Use of citron or its extract in the preparation of a drug for treating ulcerative colitis, and a pharmaceutical composition for treating ulcerative colitis

Abstract

The present invention provides the use of Citrus medica or its extract in the preparation of a medicament for treating ulcerative colitis. The present invention also provides the use of Citrus medica or its extract in combination with Akkermansia muciniphila (Akk bacteria) in the preparation of a medicament for treating ulcerative colitis. The present invention also provides a pharmaceutical composition for treating ulcerative colitis, which is prepared by fermentation from the following raw materials in a volume ratio: 0.5 - 1.5 parts of Citrus medica or its extract, and 0.5 - 1.5 parts of Akkermansia muciniphila (Akk bacteria). The Citrus medica or its extract of the present invention has a definite efficacy in treating ulcerative colitis. When used in combination with Akkermansia muciniphila, the co-culture of Citrus medica and Akkermansia muciniphila significantly enhances the probiotic effect of Akkermansia muciniphila; it can effectively relieve the inflammatory symptoms of ulcerative colitis, reduce intestinal damage, and the curative effect is significantly better than that of using Akkermansia muciniphila alone, achieving a synergistic effect.

Description

Technical Field

[0001] The present invention relates to the use of citron or its extract in the preparation of a drug for treating ulcerative colitis, and a pharmaceutical composition for treating ulcerative colitis. Background Art

[0002] Ulcerative Colitis (UC) is a chronic non-specific intestinal inflammatory disease that mainly affects the colon and rectum. The goals of treating UC are to control inflammation, relieve symptoms, maintain remission, and improve the quality of life. The etiology of UC is not fully understood, but it involves multiple factors such as genetics, environment, immunity, and microorganisms. Drug treatment is the main treatment method for UC, including 5-aminosalicylic acid (5-ASA), corticosteroids, immunosuppressants, biological agents (such as anti-TNF-α monoclonal antibodies, integrin antagonists, IL-12 / IL-23 antagonists), and JAK inhibitors, etc.

[0003] Akkermansia muciniphila (A. muciniphila) and its metabolite short-chain fatty acid (SCFA) are probiotics and metabolites that have been found in recent years to have a protective effect on the intestinal mucosal barrier in UC (Mao Huifang, et al., Research on the correlation between Akkermansia muciniphila and its metabolite short-chain fatty acid and the intestinal mucosal barrier in ulcerative colitis, DOI: 10.13343 / j.cnki.wsxb.20220657).

[0004] The medicinal material of citron mainly comes from the dried ripe fruits of the Rutaceae plants Citrus medica L. or Citrus wilsonii Tanaka. Citron is usually harvested when the fruits are ripe in autumn, and can be sliced while fresh and then dried in the sun or processed by low-temperature drying. It has the effects of soothing the liver and regulating qi, and resolving phlegm and relieving fullness. Currently, there is no report that citron can be used to treat ulcerative colitis. There is a large compound prescription containing citron for treating ulcerative colitis, such as patent application number: 201810118777.6, invention name: A traditional Chinese medicine composition for treating ulcerative colitis, which only utilizes the activities of citron in soothing the liver and regulating qi, and resolving phlegm and relieving fullness. Summary of the Invention

[0005] The technical solution of the present invention is to provide a new use of citron or its extract, specifically, its use in the preparation of a drug for treating ulcerative colitis, and a pharmaceutical composition for treating ulcerative colitis.

[0006] The present invention provides the use of citron or its extract in the preparation of a drug for treating ulcerative colitis.

[0007] Among them, the citron extract is a fermentation product of the water extract of citron.

[0008] The fermentation method is to ferment using a brain heart infusion - mucin culture system.

[0009] The present invention also provides the use of citron or its extract in combination with Akkermansia muciniphila (Akk bacteria) in the preparation of a medicament for treating ulcerative colitis.

[0010] Among them, the volume ratio of citron or its extract (calculated by the original drug amount) to Akkermansia muciniphila (Akk bacteria) is:

[0011] 0.5 - 1.5 parts of citron or its extract, 0.5 - 1.5 parts of Akkermansia muciniphila (Akk bacteria);

[0012] Among them, each liter of the citron extract contains 50 g of the original citron medicinal material; each liter of the Akkermansia muciniphila liquid contains 1 mole of Akkermansia muciniphila.

