Effective fraction of Sophora flavescens root extract, preparation method thereof, and application thereof in preparing medicine for preventing or treating inflammatory bowel disease

Abstract

The present invention belongs to the field of natural medicinal chemistry technology, and specifically relates to an effective part of a Radix Sophorae flavescentis extract, a preparation method thereof, and an application thereof in the preparation of a medicament for preventing or treating inflammatory bowel disease (IBD). The effective part of the Radix Sophorae flavescentis extract is mainly a mixture of daurisoline (DAU) and daurisoline methyl ester (DAU‑Me), which accounts for 75%‑95% by mass. The effective part of the Radix Sophorae flavescentis extract provided by the present invention significantly improves weight loss, colon shortening, intestinal barrier and intestinal inflammation in IBD model mice, has the prospect of being developed into a drug for preventing and treating inflammatory bowel disease, provides more treatment options for clinically preventing or treating inflammatory bowel disease, and has important social benefits and economic value.

Description

Technical Field

[0001] The present invention relates to the field of natural medicinal chemistry, and in particular to an effective part of a dried root extract of the medicinal plant Menispermum dauricum DC., a preparation method thereof, and an application of the effective part in preparing a drug for preventing or treating inflammatory bowel disease. Background Art

[0002] Inflammatory bowel disease (IBD) is a chronic, nonspecific intestinal inflammatory disease affecting the mucosa of the ileum, rectum, and colon. Its course is recurrent, with clinical manifestations primarily including abdominal pain, diarrhea, bloody stools containing mucus, pus, and tenesmus. Pathologically, it is characterized by chronic inflammation of the colonic mucosa, intestinal ulcers, or the formation of transmural granulomas. IBD, often referred to as the "green cancer," is difficult to cure and can severely deteriorate patients' quality of life. Patients often require lifelong medication or even surgery. IBD includes Crohn's disease (CD) and ulcerative colitis (UC).

[0003] The pathogenesis of IBD is unclear, and existing treatment options are generally ineffective. The main goal of treating IBD patients is to achieve long-term clinical symptom relief and microscopic healing of the intestinal mucosa. Traditional drug treatments include salicylic acid preparations, glucocorticoids, and immunosuppressants, most of which work by suppressing the body's immune response to control inflammation. Although these drugs have immediate effects, they have relatively large side effects. Therefore, these drugs are generally not suitable for long-term use, and patients are prone to relapse immediately after stopping the drug.

[0004] In recent years, the relationship between IBD and the gut microbiome has garnered widespread attention, with the gut microbiome potentially modulating IBD. The gut microbiome primarily comprises intestinal bacteria, fungi, viruses, and metabolites, with intestinal bacteria being the most extensively studied. Recent studies have confirmed that the human gut microbiome is closely linked to the development and progression of inflammatory bowel disease (IBD). Intestinal bacteria can induce or exacerbate IBD by producing inflammatory polysaccharides or disrupting the intestinal barrier. Some microorganisms can also produce anti-inflammatory substances, potentially ameliorating IBD. However, traditional microbial agents have poor host colonization and limited efficacy. Recent studies have found that bacterial DPP4 can mimic host DPP4 function, degrading active peptides, thereby impacting the host's innate immune system and systemic metabolic homeostasis, thus influencing the development and progression of inflammatory diseases. Therefore, targeting bacterial DPP4 as a research target and developing corresponding microbial agents and pharmaceuticals could provide new avenues and more effective measures for the treatment and prevention of IBD.

[0005] The dried rhizome of Menispermum dauricum DC. is known as the root of Menispermum dauricum. It has a bitter taste, cold properties, and is slightly toxic. It enters the lung, stomach, and large intestine meridians. It clears heat and detoxifies, promotes dampness, and reduces swelling. It is primarily used to treat acute pharyngitis, tonsillitis, sore gums, lung-heat cough, damp-heat, jaundice, and constipation. Currently, no research has shown that extracts or structural analogs of Menispermum dauricum DC. can alleviate or treat inflammatory bowel disease. Summary of the Invention

[0006] To address the current shortage of or ineffective clinical treatments for IBD, the present invention provides an effective fraction of a radix sibiricum extract, its preparation method, and its use in the preparation of a medicament for the prevention or treatment of inflammatory bowel disease (IBD). This invention provides a new approach for the clinical prevention or treatment of IBD and for the modernization of Traditional Chinese Medicine.

