Artichoke and Hovenia dulcis fermented composition and its application in protecting liver

Abstract

The present invention relates to the field of drug fermentation, and in particular to an artichoke and Hovenia dulcis fermentation composition and its liver protection application. The present invention relates to a Chinese medicine fermentation composition with medicinal and edible properties, as well as a preparation method and liver protection application of the fermentation composition. The Chinese medicine fermentation composition with medicinal and edible properties of the present invention mainly comprises Hovenia dulcis, dried shiitake mushrooms, kudzu root, Poria cocos, wolfberry, turmeric, chrysanthemum, ginger and probiotics. The fermentation strains used include probiotics. The present invention solves the problem of poor efficacy of Chinese medicine prescriptions with medicinal and edible properties. The present invention provides a Chinese medicine fermentation composition with medicinal and edible properties, and the efficacy of the composition is improved by about 2 times compared with that before fermentation; the present invention provides a preparation method of the fermentation composition, including an optimal extraction process, an optimal strain, an optimal strain combination and an optimal fermentation process; through prescription screening research, an optimal Chinese medicine prescription with medicinal and edible properties is invented.

Description

Technical Field

[0001] The invention relates to the field of drug fermentation, and in particular to an artichoke and Hovenia dulcis fermentation composition and a liver-protecting application thereof. Background Art

[0002] With the change of people's lifestyle, the incidence of liver disease is getting higher and higher, especially non-viral liver diseases such as drug-induced liver injury, alcoholic liver disease, non-alcoholic fatty liver disease and autoimmune liver disease, which have an obvious overall upward trend and gradually develop into fatty liver, hepatitis, liver fibrosis and liver cancer. Factors that cause liver disease include chemical liver injury such as alcohol, hyperlipidemia, hepatitis viruses such as hepatitis B virus. According to the disease, there are mainly alcoholic liver disease, non-alcoholic liver disease, hepatitis B, hepatitis C, etc. According to a survey report of 5,757,335 participants, the prevalence of hepatic steatosis, severe hepatic steatosis, advanced liver fibrosis and cirrhosis were 44.4%, 10.6%, 2.9% and 0.87% respectively. These surveys and studies highlight the high prevalence and heavy public health burden of chronic liver disease in China, and effective prevention and control measures are urgently needed. However, there is still a lack of specific therapeutic drugs, which leads to the high incidence of the disease and the continuous development of cirrhosis and liver cancer. Moreover, the side effects of current liver disease treatment drugs are serious, resulting in "disease A is not cured, and disease B is added". Traditional Chinese medicine with the same origin as medicine and food is highly safe and very suitable for the treatment of liver disease, liver protection and hangover. However, the main problem with traditional Chinese medicine with the same origin as medicine and food is that the formula is unscientific or the efficacy is low. Therefore, the invention of the best traditional Chinese medicine formula with the same origin as medicine and food for liver disease and improving the efficacy are urgently needed products. Summary of the invention

[0003] Based on the above, the effect of Chinese medicine compound preparations with medicinal and edible properties is poor. The present invention solves the problem of poor efficacy of Chinese medicine prescriptions with medicinal and edible properties. On the one hand, the present invention improves the efficacy of Chinese medicine with medicinal and edible properties in treating liver diseases, protecting the liver and sobering up by about 2 times through fermentation technology. On the other hand, through prescription screening research, the best Chinese medicine prescription with medicinal and edible properties is invented.

[0004] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0005] The present invention relates to a Chinese medicine fermentation composition that is both medicinal and edible, as well as a preparation method and liver protection application of the fermentation composition. The liver protection application includes preventing liver disease, treating liver disease, protecting the liver, managing chronic liver disease, and protecting the liver after alcohol detoxification, etc. The liver protection application further includes reducing transaminase, increasing acetaldehyde dehydrogenase activity, and anti-oxidation, etc. The transaminase includes alanine aminotransferase, aspartate aminotransferase, etc., and the anti-oxidation includes scavenging hydroxyl free radicals, DPPH, etc. The liver disease includes fatty liver, hepatitis, liver fibrosis, liver abscess, cirrhosis, liver cancer, etc., and the liver disease further includes alcoholic liver disease, non-alcoholic liver disease, immune liver disease, chemical liver damage, viral liver disease, genetic metabolic liver disease, etc. The alcoholic liver disease includes alcoholic fatty liver, alcoholic hepatitis, alcoholic liver fibrosis, alcoholic cirrhosis, alcoholic liver cancer, etc., the non-alcoholic liver disease includes non-alcoholic fatty liver, non-alcoholic hepatitis, non-alcoholic liver fibrosis, non-alcoholic cirrhosis, non-alcoholic liver cancer, etc., chemical liver injury includes alcoholic liver injury, drug-induced liver injury, chemical substance liver injury, etc., viral liver disease includes viral hepatitis, viral liver fibrosis, viral cirrhosis, viral liver cancer, etc., and the viruses include hepatitis viruses such as A, B, C, D and E.

[0006] The Chinese medicine fermentation composition of the present invention mainly includes Hovenia dulcis, dried shiitake mushroom, kudzu root, Poria cocos, wolfberry, turmeric, chrysanthemum and ginger. Formula idea: Hovenia dulcis and kudzu root are the main drugs in the formula, which have the effects of clearing away heat and relieving restlessness, sweating and relieving muscles, and detoxifying alcohol; dried shiitake mushroom, wolfberry and chrysanthemum are the assistant drugs, which have the functions of nourishing liver and kidney, regulating endocrine, accelerating liver detoxification and repairing damaged liver cells; Poria cocos and turmeric are adjuvant drugs, which have the functions of detoxifying alcohol, promoting water and removing dampness, promoting gallbladder and protecting liver; ginger is the guiding drug, which has the effects of protecting liver, warming the middle and stopping vomiting. The fermentation strains used include probiotics.

[0007] Preferably, the above composition comprises 100-390 parts by weight of Hovenia dulcis, 80-300 parts by weight of dried shiitake mushrooms, 60-190 parts by weight of Pueraria root, 30-150 parts by weight of Poria cocos, 30-100 parts by weight of Lycium barbarum, 20-50 parts by weight of turmeric, 10-40 parts by weight of chrysanthemum, and 5-20 parts by weight of ginger.

[0008] Preferably, the above composition comprises 110-390 parts by weight of Hovenia dulcis, 80-290 parts by weight of dried shiitake mushrooms, 60-190 parts by weight of Pueraria root, 38-140 parts by weight of Poria cocos, 30-90 parts by weight of wolfberry, 20-50 parts by weight of turmeric, 13-40 parts by weight of chrysanthemum, and 6-19 parts by weight of ginger.

[0009] Preferably, the above composition comprises 150-300 parts by weight of Hovenia dulcis, 100-200 parts by weight of dried shiitake mushrooms, 75-150 parts by weight of Pueraria root, 50-130 parts by weight of Poria cocos, 30-80 parts by weight of Lycium barbarum, 20-50 parts by weight of turmeric, 25-40 parts by weight of chrysanthemum, and 8-15 parts by weight of ginger.

[0010] Preferably, the composition comprises 200 parts by weight of Hovenia dulcis, 150 parts by weight of dried shiitake mushrooms, 100 parts by weight of Pueraria root, 75 parts by weight of Poria cocos, 50 parts by weight of Lycium barbarum, 30 parts by weight of turmeric, 25 parts by weight of chrysanthemum, and 10 parts by weight of ginger.

[0011] Preferably, the composition comprises 300 parts by weight of Hovenia dulcis, 100 parts by weight of dried shiitake mushrooms, 150 parts by weight of Pueraria root, 50 parts by weight of Poria cocos, 30 parts by weight of Lycium barbarum, 20 parts by weight of Curcuma longa, 20 parts by weight of Chrysanthemum and 8 parts by weight of ginger.

[0012] Preferably, the composition comprises 150 parts by weight of Hovenia dulcis, 200 parts by weight of dried shiitake mushrooms, 75 parts by weight of Pueraria root, 130 parts by weight of Poria cocos, 80 parts by weight of Lycium barbarum, 50 parts by weight of Curcuma longa, 40 parts by weight of Chrysanthemum and 15 parts by weight of ginger.

[0013] The probiotics of the present invention include Lactobacillus, Lactobacillus, Lactobacillus and Lactobacillus mucilaginosus.

[0014] The Lactobacillus genus described in the present invention includes Lactobacillus casei, Lactobacillus paracasei and Lactobacillus rhamnosus.

[0015] The Lactobacillus genus of the present invention includes Lactobacillus plantarum.

[0016] The Lactobacillus species of the present invention include Lactobacillus acidophilus, Lactobacillus delbrueckii subspecies bulgaricus, and Lactobacillus helveticus.

[0017] The genus Lactobacillus mucilaginosus of the present invention includes fermentative Lactobacillus mucilaginosus.

[0018] The inoculation amount of the probiotics of the present invention is 1-10%, preferably 2-5%, and more preferably 2%, 3%, 4% or 5%.

[0019] Preferably, the OD of the probiotics 600 The value is 1-8, preferably 3-6.

[0020] The pH value of the probiotics during inoculation of the present invention is 5-7, preferably 5-6.

[0021] The fermentation time of the probiotic fermentation of the present invention is 16 to 42 hours, and more preferably 20 hours, 24 hours, 28 hours and 32 hours.

[0022] The total flavonoids in the edible and medicinal Chinese medicine fermentation composition of the present invention are increased after fermentation.

[0023] Preferably, the above-mentioned Chinese medicinal fermentation composition which is both medicine and food also includes one or more of artichoke, corn oligopeptide, oyster, oligofructose, and vitamin B6, the artichoke includes artichoke juice powder, the corn oligopeptide includes corn oligopeptide powder, the oyster includes oyster peptide powder, and the oligofructose includes oligofructose powder or oligofructose liquid. The invention further comprises 10 to 40 parts by weight of artichoke juice powder, 5 to 20 parts by weight of corn oligopeptide powder, 2 to 10 parts by weight of oyster peptide powder, 100 to 500 parts by weight of oligofructose, and 0.003 to 0.012 parts by weight of vitamin B6, and further preferably 20 to 30 parts by weight of artichoke juice powder, 5 to 15 parts by weight of corn oligopeptide powder, 2 to 10 parts by weight of oyster peptide powder, 150 to 450 parts by weight of oligofructose, and 0.003 to 0.012 parts by weight of vitamin B6, and further preferably 20 parts by weight of artichoke juice powder, 10 parts by weight of corn oligopeptide powder, 5 parts by weight of oyster peptide powder, 225 parts by weight of oligofructose, and 0.0064 parts by weight of vitamin B6, and further preferably 30 parts by weight of artichoke juice powder, 15 parts by weight of corn oligopeptide powder, 5 parts by weight of oyster peptide powder, 300 parts by weight of oligofructose, and 0.003 to 0.012 parts by weight of vitamin B6. 0.005 weight part, further preferably 10 weight parts of artichoke juice powder, 15 weight parts of corn oligopeptide powder, 10 weight parts of oyster peptide powder, 150 weight parts of oligofructose, and 0.005 weight part of vitamin B6.