[0013] Further preferably, the volume ratio of citron or its extract to Akkermansia muciniphila (Akk bacteria) is:

[0014] 1 part of citron or its extract, 1 part of Akkermansia muciniphila (Akk bacteria).

[0015] The medicament provided by the present invention is prepared by co - fermenting the water extract of citron and Akkermansia muciniphila, and the fermentation method is to ferment using a brain heart infusion - mucin culture system.

[0016] The present invention provides a pharmaceutical composition for treating ulcerative colitis, which is prepared by fermentation from raw materials in the following volume ratio:

[0017] 0.5 - 1.5 parts of citron or its extract, 0.5 - 1.5 parts of Akkermansia muciniphila (Akk bacteria);

[0018] Among them, each liter of the citron extract contains 50 g of the original citron medicinal material; each liter of the Akkermansia muciniphila liquid contains 1 mole of Akkermansia muciniphila.

[0019] Further preferably, it is prepared by fermentation from raw materials in the following volume ratio:

[0020] 1 part of citron or its extract, 1 part of Akkermansia muciniphila (Akk bacteria).

[0021] The pharmaceutical composition of the present invention uses the co - fermented product of citron or its extract and Akkermansia muciniphila (Akk bacteria) liquid as the active ingredient, and is prepared into a commonly used oral preparation in pharmacy by adding pharmaceutically acceptable excipients or auxiliary components. The fermentation method is to ferment using a brain heart infusion - mucin culture system.

[0022] The citron or the extract thereof of the present invention is used to treat ulcerative colitis, has clear medicinal efficacy, and is used in combination with Akkermansia muciniphila. The co-culture of citron and Akkermansia muciniphila significantly improves the probiotic effect of Akkermansia muciniphila; the inflammatory symptoms of ulcerative colitis can be effectively alleviated, and intestinal damage can be reduced. The therapeutic effect is significantly better than that of using Akkermansia muciniphila alone, and a synergistic effect is achieved. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 .The weight change rate and recovery rate of mice in each group;

[0024] Figure 2 .Average colon length of mice in each group;

[0025] Figure 3 . Colon images of mice in each group;

[0026] Figure 4 .The colon mucosal injury status score per unit length of mice in each group;

[0027] Figure 5 .Comparison of scores of different pathological indicators in each group of mice;

[0028] Figure 6 .Pathological sections of mice in each group. DETAILED DESCRIPTION

[0029] Example 1 Preparation of Citron Water Extract of the Present Invention

[0030] Weigh 5g of Citron citron raw material accurately into a round-bottom flask, add 50ml ultrapure water, heat and reflux with an electric heating mantle for 2h, cool to room temperature, take the filtrate and temporarily store it in a conical flask, seal it and store it at 4℃. Refill the remaining drug residue into a round-bottom flask, add 50ml ultrapure water again, heat and reflux with an electric heating mantle for 2h, cool to room temperature, filter out the drug residue, and combine the two filtrates. Pass the filtrate through a bacteria filter, dispense it into 10ml EP tubes, seal it and freeze it at -20℃ for later use.

[0031] Example 2 Preparation of the fermented citron water extract of the present invention

[0032] A citron culture system was constructed according to the volume ratio of 50 g / L citron water extract of citron native medicinal materials: brain heart infusion-mucin culture medium = 1:9. After anaerobic culture for 24 hours, centrifuge at 4°C, 12000 rpm for 5 minutes, take the supernatant with a 5 ml syringe, filter it through a bacteria filter, inject it into a sterilized 10 ml EP tube, seal it, and store it at 4°C for later use.

[0033] Brain Heart Infusion-Agar (BHIA) is a microbial culture medium that combines the characteristics of Brain Heart Infusion Broth (BHI) and Agar, and is used to culture various microorganisms, especially those bacteria with relatively high nutritional requirements. This culture system provides a nutrient-rich environment to support the growth and reproduction of microorganisms.

[0034] Among them, the Brain Heart Infusion Broth was purchased from solarbio, and the Agar was purchased from sigma. The content of Brain Heart Infusion-Agar medium is stable throughout the culture system.

