Preparation method and application of active components of adzuki bean

Abstract

The application belongs to the technical field of natural medicinal chemistry and enzyme inhibitor development, and particularly relates to a preparation method and application of an active ingredient of red bean. The active ingredient is (+) catechin 7-O-beta-D-glucopyranoside accurately separated from red bean. In vitro experiments prove that the active ingredient has strong inhibitory effect on alpha-glucosidase and pancreatic lipase. Cell and animal experiments further prove that the active ingredient can significantly inhibit the differentiation and lipid accumulation of 3T3-L1 preadipocytes, and can effectively reduce the body weight, body fat and blood lipid level of high-fat diet induced obese mice, and improve liver steatosis. The application plays a multi-target anti-obesity effect by synergistically inhibiting the digestion and absorption of sugar and fat, and has a wide application prospect in the preparation of medicines and functional foods for preventing and treating obesity, type II diabetes and other metabolic diseases.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of natural medicinal chemistry and enzyme inhibitor development, and specifically relates to a new medical use and extraction method of an active ingredient in red bean (Vigna angularis), (+)-catechin 7-O-beta-D-glucopyranoside. In particular, the present application relates to the application of (+)-catechin 7-O-beta-D-glucopyranoside in preparing a dual inhibitor for simultaneously inhibiting alpha-glucosidase and pancreatic lipase, and a process for efficiently separating and purifying the active ingredient from red beans. BACKGROUND

[0002] Red bean (Vigna angularis) is a diploid legume in the family Fabaceae, also known as adzuki bean, red bean or red bean. Red bean has medicinal value of diuresis, detumescence, detoxification and pus discharge. Historically, red bean has been used in ancient China to treat edema, jaundice, edema of the feet, joint swelling, skin erythema and abdominal pain, etc. Like other legume seeds, red bean is rich in protein, polyphenols and dietary fiber, with low fat content and glycemic index. Red bean also contains carbohydrates, fatty acids, flavonoids, saponins and micronutrients, as well as a small amount of anti-nutritional factors. Due to its nutritional composition, red bean has various health benefits, including antioxidant, anti-obesity, neuroprotective, cholesterol-lowering, anticancer, antibacterial, anti-diabetic, liver and kidney protection, blood pressure reduction, and reduction of vascular oxidative stress and inflammation. In addition, red bean can alleviate metabolic syndrome such as obesity by regulating gut microbiota. For example, red bean can improve metabolic function, reduce inflammation and prevent fatty degeneration by reversing gut dysbiosis. As a nutritious food, red bean is safe and harmless, providing potential resources for the development of lead compounds for the prevention and treatment of chronic diseases and health supplements.

[0003] Medicinal value and research status of red bean: Red bean (Vigna angularis) is an important crop of food and medicine, which has been cultivated and applied in China for thousands of years. Modern research shows that red bean is rich in various physiologically active chemical components, including flavonoids (such as catechin and its derivatives), triterpenoid saponins (such as yunganoside B1), polyphenols and dietary fiber, etc. These active ingredients endow red bean with various pharmacological activities such as antioxidant, anti-inflammatory, regulation of sugar and lipid metabolism, etc.

[0004] In recent years, with the in-depth study of active ingredients from plant sources, the application potential of red bean in the intervention of metabolic diseases has become increasingly prominent. However, there is still a lack of systematic study on the components in red bean with specific metabolic enzyme inhibition activity, especially in the identification of dual enzyme inhibition active components, mechanism of action and efficient preparation process, etc.

[0005] Research progress of metabolic enzyme inhibitors: Metabolic enzymes play a key role in the process of nutrient digestion and absorption. Among them, pancreatic lipase (PDB ID 1GPL) is responsible for the decomposition of dietary fat, and alpha-glucosidase (PDB ID 3A4A) is involved in the digestion of carbohydrates. The excessive activity of these two enzymes is closely related to the occurrence and development of metabolic diseases such as obesity and diabetes. Although the currently used synthetic enzyme inhibitors (such as orlistat and acarbose) have certain efficacy, they generally have adverse reactions such as gastrointestinal irritation and liver toxicity. In contrast, enzyme inhibitors from natural sources are becoming an important direction for new drug research and development due to their high safety and small side effects. It is worth noting that multifunctional inhibitors targeting multiple metabolic enzymes can more comprehensively regulate metabolic disorders, but the development of such inhibitors still faces great challenges.