[0007] The technical solutions for achieving the purpose of the present invention are as follows:

[0008] In a first aspect, the present invention provides a method for preparing an effective fraction of a Radix Sophorae flavescentis extract having the effect of preventing or treating inflammatory bowel disease, comprising the following steps:

[0009] (1) Take the clean rhizome of the root of the northern bean root, dry it thoroughly and then extract it;

[0010] (2) preparing an ethanol:methanol:water extract in a volume ratio of (0.5-3):(0.5-2):(1-2), adjusting the temperature to 10-100°C, mixing the extract with the root of the northern bean in (1) and shaking for 1-12 hours;

[0011] (3) After the extraction in step (2) is completed, solid-liquid separation is performed to obtain a crude extract, which is then rotary evaporated at 30-60° C. for 1-6 hours to fully evaporate the organic solvent to obtain a concentrated solution. At this time, the extract still contains a large amount of water;

[0012] (4) vacuum freeze-drying the concentrated solution obtained in step (3) at -1.0 MPa to -0.10 MPa for 1 to 18 hours to obtain a dry light yellow to black powder;

[0013] (5) dissolving the powder obtained in step (4) in ethanol, filtering to obtain a clear solution, concentrating the solution, and evaporating to dryness to obtain a crude extract of the effective part of the target Rhizoma Coptidis extract;

[0014] (6) using high performance liquid chromatography to track the activity of the crude extract of the effective fraction in step (5), and the effective component obtained by elution for 6 to 7 minutes has the best effect, and the effective fraction of the Rhizoma Coptidis extract is obtained after concentration;

[0015] (7) Liquid chromatography-tandem mass spectrometry was performed to analyze that the effective part of the northern bean root extract was mainly a mixture of daurisoline (DAU) and daurisoline methyl ester (DAU-Me), accounting for 75%-95% by mass percentage.

[0016] As an optional manner, in the above-mentioned method for preparing the effective fraction of the Rhizoma Coptidis extract, the Rhizoma Coptidis in step (1) should be chopped into small segments of 2 mm.

[0017] As an optional method, in the above-mentioned method for preparing the effective part of the northern bean root extract, the volume ratio of ethanol:methanol:water in step (2) is 3:1:2, the material-liquid ratio is 1g:6-10mL, the extraction temperature is 50-60°C, and the oscillation extraction time is 4-5 hours.

[0018] As an optional manner, in the above-mentioned method for preparing the effective fraction of the Rhizoma Coptidis extract, the rotary evaporation at 50° C. is performed for 4 to 6 hours in step (3).

[0019] As an optional manner, in the above-mentioned method for preparing the effective fraction of the Rhizoma Coptidis extract, in step (4), the product is freeze-dried at -0.70 MPa for 12 to 18 hours.

[0020] As an optional manner, in the above-mentioned method for preparing the effective fraction of the Rhizoma Coptidis extract, a 0.22 μm filter membrane is used for filtration in step (5).

[0021] As an optional manner, in the above-mentioned method for preparing the effective part of the northern bean root extract, the chromatographic column used in step (6) is: a 5 μm particle size C18 chromatographic column, 4.6×250 mm; the mobile phase is: in volume percentage, A: 80% water + 20% acetonitrile, B: 80% acetonitrile + 20% isopropanol, and the liquid phase conditions are: 0-4 min, 30% B; 4-10 min, 30% B-90% B; 10-12 min, 90% B; 12-13 min, 90% B-30% B, 13-15 min, 30% B.

[0022] As an optional method, in the above-mentioned method for preparing the effective part of the northern bean root extract, the northern bean root is selected from the dried rhizome of the Menispermum dauricum DC. of the Menispermaceae family, and the collection time is limited to spring and autumn, and the production area is one or more regions in Northeast China, North China, East China, Shaanxi, Ningxia, Gansu, and Shandong.

[0023] In a second aspect, the present invention provides an effective fraction of a Radix Sophorae flavescentis extract having the effect of preventing or treating inflammatory bowel disease, obtained by the preparation method described in the first aspect. The effective fraction of the Radix Sophorae flavescentis extract is primarily a mixture of dauricine (DAU) and dauricine methyl ester (DAU-Me), accounting for 75% to 95% by mass.

