3) spraying the spore suspension prepared in step (1) onto the sterilized peel residue, stirring and mixing, adjusting the moisture content of the peel residue to 60%-90%, and then performing pile fermentation at a temperature of 45-55° C. for 7-14 days;
[0015] (4) Collect the fermented tangerine peel residue, dry it at low temperature after fermentation, and detect the total flavonoid content.
[0016] Furthermore, in step (1), the concentration of the spore suspension is 10 6 -10 8 CFU / mL.
[0017] Furthermore, in step (3), the spraying amount of the spore suspension is 1-3% of the weight of the tangerine peel residue.
[0018] Furthermore, in step (3), the spore suspension is sprayed into the tangerine peel residue to adjust the moisture content of the peel residue to 70%-80%.
[0019] Furthermore, in step (3), the fermentation temperature is 45°C.
[0020] Culture deposit information:
[0021] Aspergillus niger SFCC-M19 was deposited in the China Center for Type Culture Collection on January 10, 2025, address: Wuhan University, Wuhan, China, Postal Code: 430072; the deposit number is CCTCC NO: M 2025088; the classification name is: Aspergillus niger.
[0022] Coprinellus radians SFCC-M18 was deposited in the China Center for Type Culture Collection on January 10, 2025, address: Wuhan University, Wuhan, China, Postal Code: 430072; the deposit number is CCTCC NO: M 2025089; the classification name is: Coprinellus radians.
[0023] Stagonosporopsis sp. SFCC-M17 was deposited in the China Center for Type Culture Collection on January 10, 2025, address: Wuhan University, Wuhan, China, Postal Code: 430072; the deposit number is CCTCC NO: M 2025090; the classification name is: Stagonosporopsis sp.
[0024] The beneficial effects of the present invention compared to the prior art are:
[0025] The present invention utilizes the efficient conversion ability of specific strains (Aspergillus niger SFCC-M19, Coprinus radiatus SFCC-M18, and Mycopolysaccharides SFCC-M17) screened from tangerine peel to flavonoids to promote the aging of tangerine peel residue. The tangerine peel can be converted into aged tangerine peel with typical tangerine peel appearance characteristics in a very short time (7-14 days). After 14 days of fermentation, the total flavonoid content in the aged tangerine peel residue is increased by up to 94.16% compared with naturally fermented peel residue and by 115.94% compared with unfermented peel residue. Thus, the nutritional value and functional properties of the tangerine peel residue are improved, and the invention has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS
[0026] The present invention will be further described below with reference to the accompanying drawings and examples:
[0027] Figure 1 Comparison of characteristics of samples on the 7th day of fermentation (left) and the 14th day of fermentation (right);
[0028] Figure 2 It is the change of total flavonoid content in the sample on the 7th day and the 14th day of fermentation. DETAILED DESCRIPTION
[0029] The present invention is not limited to the following examples; any modifications and / or alterations to the present invention fall within the scope of protection of the present invention. Unless otherwise specified, all equipment and raw materials are commercially available or commonly used in the industry. Unless otherwise specified, the methods used in the examples are generally known in the art.
[0030] For numerical ranges, the endpoints of each range, the endpoints of each range and the individual points, and the individual points can be combined with each other to obtain one or more new numerical ranges, which should be considered to be specifically disclosed herein.
[0031] Unless otherwise specified, all raw materials, reagents, instruments and equipment used in the present invention can be obtained commercially or prepared by existing methods. The experimental methods in each example are conventional methods unless otherwise specified.
[0032] Example 1 - strain screening
[0033] In this example, strains with potential functions were isolated and screened from dried tangerine peels of different aging years. The specific method was as follows:
[0034] (1) Weigh 10 g of tangerine peel (commercially available) produced by traditional methods with different aging years and place them in 250 mL conical flasks filled with 50 mL of normal saline. Incubate at 30 °C and 210 rpm for 30 min to obtain bacterial suspension.
[0035] (2) The bacterial suspension was diluted in a gradient manner. In a clean bench, 100 μL of each gradient bacterial suspension was taken and spread on PDA culture medium, marked, and cultured in a constant temperature incubator at 28°C for 3-5 days. At regular intervals, colonies with different morphologies were picked and streaked on a new culture medium. After continuous culture at 28°C for 3-5 days, the growth of the colonies, morphological characteristics, etc. were observed. The above operation was repeated until the size and morphological characteristics of the colonies on the plate were consistent, and a pure culture was obtained. The obtained pure culture was inoculated into LB culture medium and cultured for 2 days. After continuous subculture, the strains with good growth conditions were preserved in glycerol.
[0036] (3) Three strains with the ability to transform characteristic components from tangerine peel were screened and identified as: Aspergillus niger SFCC-M19, Coprinellus radians SFCC-M18 and Stagonosporopsis sp. SFCC-M17.
[0037] Example 2 - Acceleration of aging and fermentation of red tangerine peel
[0038] This embodiment provides a method for increasing the total flavonoid content in tangerine peel by microbial fermentation, comprising the following steps:
[0039] (1) Select fresh, good-quality, and pest-free red tangerines as raw materials, squeeze the juice to obtain the peel residue, and then dry the peel residue.