[0024] Preferably, the above-mentioned Chinese medicinal fermentation composition which is both medicine and food also includes erythritol, concentrated pomegranate juice, concentrated pear juice, concentrated mango juice, etc., and further preferably includes 50-200 parts by weight of erythritol, 50-200 parts by weight of concentrated pomegranate juice, 150-500 parts by weight of concentrated pear juice, 50-200 parts by weight of concentrated mango juice, etc., and further preferably includes 100 parts by weight of erythritol, 100 parts by weight of concentrated pomegranate juice, 318 parts by weight of concentrated pear juice, and 100 parts by weight of concentrated mango juice.

[0025] The preparation process of the edible and medicinal Chinese medicine fermentation composition of the present invention comprises extraction, concentration, fermentation, preparation and filling.

[0026] The extraction process is preferably one extraction or two extractions; the amount of water added in the one extraction is preferably 5 to 20 times, more preferably 10 to 16 times, and more preferably 14 times; the amount of water added in the two extractions is preferably 5 to 20 times, more preferably 8 to 15 times for the first extraction and 6 to 12 times for the second extraction, more preferably 12 or 14 times for the first extraction and 8 times for the second extraction.

[0027] In the extraction process, the extraction time for one extraction is preferably 1 to 3 hours, more preferably 1 hour, 2 hours or 3 hours, and more preferably 1 hour; in the extraction process, the extraction time for two extractions is preferably 1 to 3 hours, more preferably 1 to 2 hours for the first extraction and 1 to 2 hours for the second extraction, more preferably 1 or 2 hours for the first extraction and 1 hour for the second extraction.

[0028] In one embodiment, the composition of the present invention is a simple mixture. The preparation process is to simply mix the raw materials.

[0029] In another embodiment, the composition of the present invention can also be made into a Chinese patent medicine. The dosage form of Chinese patent medicine includes but is not limited to pills, powders, ointments, tablets, granules (granules), lozenges, capsules, liquid preparations, and tinctures.

[0030] (1) Pills

[0031] Pills are spherical or quasi-spherical solid preparations made from fine powder or extracts of medicinal materials plus suitable adhesives or excipients. They are one of the oldest dosage forms of Chinese patent medicines. Depending on the different adhesives, pills are divided into honey pills, water-honey pills, water pills, paste pills, concentrated pills, micro-pills and other types.

[0032] Concentrated pills are preferably used. Concentrated pills are made of decoction or extract of all or part of the medicinal materials, suitable excipients or fine powder of the medicine, and suitable adhesives. According to the different adhesives, they are divided into concentrated honey pills, concentrated water pills, and concentrated water-honey pills. Concentrated pills are small in size, high in the content of active ingredients of the medicine, easy to take, and slowly dissolved and absorbed in the body.

[0033] In a preferred embodiment, the decoction or extract of all or part of the medicinal materials, adjuvants or fine powder of the medicine, and adhesive are added to form concentrated pills. Preferably, the decoction or extract of all the medicinal materials, adhesive and optional adjuvants are added to form concentrated pills.

[0034] (2) Powder

[0035] Powder is a powdered preparation made by mixing one or more medicinal materials. It is divided into powder for internal use and powder for external use.

[0036] One of the ancient dosage forms. Powders have a wide range of therapeutic applications, disperse quickly after taking, and take effect quickly. They are also easy to make, easy to carry, and save on medicinal materials.

[0037] (3) Ointment

[0038] Ointment is one of the drug dosage forms. Ointment is a dosage form made by boiling and concentrating drugs with water or vegetable oil. There are two types: oral and external use. Oral ointment is usually divided into three types: extract, extract, and decoction; external ointment is divided into two types: ointment and hard plaster. Among them, oral decoction and external ointment are common dosage forms of ointment. Oral decoction is also called ointment. It is made by boiling the drug with water, filtering the residue, adding rock sugar, honey, etc., and boiling it into a thick paste. It can be taken for a long time. It is often used for chronic diseases or weak people. External ointment is called ointment. Generally, ointment is made by adding beeswax to cottonseed oil or peanut oil, heating and melting, adding the applied drug powder while hot, stirring continuously, and condensing. Drugs that are easily volatile, such as borneol and camphor, can be added and stirred after the ointment cools. External ointment is generally used for external application on skin sores, scabies, etc.

[0039] The present invention preferably uses a decoction paste, which is a semi-solid preparation made by boiling medicinal materials with water, removing residues and concentrating, and adding a sweetener, also known as a paste. The sweetener can be refined honey or sugar, xylitol, preferably xylitol. The decoction paste has the characteristics of fast absorption, high concentration, small volume, easy storage, and can be taken for a long time. It has the effect of nourishing and conditioning, and is used to treat chronic

[0040] Those with sexually transmitted diseases and chronic illness.

[0041] (4) Tablets

[0042] Tablets are sheet preparations compressed from fine powder or extracts of medicinal materials and suitable excipients or fine powder of medicinal materials. They are divided into extract tablets, semi-extract tablets and whole powder tablets, etc., and are one of the commonly used modern dosage forms. Tablets are small in size, accurate in dosage, easy to disintegrate and take effect quickly, and have the advantages of high production efficiency, low cost, and convenient to take, store and transport. Tablets are suitable for various diseases. The present invention preferably uses extract tablets.

[0043] (5) Granules

[0044] Granules (powders) are granular preparations made from medicinal extracts and appropriate excipients or medicinal powder. They are new dosage forms developed on the basis of decoctions, powders and syrups. There are two types: granules and blocks, and they are divided into different types such as soluble, suspension, effervescent, sugar-containing and sugar-free. Granules are small in size, light in weight, easy to take, good in taste, and quick in effect. They are mostly used for tonic, cough-relieving, heat-clearing and other drugs.

[0045] Preferably, a water extract (decoction) is used in the preparation.

[0046] (6) Tablets

[0047] Tablets are solid preparations made of fine powder of medicinal materials and appropriate amount of adhesive such as honey, glutinous rice flour or the viscosity of the medicinal materials themselves into a prescribed shape. They can be taken orally or externally. The effect of oral administration is similar to that of paste pills. For external use, they are often ground into juice with water or vinegar and applied to the affected area. Tablets are mostly used for chewing.

[0048] Preferably, the tablet of the present invention is an oral tablet.

[0049] (7) Capsules

[0050] Capsules include hard capsules and soft capsules.

[0051] Hard capsules are a dosage form made by filling a proper amount of medicinal extracts, medicinal extracts and powders or excipients into uniform powder or granules in hard capsules. They are mainly taken orally. Hard capsules are neat and beautiful in appearance, easy to swallow, can cover up the bad smell of drugs, disintegrate quickly and are well absorbed. They are suitable for drugs that are sensitive to light, unstable, or unstable when exposed to moisture or heat, or drugs with specific odors, or drugs that need to be released at a fixed time. Drugs for children and drugs that are highly irritating to the gastric mucosa should not be made into capsules.

[0052] Soft capsules are a dosage form made by sealing oils or liquid drugs or suspensions that have no dissolving effect on gelatin and other capsule materials in capsule materials. The characteristics are similar to hard capsules. Hard capsules and soft capsules are treated by appropriate methods or processed with other pharmaceutical polymer materials to make the capsule shell insoluble in gastric juice, but disintegrate in intestinal juice to release active ingredients, which are enteric-coated capsules.

[0053] Preferably, the capsule of the present invention is prepared by adding auxiliary materials to the medicinal material extract obtained by decoction or extraction to form a uniform powder.

[0054] The dosage form is made by filling powder or granules into hard capsules.

[0055] (8) Aqueous solution

[0056] Aqueous preparations are aqueous solutions of Chinese medicine compositions. The medicine is evenly dispersed in water in the form of ions or molecules. The concentration of the medicine depends on the water solubility of the original medicine, which is generally its maximum solubility. It is diluted with water when used. In order to completely release the active ingredients in the medicine, the aqueous preparation can be prepared by hot water decoction and the number of decoctions can be 1, 2 or more times.

[0057] (9) Tincture

[0058] Tincture is a clear liquid preparation made by leaching or dissolving a drug with a specified concentration of ethanol, and can also be made by diluting a fluid extract. It is divided into two types: oral and external. Tincture preparation does not require heating, the ingredients are relatively pure, the content of active ingredients is high, the dosage is accurate, and the absorption is rapid. It is suitable for preparing preparations containing volatile ingredients or heat-labile ingredients. Preferably, the tincture of the present invention is an oral tincture.

[0059] Of course, Chinese patent medicine dosage forms can also be taken. Based on the above decoction dosage, the dosage of Chinese patent medicine dosage forms can be determined through clinical trials in this field.

[0060] The product of the present invention is preferably prepared according to the weight ratio of the present invention and processed according to the preparation process of the present invention into 50 mL per bottle, and 1 to 2 bottles per day are recommended.

[0061] The present invention relates to a Chinese medicine fermentation composition with medicinal and edible properties, as well as a preparation method and liver protection application of the fermentation composition. The Chinese medicine fermentation composition with medicinal and edible properties mainly comprises Hovenia dulcis, dried shiitake mushrooms, kudzu root, Poria cocos, wolfberry, turmeric, chrysanthemum, ginger and probiotics. The fermentation strains used include probiotics.

[0062] The invention solves the problem of poor efficacy of traditional Chinese medicine prescriptions with the same origin of medicine and food.

[0063] 1. The present invention provides a Chinese medicine fermentation composition that is both medicine and food, and the efficacy of the composition is improved by about 2 times compared with that before fermentation; 2. The present invention provides a preparation method of the fermentation composition, including an optimal extraction process, an optimal bacterial strain, an optimal bacterial strain combination and an optimal fermentation process; 3. Through formula screening research, the optimal Chinese medicine formula that is both medicine and food is invented.