[0024] In a third aspect, the present invention provides use of the effective fraction of the Rhizoma Coptidis extract according to the second aspect in the preparation of a dipeptidyl peptidase-4 (DPP4) inhibitor.

[0025] In a fourth aspect, the present invention provides use of the effective fraction of the Rhizoma Coptidis extract described in the second aspect in the preparation of a medicament for preventing or treating inflammatory bowel disease.

[0026] As an option, in the above application, the inflammatory bowel disease is ulcerative colitis or Crohn's disease.

[0027] As an optional mode, in the above application, the dosage form of the drug is selected from granules, capsules, tablets, powders, oral solutions, suspensions or emulsions.

[0028] As an optional mode, in the above application, the dosage form of the drug is an enteric-coated tablet.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] The effective fraction of the Rhizoma Coptidis extract provided by the present invention significantly improves weight loss, colon shortening, intestinal barrier and intestinal inflammation in IBD model mice, and has the prospect of being developed into a drug for the prevention and treatment of inflammatory bowel disease, providing more treatment options for the clinical prevention or treatment of inflammatory bowel disease, and has important social benefits and economic value. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 Effects of different components of Radix Sophorae flavescentis on body weight and disease activity index in mice model of DSS. Figures are presented as mean ± SD. One-way ANOVA was used to evaluate the significance of each group. * Compared with the model group, P < 0.05. ** Compared with the model group, P < 0.01. Component 7 had the most significant improvement effect.

[0032] Figure 2 Effects of the active fraction (fraction 7) of the extract of Sophora flavescens root and the positive drug 5-ASA on the body weight of mice modeled with DSS. The figures are presented as mean ± SD. One-way analysis of variance was used to assess the significance of each group. * Compared with the model group, P < 0.05. **Compared with the model group, P < 0.01. Component 7 showed a better improvement on DSS-induced ulcerative colitis than the positive drug 5-ASA.

[0033] Figure 3 Effects of the active fraction (fraction 7) of the extract of Sophora flavescens root and the positive drug 5-ASA on the disease activity index of DSS model mice. Figures are presented as mean ± SD. One-way analysis of variance was used to evaluate the significance of each group. * Compared with the model group, P < 0.05. ** Compared with the model group, P < 0.01. Component 7 showed a better improvement on DSS-induced ulcerative colitis than the positive drug 5-ASA.

[0034] Figure 4 : In vivo imaging of the effective fraction (component 7) of the extract of Sophora flavescens root and the positive drug 5-ASA in improving colitis in DSS mice.

[0035] Figure 5 : Representative pathological sections of the effective fraction (fraction 7) of the extract of Sophora flavescens root and the positive drug 5-ASA in improving colitis in DSS mice, upper panel: 10×; lower panel: 400×.

[0036] Figure 6 : The effective fraction (fraction 7) of the extract of Sophora flavescens root and the positive drug 5-ASA improve colon length and intestinal barrier function in DSS mice. Left: Colon length statistics; Right: Intestinal permeability test. Figures are presented as mean ± SD. One-way analysis of variance was used to assess the significance of each group. * Compared with the model group, P < 0.05. ** Compared with the model group, P < 0.01. Component 7 showed a better improvement on DSS-induced ulcerative colitis than the positive drug 5-ASA. DETAILED DESCRIPTION

[0037] After extensive and in-depth research and experiments, the inventors discovered that a specific effective fraction of a Rhizoma Coptidis extract can inhibit bacterial DPP4 activity, thereby preventing and treating inflammatory bowel disease. Feeding this effective fraction to experimental subjects revealed that it improved weight loss, colon length reduction, disease activity scores, and intestinal barrier disruption in a DSS-induced inflammatory bowel disease mouse model. This work was based on these findings and led to the completion of the present invention.

[0038] As used herein, the term "comprising" indicates that various ingredients can be used together in the mixture or composition of the present invention. Therefore, the terms "consisting essentially of" and "consisting of" are encompassed by the term "comprising".

