Gastrodia elata fermentation product and application thereof
Through the fermentation products of Gastrodia elata fermented by Guantusan Compass fermentation, the problems of low cure rate, high drug price and increased antibiotic resistance in the treatment of ulcerative colitis were solved, and significant prevention and treatment effects and safety were achieved.
Patent Information
- Application Number
- CN202411814059.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-10
- Publication Date
- 2025-02-18
AI Technical Summary
The prior art has problems with low cure rates, high drug prices and increased drug resistance in the treatment of ulcerative colitis.
Through the fermentation products of Gastrodia elata fermentation products of Guantusan Compass fermentation, specific fermentation methods and steps are adopted to prepare Gastrodia elata fermentation products with significant effects on preventing and treating ulcerative enteritis.
The fermentation products of Gastrodia elata can significantly reduce ulcerative enteritis induced by sodium glucan sulfate, have good anti-inflammatory effects and no adverse reactions, providing a new biological source substance and reducing the use of antibiotics.
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Figure CN119454853A_ABST
Abstract
Description
Technical Field
[0001] The invention relates to the field of biomedicine, and in particular to a fermentation product of Gastrodia elata and application thereof. Background Art
[0002] Ulcerative colitis (UC) is a chronic intestinal inflammatory disease, characterized by recurrent colorectal mucosal inflammation, with an increasing incidence rate year by year worldwide. Its clinical symptoms are usually bloody stools and diarrhea, which seriously affect the quality of life of patients. The pathogenesis of UC is not yet fully understood, mainly including impaired intestinal barrier function, intestinal mucosal inflammatory response, and intestinal flora imbalance. Although biological agents have brought relief effects to the treatment of UC in clinical practice in recent years, the long-term cure rate is still low, and biological drugs are expensive, bringing a huge economic burden to the public health system. Therefore, the development of drugs that are suitable for my country's national conditions and effective in treating UC is crucial for patients and the national health system.
[0003] In animal husbandry, the current clinical prevention and treatment of ulcerative enteritis mostly relies on antibiotics, but the use of antibiotics leads to increased drug resistance and drug residues, which has become a major hidden danger in the dairy farming industry and is also the purpose of the present invention. Summary of the invention
[0004] The object of the present invention is to provide a fermentation product of Gastrodia elata to solve the problems raised in the above background technology.
[0005] To achieve the above objectives, the embodiments of the present invention provide the following technical solutions:
[0006] A fermentation product of Gastrodia elata, which is a fermentation product produced by fermenting Gastrodia elata with Eurotium cristatum.
[0007] Preferably, the method for preparing the fermentation product of Gastrodia elata comprises the following steps:
[0008] Place the Eurotium cristatum on a maltose agar plate for activation culture;
[0009] After the spores of the Eurotium cristatum are formed, the spores are scraped and diluted to obtain a spore suspension;
[0010] Take fresh Gastrodia elata, wash it, add distilled water, crush it into juice, and sterilize it to obtain a fermentation substrate;
[0011] The spore suspension is inoculated into a fermentation substrate for culturing and fermentation to obtain a fermentation product of Gastrodia elata.
[0012] Preferably, the concentration of the spore suspension is (2-3)×10 8 Spores / mL.
[0013] Preferably, the mass ratio of Gastrodia elata to distilled water is 1:(1-1.5).
[0014] Preferably, the number of spores inoculated per gram of Gastrodia elata is (1-2)×10 7 Spores / mL.
[0015] Another object of the present invention is to provide an application of the fermentation product of Gastrodia elata in the preparation of a medicine for preventing and treating ulcerative enteritis.
[0016] Another object of the present invention is to provide a medicine for preventing and treating ulcerative enteritis, comprising a carrier and an active ingredient, wherein the active ingredient comprises the fermentation product of the above-mentioned Gastrodia elata.