[0035] Example 3 Preparation of the test drug from the co-fermentation supernatant of Citrus medica and Akkermansia muciniphila of the present invention

[0036] Construct a co-fermentation system of Citrus medica and Akkermansia muciniphila extract bacteria according to the volume ratio of 1:1:8 of Citrus medica raw medicinal material 50g / L Citrus medica water extract: Akkermansia muciniphila 24h culture broth: Brain Heart Infusion-Agar medium. After anaerobic culture for 24h, centrifuge at 4°C and 12000rpm for 5min. Take the supernatant with a 5ml syringe, filter through a bacteria filter and inject it into a sterilized 10ml EP tube and seal it, and store it at 4°C for later use.

[0037] The beneficial effects of the present invention are demonstrated by the following pharmacodynamic experiments.

[0038] Test Example 1 Pharmacodynamic experiments on the fermented product of the water extract of Citrus medica and the co-fermentation of Citrus medica and Akkermansia muciniphila of the present invention

[0039] 1. Experimental purpose

[0040] To explore the synergistic effect of the co-fermentation product of Citrus medica and Akkermansia muciniphila on the treatment and improvement of ulcerative colitis compared with using the drugs alone.

[0041] 2. Materials and methods

[0042] 2.1 Preparation of the test drug

[0043] Prepare a Citrus medica water extract solution containing 50g / L of Citrus medica raw medicinal material by heating under reflux for 2h. Prepare a Codonopsis pilosula water extract solution containing 50g / L of Codonopsis pilosula raw medicinal material by heating under reflux for 2h. Prepare a 1 McFarland concentration of Akkermansia muciniphila (AKK) bacterial solution, and culture it in Brain Heart Infusion-Agar medium by inoculating 100ul of the bacterial solution per ml for 24h for later use.

[0044] Preparation of the test drug from the supernatant of the co-fermentation of Citrus medica and Akkermansia muciniphila: A co-fermentation system of Citrus medica and Akkermansia muciniphila was constructed according to the volume ratio of Citrus medica crude drug 50 g / L Citrus medica water extract: Akkermansia muciniphila 24-hour culture broth: Brain Heart Infusion-Mucin Medium = 1:1:8. After anaerobic culture for 24 hours, it was centrifuged at 4°C and 12,000 rpm for 5 minutes. The supernatant was taken with a 5-ml syringe, filtered through a bacterial filter, and then injected into a sterilized 10-ml EP tube and sealed, and stored at 4°C for later use.

[0045] Preparation of the test drug of Citrus medica: A Citrus medica culture system was constructed according to the volume ratio of Citrus medica crude drug 50 g / L Citrus medica water extract: Brain Heart Infusion-Mucin Medium = 1:9. After anaerobic culture for 24 hours, it was centrifuged at 4°C and 12,000 rpm for 5 minutes. The supernatant was taken with a 5-ml syringe, filtered through a bacterial filter, and then injected into a sterilized 10-ml EP tube and sealed, and stored at 4°C for later use.

[0046] Preparation of the test drug from the supernatant of the co-fermentation of Codonopsis pilosula and Akkermansia muciniphila: A co-fermentation system of Codonopsis pilosula and Akkermansia muciniphila was constructed according to the volume ratio of Codonopsis pilosula crude drug 50 g / L Codonopsis pilosula water extract: Akkermansia muciniphila 24-hour culture broth: Brain Heart Infusion-Mucin Medium = 1:1:8. After anaerobic culture for 24 hours, it was centrifuged at 4°C and 12,000 rpm for 5 minutes. The supernatant was taken with a 5-ml syringe, filtered through a bacterial filter, and then injected into a sterilized 10-ml EP tube and sealed, and stored at 4°C for later use.

[0047] Preparation of the test drug of Codonopsis pilosula: A Codonopsis pilosula culture system was constructed according to the volume ratio of Codonopsis pilosula crude drug 50 g / L Codonopsis pilosula water extract: Brain Heart Infusion-Mucin Medium = 1:9. After anaerobic culture for 24 hours, it was centrifuged at 4°C and 12,000 rpm for 5 minutes. The supernatant was taken with a 5-ml syringe, filtered through a bacterial filter, and then injected into a sterilized 10-ml EP tube and sealed, and stored at 4°C for later use.