[0039] The present invention provides the use of an effective portion of a Radix Sophorae flavescentis extract in preventing or treating inflammatory bowel disease. Mice are given DSS drinking water to create an inflammatory bowel disease model, and the effective portion of the Radix Sophorae flavescentis extract has the ability to improve weight loss, shortened colon length, disease activity score, and intestinal barrier damage in the model animals. According to a preferred embodiment of the present invention, DSS-induced inflammatory bowel disease model mice treated with the effective portion of the Radix Sophorae flavescentis extract can maintain normal body weight, colon length, and intestinal barrier compared to an untreated control group. Therefore, the effective portion of the Radix Sophorae flavescentis extract can be used to prevent and treat inflammatory bowel disease.

[0040] The drug of the present invention can be administered in the form of any conventional pharmaceutical preparation in the art, wherein the pharmaceutical preparation includes excipients, pharmaceutically acceptable media and carriers, which can be selected according to the route of administration. The drug of the present invention may further include auxiliary components.

[0041] The pharmaceutically acceptable excipients of the present invention include conventional solvents in the pharmaceutical field (such as water, ethanol, propylene glycol, injection oil, etc.), diluents (such as starch, powdered sugar, dextrin, lactose, pregelatinized starch, microcrystalline cellulose, inorganic calcium salts (such as calcium sulfate, calcium hydrogen phosphate, pharmaceutical calcium carbonate, etc.), mannitol, etc., vegetable oil, polyethylene glycol, etc.), binders (such as water, ethanol, starch slurry, sodium carboxymethyl cellulose, hydroxypropyl cellulose, methyl cellulose and ethyl cellulose, hydroxypropyl methylcellulose, etc.), disintegrants (such as dry starch, sodium carboxymethyl starch, low-substituted hydroxypropyl cellulose, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose, etc.), lubricants (such as magnesium stearate, micronized silica gel, talc, hydrogenated vegetable oil, polyethylene glycol, magnesium lauryl sulfate, etc.), absorption enhancers (such as surfactants, Azone (lauroamphetamine), EDTA, salicylic acid, amino acid ethylamine derivatives, acetoacetic acid esters, β-dicarboxylates, aromatic acid compounds, aliphatic acids, etc.), preservatives (such as benzoic acid, butyl hydroxypropyl ester, methyl hydroxypropyl ester, phenol, m-cresol, etc.), flavoring agents (such as sucrose, stevia, etc.), etc.

[0042] In addition, as a preferred embodiment, the drug of the present invention can be produced as an enteric coating preparation by various well-known methods so that the active ingredients of the drug can pass through the stomach smoothly without being destroyed by gastric acid.

[0043] The pharmaceutical composition of the present invention can be prepared as enteric-coated tablets for oral administration. The term "enteric coating" as used herein includes all conventional pharmaceutical coatings that are not degraded by gastric acid but fully decompose in the small intestine to rapidly release the active ingredient of the present invention. The enteric coating of the present invention can be maintained at 36-38°C for more than 2 hours in synthetic gastric acid, such as a pH 1 HCl solution, and preferably decomposes within 1.0 hour in synthetic intestinal fluid, such as a pH 7.0 buffer.

[0044] The enteric coating of the present invention is applied to each tablet at a weight of about 16 to 30 mg, preferably 16 to 25 mg, and more preferably 16 to 20 mg. The thickness of the enteric coating is 5 to 100 μm, preferably 20 to 80 μm. The enteric coating is composed of conventional polymers used in pharmaceutical preparations.

[0045] The present invention is described below with reference to specific examples. Those skilled in the art will appreciate that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention in any way.

[0046] The experimental methods in the following examples are conventional methods unless otherwise specified. The raw materials, reagents, etc. used in the following examples are commercially available products unless otherwise specified. The purchase information of some raw materials and reagents is as follows:

[0047] The radix dauricum used in the present invention is selected from the dried rhizomes of Menispermum dauricum DC., a plant of the Menispermaceae family. The collection time is limited to spring and autumn, and the production area is one or more regions in Northeast China, North China, East China, Shaanxi, Ningxia, Gansu, and Shandong. Example 1: Preparation of radix dauricum extract