[0006] Traditional active ingredient extraction methods of red beans mainly include: (1) water extraction method: simple operation but low extraction efficiency (target component yield is usually <0.5%), and it is difficult to effectively retain heat-labile components; (2) organic solvent extraction method: although the extraction efficiency is improved (ethanol extraction yield can reach 1.2-1.8%), there is a risk of solvent residue; (3) auxiliary extraction technology: including ultrasonic-assisted (extraction time is shortened by 40-60%) and microwave-assisted (extraction efficiency is improved by 20-30%) methods, but there are problems such as high equipment cost and poor process stability when industrialized. It is particularly important to note that the existing technology lacks the ability to selectively enrich specific active ingredients in red beans, making it difficult to meet the preparation needs of high-purity active ingredients.

[0007] Red beans (Vigna angularis) have a long history as a medicinal and edible crop with high safety. Modern research shows that they are rich in active ingredients such as flavonoids and triterpenoid saponins, and have various physiological activities. However, the existing technology has the following shortcomings: 1. The identification of specific high-activity enzyme inhibiting components in red beans is not clear enough, and there is a lack of precise mechanism research on monomer compounds; 2. Traditional extraction methods are inefficient and lack selectivity, making it difficult to obtain high-purity active ingredients, and there is no mention of the construction of active ingredient combinations; 3. The research on the weight loss efficacy of red bean active ingredients mostly stays at the level of crude extract, and there is a lack of systematic efficacy verification of monomer components or specific combinations in cell and animal models, especially a lack of direct evidence of inhibiting adipocyte differentiation and lipid accumulation. SUMMARY

[0008] The first object of the present application is to provide a preparation method of red bean active ingredients.

[0009] The present application further provides the use of the red bean active ingredients in the preparation of a drug for treating metabolic syndrome.

[0010] To achieve the above-mentioned technical objectives, the present invention adopts the following technical solution: This invention first provides a method for preparing active ingredients from red beans, comprising the following steps: 1) Crude extract extraction: Weigh red bean seeds, crush and sieve them, and then extract them 2-3 times with an ethanol aqueous solution at room temperature; combine the extracts, concentrate them under reduced pressure, and then extract them with petroleum ether and n-butanol in sequence to obtain the crude extract; 2) Macroporous adsorption resin enrichment: The crude extract obtained in step 1) was subjected to macroporous resin chromatography to obtain five components: A, B, C, D, and E. 3) Silica gel column chromatography separation: Take the C component obtained in step 2) and separate the C component by silica gel column chromatography. The C component is separated by silica gel column chromatography to obtain four sub-components: C1, C2, C3 and C4. 4) Recrystallization purification: The C1 sub-fraction of the C component obtained in step 3) is recrystallized and purified to obtain the active ingredients of red beans.

[0011] Specifically, in step 1), after the red bean seeds are crushed, they are extracted 2-3 times with 70-80% (volume fraction) ethanol aqueous solution at 2-3 times their weight, each time for 12-24 hours.

[0012] Specifically, in step 1), the vacuum concentration process parameters are -0.090~-0.098 MPa and heating temperature 40-50℃.

[0013] Specifically, in step 1), when using petroleum ether and n-butanol for extraction, each extraction lasts 0.5 to 1 hour, and is performed 3 to 5 times.

[0014] Specifically, in step 2), the macroporous resin chromatography step is as follows: the n-butanol extract obtained in step 1) is placed on a macroporous resin and eluted sequentially with pure water and 20%, 50%, 70%, and 95% ethanol aqueous solutions (all ethanol aqueous solutions are volume fractions), and then concentrated under reduced pressure.