[0048] Preparation of the test drug from the supernatant of the fermentation of Akkermansia muciniphila: A fermentation system of Akkermansia muciniphila was constructed according to the volume ratio of Akkermansia muciniphila 24-hour culture broth: Sterile ultrapure water: Brain Heart Infusion-Mucin Medium = 1:1:8. After anaerobic culture for 24 hours, it was centrifuged at 4°C and 12,000 rpm for 5 minutes. The supernatant was taken with a 5-ml syringe, filtered through a bacterial filter, and then injected into a sterilized 10-ml EP tube and sealed, and stored at 4°C for later use.

[0049] Preparation of the test drug of viable Akkermansia muciniphila fermented bacteria: A fermentation system of Akkermansia muciniphila was constructed according to the volume ratio of Akkermansia muciniphila 24-hour culture broth: Sterile ultrapure water: Brain Heart Infusion-Mucin Medium = 1:1:8. After anaerobic culture for 24 hours, it was mixed evenly for later use (this test drug needs to be freshly prepared every day).

[0050] Source of materials:

[0051] The raw medicinal materials of Citrus medica and Codonopsis pilosula were both purchased from Sichuan New Lotus Chinese Herbal Pieces Co., Ltd.; Dextran sulfate sodium (DSS) was purchased from Seebio Technology Co., Ltd., batch number: DCJ1606A; Akkermansia muciniphila (AKK) was purchased from Beina Biological, number: BNCC323275; Brain heart infusion medium was purchased from solarbio, batch number: 2942479; Porcine gastric mucin was purchased from sigma, batch number: SLCB4590.

[0052] 2.2 Study on the synergistic effect of Citrus medica and Akkermansia muciniphila in the treatment of ulcerative colitis

[0053] 2.2.1 Establishment of ulcerative colitis mouse model by free drinking of DSS

[0054] SPF-grade female mice were adaptively fed for 3 days. After confirming the health of the mice, 54 female mice were randomly divided into 9 groups: blank group (K), model group (M), brain heart infusion medium group (B), Akkermansia muciniphila supernatant group (AS), Akkermansia muciniphila live bacteria group (A), co-fermentation group of Akkermansia muciniphila and Citrus medica (XA), Citrus medica group (X), co-fermentation group of Akkermansia muciniphila and Codonopsis pilosula (XA), Codonopsis pilosula group (X), and finally, at least 5 mice were ensured in each group. Except for the mice in the blank group, the mice in the other groups were given 3% DSS for free drinking for 7 days. The mouse bedding was changed on time every day, the body weight of the mice was weighed, their diet and water intake were observed, and the excreta of the mice were observed. Loose stools or bloody stools in the mice indicated successful modeling.

[0055] 2.2.2 Drug intervention in ulcerative colitis model mice

[0056] After the mice were successfully modeled, the model group (group M) was given 0.2 ml of normal saline daily; the medium group (group B) was given 0.2 ml of brain heart infusion-mucin medium daily; the Akkermansia muciniphila supernatant group (group AS) was given 0.2 ml of the 24-hour fermentation supernatant of Akkermansia muciniphila daily; the Akkermansia muciniphila live bacteria group (group A) was given 0.2 ml of the 24-hour culture mixture of Akkermansia muciniphila daily; the co-fermentation group of Akkermansia muciniphila and Citrus medica (group XA) was given 0.2 ml of the supernatant of the 24-hour fermentation broth of Akkermansia and Citrus medica daily; the Citrus medica group (group X) was given 0.2 ml of the supernatant of the 24-hour culture broth of Citrus medica daily, the co-fermentation group of Akkermansia muciniphila and Codonopsis pilosula (group DA) was given 0.2 ml of the supernatant of the 24-hour fermentation broth of Akkermansia and Codonopsis pilosula daily; the Codonopsis pilosula group (group D) was given 0.2 ml of the supernatant of the 24-hour culture broth of Codonopsis pilosula daily. The above drugs were all administered by gavage, once a day for 7 consecutive days.