[0064] The unexpected effects achieved by the present invention are:

[0065] 1. Not all strains can ferment, and thermophilic Streptococcus is not suitable for growth (see the example strain research);

[0066] 2. Traditional Chinese medicine fermentation formula: Compared with before fermentation, the DPPH free radical scavenging rate after fermentation is significantly improved (see Example);

[0067] 3. Traditional Chinese medicine fermentation formula: Through zebrafish experiments, compared with before fermentation, the function of reducing the activity of alanine aminotransferase and aspartate aminotransferase has increased by about 2 times, and not all bacterial combinations have improved. After fermentation, the activity of some bacterial combinations has decreased (see Example).

[0068] 4. Traditional Chinese medicine fermentation formula + preparation efficacy formula: The present invention selects traditional Chinese medicines that are both medicinal and edible, but through zebrafish experiments, the liver protection effect of reducing aspartate aminotransferase activity is higher than that of clinical treatment drugs: 2 bags (bottles) of the present invention's product / day> 4 capsules of Metadoxine Capsules / day> 1 bottle of marketed health food / day. Metadoxine Capsules: clinically used drugs for the treatment of alcoholic liver disease; marketed health food: blue hat health food, function: auxiliary protection of chemical liver damage and relief of physical fatigue (see examples).

[0069] 5. Traditional Chinese medicine fermentation formula + preparation of efficacy formula: through the efficacy of zebrafish, the activity of acetaldehyde dehydrogenase is improved. The product of the present invention is > the health food that has been on the market. The health food that has been on the market is the product that has been promoted as a hangover-relieving product on the market (see the examples).

[0070] 6. Traditional Chinese medicine fermentation formula + preparation of efficacy formula: The dosage of the main drug of the product of the present invention is increased by 2 times, and fermentation is carried out according to the process of the present invention. The liver protection effect is not only not improved, but significantly reduced, the dosage of medicinal materials is saved, the best ratio is found, and unexpected effects are achieved (see Examples).

[0071] 7. Traditional Chinese medicine fermentation formula: Through liver cell experiments, the liver protection effect within the scope of the present invention is the best, which is significantly better than the formula ratio outside the scope of the present invention. DETAILED DESCRIPTION

[0072] The present invention discloses an artichoke and Hovenia dulcis fermentation composition and its liver protection application. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve it. It is particularly important to point out that all similar substitutions and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0073] In the artichoke and Hovenia dulcis fermentation composition and liver protection application thereof provided by the invention, the raw materials and reagents used can be purchased from the market.

[0074] The present invention will be further described below in conjunction with embodiments:

[0075] Example 1 Extraction process study

[0076] 1. Formulation

[0077]

[0078] 2. Research on extraction process of effective ingredients

[0079] (1) Study on water addition amount and extraction time

[0080] 1) Amount of water added in two extractions: 14 / 10, 14 / 8, 12 / 10, 12 / 8, 10 / 10, 10 / 8 Time: 1 / 1, 1.5 / 1, 2 / 1.

[0081] 2) Amount of water added in one extraction: According to the volume of the laboratory and pilot extraction tanks, the amount of water added in one extraction cannot exceed 16 times the dosage of the drug, so the selected extraction ratio is 12 / 14 / 16.

[0082] 3) Detection indicators: total flavonoids (detection method: 2020 Method 1 for the determination of total flavonoids in health foods) and total crude polysaccharides (detection method: 2015- Determination of crude polysaccharides in exported plant-based foods - phenol-sulfuric acid method).

[0083] 3. Results

[0084] (1) Results of two extractions

[0085]

[0086] Results: According to the above test results, it can be seen that the No. 6 combination of two extractions has the highest total flavonoids and crude polysaccharides in the extract after extraction, so the No. 6 combination was finally selected as the best process for two extractions.

[0087] (2) One-time extraction results

[0088] Results: From the above extraction results, it can be seen that after extraction with combination No. 4, the content of total flavonoids and crude polysaccharides is the highest, so extraction combination No. 4 is selected as the one-time extraction process.

[0089] 4. Determination of extraction process

[0090] According to the above, the best extraction process obtained by two extractions and the best extraction process obtained by one extraction, after comparing the two processes, the final extraction process selected is: two extractions, the first water addition amount is 14 times the total amount of medicine, soaked at 45℃ for 1 hour, and then boiled for 2 hours; the second water addition amount is 8 times the total amount of medicine, boiled for 1 hour. The theoretical concentration multiple is determined according to the preparation process (generally 2-6 times).

[0091] Example 2 Bacterial strain research

[0092] 1. Find relevant literature and conduct experiments (to detect total flavonoids) based on the bacterial strains in the laboratory.

[0093] 1.1 Single factor strain screening to select better strains Nine strains (Lactobacillus plantarum, Lactobacillus casei, Lactobacillus paracasei, Streptococcus thermophilus, Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus rhamnosus, and Lactobacillus mucosa fermentum) were initially used for the test. The fermentation was allowed to stand at 37°C for 72 h, and samples were taken every 24 h for testing of pH, OD600, and active ingredients (total flavonoids). The results are shown in Table 1.

[0094] 1.2 Results Table 1 pH values ​​of fermentation broths of various strains at different fermentation time periods

[0095]

[0096] Results: According to Table 1, the pH values ​​of 9 experimental bacteria were measured at different time periods during the fermentation process. Among them, the pH values ​​of the fermentation broth inoculated with thermophilic Streptococcus, Lactobacillus delbrueckii subspecies bulgaricus and fermented Lactobacillus mucosa were all above 4.20, which did not meet the fermentation requirements. The other 6 bacteria, Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus paracasei, Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus casei, grew well, and the pH values ​​were all below 4.0, which met the fermentation requirements. Among them, Lactobacillus paracasei, Lactobacillus rhamnosus and Lactobacillus casei had the lowest pH during the fermentation process. The lower the pH during the fermentation process, the stronger the ability of the bacteria to utilize the carbon source during the fermentation process, and the better the ability to decompose the effective components of traditional Chinese medicine. Therefore, it shows that the bacterial combination of the experimental group fermented the extract more thoroughly and was more conducive to fermentation.

[0097] Table 2 OD600 values ​​of biological activity of each strain at different fermentation stages

[0098]

[0099] Graphpad Prism 9.5.1 software was used for statistical analysis of the data, and all experimental data were expressed as mean + SD.

[0100] result:

[0101] According to Table 2, OD600 value determination found 9 kinds of test bacteria in each time period of fermentation process, wherein the fermentation liquid OD600 value of inoculation of thermophilic streptococcus and Lactobacillus delbrueckii subspecies bulgaricus was all below 1.5, indicating that bacterial species did not grow substantially in the whole fermentation process, and could not meet the fermentation requirement. Among the remaining 7 kinds of bacteria, after fermentation for 48h, Lactobacillus paracasei, Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus casei grew better, and the OD600 value reached about 6.0, indicating that bacterial species can make full use of the nutrients in the culture medium, and then illustrating that bacterial species can effectively decompose or transform the Chinese medicine components in the fermentation medium.

[0102] Table 3 Total flavonoid content of fermentation broth of each strain at different fermentation time periods (unit: mg / mL)

[0103]

[0104] Graphpad Prism 9.5.1 software was used for statistical analysis of the data, and all experimental data were expressed as mean + SD.

[0105] result:

[0106] According to Table 3, it can be seen that the total flavonoid content during the fermentation process is the highest from the beginning to 48h of fermentation, and then gradually begins to decline from 48h. The total flavonoids determination found that the total flavonoids content of the four bacteria, Lactobacillus paracasei, Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus casei, during the fermentation process was better than that of other bacteria.

[0107] 1.3 Conclusion:

[0108] In summary, based on the pH, OD600 value and total flavonoids content of the fermentation process, the preliminary screening showed that the fermentation bacteria were Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus rhamnosus and Lactobacillus casei.

[0109] 2. Research on bacterial strain combinations: Combined fermentation is carried out based on the bacterial strains obtained in the initial screening.

[0110] 2.1 Strain combination method:

[0111] 1) Combination of bacterial strains: Four bacterial strains were used for combined fermentation experiments: No. 1: Lactobacillus paracasei and Lactobacillus rhamnosus; No. 2: Lactobacillus paracasei and Lactobacillus casei; No. 3: Lactobacillus rhamnosus and Lactobacillus casei; No. 4: Lactobacillus paracasei, Lactobacillus rhamnosus and Lactobacillus casei; No. 5: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei.

[0112] 2) Conduct the test with a single bacterial strain at 1% inoculation, natural pH, 37°C, and ferment for 72 hours. Take samples every 24 hours to test the pH, OD600, and total flavonoids content of the fermentation liquid. Select the best bacterial strain combination.

[0113] 2.2 Results, see Table 4-6.

[0114] Table 4 pH value of fermentation liquid of each bacterial combination at different fermentation time periods

[0115]

[0116] According to Table 4, the pH values ​​at different time periods during the fermentation process were measured and it was found that the fermentation pH of the five bacterial combinations was around 3.50, and the bacterial growth effects were all very good, with no significant differences.

[0117] Table 5 OD600 values ​​of biological activity of each bacterial combination at different fermentation time periods

[0118]

[0119] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD. All experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. The strain combinations of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei were fermented for 24h, 48h and 72h compared with other strain combinations, *P < 0.05, **P < 0.01. Results: According to Table 5, according to the OD600 values ​​of biological activity of each strain combination at different fermentation time periods, it can be concluded that the strain combinations of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei had higher OD values ​​than other combinations during the entire fermentation stage, and the OD600 values ​​of fermentation for 24h, 48h and 72h were significantly different from those of other strain combinations, indicating that the synergistic effect of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei in the fermentation process was better than that of other strain combinations.

[0120] Table 6 Total flavonoid content of fermentation broth of each bacterial strain combination at different fermentation time periods (unit: mg / mL)

[0121] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. The bacterial strain combination was Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei. The total flavonoids of fermentation at 24h, 48h and 72h were compared with other fermentations. *P < 0.05, **P < 0.01.