[0015] Specifically, in step 2), the vacuum concentration process parameters are -0.090~-0.098 MPa and heating temperature 40-50℃.

[0016] Specifically, in step 3), the separation of component C using silica gel column chromatography involves thoroughly grinding and mixing component C with 80-100 mesh silica gel, using 100-200 mesh silica gel as the stationary phase, and packing the column using a wet method. During the elution stage, mobile phase I (dichloromethane-methanol-water system) is used for stepwise gradient elution, with gradient ratios (v / v / v) sequentially as follows: 9:2:0.1 → 8:2:0.2 → 7:3:0.5. Each ratio is maintained until the target component elutes before switching to the next ratio. During elution, each fraction is qualitatively analyzed using thin-layer chromatography (TLC), similar fractions are combined, and the combined fractions are concentrated under reduced pressure. Finally, four sub-components are separated from component C and named HXD-50%-1 (C1), HXD-50%-2 (C2), HXD-50%-3 (C3), and HXD-50%-4 (C4), respectively.

[0017] Specifically, in step 3), the vacuum concentration process parameters are -0.090~-0.098 MPa and heating temperature 40-50℃.

[0018] Specifically, in step 4), the recrystallization purification step is to dissolve the C1 sub-fraction in component C by heating it with methanol at 40~50℃ for 1~2 hours, then let it stand for 0.5~2 hours to crystallize, filter it, and repeat the same dissolution-crystallization-filtration procedure 3~5 times to obtain a white solid, which is the active ingredient of red bean.

[0019] Specifically, based on spectral analysis including NMR and mass spectrometry, the active ingredient in red beans is identified as: (+)-catechin 7-O-β-D-glucopyranoside ((+)-catechin 7-O-β-D-glucopyranoside), with the following structural formula:

[0020] Furthermore, based on a general inventive concept, the present invention also provides the application of the said red bean active ingredients in the preparation of a drug for the treatment of metabolic syndrome.

[0021] Furthermore, based on a general inventive concept, the present invention also provides the use of the said red bean active ingredient in the preparation of pancreatic lipase and / or α-glucosidase inhibitors.

[0022] Furthermore, based on a general inventive concept, the present invention also provides the application of the said red bean active ingredient in the preparation of a dual inhibitor that simultaneously inhibits α-glucosidase and pancreatic lipase.

[0023] Furthermore, based on a general inventive concept, the present invention also provides the application of the said red bean active ingredients in the preparation of drugs or functional foods for the prevention or treatment of obesity.

[0024] Furthermore, based on a general inventive concept, the present invention also provides the application of the said red bean active ingredients in the preparation of drugs or functional foods for the prevention or treatment of type II diabetes.

[0025] Furthermore, based on a general inventive concept, the present invention also provides the application of the said red bean active ingredient in the preparation of an adipocyte proliferation inhibitor.

[0026] Furthermore, based on a general inventive concept, the present invention also provides the application of the active ingredient of red bean in inhibiting lipid droplet accumulation during the differentiation of 3T3-L1 preadipocytes.

[0027] Furthermore, based on a general inventive concept, the present invention also provides the application of the active ingredients of red beans in the preparation of drugs or foods for regulating blood lipid levels.

[0028] Furthermore, based on a general inventive concept, the present invention also provides the application of the active ingredients of red beans in the preparation of drugs or foods that regulate liver function. Specifically, the active ingredients of red beans can regulate alanine aminotransferase (ALT) and aspartate aminotransferase (AST).

[0029] Furthermore, based on a general inventive concept, the present invention also provides the application of the active ingredients of red beans in the preparation of weight-loss drugs or weight-loss foods.

[0030] Furthermore, based on a general inventive concept, this invention also provides the application of the active ingredients of red beans in the preparation of health products. Specifically, the health products are weight-loss health care drugs or weight-loss functional foods.