[0017] The present invention provides a new method for fermenting Gastrodia elata with Eurotium cristatum, and finds that the fermentation product has a significant preventive and therapeutic effect on ulcerative enteritis and has a significant medical effect. Animal experiments have verified that the fermentation product of Gastrodia elata fermented by Eurotium cristatum can effectively alleviate ulcerative enteritis induced by sodium dextran sulfate. This discovery provides a new biological source material for the prevention and treatment of ulcerative enteritis, which is expected to reduce the use of antibiotic drugs and thus reduce related adverse reactions. Although the fermentation product of Gastrodia elata fermented by Eurotium cristatum shows good effects in the treatment of ulcerative enteritis, its specific mechanism of action still needs further study. The present invention provides an important basis for the subsequent pharmacological exploration of Gastrodia elata fermented by Eurotium cristatum, and also provides a valuable research idea for the development of new functions of biological autologous substances, and has the potential to promote the modernization of the pharmaceutical industry. BRIEF DESCRIPTION OF THE DRAWINGS
[0018] Figure 1 The representative visual images of the effects of Gastrodia elata fermentation products on ulcerative colitis induced by sodium dextran sulfate;
[0019] Figure 2 This is a picture of H&E stained intestinal tissue sections showing the effect of Gastrodia elata fermentation products on ulcerative enteritis induced by sodium dextran sulfate;
[0020] Figure 3 This is a picture of intestinal tissue Alcian blue staining section showing the effect of Gastrodia elata fermentation products on ulcerative enteritis induced by sodium dextran sulfate;
[0021] Figure 4 This is the result graph of the effect of Gastrodia elata fermentation products on the intestinal tissue length of ulcerative enteritis induced by sodium dextran sulfate;
[0022] Figure 5 This is a graph showing the disease activity index (DAI) results of the effect of Gastrodia elata fermentation products on ulcerative enteritis induced by dextran sulfate sodium;
[0023] Figure 6This is a schematic diagram showing the results of the fermentation product of Gastrodia elata reducing the expression level of the pro-inflammatory cytokine IL-1β in the colon;
[0024] Figure 7 This is a schematic diagram of the results of Gastrodia elata fermentation products reducing the expression levels of the inflammatory cytokine TNF-α in the colon. DETAILED DESCRIPTION
[0025] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.
[0026] Eurotium cristatum is a common edible fungus with multiple biological activities, including antioxidant, immunomodulatory and anti-inflammatory effects. As a traditional Chinese medicine, Gastrodia elata is widely used in clinical practice for its sedative, anti-epileptic and improvement of nervous system function. In recent years, studies have shown that the combination of Eurotium cristatum and Gastrodia elata may produce a synergistic effect and further enhance their pharmacological activities. However, there are no relevant studies or reports on Gastrodia elata products fermented with Eurotium cristatum. Therefore, exploring the potential efficacy of Gastrodia elata fermented with Eurotium cristatum, especially in the treatment of diseases such as ulcerative colitis, is still a research direction worthy of attention.
[0027] Specifically, in one embodiment of the present invention, a fermentation product of Gastrodia elata is provided, wherein the fermentation product of Gastrodia elata is a fermentation product produced by fermenting Gastrodia elata with Eurotium cristatum.
[0028] Specifically, the preparation method of the fermentation product of Gastrodia elata comprises the following steps:
[0029] S1. Place the Eurotium cristatum on a maltose agar plate for activation culture;
[0030] S2, after the Eurotium cristatum forms spores, scrape the Eurotium cristatum and dilute it to obtain a spore suspension;
[0031] S3, taking fresh Gastrodia elata, washing it, adding distilled water, crushing it into juice, and sterilizing it to obtain a fermentation substrate;
[0032] S4, inoculating the spore suspension into a fermentation substrate, culturing and fermenting, and obtaining a fermentation product of Gastrodia elata.
[0033] The concentration of spore suspension is (2-3)×10 8spores / mL; the mass ratio of Gastrodia elata to distilled water is 1:(1-1.5); the number of spores inoculated per gram of Gastrodia elata is (1-2)×10 7 Spores / mL.
[0034] In another embodiment of the present invention, a medicine for preventing and treating ulcerative enteritis is provided, comprising a carrier and an active ingredient, wherein the active ingredient comprises the fermentation product of the Gastrodia elata mentioned above.
[0035] In the present invention, a mouse ulcerative enteritis model was induced by administering dextran sulfate sodium (DSS) in drinking water, and the preventive and therapeutic effects of Gastrodia elata on the model were observed.
[0036] In the present invention, ulcerative enteritis was induced in mice by giving them sodium dextran sulfate in drinking water, and the changes in colon tissue appearance, pathological change sections, disease activity index (DAI) were observed, and changes in the transcription level of the inflammatory cytokine IL-1β were detected.
[0037] The fermented product of Gastrodia elata prepared in the embodiment of the present invention has good anti-inflammatory effect, has no adverse reaction, and can be used for preventing and treating ulcerative enteritis.
[0038] The following examples are some specific implementation cases and application cases of the present invention in practical applications, but are not limited thereto. Unless otherwise defined, the technical terms used in the following examples have the same meanings as those generally understood by technicians in the field to which the present invention belongs. The test reagents used in the following examples, unless otherwise specified, are all conventional biochemical reagents available from commercial companies; the experimental methods, unless otherwise specified, are all conventional methods.