[0048] The clean rhizomes of the root of the northern bean root produced in Heilongjiang Province were taken, thoroughly dried, and then chopped for extraction. An ethanol:methanol:water extract was prepared in a volume ratio of 3:1:2, the temperature was adjusted to 60°C, and the extract was mixed with the northern bean root and shaken for 5 hours. The crude extract was obtained by solid-liquid separation. The organic solvent was fully evaporated at 50°C for 4 hours to obtain a concentrated solution, at which point a large amount of water was still present in the extract. The concentrated solution was vacuum freeze-dried at -0.7MPa for 18 hours to obtain a dry light yellow to black powder. This powder was dissolved in ethanol and filtered to obtain a clear solution. The solution was concentrated and evaporated to dryness to obtain the crude extract of the effective part of the target northern bean root extract.

[0049] The crude extract was isolated using high-performance liquid chromatography (HPLC) to identify key active components. The chromatographic column used was a 5μm particle size C18 column, 4.6×250mm. The mobile phase consisted of, by volume, 80% water + 20% acetonitrile (A) and 80% acetonitrile + 20% isopropanol (B). The liquid phase conditions were: 30% B (0-4 min); 30% B to 90% B (4-10 min); 90% B (10-12 min); 90% B to 30% B (12-13 min); and 30% B (13-15 min). The liquid phase was run for 15 minutes, with effluent collected every minute, yielding 15 fractions. Each fraction was rotary evaporated to dryness at 50°C, weighed, and reconstituted in methanol to yield fractions 1-15. Equal amounts of the dried crude extract of Rhizoma Caulis Sophorae flavescentis were dissolved in methanol to yield fraction 16.

[0050] Example 2: Inhibitory activity of the extract of Sophora flavescens root and its components against bacterial DPP4

[0051] DPP4 is an important protein that regulates the levels of many endogenous polypeptides. It regulates intestinal cell proliferation and inflammation by affecting the levels of functional polypeptides such as GLP-1, GLP-2, and GM-CSF. Recent studies have found that increased levels of bacterial DPP4 are associated with the occurrence of IBD. Inhibiting the activity of bacterial DPP4 may contribute to the recovery of IBD. However, it is not yet clear what the main component in the extract of the root of northern soybean is that inhibits the activity of bacterial DPP4. By separating the components of the root of northern soybean by liquid chromatography, 16 components were obtained (see Example 1), and then the activity tracking strategy was used to evaluate the inhibitory effect of each component on DPP4. The bacterial DPP4 was purified from an in vitro expression system independently constructed by this team. The inhibitory effects of the 16 components on bacterial DPP4 are shown in Table 1.

[0052] Table 1: Inhibitory activity of the extract from the root of Sophora flavescens and its components against bacterial DPP4

[0053]

[0054]

[0055] Among them, fraction 16 is a crude extract, and fractions 1-15 are fractions separated by liquid chromatography according to retention time. Fraction 7 shows the most significant DPP4 inhibitory activity.

[0056] Example 3: Analysis of the main components of the effective part of the extract of the root of Sophora flavescens (Component 7)

[0057] LC-MS was used to analyze the main components and content of fraction 7, and the inhibitory effect of each component on DPP4 was evaluated. The results are shown in Table 2.

[0058] Table 2: Analysis of the main components of component 7 and the inhibitory effect of each component on DPP4

[0059]

[0060] LC-MS analysis determined that the major component of Fraction 7 was daurine (DAU), accounting for approximately 60.3%, and the minor component was daurine methyl ester (DAU-Me), accounting for approximately 23.2%. The remaining components were daurine analogs, all of which exhibited DPP4 inhibitory activity. Furthermore, Fraction 7 exhibited greater activity than any of the individual components, indicating a synergistic effect among the components.

[0061] Example 4: Comparison of the improvement effects of the single active ingredient in the extract of Sophora flavescens root, component 7 and crude extract on the IBD mouse model induced by dextran sodium sulfate (DSS)

[0062] Materials: The test substance was a powder obtained from the extracts of various components of the Rhizoma Coptidis (Psoralea corylifolia) collected from North China, purified, and dried. The drug was administered orally at a dose of 10 mg / kg body weight once daily.