[0057] 2.2.3 Indexes for studying the synergistic effect of Citrus medica and Akkermansia muciniphila in the treatment of ulcerative colitis

[0058] 2.2.3.1 General status and disease activity index

[0059] At 9:00 am every day, observe the fur, spirit, diet, drinking water, loose stools and bloody stools of mice in each group, and record the body weight of mice.

[0060] 2.2.3.2 Colon length

[0061] After continuously intragastrically administering drugs to mice for 7 days, anesthetize them with sodium pentobarbital and then decapitate them. Isolate and excise the colon, and measure the colon length.

[0062] 2.2.3.3 Score of the damage status of unit colon mucosa

[0063] Longitudinally cut the remaining 2 / 3 length of the colon along the mesentery. After collecting the feces of mice in a 2 ml EP tube for backup, rinse it thoroughly with physiological saline, take pictures, observe the mucosal damage situation with the naked eye, and perform CMDI scoring. The scoring criteria are shown in the table.

[0064] Table 1. Scoring criteria for colon mucosal morphology

[0065]

[0066] 2.2.3.4 Pathological status of colon tissue

[0067] Cut a 1 / 3 length of the colon along the enlarged end of the colon, rinse the inside of the colon thoroughly with PBS, place it in 4% paraformaldehyde for fixation, and send it to Sichuan Yangkest Technology Co., Ltd. for pathological detection.

[0068] 3. Experimental results

[0069] 3.1 Effects of the co-fermentation product of Citrus medica and Akkermansia muciniphila on the body weight of ulcerative colitis mice

[0070] From the start of modeling, the body weights of mice in each group decreased significantly, and UC clinical symptoms such as dull and messy hair color, decreased food and water intake, loose stools and bloody stools all appeared.

[0071] Through Figure 1 .A It can be seen from the effects of each test drug administration on the body weight change rate of UC mice that after the corresponding drug intervention was given to mice in each group, the body weights all showed a recovery phenomenon. By comparing between groups of each treatment group, it was found that when using the co-fermentation product of Citrus medica and Akkermansia muciniphila, compared with using Akkermansia muciniphila and Citrus medica alone, the improvement of body weight was significantly increased. However, when using the co-fermentation product of Codonopsis pilosula and Akkermansia muciniphila, there was no significant difference in the improvement of body weight compared with using Codonopsis pilosula alone. Therefore, it can be judged that the co-fermentation of Codonopsis pilosula and Akkermansia muciniphila has no synergistic effect on the treatment and improvement of ulcerative colitis compared with single drug use.

[0072] Through Figure 1.B From the effects of each test drug administration on the weight recovery rate of UC mice, it can be seen that the co-ferment of Citrus medica and Akkermansia muciniphila has a significantly higher weight recovery rate than other treatment groups. The ranking of the weight recovery rates of each treatment group from high to low is as follows: the co-fermentation group of Citrus medica and Akkermansia muciniphila (XA group), the co-fermentation group of Codonopsis pilosula and Akkermansia muciniphila (DA group), the Codonopsis pilosula group (D group), the Citrus medica group (X group), the supernatant group of Akkermansia muciniphila (AS group), the heart-brain infusion-mucin medium group (B group), and the live bacteria group of Akkermansia muciniphila (A group). Although the co-fermentation group of Codonopsis pilosula and Akkermansia muciniphila (DA group) and the Codonopsis pilosula group (D group) have a relatively high ranking in terms of weight recovery rate, there is no significant difference in the weight recovery rate between the co-fermentation group of Codonopsis pilosula and Akkermansia muciniphila (DA group) and the Codonopsis pilosula group (D group). Therefore, it can be judged that the co-fermentation of Codonopsis pilosula and Akkermansia muciniphila has no synergistic effect on the treatment and improvement of ulcerative colitis compared with single drug use. The co-fermentation group of Citrus medica and Akkermansia muciniphila (XA group) has a significantly higher weight recovery rate compared with the Citrus medica group (X group) and the supernatant group of Akkermansia muciniphila (AS group). Therefore, it can be judged that the co-fermentation of Citrus medica and Akkermansia muciniphila has a synergistic effect on the treatment and improvement of ulcerative colitis compared with single drug use.