[0031] Furthermore, the present invention also provides the application of the active ingredient of red bean (i.e., compound CG) in the preparation of weight loss preparations, specifically, the preparations are any one of tablets, capsules, oral preparations, and injections.

[0032] Furthermore, the present invention also provides a tablet comprising the active ingredient of the red bean.

[0033] Specifically, the preparation is any one of tablets, capsules, oral preparations, and injections. When the preparation is a tablet, the tablet includes the red bean active ingredient, microcrystalline cellulose, pregelatinized starch, crospovidone, and magnesium stearate.

[0034] Specifically, the mass of each component in the tablet is as follows: 40-60g of red bean active ingredient, 70-90g of microcrystalline cellulose, 40-60g of pregelatinized starch, 5-15g of crospovidone, and 1-5g of magnesium stearate.

[0035] Specifically, the tablets are prepared through the following steps: (1) The active ingredients of red beans (i.e., compound CG in this application), microcrystalline cellulose, pregelatinized starch and cross-linked polyvinyl ether are sieved, then mixed for 10 to 30 minutes until uniform, and then magnesium stearate is added and mixed for another 5 to 10 minutes to obtain a mixture.

[0036] (2) The mixture is compressed into tablets using a single punch tablet press to obtain tablets with smooth surface and uniform color.

[0037] Specifically, when compressing tablets, adjust the tablet weight to 0.1~0.3g / tablet.

[0038] Specifically, each tablet prepared contains 80-150 mg of active ingredient.

[0039] Furthermore, the present invention also provides a functional food comprising the active ingredients of red beans, mainly used for weight loss, prevention or treatment of type II diabetes, or treatment of metabolic syndrome.

[0040] Furthermore, the present invention also provides a health care product comprising the active ingredients of red beans, mainly used for weight loss, prevention or treatment of type II diabetes, or treatment of metabolic syndrome.

[0041] The (+)-catechin 7-O-β-D-glucopyranoside described in this invention can be used, either alone or in combination, to prepare drugs or health foods for the treatment of diabetes, obesity, and their complications. This invention overcomes the technical bottleneck of unknown active ingredients in existing plant extracts, obtaining high-purity monomers (≥98%) through a specific separation and purification process, providing a clear material basis for the development of multi-target, low-toxicity, naturally derived anti-metabolic syndrome drugs.

[0042] The active ingredient of this invention is (+)-catechin 7-O-β-D-glucopyranoside, precisely isolated from red beans. In vitro experiments have demonstrated that this active ingredient has potent inhibitory effects on both α-glucosidase and pancreatic lipase. Further cell and animal experiments have shown that this active ingredient can significantly inhibit the differentiation and lipid accumulation of 3T3-L1 preadipocytes, and can effectively reduce the body weight, body fat, and blood lipid levels in high-fat diet-induced obese mice, and improve hepatic steatosis. This invention exerts a multi-target anti-obesity effect by synergistically inhibiting the digestion and absorption of sugars and fats, and has broad application prospects in the preparation of drugs and functional foods for the prevention and treatment of metabolic diseases such as obesity and type II diabetes.

[0043] Compared with the prior art, the advantages of the present invention are: 1. This invention obtains the key active ingredient from red beans: (+)-catechin 7-O-β-D-glucopyranoside (flavonoid glycoside), and for the first time clearly identifies that this active ingredient has dual metabolic enzyme inhibitory properties. In vitro enzyme activity experiments confirm that (+)-catechin 7-O-β-D-glucopyranoside has a half-maximal inhibitory concentration (IC50) against α-glucosidase. 50 The concentration of [unspecified substance] was 0.43 ± 0.06 mmol / L, indicating a strong inhibitory effect on glucose absorption; the IC50 value for pancreatic lipase was [unspecified]. 50 Reaching 0.41 ± 0.07 mmol / L (equivalent to orlistat (IC50)). 50 It showed 2.49 times the activity of the standard (1.02±0.25 mmol / L), exhibiting significant inhibitory activity.