[0063] C57 mice were purchased from Beijing Weitonglihua Laboratory Animal Technology Co., Ltd. and housed at a temperature of 20-24°C, a constant humidity of 50-60%, a 12-hour light cycle (8:00-20:00), soundproofed, and free access to food and water. Experiments were performed after one week of acclimatization. DSS was purchased from MP Company, USA.

[0064] Methods: 36 male C57 mice aged 8 weeks, weighing 20±1g, were randomly divided into (1) model control group, (2) DAU group, (3) DAU-Me group, (4) other DAU analogs in component 7 group, (5) component 7 group, and (6) crude extract group, with 6 mice in each group. Drinking water was replaced with 2% DSS. Mice in groups (2)-(6) were gavaged with 10 mg / kg body weight of the corresponding drug every day for 1 week. The model control group was given an equal amount of normal saline. The body weight of the animals was measured daily and the fecal bleeding was observed. The body weight of the mice in the model group was significantly reduced, and obvious bloody and loose stools were observed, which proved that the model was successfully established. After the last administration, the body weight changes and disease activity index of each group were evaluated to clarify the effects of each component of the extract of the root of the northern bean on inflammatory bowel disease. The evaluation of the disease activity index was mainly based on the quantitative measurement of weight loss, bloody stools and loose stools. Specifically, the assessment criteria are: weight loss (0 points, weight loss <1%; 1 point, weight loss ≥1% but <5%; 2 points, weight loss ≥5% but <10%; 3 points, weight loss ≥10% but <20%; 4 points, weight loss ≥20%); blood in stool (0 points, no blood in stool; 2 points, occult blood; 4 points, obvious blood in stool); loose stool (0 points, normal stool; 2 points, loose and soft stool; 4 points, stool adhering to the anus). The sum of the three scores divided by 3 is the disease activity index.

[0065] Results: Compared with the model group, each component of the extract of the root of Sophora flavescens could significantly improve the related indicators of inflammatory bowel disease, restore body weight, and reduce the disease activity index. Among them, component 7 had the best improvement effect. Figure 1 .

[0066] Example 5: Improvement of the Effective Fraction of the Rhizoma Coptidis Extract (Component 7) on the DSS-Induced IBD Mouse Model

[0067] Materials: The test substance was Fraction 7 of the Radix Sophorae flavescentis extract collected from North China. After extraction, purification, and drying, a light yellow to dark yellow powder was obtained. 5-Aminosalicylic acid (5-ASA) was used as a positive control. Fraction 7 was administered orally at a dose of 10 mg / kg body weight, and 5-ASA was administered orally at a dose of 100 mg / kg body weight (this dose of 5-ASA has been shown to effectively ameliorate DSS-induced colitis). Both doses were administered orally once daily.

[0068] C57 mice were purchased from Beijing Weitonglihua Laboratory Animal Technology Co., Ltd. and housed at a temperature of 20-24°C, a constant humidity of 50-60%, a 12-hour light cycle (8:00-20:00), soundproofed, and free access to food and water. Experiments were performed after one week of acclimatization. DSS was purchased from MP Company, USA.

[0069] Methods: Eighteen male C57 mice, 8 weeks old and weighing 20±1 g, were randomly divided into (1) model control group; (2) bean root extract group; and (3) 5-ASA positive drug group. Each group had 6 mice, and drinking water was replaced with 2% DSS. Mice in group (2) were gavaged with 10 mg / kg body weight of component 7 daily, and mice in group (3) were gavaged with 100 mg / kg body weight of 5-ASA daily for 1 week. The model group was given an equal amount of normal saline. The body weight of the animals was measured daily, and fecal bleeding was observed. The body weight of the mice in the model group was significantly reduced, and obvious bloody and loose stools were observed, indicating that the model was successfully established. After the last administration, the mice were intraperitoneally injected with L-012 chemiluminescent probe for in vivo imaging to assess the level of intestinal inflammation. The darker the blue and the larger the range, the more severe the inflammation. After fasting for 4 hours, the intestinal permeability of the mice was assessed by gavage with FITC-dextran (4kD, 600 mg / kg). Finally, the mice were killed and the colon length of the mice was measured. About 1 cm of colon was placed in tissue fixative, dehydrated, paraffin-embedded, sectioned, and then stained with H&E. The level of inflammatory infiltration and barrier damage in the mouse colon were evaluated by histopathology to clarify the effect of component 7 of the northern bean root extract on inflammatory bowel disease.