[0073] 3.2 Effects of the co-fermentation product of Citrus medica and Akkermansia muciniphila on the colon length of ulcerative colitis mice

[0074] After 7 days of drug administration, the mice were sacrificed and the colon tissues were taken out to measure the length. It was found that the colon length of the mice in the model group (M group) was significantly shorter than that of the mice in the blank group (K group), indicating that the UC mouse model was successfully constructed. After each treatment group was given the corresponding test drug, the colon length showed a significant recovery, and the live bacteria group of Akkermansia muciniphila (A group) was the most significant. The improvement of the colon length in the co-fermentation group of Citrus medica and Akkermansia muciniphila (XA group) was more obvious compared with the supernatant group of Akkermansia muciniphila (AS group) alone. However, the co-fermentation group of Codonopsis pilosula and Akkermansia muciniphila (DA group) did not show obvious improvement in the colon length compared with the supernatant group of Akkermansia muciniphila (AS group) alone and the Codonopsis pilosula group alone, and the co-fermentation group of Codonopsis pilosula and Akkermansia muciniphila (DA group) was even worse than the Codonopsis pilosula group alone. Therefore, in terms of the colon length index, the co-fermentation group of Citrus medica and Akkermansia muciniphila (XA group) has a significant synergistic effect on the treatment and improvement of ulcerative colitis compared with single drug use, while the co-fermentation group of Codonopsis pilosula and Akkermansia muciniphila (DA group) does not have this effect.

[0075] 3.3 Effects of the co-fermentation product of Citrus medica and Akkermansia muciniphila on the damage status of the colon mucosa in ulcerative colitis mice

[0076] By scoring the damage status of the colonic mucosa of each group of mice, it was found that compared with the blank group (K group), the mucosa in the model group (M group) showed severe damage (**<0.01). After seven days of administration of the corresponding test drugs to each treatment group, except for the group treated with only Citrus medica (X group) and the group treated with only Codonopsis pilosula (D group), the damage status of the colonic mucosa of mice in the other treatment groups was significantly improved. And compared with the model group (M group), there were significant differences in the Citrus medica and Akkermansia muciniphila co-fermentation group (XA group), the Codonopsis pilosula and Akkermansia muciniphila co-fermentation group (DA group), and the supernatant of Akkermansia muciniphila (AS group), among which the Citrus medica and Akkermansia muciniphila co-fermentation group (XA group) was the most significant (**<0.001). Compared with the group treated with only live Akkermansia muciniphila (A group), the supernatant of Akkermansia muciniphila (AS group), and Citrus medica (X group), the improvement status of the Citrus medica and Akkermansia muciniphila co-fermentation group (XA group) was more obvious. Therefore, the co-fermentation of Citrus medica and Akkermansia muciniphila has a synergistic effect on the improvement of ulcerative colitis compared with the use of only Akkermansia muciniphila and Citrus medica.

[0077] 3.4 Effect of the co-fermentation product of Citrus medica and Akkermansia muciniphila on the colonic pathological state of mice with ulcerative colitis

[0078] By comparing the scores of the colonic pathological sections of each group of mice, it was found that compared with the blank group (K group), the mice in the model group (M group) showed significant differences in four indicators: inflammatory response, degree of inflammation, crypt damage, and damage area (***<0.001), indicating that the ulcerative colitis mouse model in this experiment was successfully established. Compared with the mice in the model group (M group), except for the heart-brain infusion-mucin medium group (B group), the other treatment groups had obvious improvement effects on each index compared with the model group (M group). Among the improvement indexes of the degree of inflammation and inflammatory response, the Citrus medica and Akkermansia muciniphila co-fermentation group (XA) was significantly higher than the group treated with only Akkermansia muciniphila (A group and AS group) and the group treated with only Citrus medica (X group) (*<0.05), and there were significant differences. In terms of the indexes of crypt damage and damage area, the Citrus medica and Akkermansia muciniphila co-fermentation group (XA) also had certain improvements compared with the group treated with only Akkermansia muciniphila (A group and AS group) and the group treated with only Citrus medica (X group). On the contrary, the Codonopsis pilosula and Akkermansia muciniphila co-fermentation group (DA) had a slight improvement effect on each pathological index compared with the group treated with only Akkermansia muciniphila (A group and AS group), but the improvement effects of the Codonopsis pilosula and Akkermansia muciniphila co-fermentation group (DA) and the group treated with only Codonopsis pilosula (D group) on each index were basically the same. Therefore, we can judge that the Codonopsis pilosula and Akkermansia muciniphila co-fermentation group (DA group) has no synergistic effect on the treatment and improvement of ulcerative colitis compared with single drug use, while the Citrus medica and Akkermansia muciniphila co-fermentation (XA group) has an obvious synergistic effect on the improvement of the pathological indexes of ulcerative colitis compared with the group treated with only Akkermansia muciniphila (A group and AS group) and the group treated with only Citrus medica (X group).