[0122] Results: According to Table 6, among the five bacterial combinations, the combination of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei had significant differences with the bacterial combinations of Lactobacillus casei and Lactobacillus rhamnosus after 24 hours of fermentation, and had extremely significant differences with Lactobacillus paracasei and Lactobacillus rhamnosus, and had no statistical differences with Lactobacillus casei and Lactobacillus paracasei, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus paracasei;

[0123] The total flavonoid content of the fermentation 48h group was significantly different from that of the other four groups of bacterial combinations;

[0124] The bacterial strain combination fermented for 72 hours had extremely significant differences with Lactobacillus paracasei and Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus rhamnosus and Lactobacillus paracasei, but had no statistical difference with Lactobacillus casei and Lactobacillus rhamnosus, Lactobacillus casei and Lactobacillus paracasei.

[0125] In summary, during the entire fermentation process, the combination of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei had the highest total flavonoids content, which was the best compared with other combinations.

[0126] 2.3 Conclusion: The fermentation strain combination is determined

[0127] (1) The five bacterial strains were combined and cultured for 72 hours, and the pH values ​​were measured and the results showed that there was little difference.

[0128] (2) The OD600 value of biological activity was determined and it was found that the OD600 value was the highest during the fermentation of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei.

[0129] (3) The results of total flavonoid content determination showed that the total flavonoid content was the highest during the fermentation process of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei.

[0130] In summary, the final fermentation bacteria combination selected was Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei.

[0131] Example 3 Fermentation study

[0132] 1. Fermentation process research: inoculation amount (ratio of bacterial strains, total inoculation amount), pH value, fermentation time

[0133] 1.1 Study on the ratio of bacterial strain combination inoculation

[0134] 1.1.1 Vaccination ratio: 7 groups in total

[0135]

[0136] 1.1.2 Results

[0137] (1) Fermentation process data

[0138] 1) The OD600 values ​​of biological activity at different fermentation times and inoculation ratios of various strains are shown in Table 7.

[0139] Table 7 OD600 values ​​of biological activity at different fermentation times and inoculation ratios of various strains

[0140]

[0141] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. The inoculation ratio was 1:1:1. The OD600 values ​​of fermentation 24h, 48h and 72h were compared with other fermentation values. *P < 0.05, **P < 0.01.

[0142] result:

[0143] According to Table 7, when the strain ratio is 1:1:1, the OD600 value is the highest during the whole fermentation process, and the OD600 values ​​of fermentation 24h, 48h and 72h are significantly different from those of other inoculation ratios, which proves that the inoculation ratio of 1:1:1 for Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei is optimal.

[0144] 2) The total flavonoid content of the fermentation liquid at different fermentation time periods and inoculation ratios of various strains is shown in Table 8.

[0145] Table 8 Total flavonoids content in fermentation broth at different fermentation time periods and inoculation ratios of various strains (unit: mg / mL)

[0146]

[0147] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. The inoculation ratio was 1:1:1. The fermentation time was 24h, 48h and 72h, and the total flavonoids were compared with other fermentations. *P < 0.05, **P < 0.01.

[0148] result:

[0149] According to Table 8, when the inoculation ratio is 1:1:1, the total flavonoids of fermentation 24h are significantly different from those of inoculation ratios 3:1:2 and 1:3:2, and are extremely significantly different from those of 2:2:1 and 2:2:3, and are not statistically different from those of 1:2:3 and 2:1:3; the total flavonoids of fermentation 48h and other inoculation ratios are significantly different; the total flavonoids of fermentation 72h are significantly different from those of 2:1:3 and 1:3:2, and are not statistically different from other ratios. When the inoculation ratio is 1:1:1, the total flavonoids of the fermentation broth are the highest during the entire fermentation process, so the optimal inoculation ratio is determined to be 1:1:1.

[0150] 1.1.3 Conclusion

[0151] The inoculation ratio of Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei was 1:1:1.

[0152] 1.2 Total inoculum study

[0153]

[0154] 1) The OD600 values ​​of biological activity at different inoculation amounts and fermentation times are shown in Table 9.

[0155] Table 9 OD600 values ​​of biological activity at different inoculation amounts and fermentation times

[0156]

[0157] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. When the 9% inoculation volume was fermented for 24h, 48h and 72h, *P < 0.05, **P < 0.01 compared with other fermentation pH.

[0158] result:

[0159] According to Table 9, when the total inoculation amount is 9%, the OD600 value of the entire fermentation process is the highest, and the OD600 values ​​at 24h, 48h and 72h of fermentation are significantly different from those of other fermentation inoculation amounts. Therefore, the total inoculation amount is determined to be 9%.

[0160] 2) The total flavonoid content of the fermentation broth at different inoculation amounts and different fermentation time periods is shown in Table 10.

[0161] Table 10 Total flavonoid content in fermentation broth at different inoculation amounts and different fermentation time periods (unit: mg / mL)

[0162]

[0163] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. When the 9% inoculation volume was fermented for 24h, 48h and 72h, *P < 0.05, **P < 0.01 compared with other fermentation pH.

[0164] Statistically significant, *P<0.05, **P<0.01 when fermenting at 9% inoculum for 24h, 48h and 72h compared with other fermentation pH.

[0165] result:

[0166] According to Table 10, when the total inoculum size is 9%, the fermentation time of 24 h is significantly different from that of 5%, and is extremely significantly different from that of 3%, 4%, 7%, and 8%, but has no statistical difference from that of 6% and 10%; the total flavonoid values ​​of fermentation time of 48 h are significantly different from those of other fermentation inoculum sizes; the fermentation time of 72 h is extremely significantly different from that of 3%, 4%, and 8%, but has no statistical difference from other inoculum sizes; when the total inoculum size is 9%, the content of total flavonoids in the entire fermentation process is the highest, so the total inoculum size is determined to be 9%.

[0167] in conclusion

[0168] According to the above test results, based on the soluble solid content and total flavonoids content, it was found that when the inoculation amount was 3%-9%, the more the inoculation amount, the better the fermentation effect of the strain. When it exceeded 9%, the OD600 value and total flavonoids content decreased, and the growth effect of the strain became worse. Finally, the optimal inoculation amount was selected as 9%.

[0169] 1.3 Study on the initial pH of fermentation

[0170]

[0171] result

[0172] (1) The OD600 of biological activity at different fermentation times and initial pH after inoculation is shown in Table 11.

[0173] Table 11 OD600 values ​​of biological activity at different fermentation times after inoculation at different initial pH

[0174]

[0175] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. The pH natural fermentation 24h, 48h and 72h was compared with other fermentation pHs, *P < 0.05, **P < 0.01.

[0176] result:

[0177] According to Table 11, the OD600 value of biological activity at different fermentation times after inoculation is optimal, and the OD600 values ​​at 24h, 48h and 72h of fermentation are significantly different from those at other fermentation pHs. Therefore, the natural pH after inoculation is selected for fermentation.

[0178] (2) The total flavonoid content of the fermentation broth at different fermentation time periods after each initial pH after inoculation is shown in Table 12. Table 12 Total flavonoid content of the fermentation broth at different fermentation time periods after each initial pH after inoculation (unit: mg / mL)

[0179] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. The total flavonoids of natural fermentation at pH 24h, 48h and 72h were compared with other fermentations, *P < 0.05, **P < 0.01.

[0180] result:

[0181] According to Table 12, the content of total flavonoids in the fermentation broth at different fermentation times after inoculation is optimal, and the highest value at 48h of fermentation is significantly different from the OD600 values ​​of other fermentation pHs. Therefore, the natural pH after inoculation is selected for fermentation.

[0182] in conclusion:

[0183] Based on the above data test results, it can be concluded that keeping the pH natural (pH 5.5) without control after inoculation is most conducive to bacterial fermentation.

[0184] 1.4 Fermentation time study

[0185] 1.4.1 Research plan (1) Fermentation strains and inoculation amount: 3% Lactobacillus plantarum, 3% Lactobacillus rhamnosus and 3% Lactobacillus paracasei. (2) Fermentation conditions: 37°C, natural pH after inoculation, static culture and fermentation for 72 hours. (3) Detection indicators: Samples were taken every 8 hours to detect the OD600 and total flavonoids of the fermentation liquid.

[0186] 1.4.2 Fermentation process data test results (1) Bioactivity OD600 values ​​at different fermentation times are shown in Table 13. Table 13 Bioactivity OD600 values ​​at different fermentation times

[0187]

[0188] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. Compared with other fermentation time periods, *P < 0.05 and **P < 0.01 for 32h fermentation.

[0189] Results: According to the data in Table 13, the following conclusions were drawn: 1) There was a very significant difference between fermentation 24h, 8h and 16h, P < 0.01; 2) The OD600 value was the highest from the beginning of fermentation to 32h, and then gradually decreased; the OD600 value at 32h of fermentation was compared with other time periods (except 40h), P < 0.01, which was a very significant difference.

[0190] This indicates that controlling the fermentation time to about 32 h is more conducive to fermentation. (2) Total flavonoid content of fermentation liquid at different fermentation time periods, see Table 14

[0191] Table 14 Total flavonoid content of fermentation broth at different fermentation time periods (mg / mL)

[0192]

[0193] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. The experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. Compared with fermentation before fermentation and other fermentation time periods at 32h, *P < 0.05, **P < 0.01. Results: According to the data in Table 14:

[0194] 1) The total flavonoids content at 24h fermentation was significantly different from that at 8h and 16h fermentation (P<0.01), but there was no statistical difference with that at 48h, 56h, 64h and 72h fermentation.

[0195] 2) From the beginning of fermentation to 16h, the total flavonoid content gradually decreased, and from 16h to 32h, the total flavonoid content gradually increased to the highest value, and then gradually decreased; the total flavonoid content at 32h of fermentation was significantly different from that at other time periods (except 40h), with a P<0.01, indicating that controlling the fermentation time at around 32h is more conducive to fermentation.

[0196] 1.4.3 Conclusion

[0197] Based on the changing trends of OD600 value and total flavonoids content during the fermentation process, it can be concluded that the optimal fermentation time is about 32 hours. However, in order to control production costs, it is best to control the fermentation time between 24 and 32 hours.