[0039] Example 1: This example provides a method for preparing a fermentation product of Gastrodia elata, comprising the following steps:
[0040] S1, press 1×10 6 The amount of CFU was placed on a maltose agar plate and placed upside down in an incubator at 30°C for activation culture for 3-5 days;
[0041] S2. After the spores of Eurotium cristae are formed, use a sterile cell scraper to scrape the spores on the flat surface of the plate, count them with a hemocytometer, and then dilute to 2.5×10 8 spores / mL to obtain a spore suspension for further use;
[0042] S3, take fresh Gastrodia elata, wash it thoroughly, add distilled water according to a mass ratio of 1:1, place it in a grinder to crush it into juice, and place it at 120 ° C for high-pressure steam sterilization for 20 minutes to obtain a fermentation substrate, set aside, and cool to room temperature before fermentation;
[0043] S4, the above spore suspension is used to correspond to 1×10 7 The amount of spores was inoculated into the above fermentation substrate and fermented in an incubator at 30°C for 15 days under sterile conditions. The fermented product after 15 days of fermentation was collected and stored at -80°C. The fermented product was then freeze-dried into powder using an edible freeze dryer to obtain the fermented product of Gastrodia elata.
[0044] Example 2: This example provides a method for preparing a fermentation product of Gastrodia elata, comprising the following steps:
[0045] S1, press 1×10 6 The amount of CFU was placed on a maltose agar plate and placed upside down in an incubator at 30°C for activation culture for 3-5 days;
[0046] S2. After the spores of Eurotium cristae are formed, use a sterile cell scraper to scrape the spores on the flat surface of the plate, count them with a hemocytometer, and then dilute to 2×10 8 spores / mL to obtain a spore suspension for further use;
[0047] S3, take fresh Gastrodia elata, wash it thoroughly, add distilled water according to the mass ratio of 1:1.2, place it in a grinder to crush and squeeze it into juice, and place it at 120 ° C for high pressure steam sterilization for 20 minutes to obtain a fermentation substrate, set aside, and cool to room temperature before fermentation;
[0048] S4, the above spore suspension is used to treat 1.5×10 7 The amount of spores was inoculated into the above fermentation substrate and fermented in an incubator at 30°C for 15 days under sterile conditions. The fermented product after 15 days of fermentation was collected and stored at -80°C. The fermented product was then freeze-dried into powder using an edible freeze dryer to obtain the fermented product of Gastrodia elata.
[0049] Example 3: This example provides a method for preparing a fermentation product of Gastrodia elata, comprising the following steps:
[0050] S1, press 1×10 6 The amount of CFU was placed on a maltose agar plate and placed upside down in an incubator at 30°C for activation culture for 3-5 days;
[0051] S2. After the spores of Eurotium cristata are formed, use a sterile cell scraper to scrape the spores on the flat surface of the plate, count them with a hemocytometer, and then dilute to 3×10 8 spores / mL to obtain a spore suspension for further use;
[0052] S3, take fresh Gastrodia elata, wash it thoroughly, add distilled water according to the mass ratio of 1:1.5, place it in a grinder to crush and squeeze it into juice, and place it at 120 ° C for high pressure steam sterilization for 20 minutes to obtain a fermentation substrate, set aside, and cool to room temperature before fermentation;
[0053] S4, the above spore suspension is used to correspond to 2×10 7 The amount of spores was inoculated into the above fermentation substrate and fermented in an incubator at 30°C for 15 days under sterile conditions. The fermented product after 15 days of fermentation was collected and stored at -80°C. The fermented product was then freeze-dried into powder using an edible freeze dryer to obtain the fermented product of Gastrodia elata.
[0054] Example 4: This example provides a method for preparing a fermentation product of Gastrodia elata, comprising the following steps:
[0055] S1, press 1×10 6 The amount of CFU was placed on a maltose agar plate and placed upside down in an incubator at 30°C for activation culture for 3-5 days;
[0056] S2. After the spores of Eurotium cristata are formed, use a sterile cell scraper to scrape the spores on the flat surface of the plate, count them with a hemocytometer, and then dilute to 2.8×10 8 spores / mL to obtain a spore suspension for further use;
[0057] S3, take fresh Gastrodia elata, wash it thoroughly, add distilled water according to the mass ratio of 1:1.3, put it in a grinder to crush it into juice, and place it at 120 ° C for high pressure steam sterilization for 20 minutes to obtain a fermentation substrate, set aside, and cool to room temperature before fermentation;
[0058] S4, the above spore suspension is used to correspond to 1.2×10 7 The amount of spores was inoculated into the above fermentation substrate and fermented in an incubator at 30°C for 15 days under sterile conditions. The fermented product after 15 days of fermentation was collected and stored at -80°C. The fermented product was then freeze-dried into powder using an edible freeze dryer to obtain the fermented product of Gastrodia elata.