[0040] (2) Weigh 37 g of potato dextrose agar (PDA) medium, dissolve it in 1000 mL of water, sterilize it at 121° C. for 15 min, and pour it into a plate to solidify it to obtain a microbial solid culture medium. The three strains with characteristic component transformation ability derived from tangerine peel described in Example 1 (hereinafter referred to as: strain 1, strain 2, strain 3) were placed in a 28° C. incubator for 3-5 days, and then spores were taken with an inoculating loop and added to sterile water to prepare a concentration of 10 6 CFU / mL of three spore suspensions.
[0041] (3) Spray the spore suspension on the dried peel residue at a spraying amount of 1% of the peel residue weight, stir and mix while spraying, then spray sterile water to control the moisture content of the peel residue to 70%, and then ferment at a temperature of 45°C for 7-14 days. Turn and ventilate regularly during the fermentation process.
[0042] (4) After fermentation, dry the skin residue at 50-60°C until the moisture content is less than 10%. Package the dried product and store it in a cool and dry place.
[0043] Example 3
[0044] like Figure 1 As shown, the fermented peel residue and the unfermented peel residue were ground into powder, and the appearance was observed and the indicators were measured. The total flavonoid content was tested according to the method provided in the national standard GB / T 19416-2003.
[0045] 0.8015 g of aged tangerine peel powder and naturally fermented and unfermented tangerine peel powder after fermentation with the three strains on the 7th and 14th days were weighed respectively. Each sample was randomly sampled three times, 60 mL of methanol was added, and ultrasonic extraction was performed for 60 min before filtration. The residue was washed with an appropriate amount of methanol and filtered. The filtrate was collected and diluted to 100 mL with methanol. After treatment with aluminum nitrate colorimetry, the absorbance was measured, and the total flavonoid content was obtained from the standard curve (Tables 1 and 2).
[0046] Table 1 Total flavonoid content of samples on the 7th day of fermentation
[0047]
[0048] Table 2 Total flavonoid content of samples on day 14 of fermentation
[0049]
[0050] Depend on Figure 2The total flavonoid content of tangerine peel samples fermented with the three strains was significantly higher than that of naturally fermented and unfermented samples. On the seventh day of fermentation, the total flavonoid content of samples fermented with strains 2 and 3 was significantly higher than that of strain 1. On the 14th day of fermentation, the total flavonoid content of the sample fermented with strain 1 was significantly higher than that of the other four samples, with the fastest increase in total flavonoid content. Overall, microbial fermentation can accelerate the aging of tangerine peel and increase the total flavonoid content.
[0051] Finally, it should be noted that the above is only used to illustrate the technical solution of the present invention and is not limiting. Although the present invention is described in detail with reference to the preferred arrangement scheme, ordinary technicians in this field should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the spirit and scope of the technical solution of the present invention.
Claims
1. A microbial agent, characterized in that: The microbial agent is a mixture of one or more strains of Aspergillus niger SFCC-M19, Coprinellus radians SFCC-M18, and Stagonosporopsis sp. SFCC-M17; Aspergillus niger SFCC-M19 was deposited in China Center for Type Culture Collection, Wuhan University, Wuhan, China, 430072, on January 10, 2025; the deposit number is CCTCC NO: M 2025088; Coprinus radiatus SFCC-M18 was deposited on January 10, 2025, at China Center for Type Culture Collection, Wuhan University, Wuhan 430072, China; the deposit number is CCTCC NO: M 2025089. Sporocystis SFCC-M17 was deposited in the China Center for Type Culture Collection on January 10, 2025, address: Wuhan University, Wuhan, China, Postal Code: 430072; the deposit number is CCTCC NO: M 2025090.
2. Use of the microbial agent as claimed in claim 1 in accelerating the aging and fermentation of red tangerine peel.
3. A method for increasing the total flavonoid content in tangerine peel by microbial fermentation, characterized in that: The method uses the microbial agent according to claim 1, comprising the following steps: (1) Cultivating the microbial agent according to claim 1 in a culture medium, then taking spores and adding them to sterile water to prepare a spore suspension; (2) selecting fresh tangerines, squeezing the juice to obtain the tangerine peel residue, and then drying and sterilizing the peel residue; (3) spraying the spore suspension prepared in step (1) onto the sterilized peel residue, stirring and mixing, adjusting the moisture content of the peel residue to 60%-90%, and then performing pile fermentation at a temperature of 45-55° C. for 7-14 days; (4) Collect the fermented tangerine peel residue, dry it at low temperature after fermentation, and detect the total flavonoid content.
4. The method according to claim 3, characterized in that In step (1), the concentration of the spore suspension is 10 6 -10 8 CFU / mL.
5. The method according to claim 3, characterized in that In step (3), the spraying amount of the spore suspension is 1-3% of the weight of the tangerine peel residue.
6. The method according to claim 3, characterized in that In step (3), the spore suspension is sprayed into the tangerine peel residue to adjust the moisture content of the peel residue to 70%-80%.
7. The method according to claim 3, characterized in that In step (3), the fermentation temperature is 45°C.