[0070] Results: Compared with the model group, component 7 could significantly improve the related indicators of inflammatory bowel disease and restore body weight ( Figure 2 ), reduce disease activity score ( Figure 3 ), reduce inflammation levels in the body ( Figure 4 ), reduced intestinal ulcers and inflammatory infiltration ( Figure 5 ), significantly increased the length of the mouse intestine and improved the intestinal barrier of the mouse ( Figure 6 ), and the degree of improvement was better than that of 5-ASA.

[0071] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if such changes and modifications fall within the scope of the claims and their equivalents, the present invention is intended to include such changes and modifications.

Claims

1. A method for preparing an effective fraction of a Radix Sophorae flavescentis extract having the effect of preventing or treating ulcerative colitis, characterized in that: The following steps are involved: (1) Take the clean rhizome of the root of the northern bean root, dry it thoroughly and then extract it; (2) Prepare an ethanol:methanol:water extract in a volume ratio of (0.5-3):(0.5-2):(1-2), adjust the temperature to 10-100°C, mix the extract with the root of the northern bean in (1) and shake for 1-12 hours; (3) After the extraction in step (2) is completed, solid-liquid separation is performed to obtain a crude extract, which is then rotary evaporated at 30-60°C for 1-6 hours to fully evaporate the organic solvent to obtain a concentrated solution. At this time, the extract still contains a large amount of water; (4) freeze-drying the concentrated solution obtained in step (3) under vacuum at -1.0 MPa to -0.10 MPa for 1 to 18 hours to obtain a dry light yellow to black powder; (5) dissolving the powder obtained in step (4) in ethanol, filtering to obtain a clear solution, concentrating the solution, and evaporating to dryness to obtain a crude extract of the effective part of the target Rhizoma Coptidis extract; (6) The activity of the crude extract of the effective part in step (5) was tracked by high performance liquid chromatography. The effective component obtained by elution for 6 to 7 minutes had the best effect. After concentration, the effective part of the root extract of Sophora flavescens was obtained. The chromatographic column used was: 5 μm particle size C18 chromatographic column, 4.6×250 mm; the mobile phase was: in volume percentage, A: 80% water + 20% acetonitrile, B: 80% acetonitrile + 20% isopropanol, and the liquid phase conditions were: 0-4 min, 30% B; 4-10 min, 30% B-90% B; 10-12 min, 90% B; 12-13 min, 90% B-30% B, 13-15 min, 30% B; (7) Liquid chromatography-tandem mass spectrometry was performed to analyze that the effective fraction of the extract of the Radix Sophorae Flavescentis was mainly a mixture of daurisoline and daurisoline methyl ester, accounting for 75%-95% by mass.

2. The method for preparing the effective fraction of the Rhizoma Coptidis extract according to claim 1, wherein In step (1), chop the bean roots into 2 mm pieces.

3. The method for preparing the effective fraction of the Rhizoma Coptidis extract according to claim 1, wherein: In step (2), the volume ratio of ethanol:methanol:water is 3:1:2, the material-liquid ratio is 1 g:6-10 mL, the extraction temperature is 50-60°C, and the oscillation extraction time is 4-5 hours.

4. The method for preparing the effective fraction of the Rhizoma Codonopsis pilosula extract according to claim 1, wherein In step (3), the mixture is subjected to rotary evaporation at 50° C. for 4 to 6 hours; in step (4), the mixture is freeze-dried at −0.70 MPa for 12 to 18 hours; and in step (5), the mixture is filtered using a 0.22 μm filter membrane.

5. The effective fraction of the Rhizoma Coptidis extract having the effect of preventing or treating ulcerative colitis, prepared by the method according to any one of claims 1 to 4, characterized in that: The effective part of the northern bean root extract is mainly a mixture of daurisoline and daurisoline methyl ester, which accounts for 75%-95% by mass.

6. Use of the effective fraction of the Rhizoma Coptidis extract according to claim 5 in the preparation of a medicament for preventing or treating ulcerative colitis.

7. The use according to claim 6, characterized in that The dosage form of the drug is selected from granules, capsules, tablets, powders, oral solutions, suspensions or emulsions.

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