[0079] 4. Experimental conclusions:

[0080] Citrus medica is a qi-regulating medicine, which has the functions of soothing the liver and regulating qi, regulating the middle energizer, and resolving phlegm. It is mainly used for liver-stomach qi stagnation, chest and hypochondrium distending pain, abdominal fullness and discomfort, vomiting and excessive qi, and excessive phlegm with cough. Codonopsis pilosula is a qi-tonifying medicine, which has the functions of strengthening the spleen and replenishing the lung, nourishing blood and promoting the production of body fluid. It can be used for spleen and lung qi deficiency, anorexia and fatigue, cough with deficiency asthma, insufficient qi and blood, sallow complexion, palpitation and shortness of breath, thirst due to fluid injury, and internal heat with consumptive thirst. Both of them have the efficacy of regulating the middle energizer and tonifying the spleen. Codonopsis pilosula is good at tonifying, while Citrus medica is good at regulating qi. In this invention, Codonopsis pilosula is selected as a reference object to compare the obvious differences in the efficacy of the products after the co-fermentation of qi-tonifying and qi-regulating medicines with Akkermansia muciniphila. The co-fermentation product of Citrus medica and Akkermansia muciniphila has a synergistic effect on the improvement of ulcerative colitis treatment compared with the single medicine use, while Codonopsis pilosula has no such effect.

Claims

1. Use of Citrus medica or its extract in combination with Akkermansia muciniphila in the preparation of a medicament for treating ulcerative colitis, wherein the volume ratio of Citrus medica or its extract to Akkermansia muciniphila in terms of the original drug amount is as follows: 0.5 - 1.5 parts of Citrus medica or its extract, 0.5 - 1.5 parts of Akkermansia muciniphila; Among them, Each liter of the Citrus medica extract contains 50 g of the original Citrus medica medicinal material; each liter of the Akkermansia muciniphila liquid contains 1 mole of Akkermansia muciniphila; the said medicament is prepared by co-fermenting Citrus medica or its extract with Akkermansia muciniphila, the said Citrus medica extract is an aqueous extract of Citrus medica, and the said fermentation method is carried out using a brain heart infusion - mucin culture system.

2. The use according to claim 1, characterized in that: The volume ratio of Citrus medica or its extract to Akkermansia muciniphila is as follows: 1 part of Citrus medica or its extract, 1 part of Akkermansia muciniphila.

3. A pharmaceutical composition for treating ulcerative colitis, characterized in that: It is prepared by fermentation from raw materials with the following volume ratio: 0.5 - 1.5 parts of Citrus medica or its extract, 0.5 - 1.5 parts of Akkermansia muciniphila; the said Citrus medica extract is an aqueous extract of Citrus medica; Among them, each liter of the Citrus medica extract contains 50 g of the original Citrus medica medicinal material; each liter of the Akkermansia muciniphila liquid contains 1 mole of Akkermansia muciniphila.

4. The pharmaceutical composition for treating ulcerative colitis according to claim 3, wherein: It is prepared by fermentation from raw materials with the following volume ratio: 1 part of Citrus medica or its extract, 1 part of Akkermansia muciniphila.

5. The pharmaceutical composition for treating ulcerative colitis according to any one of claims 3 or 4, characterized in that: It is prepared by using the co-fermentation product of Citrus medica or its extract and Akkermansia muciniphila liquid as the active ingredient, adding pharmaceutically acceptable excipients or auxiliary ingredients to prepare a commonly used oral preparation in pharmacy, and the said fermentation method is carried out using a brain heart infusion - mucin culture system.

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