[0198] Example 4 Preparation of Extract, Fermentation Broth and Traditional Chinese Medicine Composite Fermented Drink

[0199] Step 1: Preparation of extract Step 1: Preferably, the fermented beverage composition with the function of sobering up and protecting the liver, the weight components of Chinese medicinal materials are 200 parts of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger, which are mixed and packed in a Chinese medicine extraction filter bag. After adding 12 times the volume ratio of drinking water to the Chinese medicine mixture for rinsing for 10 minutes, then adding 12 times the volume ratio of drinking water for rinsing for 3 minutes, after rinsing, adding 14 times the volume ratio of drinking water at 45°C for 1 hour, then extracting at 100°C for 2 hours, filtering through a double pipeline filter with 100 mesh, adding 8 times the volume ratio of drinking water to the residue for extraction at 100°C for 1 hour, filtering through a double pipeline filter with 100 mesh, and combining the two filtrates.

[0200] Step 2: Double-effect concentration is stopped when the volume ratio reaches 4.09 times to obtain an extract.

[0201] Step 2; Preparation of fermentation broth Step 1: Primary seed liquid culture. 1.1 Prepare primary seed culture medium (common culture medium such as MRS medium), 600mL / bottle, 3 bottles, pH not controlled, sterilize at 115℃ for 30min. 1.2 Thaw the three strains of frozen Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei, inoculate them into liquid culture medium and culture them in a 37℃ constant temperature incubator for 16-18h. 1.3 Detection: Take a sample of the primary seed liquid for microscopic detection to ensure that there are no contaminants, and detect the seed liquid OD600 (Lactobacillus plantarum OD600 3.0-5.0; Lactobacillus rhamnosus OD600 3.5-5.5; Lactobacillus paracasei OD600 3.5-5.5). Step 2: Secondary seed tank culture 2.1 The seed tank is first sterilized at 121℃ for 30min, and then the secondary culture medium (common culture medium such as MRS culture medium) is prepared, 20L / tank, 3 tanks, pH is not controlled, and sterilized at 115℃ for 30min. 2.2 The cultured primary seed liquid (inoculum amount: 3% of Lactobacillus plantarum, 3% of Lactobacillus rhamnosus and 3% of Lactobacillus paracasei, V / V / V) is inoculated into 3 seed tanks respectively, and the fermentation parameters (temperature 37℃; stirring speed 50rpm; tank pressure 0.04-0.06MPa) are maintained. The pH is not controlled and cultured for 10-14h. 2.3 Detection: Sample the secondary seed liquid for microscopic detection to ensure that there is no contamination of foreign bacteria, and detect the OD600 of the seed liquid (OD600 of Lactobacillus plantarum 3.0-5.0; OD600 of Lactobacillus rhamnosus 3.5-5.5; OD600 of Lactobacillus paracasei 3.5-5.5).

[0202] Step 3: Fermentation

[0203] 3.1 Sterilization of fermentation medium: The fermentation tank was sterilized in advance at 121°C for 30 min, and then the Chinese medicine extract concentrate was sterilized at 115°C for 0.5 h in a (300L) fermentation tank and cooled to 37°C. 3.2 Fermentation inoculation: Preparation of fermentation broths of 3 groups of different bacterial strains

[0204] 3.2.1 Inoculate the cultured secondary seed solution (total inoculation volume of 9%: 3% Lactobacillus plantarum, 3% Lactobacillus rhamnosus and 3% Lactobacillus paracasei, V / V / V) into the fermentation tank. 3.2.2 Inoculate the cultured secondary seed solution (total inoculation volume of 6%: 3% Lactobacillus plantarum and 3% Lactobacillus rhamnosus, V / V) into the fermentation tank.

[0205] 3.2.3 Inoculate the cultured secondary seed liquid (total inoculation amount is 9%: 3% Lactobacillus plantarum, 3% Lactobacillus rhamnosus and 3% Lactobacillus casei, V / V / V) into the fermentation tank. 3.3 Adjust the fermentation conditions, and culture for 24-32 hours under the conditions of temperature of 37°C, rotation speed of 50rpm, natural initial pH, tank pressure of 0.04-0.06MPa. Detect the pH of the fermentation liquid every 8 hours. Fermentation ends when the pH of the fermentation liquid is in the range of 3.3-3.8.

[0206] 3.4 After the fermentation is completed, sterilize at 72-75℃ for 0.5h, homogenize under high pressure for 0.5h, and filter through 10μm or centrifuge on a disc to obtain the fermentation liquid.

[0207] Step 3: Preparation of a compound fermented beverage of a Chinese medicine composition

[0208] The preparation process is as follows: the fermentation liquid prepared in the second step (the fermentation liquid is the fermentation liquid obtained by fermenting the bacterial combination in the above step 3.2.1) is added with seasoning materials: 20 parts by weight of artichoke juice powder, 10 parts by weight of corn oligopeptide powder, 5 parts by weight of oyster peptide powder, 225 parts by weight of oligofructose, 0.0064 parts by weight of vitamin B6, 100 parts by weight of erythritol, 100 parts by weight of concentrated pomegranate juice, 318 parts by weight of concentrated pear juice, and 100 parts by weight of concentrated mango juice; and the mixture is prepared by sugaring, filling, and sterilization.

[0209] Example 5 Antioxidant

[0210] Antioxidant activity test

[0211] 1 DPPH free radical scavenging rate determination

[0212] 1.1 Purpose of the experiment

[0213] The DPPH radical scavenging rate of the extract obtained in Example 4 and the fermentation broth (prepared in 3.2.1 of Example 4) was investigated.

[0214] 1.2 Research content

[0215] 1.2.1 Preparation of experimental reagents

[0216] 0.1mmol / L DPPH solution: Accurately weigh 0.004g of DPPH into a beaker, add a small amount of anhydrous ethanol to fully dissolve it, place it in a 100mL volumetric flask and dilute to the mark with anhydrous ethanol to prepare a 0.1mmol / L DPPH solution. The solution should be used immediately after preparation and stored away from light.

[0217] 1.2.2 Experimental procedures

[0218] 1.2.2.1 Extraction solution determination

[0219] Sample group A1: Take 2 mL of the extract and 2 mL of the DPPH solution in a test tube and shake them thoroughly to mix them evenly;

[0220] Control group A2: Take 2 mL of the extract and 2 mL of anhydrous ethanol in a test tube and shake them thoroughly to mix them evenly;

[0221] Blank group A3: Take 2 mL of ultrapure water and 2 mL of DPPH solution in a test tube and shake them thoroughly to mix them evenly;

[0222] A1, A2 and A3 were reacted at room temperature in the dark for 30 min, and anhydrous ethanol was used as a blank control. The absorbance was measured at 517 nm, and three parallel experiments were performed.

[0223] The calculation formula is as follows:

[0224] DPPH free radical scavenging rate (%) = ×100%

[0225] 1.2.2.2 Fermentation broth determination The determination method is the same as 1.2.2.1 Extraction solution determination.

[0226] 1.3 Test results The test results are shown in Table 15.

[0227] Table 15 Comparative test of DPPH free radical scavenging rate between extract and fermentation broth

[0228]

[0229] Graphpad Prism 9.5.1 software was used for statistical analysis of the data, and all experimental data were expressed as mean + SD.

[0230] The results in Table 15 show that the DPPH radical scavenging rate of the fermentation broth is more than 1.5 times that of the extract, and the antioxidant effect after fermentation is better. 2. Determination of hydroxyl radical scavenging rate 2.1 Experimental purpose To explore the hydroxyl radical scavenging rate of the extract prepared in Example 4 and the fermentation broth (prepared in 3.2.1 of Example 4). 2.2 Research content 2.2.1 Preparation of experimental reagents (1) 9mmol / L ferrous sulfate solution: Accurately weigh 0.25g ferrous sulfate in a beaker, dissolve it fully with a small amount of pure water, and place it in a 100mL volumetric flask to the mark. (2) 9mmol / L salicylic acid-ethanol solution: Accurately weigh 0.1243g salicylic acid in a beaker, dissolve it fully with a small amount of anhydrous ethanol, and place it in a 100mL volumetric flask to the mark. (3) 8.8mmol / L H2O2 solution: Take 1mL 30% H2O2 and add pure water to 1L. 2.2.2 Experimental steps 2.2.2.1 Extraction solution determination Sample group A1: Take 1mL of the extraction solution in a test tube, add 1mL of 9mmol / L ferrous sulfate solution and 1mL of 9mmol / L salicylic acid-ethanol solution in sequence, and shake it thoroughly to mix it evenly; then add 1mL of 8.8mmol / L H2O2 solution to start the reaction, and shake it thoroughly to mix it evenly. Control group A2: Take 1mL of the extraction solution in a test tube, add 1mL of 9mmol / L ferrous sulfate solution and 1mL of 9mmol / L salicylic acid-ethanol solution in sequence, and shake it thoroughly to mix it evenly; then add 1mL of ultrapure water, and shake it thoroughly to mix it evenly. Blank group A3: Take 1mL of ultrapure water in a test tube, add 1mL of 9mmol / L ferrous sulfate solution and 1mL of 9mmol / L salicylic acid-ethanol solution in sequence, and shake it thoroughly to mix it evenly; then add 1mL of 8.8mmol / L H2O2 solution to start the reaction, and shake it thoroughly to mix it evenly. A1, A2 and A3 were placed in a constant temperature water bath at 37°C for reaction for 30 min. Ultrapure water was used as a blank control. The absorbance was measured at 510 nm. Three parallel experiments were performed.

[0231] The calculation formula is as follows: Hydroxyl radical scavenging rate (%) = ×100%

[0232] 2.2.2.2 Fermentation broth determination The determination method is the same as that of 2.2.2.1 Extraction solution determination. 2.3 Test results The test results are shown in Table 16.

[0233] Table 16 Comparative test of hydroxyl radical scavenging rate between extract and fermentation broth

[0234]

[0235] Graphpad Prism 9.5.1 software was used for statistical analysis of the data, and all experimental data were expressed as mean + SD.

[0236] The results in Table 16 show that the hydroxyl radical scavenging rate of the fermentation broth is more than twice that of the extract, and the effect is optimal after fermentation.

[0237] Based on the results of the DPPH radical scavenging rate and hydroxyl radical scavenging rate of the above-mentioned extract and fermentation liquid, it is shown that the weight components of the traditional Chinese medicine preferably obtained by the present invention and the fermentation treatment of the extract using mixed bacteria have positive significance for improving the effect of protecting the liver.