[0044] 2. This invention develops a new extraction and purification process: adopting an integrated technical route of "ethanol gradient extraction - macroporous resin enrichment - preparative chromatographic purification", which enables the purity of the target component to reach more than 98% and the yield to be 3-5 times higher than that of traditional methods.

[0045] 3. This invention establishes a standardized quality control system, including a dual-enzyme activity evaluation method, providing a reliable basis for product quality control.

[0046] 4. The compounds obtained in this invention can effectively block the intestinal absorption of carbohydrates and fats in the diet by synergistically inhibiting the action of carbohydrate hydrolases and lipases, and have significant application prospects in the prevention and treatment of obesity, type II diabetes and related metabolic disorders.

[0047] 5. This invention not only provides new candidate active ingredients for the prevention and treatment of metabolic syndrome, but also promotes the high-value utilization of medicinal and edible plants. The standardized extract obtained can be widely used in functional foods, dietary supplements, and drug development, and has significant research value and industrialization potential. Attached Figure Description

[0048] Figure 1 Mass spectra of compound (+)-catechin 7-O-β-D-glucopyranoside (CG); Figure 2 The 1H NMR spectrum of compound (+)-catechin 7-O-β-D-glucopyranoside (CG); Figure 3 The carbon NMR spectrum of compound (+)-catechin 7-O-β-D-glucopyranoside (CG); Figure 4 The inhibitory activity of compound CG against α-glucosidase; Figure 5 The inhibitory activity of compound CG on pancreatic lipase; Figure 6 The change in cell viability of 3T3-L1 cells after co-culturing with sample (+)-catechin 7-O-β-D-glucopyranoside (HXD-50-1) for 48 h; Figure 7 Quantitative analysis of lipid droplet accumulation in 3T3-L1 adipocytes using CG; Figure 8 Oil Red O staining (200x) showed that (+)-catechin 7-O-β-D-glucopyranoside (HXD-50-1) inhibited lipid droplet accumulation in 3T3-L1 adipocytes; where A is blank, B is model, C is hxd-50-1 (1.25), and D is hxd-50-1 (2.5). Figure 9 The effect of (+)-catechin 7-O-β-D-glucopyranoside (CG) on mouse body weight; Figure 10 The effect of CG on fat index and liver index in obese mice; Figure 10 The left-middle chart shows the effect on the liver coefficient, and the right-middle chart shows the effect on the fat coefficient. Figure 11 The effect of CG on blood lipid levels in obese mice; Figure 11 In the table, A represents the triglyceride (TG) level of each group (NC: normal diet group, HFD: high-fat diet induction group, OT: orlistat positive drug group, CG-L, CG-M, CG-H: low, medium, and high dose CG administration groups); B represents the total cholesterol (TG) level of each group; C represents the low-density lipoprotein (LDL) level of each group; and D represents the high-density lipoprotein (HDL) level of each group. Figure 12 The effect of CG on alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in obese rats; Figure 12 Left image: Liver function indicator alanine aminotransferase (ALT) level; Right image: Aspartate aminotransferase (AST) level. Figure 13 The impact on the morphological and pathological analysis of liver tissue in obese mice; Figure 14 The impact on the morphological and pathological analysis of brown adipose tissue in obese mice; Figure 15 The effect of white adipose tissue morphology on obese mice. Detailed Implementation

[0049] The technical solutions in the embodiments of this application will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments in this application, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of this application. Unless otherwise specified, the instruments and equipment used in the following embodiments are all commercially available conventional instruments and equipment; unless otherwise specified, the reagents and materials used in the following embodiments are all commercially available conventional reagents and materials.

[0050] In the following examples, room temperature or normal temperature refers to 25±5℃.