[0059] Example 5: This example provides a method for preparing a fermentation product of Gastrodia elata, comprising the following steps:
[0060] S1, press 1×10 6 The amount of CFU was placed on a maltose agar plate and placed upside down in an incubator at 30°C for activation culture for 3-5 days;
[0061] S2. After the spores of Eurotium cristae are formed, use a sterile cell scraper to scrape the spores on the flat surface of the plate, count them with a hemocytometer, and then dilute to 2.5×10 8 spores / mL to obtain a spore suspension for further use;
[0062] S3, take fresh Gastrodia elata, wash it thoroughly, add distilled water according to the mass ratio of 1:1.4, place it in a grinder to crush and squeeze it into juice, and place it at 120 ° C for high pressure steam sterilization for 20 minutes to obtain a fermentation substrate, set aside, and cool to room temperature before fermentation;
[0063] S4, the above spore suspension is used to correspond to 1.8×10 7 The amount of spores was inoculated into the above fermentation substrate and fermented in an incubator at 30°C for 15 days under sterile conditions. The fermented product after 15 days of fermentation was collected and stored at -80°C. The fermented product was then freeze-dried into powder using an edible freeze dryer to obtain the fermented product of Gastrodia elata.
[0064] Example 6: This example is an experiment on the effect of the fermentation product of Gastrodia elata on ulcerative enteritis in mice induced by sodium dextran sulfate, as follows:
[0065] Preparation of dextran sulfate sodium-induced mouse ulcerative colitis model: C57 mice (8 weeks old) were given drinking water containing 1.5% DSS for 7 days.
[0066] Grouping: The control group (CON group) was given the same volume of distilled water for one week; the modeling group (DSS group) was given 1.5% DSS drinking water for one week; the 5-aminosalicylic acid group (5-ASA group) was given 1.5% DSS drinking water for one week, and at the same time, 50 mg / kg 5-ASA 200 μL (dissolved in 0.9% NaCl solution) was gavaged for one week; the Gastrodia elata fermentation product group (E.YG group) was given 1.5% DSS drinking water for one week, and at the same time, 200 mg / kg of Gastrodia elata fermentation product 200 μL (dissolved in 0.9% NaCl solution) was gavaged for one week. On the eighth day, the mice were euthanized and colon tissues were taken.
[0067] Visual observation of tissue morphology: Observe the colon morphology among the groups.
[0068] Colon length measurement: The colon length of each group was measured and compared.
[0069] Mouse body weight changes: The body weight of each mouse was weighed and recorded every day.
[0070] Disease index score: Starting from the first day of modeling, DAI score test was performed twice a day. The DAI score details are shown in Table 1:
[0071] Table 1
[0072]
[0073] Detection of the expression level of inflammatory cytokines: The colon tissues of each group were ground into a 10% solution prepared with PBS to prepare tissue homogenate, and the corresponding Elisa kit was used to detect the level changes.
[0074] The above experimental results are as follows Figure 1-Figure 7 As shown. Figure 1 It can be seen that the colon of the mice in the normal group was in good condition, without symptoms such as congestion and edema. However, the colon of the mice in the model group showed obvious symptoms such as edema, congestion, adhesion, etc., and the colon length was significantly lower than that of the normal group. The 5-ASA group and the Gastrodia elata fermentation product experimental group showed significantly less edema, congestion, and adhesion symptoms in the colon than the model group, and the symptoms of the positive control group and the Gastrodia elata fermentation product group were similar. In other words, daily administration of Gastrodia elata fermentation products for one week can relieve ulcerative colitis in mice.
[0075] Depend on Figure 2 It can be seen that the colon tissue structure of the normal control group is intact, the goblet cells are intact and the number is normal. The colon tissue structure of the mice in the model group was severely damaged, the villus structure was messy and damaged, the muscle layer structure was incomplete, and the inflammatory cell infiltration was serious. The intestinal mucosal morphology of the 5-ASA group and the Gastrodia elata fermentation product experimental group was significantly improved, the intestinal villus boundaries were clear, the mucosal morphology was relatively complete, and the number of goblet cells was normal. Daily administration of Gastrodia elata fermentation products for one week can relieve ulcerative enteritis in mice to varying degrees.