[0238] Example 6 Comparative experiment on liver protection effect of extract before and after fermentation and alcohol detoxification and liver protection

[0239] 2.1 Experimental purpose: to detect the changes of alanine aminotransferase and aspartate aminotransferase before and after fermentation

[0240] 2.2 Experimental materials 2.2.1 Experimental animals Zebrafish 2.2.2 Main reagents Anhydrous ethanol, pure water, AST and ALT activity detection kit

[0241] 2.2.3 Experimental instruments: ELISA reader

[0242] 2.2.4 Experimental methods and steps (1) Fish selection: Select 5df juvenile fish and add them to a 6-well plate, 40 per well. Aspirate the fish culture water and add 3ml of prepared standard water per well. (2) Modeling: Add anhydrous ethanol to the model group and experimental group, 60ul per well, with a concentration of 2%, and do not add it to the blank group. (3) Drug addition: Dilute the extract group, fermentation liquid group 1, fermentation liquid group 2, and fermentation liquid group 3 with standard water 60 times and then add the drug and mix well. Seal the 6-well plate with plastic wrap, then wrap it with aluminum foil and place it in a 28℃ incubator for 30h. (4) Pre-test treatment: Aspirate the culture medium, rinse the juvenile fish with pure water once, aspirate all the fish in one well into a 1.5ml centrifuge tube, aspirate as much water as possible, add 400ul of extract solution, grind it thoroughly on ice for 1min with a grinder, and place it on ice for testing. (5) Detection: Perform the test according to the instructions of the alanine aminotransferase and aspartate aminotransferase activity detection kit. 2.3 Experimental results 2.3.1 Aspartate aminotransferase test results The aspartate aminotransferase test results are shown in Table 17.

[0243] Table 17 Comparative experimental results of reducing aspartate aminotransferase activity before and after fermentation of extract GOT activity, u / mgprot

[0244]

[0245] Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant, compared with the blank group # P < 0.05, # # P < 0.01, compared with the model group * P < 0.05, ** P < 0.01.

[0246] It can be seen from Table 17 that: (1) The extract (200 parts of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, 10 parts of ginger) reduced the activity of aspartate aminotransferase in the model group from 6.857u / mg prot to 5.975u / mg prot, a decrease of 12.86% in the same period of time; (2) Fermentation broth 1 (200 parts of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, 10 parts of ginger, and the bacterial strain combination was: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei) reduced the activity of aspartate aminotransferase from 6.857u / mg prot to 5.255u / mg in the same period of time. prot, decreased by 23.36%. The ability of fermentation liquid 1 to reduce the activity of aspartate aminotransferase was 1.82 times that of the extract. (3) Fermentation liquid 2 (200 parts of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, 10 parts of ginger, and the bacterial strain combination was: Lactobacillus plantarum and Lactobacillus rhamnosus) reduced the activity of aspartate aminotransferase from 6.857u / mg prot to 5.344u / mg in the same period of time. prot, decreased by 22.07%. The ability of fermentation liquid 2 to reduce the activity of alanine aminotransferase was 1.72 times that of the extract. (4) Fermentation liquid 3 (200 parts of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of Curcuma longa, 25 parts of Chrysanthemum, 10 parts of ginger, bacterial strain combination: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus casei) reduced the activity of aspartate aminotransferase from 6.857u / mg prot to 6.742u / mg prot in the same period of time, a decrease of 1.68%. The ability of fermentation liquid 3 to reduce the activity of aspartate aminotransferase was not as good as that of the extract. Results: According to the above results, fermentation liquid 1 (bacterial strain combination: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei) had the strongest ability to reduce the activity of aspartate aminotransferase, which also showed that the extract had better liver protection effect and faster onset time after probiotic fermentation.

[0247] 2.3.2 Alanine aminotransferase test results The results of alanine aminotransferase test are shown in Table 18.

[0248] Table 18 Comparative experimental results of reducing alanine aminotransferase activity before and after fermentation of extract GPT activity, u / mgprot

[0249]

[0250] Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant, compared with the blank group # P < 0.05, # # P < 0.01, compared with the model group * P < 0.05, ** P < 0.01.

[0251] It can be seen from Table 18 that: (5) The extract reduced the activity of alanine aminotransferase in the model group from 7.367u / mg prot to 6.688u / mg prot in the same period of time, a decrease of 9.22%; (6) Fermentation broth 1 (the bacterial strain combination is: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei) reduced the activity of alanine aminotransferase from 7.367u / mg prot to 5.646u / mg prot in the same period of time, a decrease of 23.36%. The ability of fermentation broth 1 to reduce the activity of alanine aminotransferase is 2.53 times that of the extract; (7) Fermentation broth 2 (the bacterial strain combination is: Lactobacillus plantarum and Lactobacillus rhamnosus) reduced the activity of alanine aminotransferase from 7.367u / mg prot to 6.543u / mg in the same period of time. prot, decreased by 12.59%. The ability of fermentation broth 2 to reduce the activity of alanine aminotransferase is 1.37 times that of the extract. (8) Fermentation broth 3 (the bacterial strain combination is: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus casei) reduced the activity of alanine aminotransferase from 7.367u / mg prot to 6.625u / mg prot in the same period of time, a decrease of 11.2%. The ability of fermentation broth 3 to reduce the activity of alanine aminotransferase is 1.09 times that of the extract. Results: According to the above results, fermentation broth 1 (the bacterial strain combination is: Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei) has the best ability to reduce the activity of alanine aminotransferase, which also shows that the extract has a better liver protection effect after probiotic fermentation and a faster onset time.

[0252] 3. Research on the effect of fermented drinks on sobering up and protecting liver

[0253] 3.1 Experimental purpose: To detect the effect of Chinese herbal medicine compound fermented beverage on sobering up and protecting the liver.

[0254] 3.2 Experimental materials and equipment 3.2.1 Experimental animals Zebrafish were provided by Qingdao Kangmaichen Biotechnology Co., Ltd.

[0255] 3.2.2 Test drugs The Chinese medicine composition prepared in Example 4, the compound fermented beverage, Metadoxine capsules (batch number: 220501), and the marketed health food (batch number: 20230104 174)

[0256] 3.2.3 Main reagents: anhydrous ethanol, pure water, aspartate aminotransferase and acetaldehyde dehydrogenase activity detection kit.

[0257] 3.2.4 Experimental Instruments ELISA Analyzer 3.2.5 Experimental Methods and Procedures (1) Fish Selection: Select 5df juvenile fish and add them to a 6-well plate, 40 per well. Aspirate the fish culture water and add 3ml of prepared standard water per well. (2) Modeling: Add anhydrous ethanol to the model group and the experimental group, 60ul per well, at a concentration of 2%, and the blank group does not add any. (3) Drug Addition: Add drugs to the Chinese medicine compound fermented beverage group (2 bottles / day), the metadoxine capsule group (4 capsules / day), and the marketed health food group (1 bottle / day) according to the calculated experimental concentration and mix well. Seal the 6-well plate with plastic wrap, wrap it with aluminum foil, and place it in a 28℃ incubator for 30h. (4) Pre-test treatment: drain the culture medium, rinse the fry with pure water once, transfer all the fish in one well to a 1.5ml centrifuge tube, drain the water as much as possible, add 400ul of extraction solution, grind thoroughly on ice for 1min with a grinder, and place on ice for testing. (5) Detection: perform the test according to the instructions of the aspartate aminotransferase and acetaldehyde dehydrogenase activity test kit.

[0258] 3.3 Experimental results 3.3.1 Aspartate aminotransferase activity determination results Aspartate aminotransferase activity determination results, see Table 19

[0259] Table 19 Effect of Chinese herbal medicine compound fermented beverage on aspartate aminotransferase activity GOT activity, u / mg prot

[0260]

[0261] Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant, compared with the blank group # P < 0.05, # # P < 0.01, compared with the model group * P < 0.05, ** P < 0.01.

[0262] As shown in Table 19, compared with the model group, the efficacy of the Chinese medicine compound drink (2 bottles / day) in reducing the activity of aspartate aminotransferase is better than that of metadoxine capsules (4 capsules / day) and marketed health foods (1 bottle / day).

[0263] 3.3.2 Results of acetaldehyde dehydrogenase activity assay The results of acetaldehyde dehydrogenase activity assay are shown in Table 20.

[0264] Table 20 Effect of Chinese herbal medicine compound beverage on acetaldehyde dehydrogenase activity ALDH activity, u / mg prot

[0265]

[0266] Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant, compared with the blank group # P < 0.05, # # P < 0.01, compared with the model group * P < 0.05, ** P < 0.01.

[0267] As shown in Table 20, compared with the model group, the effect of the Chinese medicine compound beverage (2 bottles / day) in improving the activity of acetaldehyde dehydrogenase is better than that of the marketed health food (1 bottle / day). The above results of the determination of aspartate aminotransferase and acetaldehyde dehydrogenase activities show that the Chinese medicine compound fermented beverage has good alcohol sobering and liver protection effects verified by animal experiments.

[0268] Example 7 Comparative experiment on the efficacy of different formulas on zebrafish

[0269] 7.1 Formula and preparation method 7.1.1 The formula of the present invention: 200 parts by weight of Hovenia dulcis, 150 parts by weight of dried shiitake mushrooms, 100 parts by weight of Pueraria root, 75 parts by weight of Poria cocos, 50 parts by weight of wolfberry, 30 parts by weight of turmeric, 25 parts by weight of chrysanthemum, and 10 parts by weight of ginger; the preparation process is: weigh the above-mentioned medicinal materials for extraction and concentration, fermentation with composite bacteria strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei, homogenization, filtration or disc centrifugation, add seasoning materials of 10 parts by weight of artichoke juice powder, 15 parts by weight of corn oligopeptide powder, 10 parts by weight of oyster peptide powder, 150 parts by weight of oligofructose, 0.005 parts by weight of vitamin B6, 100 parts by weight of erythritol, 100 parts by weight of concentrated pomegranate juice, 318 parts by weight of concentrated pear juice, and 100 parts by weight of concentrated mango juice, and prepare the product through sugaring, filling, and sterilization. 7.1.2 Comparative formula 1: 300 parts of Hovenia dulcis, 300 parts of dried shiitake mushrooms, 200 parts of Pueraria root, 150 parts of Poria cocos, 90 parts of wolfberry, 15 parts of chrysanthemum, and 15 parts of ginger; Preparation process: Weigh the above-mentioned medicinal materials for extraction and concentration, add seasoning materials 150 parts by weight of corn oligopeptide powder, 30 parts by weight of artichoke juice powder, 1 part by weight of oyster peptide powder, 270 parts by weight of idaxin sugar, 40 parts by weight of oligofructose powder, 2 parts by weight of stevioside, 0.5 parts by weight of sucralose, 24 parts by weight of anhydrous citric acid, and 400 parts by weight of maltodextrin, and prepare the product through sugaring, filling, and sterilization. 7.1.3 Comparative formula 2: 400 parts by weight of Hovenia dulcis, 150 parts by weight of dried shiitake mushrooms, 200 parts by weight of Pueraria root, 75 parts by weight of Poria cocos, 50 parts by weight of wolfberry, 30 parts by weight of turmeric, 25 parts by weight of chrysanthemum, and 10 parts by weight of ginger; the preparation process is as follows: the above medicinal materials are weighed for extraction and concentration, and fermented with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei, and then homogenized, filtered or centrifuged on a disc, and then seasoning materials are added, including 10 parts by weight of artichoke juice powder, 15 parts by weight of corn oligopeptide powder, 10 parts by weight of oyster peptide powder, 150 parts by weight of oligofructose, 0.005 parts by weight of vitamin B6, 100 parts by weight of erythritol, 100 parts by weight of concentrated pomegranate juice, 318 parts by weight of concentrated pear juice, and 100 parts by weight of concentrated mango juice, and the mixture is prepared by sugaring, filling, and sterilization.