[0051] The red beans used in the following examples are from: Heilongjiang Hegang City, October 2024, Heiken 290 red beans, deep red color, white hilum, thin skin, plump seeds, uniform shape and size, 100 seeds weigh 15-17 grams. Example 1

[0052] Example 1 provides a method for preparing the active ingredients of red beans, the specific steps of which are as follows: 1) Crude extract extraction: Weigh 30 kg of red bean seeds and crush them into blocks or large particles using a pulverizer. The particle size of the large particles is usually between 3-8 mm. At room temperature, extract 2-3 times with 2-3 times the weight of 70% (volume fraction) ethanol aqueous solution, 24 h each time. Combine the extracts and concentrate under reduced pressure to obtain a reddish-brown extract. The reduced pressure concentration process parameters are -0.090~-0.098 MPa and heating temperature 40-50℃. Dissolve the reddish-brown extract in 5 L of water and extract with an equal volume of petroleum ether 3-5 times, 0.5-1 h each time to remove fatty acids. Then extract with 5 L of n-butanol 3-5 times, 0.5-1 h each time to obtain 204 g of crude n-butanol extract.

[0053] 2) Macroporous adsorption resin enrichment: The n-butanol extract obtained in step 1) was subjected to macroporous resin chromatography. Specifically, the n-butanol extract obtained in step 1) was placed on a macroporous resin column (specifically, column model: RY Boro 3.3; using AB-8 macroporous resin, resin weight approximately 3000g), and eluted sequentially with pure water and 20%, 50%, 70%, and 95% ethanol aqueous solutions (all ethanol aqueous solutions are volume fractions). Then, the solution was concentrated under reduced pressure to obtain five components A, B, C, D, and E. The reduced pressure concentration process parameters were -0.090~-0.098 MPa and heating temperature 40-50℃.

[0054] 3) Silica gel column chromatography separation: Take 22 g of fraction C (i.e., the fraction eluted with 50% ethanol) for chromatographic separation; The specific steps are as follows: Component C is thoroughly ground and mixed with 80-100 mesh silica gel. A wet column is packed using 100-200 mesh silica gel as the stationary phase. During the elution stage, mobile phase I (dichloromethane-methanol-water system) is used for stepwise gradient elution. The gradient ratios (v / v / v) are: 9:2:0.1 → 8:2:0.2 → 7:3:0.5. Each ratio is maintained until the target component elutes before switching to the next ratio. During elution, each fraction is qualitatively analyzed by thin-layer chromatography (TLC). Similar fractions are combined, and the combined fractions are concentrated under reduced pressure. Finally, four sub-components are separated from component C and named HXD-50%-1 (C1), HXD-50%-2 (C2), HXD-50%-3 (C3), and HXD-50%-4 (C4), respectively. The vacuum concentration process parameters are -0.090~-0.098 MPa and heating temperature 40-50℃.

[0055] 4) Recrystallization purification: The HXD-50%-1 subfraction in component C was dissolved in methanol at 40℃ for 1-2 h, then allowed to stand for 0.5 h to crystallize, filtered, and the filter cake was repeated 3-5 times according to the same dissolution-crystallization-filtration procedure to obtain 1.4 g of white solid, namely product I (pure product, HPLC purity ≥98%). Product I was identified by NMR, mass spectrometry and other spectroscopic methods. According to the literature (Hori Yumiko, Murakoso Tomomi, Fukumura Motonori. Constituents and antioxidant activity of a hot-water extract of adzuki (Vigna angularis) beans[J]. Japanses Society Nutrition Food Science, 2009, 62(1): 3-11), product I was identified as: (+)-catechin7-O-β-D-glucopyranoside ((+)-catechin7-O-β-D-glucopyranoside). In subsequent experiments in this application, product I will be referred to as CG.

[0056] The NMR data of the compounds obtained in step 4) are shown in Table 1 below, and the corresponding NMR and mass spectra are as follows. Figure 1 , 2 As shown in Figure 3.

[0057] Table 1. NMR data of (+)-catechin 7-O-β-D-glucopyranoside.