[0076] Depend on Figure 3 It can be seen that the goblet cells in the colon of normal mice are rich in content, compact in structure, and neatly arranged, and the outside of the goblet cells are wrapped by a mucus layer. Compared with the normal group, the goblet cells in the enteritis group were severely damaged, the goblet cells in the intestinal crypts were extremely low, and only part of the intestine was protected by the mucus layer. The goblet cells and mucus layer in the 5-ASA group and the Gastrodia elata fermentation product experimental group were similar to those in the control group. Daily administration of Gastrodia elata fermentation products for one week can relieve ulcerative enteritis in mice to varying degrees.
[0077] Depend on Figure 4 It can be seen that the colon length of mice in the positive control group was significantly shorter than that of mice in the normal control group. The colon length of the 5-ASA group and the Gastrodia elata fermentation product experimental group was close to that of the normal control group. Daily administration of Gastrodia elata fermentation products for one week can alleviate ulcerative enteritis in mice to varying degrees.
[0078] Depend on Figure 5It can be seen that compared with the DAI of mice in the normal control group, the colon DAI of mice in the positive control group was significantly increased. The colon DAI of the drug control group was close to that of the normal control group. Daily administration of the fermented product of Gastrodia elata for one week can alleviate ulcerative enteritis in mice to varying degrees.
[0079] Depend on Figure 6 It can be seen that compared with the mice in the normal control group, the expression level of the inflammatory cytokine IL-1β in the modeling group was higher. The expression level of the inflammatory cytokine IL-1β in the mice in the Gastrodia elata treatment group was between the control group and the modeling group. The expression level of the inflammatory cytokine IL-1β in the drug control group was close to that in the colon of mice in the control group. Giving Gastrodia elata fermentation products daily for one week can relieve ulcerative enteritis in mice to varying degrees.
[0080] Depend on Figure 7 It can be seen that compared with the mice in the normal control group, the expression level of the inflammatory cytokine TNF-α in the modeling group was higher. The expression level of the inflammatory cytokine TNF-α in the mice treated with the fermented product of Gastrodia elata was between the control group and the modeling group. The expression level of the inflammatory cytokine TNF-α in the 5-ASA control group was close to that in the mice colon of the fermented product of Gastrodia elata group. Giving the fermented product of Gastrodia elata to the group daily for one week can relieve ulcerative enteritis in mice to varying degrees.
[0081] In summary, the fermentation product of Gastrodia elata prepared in the embodiment of the present invention has a good effect on enteritis, has no adverse reactions, can alleviate DSS-induced ulcerative enteritis inflammation in mice and repair the colon barrier, and has a good application prospect in preventing, treating or improving ulcerative enteritis.
[0082] Based on the above-mentioned ideal embodiments of the present invention, the relevant staff can make various changes and modifications without departing from the technical concept of the present invention through the above-mentioned description. The technical scope of the present invention is not limited to the contents of the specification.
Claims
1. A fermented product of Gastrodia elata, characterized in that: The fermentation product of Gastrodia elata is a fermentation product produced by fermenting Gastrodia elata with Eurotium cristatum.
2. The fermented product of Gastrodia elata according to claim 1, characterized in that: The preparation method of the fermentation product of Gastrodia elata comprises the following steps: Place the Eurotium cristatum on a maltose agar plate for activation culture; After the spores of the Eurotium cristatum are formed, the spores are scraped and diluted to obtain a spore suspension; Take fresh Gastrodia elata, wash it, add distilled water, crush it into juice, and sterilize it to obtain a fermentation substrate; The spore suspension is inoculated into a fermentation substrate for culturing and fermentation to obtain a fermentation product of Gastrodia elata.
3. The fermented product of Gastrodia elata according to claim 2, characterized in that: The concentration of the spore suspension is (2-3)×10 8 Spores / mL.
4. The fermented product of Gastrodia elata according to claim 2, characterized in that: The mass ratio of the Gastrodia elata to distilled water is 1:(1-1.5).
5. The fermented product of Gastrodia elata according to claim 2, characterized in that: The number of spores inoculated per gram of Gastrodia elata is (1-2)×10 7 Spores / mL.
6. Use of the fermentation product of Gastrodia elata as claimed in any one of claims 1 to 5 in the preparation of a medicament for preventing and treating ulcerative enteritis.
7. A drug for preventing and treating ulcerative enteritis, comprising a carrier and an active ingredient, characterized in that: The active ingredient comprises the fermentation product of Gastrodia elata as claimed in any one of claims 1 to 5.