[0270] 7.2. Alanine aminotransferase and aspartate aminotransferase determination 7.2.1.1 Alanine aminotransferase test results:

[0271] Table 21 Alanine aminotransferase assay results

[0272]

[0273] 7.2.1.2 Conclusion: (1) The efficacy of the experimental formula is better than that of comparative formula 2, which is 2.51 times that of comparative formula 2. This shows that when the dosage of the main drug is doubled, the efficacy is reduced, which changes the perception that the more the dosage, the better the effect. The optimal ratio is the most important. (2) The amount of Chinese medicinal materials in the experimental formula is similar to that in comparative formula 1. The ability of the experimental formula to reduce alanine aminotransferase is 1.27 times that of comparative formula 1. 7.2.2.1 Aspartate aminotransferase test results:

[0274] Table 22 Aspartate aminotransferase assay results

[0275]

[0276] 7.2.2.2 Conclusion: (1) The amount of medicinal materials used in comparative formula 1 is significantly higher than that in the present invention formula, but the efficacy is basically the same as that in the present invention formula, which significantly saves the amount of medicinal materials used; (2) The efficacy of comparative formula 2 is the weakest, indicating that the dosage of the main drug is increased by 1 times, but the efficacy is not increased.

[0277] 7.3 Conclusion: Based on the above conclusions, it is concluded that the formula of the present invention is superior to the formulas outside the scope of protection of the present invention in reducing transaminase.

[0278] Example 8 Cell comparison experiment 1 on the liver protection effect of different formulas

[0279] 8.1 Experimental purpose: To determine the safety of different formulations of hepatocytes and to compare the effects of protecting the liver 8.2 Experimental materials: 8.2.1 Experimental cells LO-2 cells 8.2.2 Reagents Hydrogen peroxide (3%); 1640 culture medium, fetal bovine serum (FBS), penicillin / streptomycin double antibody; MTT; trypsin (+EDTA); PBS. 8.2.3 Consumables 5mL centrifuge tubes, 1.5mL EP tubes, 1mL and 200uL pipette tips, prepared before the experiment, sterilized at 121℃ for 30min and placed in the clean bench. 96-well culture plates are disposable plastic products. 8.2.4 Instruments Clean bench, CO2 incubator, microscope, centrifuge, single-channel and eight-channel pipettes, cell counting plate, microplate reader, etc. 8.3 Experimental methods 8.3.1 Lo-2 culture Lo-2 cells were inoculated in DEME medium (containing 10% fetal bovine serum, 100 U / mL penicillin, 100 U / mL streptomycin) and cultured in a 37°C, 5% CO2 incubator. The cells adhered to the wall and grew to 70%~80% (i.e., subculture, generally 24 hours). The cell density was adjusted to 1×106 cells / mL for standby use. 8.3.2 MTT cell viability test The cultured cells were resuspended in culture medium, and after cell counting, they were inoculated into 96-well plates. The cell density in the 96-well plates was 3×104 cells / well and cultured in a carbon dioxide incubator for 12 hours. The supernatant was discarded, hydrogen peroxide was added and incubated for 20 hours, and then different concentrations of samples were added and incubated for 24 hours. Finally, 20μL MTT was added and incubated in a cell incubator for 4 hours. The cell culture medium containing MTT was removed and washed 1-2 times with PBS solution. Add 150-180 μL DMSO for lysis, and lyse on a shaker at room temperature for 10 minutes. Measure the absorbance at A570nm. 8.4 Experimental process 8.4.1 Formula and preparation method: Formula of experimental group 1 (formula of the present invention): 200 parts by weight of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of wolfberry, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei, homogenize, filter or disc centrifuge to obtain a fermentation liquid. The formula of experimental group 2: 300 parts by weight of Hovenia dulcis, 100 parts of dried shiitake mushrooms, 150 parts of Pueraria root, 50 parts of Poria cocos, 30 parts of wolfberry, 20 parts of turmeric, 20 parts of chrysanthemum, and 8 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, fermentation with composite bacteria strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei, homogenization, filtration or disc centrifugation to obtain the fermentation liquid.The formula of experimental group 3: 150 parts by weight of Hovenia dulcis, 200 parts of dried shiitake mushrooms, 75 parts of Pueraria root, 130 parts of Poria cocos, 80 parts of wolfberry, 50 parts of turmeric, 40 parts of chrysanthemum, and 15 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei to obtain the fermentation liquid. The formula of control group 1: 400 parts by weight of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 200 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of wolfberry, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei to obtain the fermentation liquid. The formula of control group 2: 100 parts by weight of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 50 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei to obtain a fermentation liquid. The formula of control group 3: 200 parts by weight of Hovenia dulcis, 100 parts of Pueraria root; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei to obtain a fermentation liquid. Formula of control group 4: 150 parts of dried shiitake mushrooms, 75 parts of Poria cocos, 50 parts of wolfberry, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, fermentation, homogenization, filtration or disc centrifugation with composite strains of Lactobacillus plantarum, Lactobacillus rhamnosus, and Lactobacillus paracasei to obtain fermentation liquid. Positive drugs: Metadoxine capsules (batch number: 220501), marketed health food (batch number: 20230104 174). 8.4.2 Experimental steps: The formula of the experimental group and the formula of the control group were diluted 60 times for the experiment.

[0280] 8.5 Experimental results: Table 23 Comparison of liver protection effects of different formulas Liver cell survival rate, %

[0281]

[0282] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD, and the experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant, compared with the blank group # P < 0.05, # # P < 0.01, experimental group 1 compared with the hydrogen peroxide group and other experimental groups and the control group * P < 0.05, ** P < 0.01.

[0283] result:

[0284] (1) According to the results of Table 23, the ability of experimental group 1 to repair liver cell damage after hydrogen peroxide injury was the strongest, and its cell survival rate was the highest. In addition, the hydrogen peroxide group and other experimental groups were compared with the control group, and P < 0.01, which was significantly different, proving that experimental group 1 (the formula of the present invention) had the best liver protection effect.

[0285] (2) The cell survival rate in experimental group 2 was higher than that in control group 2, control group 3, control group 4 and Metadoxine Capsules, and was significantly different from that in control group 3, control group 4 and Metadoxine Capsules (P < 0.01).

[0286] (3) The cell survival rate of experimental group 3 was higher than that of control group 3, control group 4 and metadoxine capsule, and was significantly different from that of control group 3 and control group 4 (P < 0.01).

[0287] Conclusion: Based on the above results, it can be concluded that the formula of the present invention has the strongest ability to repair liver cell damage and the highest cell survival rate, so its liver protection effect is the best.

[0288] Example 9 Cell comparison experiment 2 on the liver protection effect of different formulas

[0289] The experimental cells, consumables, reagents, instruments, methods and steps are the same as those in Example 8

[0290] 9.1 Formula and preparation method Formula of experimental group 1 (formula of the present invention): 200 parts by weight of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of kudzu root, 75 parts of Poria cocos, 50 parts of wolfberry, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or centrifuge with composite strains of plant lactobacillus, rhamnosus lactobacillus, and paracasei lactobacillus to obtain a fermentation liquid. Formula of experimental group 2: 300 parts by weight of Hovenia dulcis, 100 parts of dried shiitake mushrooms, 150 parts of kudzu root, 50 parts of Poria cocos, 30 parts of wolfberry, 20 parts of turmeric, 20 parts of chrysanthemum, and 8 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or centrifuge with composite strains of plant lactobacillus, rhamnosus lactobacillus, and paracasei lactobacillus to obtain a fermentation liquid. The formula of the missing prescription control group 1: 200 parts by weight of Hovenia dulcis, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of plant lactobacillus, rhamnosus lactobacillus, and paracasei lactobacillus to obtain fermentation liquid. The formula of the missing prescription control group 2: 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of Lycium barbarum, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of plant lactobacillus, rhamnosus lactobacillus, and paracasei lactobacillus to obtain fermentation liquid. The formula of the missing prescription control group 3: 500 parts by weight of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 75 parts of Poria cocos, 50 parts of wolfberries, 30 parts of turmeric, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of plant lactobacillus, rhamnosus lactobacillus, and paracasei lactobacillus to obtain a fermentation liquid. The formula of the missing prescription control group 4: 200 parts by weight of Hovenia dulcis, 150 parts of dried shiitake mushrooms, 100 parts of Pueraria root, 75 parts of Poria cocos, 50 parts of wolfberries, 30 parts of hawthorn, 25 parts of chrysanthemum, and 10 parts of ginger; the preparation process is: weigh the above medicinal materials for extraction and concentration, and ferment, homogenize, filter or disc centrifuge with composite strains of plant lactobacillus, rhamnosus lactobacillus, and paracasei lactobacillus to obtain a fermentation liquid. The company's authorized control formula: 300 parts by weight of Hovenia dulcis, 270 parts by weight of Pueraria root, 270 parts by weight of Lycium barbarum, 150 parts by weight of Poria cocos, 20 parts by weight of Chrysanthemum morifolium, 300 parts by weight of dried shiitake mushrooms, and 20 parts by weight of ginger. The preparation process is: weigh the above medicinal materials for extraction and concentration, add 150 parts by weight of corn oligopeptide powder and 75 parts by weight of mussel polysaccharide, and prepare it through sugaring, filling and sterilization. Positive drugs: Metadoxine capsules (batch number: 220501), marketed health food (batch number: 20230104174).