[0058]

[0059] The structural formula of the prepared compound is shown below:

[0060] (+)-Catechin 7-O-β-D-glucopyranoside (CG). Example 2

[0061] Example 2 describes the activity determination of the compound described in Example 1.

[0062] 1. Assay for inhibition of pancreatic lipase activity 1.1 Solution Preparation (1) Tris-HCl buffer: Weigh 6.05g of Tris base into a 50ml volumetric flask, add 40ml of distilled water, adjust the pH to 8 with hydrochloric acid and make up to 50ml with distilled water to prepare a 1mol / L solution. Then take 2.6ml from the 1mol / L solution and add 200ml of distilled water to dilute to 13mmol / L, which is the Tris-HCl buffer solution, for later use.

[0063] (2) Pancreatic lipase solution: Weigh 10 mg of pancreatic lipase (porcine pancreatic lipase S100035, 30,000 u / g; Shanghai Yuanye Biotechnology Co., Ltd.) and add 12 ml of 13 mmol / L Tris-HCl buffer to prepare a solution of 25 U / ml. Then centrifuge at 12,000 rpm and 4℃ for 20 minutes and remove the supernatant for later use.

[0064] (3) PNPB solution: Take 0.21 ml of 4-nitrophenylbutyrate PNPB and add 5.29 ml of 13 mmol / L Tris-HCl buffer to prepare a 50 mg / ml solution, then take 0.1 ml and add 4.9 ml of Tris-HCl buffer to prepare a 1 mg / ml solution.

[0065] (4) Orlistat solution: Weigh 25 mg of orlistat and prepare a 1 mg / mL solution using DMSO as solvent. Pipette 3.2 mL, 2.4 mL, 1.6 mL, 0.8 mL and 0.4 mL of the 1 mg / mL solution and dilute with DMSO to prepare 800 μg / mL, 600 μg / mL, 400 μg / mL, 200 μg / mL and 100 μg / mL solutions, respectively.

[0066] 1.2 Screening of active components of pancreatic lipase Following the method described in the reference (Lin Haisheng, Rao Mengwei, Qin Xiaoming, et al. Inhibitory effect of flavonoid extract from Hongjiang orange peel on α-glucosidase and pancreatic lipase [J]. Modern Food Science and Technology, 2023, 39(10): 89-96.), 50 μL of sample solution and pancreatic lipase solution (25 U / mL, prepared with 13 mmol / L Tris-HCl buffer) were respectively pipetted into 96-well plates using a 200 μL pipette. After mixing, the plates were incubated at 37 ℃ for 10 min, and then 50 μL of 1 mg / mL PNPB solution was added. The plates were mixed and incubated at 37 ℃ for 20 min. The absorbance value A was then measured at 405 nm using a microplate reader. Four groups were tested (as shown in Table 2), with three wells per group. The inhibition rate and IC50 were calculated. 50 The value and inhibition rate are shown in the following formula.

[0067]

[0068] Table 2. Grouping of the screening experiment for active components of pancreatic lipase.

[0069]

[0070] 2. Assay for α-glucosidase activity inhibition 2.1 Solution Preparation (1) 0.1 mol / L phosphate buffer: Take 1.39 g of dipotassium hydrogen phosphate and 0.53 g of potassium dihydrogen phosphate, add distilled water to 100 ml to prepare the solution.

[0071] (2) α-glucosidase solution: Weigh 0.4 mg of α-glucosidase (G8823-100U; α-glucosidase (77.16 U / mg), Beijing Solarbio Science & Technology Co., Ltd.) and add 11.61 ml of phosphate buffer (0.1 mol / L) to prepare a 2.5 U / ml solution.

[0072] (3) PNPG solution: Take 3 mg of PNPG (p-nitrophenyl-α-D-glucopyranoside) and use 0.1 mol / L phosphate buffer as solvent to prepare a PNPG solution with a concentration of 2.5 mmol / L.