[0291] 9.2 Experimental results Table 24 Comparison of liver protection effects of different formulas Liver cell survival rate, %

[0292]

[0293] Statistical methods Graphpad Prism 9.5.1 software was used for statistical analysis of data. All experimental data were expressed as mean + SD. All experimental group data were analyzed by one-way ANOVA. P < 0.05 was statistically significant. Compared with the blank group, #P < 0.05, ##P < 0.01. Compared with the hydrogen peroxide group and other experimental groups and the control group, *P < 0.05, **P < 0.01 were used. Results: According to the results of Table 24, it was found that: (1) Under the condition of 15-fold dilution, the experimental group 1 had the strongest ability to repair liver cell damage after hydrogen peroxide injury, and its cell survival rate was the highest. Compared with the hydrogen peroxide group and other experimental groups and the control group, P < 0.01 was significantly different, which proved that the experimental group 1 (the formula of the present invention) had the best liver protection effect. (2) The cell survival rate of the experimental group 2 was higher than that of the control group 1, the control group 2, the control group 3, the marketed health food and the metadoxine capsule, and P < 0.01 was significantly different. Conclusion: Based on the above results, it can be concluded that the formula of the present invention has the strongest ability to repair liver cell damage and the highest cell survival rate, so its liver protection effect is the best. The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A Chinese medicinal fermentation composition that is both medicinal and edible, characterized in that: Its raw materials include Hovenia dulcis, dried shiitake mushrooms, kudzu root, Poria cocos, wolfberry, turmeric, chrysanthemum, ginger and fermented bacteria, and the fermented bacteria used include probiotics; The raw materials of the edible and medicinal Chinese medicine fermentation composition include, by weight: 150-300 parts by weight of Hovenia dulcis, 100-200 parts by weight of dried shiitake mushrooms, 75-150 parts by weight of Pueraria root, 50-130 parts by weight of Poria cocos, 30-80 parts by weight of Lycium barbarum, 20-50 parts by weight of turmeric, 25-40 parts by weight of chrysanthemum, and 8-15 parts by weight of ginger; The probiotics are Lactobacillus plantarum, Lactobacillus rhamnosus and Lactobacillus paracasei; The inoculation ratio of the Lactobacillus plantarum, the Lactobacillus rhamnosus and the Lactobacillus paracasei is: 1:1:1, 1:2:3, 3:1:2 or 2:2:1; The inoculation amount of the probiotics is 1-10%; The OD of the probiotics 600 The value is 1~8; The pH value of the probiotics during inoculation is 5-7; The fermentation time of the probiotic fermentation is 16 to 42 hours.

2. The Chinese medicinal fermentation composition as claimed in claim 1, characterized in that: The raw materials of the edible and medicinal Chinese medicine fermentation composition include, by weight: 200 parts by weight of Hovenia dulcis, 150 parts by weight of dried shiitake mushrooms, 100 parts by weight of Pueraria root, 75 parts by weight of Poria cocos, 50 parts by weight of Lycium barbarum, 30 parts by weight of Curcuma longa, 25 parts by weight of Chrysanthemum and 10 parts by weight of ginger; or 150 parts by weight of Hovenia dulcis, 200 parts by weight of dried shiitake mushrooms, 75 parts by weight of Pueraria root, 130 parts by weight of Poria cocos, 80 parts by weight of Lycium barbarum, 50 parts by weight of Curcuma longa, 40 parts by weight of Chrysanthemum and 15 parts by weight of ginger.

3. The Chinese medicinal fermentation composition as claimed in claim 2, characterized in that: The inoculation amount of the probiotics is 2-5%; and / or The OD of the probiotics 600 A value of 3 to 6; and / or The pH value of the probiotics during inoculation is 5-6; and / or The fermentation time of the probiotic fermentation is 20 hours, 24 hours, 28 hours or 32 hours.

4. The Chinese medicinal fermentation composition as claimed in claim 3, characterized in that: The inoculation amount of the probiotics is 2%, 3%, 4% or 5%.

5. The Chinese medicinal fermentation composition as claimed in claim 4, characterized in that: The raw materials of the Chinese medicinal fermentation composition with the same function as food and medicine also include one or more of artichoke, corn oligopeptide, oyster, oligofructose or vitamin B6.

6. The Chinese medicinal fermentation composition as claimed in claim 5, characterized in that: The artichokes include artichoke juice powder.

7. The Chinese medicinal fermentation composition as claimed in claim 6, characterized in that: The corn oligopeptide comprises corn oligopeptide powder.

8. The Chinese medicinal fermentation composition as claimed in claim 7, characterized in that: The oysters include oyster peptide powder.

9. The Chinese medicinal fermentation composition as claimed in claim 8, characterized in that: The oligofructose includes oligofructose powder or oligofructose liquid.

10. The Chinese medicinal fermentation composition as claimed in claim 9, characterized in that: The raw materials of the edible and medicinal Chinese medicine fermentation composition also include, by weight: 10-40 parts by weight of artichoke juice powder, 5-20 parts by weight of corn oligopeptide powder, 2-10 parts by weight of oyster peptide powder, 100-500 parts by weight of oligofructose and 0.003-0.012 parts by weight of vitamin B6; or 20-30 parts by weight of artichoke juice powder, 5-15 parts by weight of corn oligopeptide powder, 2-10 parts by weight of oyster peptide powder, 150-450 parts by weight of oligofructose and 0.003-0.012 parts by weight of vitamin B6.

11. The Chinese medicinal fermentation composition as claimed in claim 9, characterized in that: The raw materials of the edible and medicinal Chinese medicine fermentation composition also include, by weight: 20 parts by weight of artichoke juice powder, 10 parts by weight of corn oligopeptide powder, 5 parts by weight of oyster peptide powder, 225 parts by weight of oligofructose and 0.0064 parts by weight of vitamin B6; 30 parts by weight of artichoke juice powder, 15 parts by weight of corn oligopeptide powder, 5 parts by weight of oyster peptide powder, 300 parts by weight of oligofructose and 0.005 parts by weight of vitamin B6.

12. The Chinese medicinal fermentation composition as claimed in claim 11, characterized in that: The raw materials of the edible and medicinal Chinese medicine fermentation composition also include: one or more of erythritol, concentrated pomegranate juice, concentrated pear juice or concentrated mango juice.

13. The Chinese medicinal fermentation composition as claimed in claim 12, characterized in that: In parts by weight, the raw materials of the edible and medicinal Chinese medicine fermentation composition also include: 50 to 200 parts by weight of erythritol, 50 to 200 parts by weight of concentrated pomegranate juice, 150 to 500 parts by weight of concentrated pear juice and 50 to 200 parts by weight of concentrated mango juice.

14. The Chinese medicinal fermentation composition as claimed in claim 13, characterized in that: In parts by weight, the raw materials of the Chinese medicinal fermentation composition that is both medicinal and edible also include: 100 parts by weight of erythritol, 100 parts by weight of concentrated pomegranate juice, 318 parts by weight of concentrated pear juice and 100 parts by weight of concentrated mango juice.

15. The method for preparing the edible and medicinal Chinese medicine fermentation composition according to any one of claims 1 to 14, characterized in that: Take the formulated amount of raw materials for extraction, concentration, fermentation, blending and filling.

16. The preparation method according to claim 15, characterized in that: The extraction is performed at least once.

17. The preparation method according to claim 15, characterized in that: The number of extractions is 1 extraction or 2 extractions.

18. The preparation method according to claim 17, characterized in that: The amount of water added for one extraction is 5 to 20 times that of the raw material.

19. The preparation method according to claim 17, characterized in that: The amount of water added in one extraction is 10 to 16 times that of the raw material.

20. The preparation method according to claim 17, characterized in that: The amount of water added in the extraction is 14 times that of the raw material.

21. The preparation method according to claim 17, characterized in that: The amount of water added in the second extraction is 5 to 20 times that of the raw material.

22. The preparation method according to claim 17, characterized in that: The amount of water added in the first extraction of the two extractions is 8 to 15 times the amount of the raw material, and the amount of water added in the second extraction of the two extractions is 6 to 12 times the amount of the raw material.

23. The preparation method according to claim 17, characterized in that: The amount of water added in the first extraction of the two extractions is 12 or 14 times the amount of the raw material, and the amount of water added in the second extraction of the two extractions is 8 times the amount of the raw material.

24. The preparation method according to claim 17, characterized in that: The time for one extraction includes 1 to 3 hours.

25. The preparation method according to claim 17, characterized in that: The extraction time is 1 hour, 2 hours or 3 hours.

26. The preparation method according to claim 17, characterized in that: The extraction time is 1 hour.

27. The preparation method according to claim 17, characterized in that: The time of the two extractions includes 1 to 3 hours.

28. The preparation method according to claim 17, characterized in that: The time for the first extraction in the two extractions is 1 to 2 hours, and the time for the second extraction in the two extractions is 1 to 2 hours.

29. The preparation method according to claim 17, characterized in that: The time for the first extraction of the two extractions is 1 hour or 2 hours, and the time for the second extraction of the two extractions is 1 hour.

30. A Chinese medicinal fermentation composition that is both edible and medicinal obtained by the preparation method according to any one of claims 15 to 29.

31. Use of the Chinese medicinal fermentation composition for food and medicine as claimed in any one of claims 1 to 14 or the Chinese medicinal fermentation composition for food and medicine as claimed in claim 30 in the preparation of a liver-protecting drug.

32. The use according to claim 31, characterized in that The liver protection is one or more of protecting the liver or protecting the liver by alcohol detoxification.

33. The use according to claim 31, characterized in that The liver protection includes one or more of reducing transaminase, increasing acetaldehyde dehydrogenase activity or anti-oxidation.

34. The use according to claim 33, characterized in that The transaminase includes alanine aminotransferase and / or aspartate aminotransferase, and the antioxidant includes scavenging hydroxyl free radicals and / or DPPH.

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