[0073] (4) Take 1 mg of acarbose (Acarbose, A129816-1g, Shanghai Aladdin Biochemical Technology Co., Ltd.) and add 1 ml of DMSO solvent to prepare a 1 mg / ml solution. Then dilute with DMSO to prepare 100 ug / ml, 50 ug / ml, 30 ug / ml, 20 ug / ml and 8 ug / ml solutions.

[0074] (5) Reaction termination solution: Weigh 1.06 g of Na2CO3 solid, dissolve it in distilled water, transfer it to a 50 mL volumetric flask and make up to volume to prepare a Na2CO3 solution with a concentration of 0.2 mol / mL.

[0075] (6) Sample solution: Same as the sample solution in 1.1.

[0076] 2.2 Screening of α-glucosidase active components The assay was performed on 96-well microplates, following the method described in the reference (Guo Qingfeng, Chen Lin, Yin Zhenhua, et al. Secondary metabolites of Chaetomium globosum H6 and their α-glucosidase inhibitory activity[J]. Acta Mycosystema Sinica, 2019, 38(1):11.DOI:10.13346 / j.mycosystema.180155.). 8 μL of DMSO or sample solution was pipetted into 96-well microplates using a 20 μL pipette. 112 μL of phosphate buffer (1 mmol / L, pH 6.8) was added using a 200 μL pipette. Then, 20 μL of α-glucosidase solution (2.5 U / mL) was added using a 20 μL pipette, mixed, and incubated at 37 ℃ for 15 min. Finally, 20 μL of PNPG (2.5 mmol / L) was added, mixed, and incubated at 37 ℃ for 15 min. After min, 80 μL of Na2CO3 (0.2 mol / L) stop agent was added to each well, mixed well, and the absorbance (A) was measured at 405 nm using a microplate reader. Five groups were set up (as shown in Table 3), with three wells per group. The inhibition rate and IC50 were calculated. 50 The value and inhibition rate are shown in the following formula.

[0077]

[0078] Table 3. Screening experiment groups for α-glucosidase active components.

[0079]

[0080] Test results, such as Figure 4 , 5 As shown: Among them, the inhibitory activity of (+)-catechin 7-O-β-D-glucopyranoside was 0.43±0.06 mmol / L (against α-glucosidase); 0.41±0.06 mmol / L (against pancreatic lipase) was equivalent to that of orlistat (IC50). 50 =1.02±0.25mmol / L) 2.49 times the activity of the standard. Example 3

[0081] Example 3 investigates the cellular-level anti-obesity effect of compound CG (inhibition of 3T3-L1 preadipocyte differentiation).

[0082] The 3T3-L1 preadipocytes (mouse embryonic fibroblasts) used in the following experiments were purchased from ATCC in the United States.

[0083] The main experimental reagents are shown in Table 4.

[0084] Table 4. Main reagents used in the experiment.

[0085]

[0086] 1. Cell proliferation assay (MTT method) The objective was to determine the effect of compound CG on the proliferation of 3T3-L1 cells and its safe concentration. The specific steps are as follows: 1.1 Remove the selected cell line (3T3-L1 cells) from liquid nitrogen and quickly place it in a 37°C water bath. Gently shake the cryovial to thaw the cryopreservation solution. After thawing, transfer the cells to a centrifuge tube containing 5 ml of culture medium, centrifuge to collect the cells, centrifuge at 1000 rpm for 5 min at room temperature, and discard the supernatant. Resuspend the cells in complete culture medium containing 10% fetal bovine serum, seed them into culture dishes, gently pipette to mix, and incubate at 37°C under saturated humidity of 5% CO2.

[0087] Take cells in the logarithmic growth phase and in good growth condition (3T3-L1), and use 6×10 3 Seeds were planted per well in 96-well cell culture plates and incubated overnight at 37°C in a 5% CO2 incubator; (100 μL of sterile PBS was added to the wells around the cells).

[0088] 1.2 Take the sample (compound CG), sterilize it by UV irradiation for 30 min, add complete culture medium to 1 mg / mL, and sonicate until completely dissolved to prepare the sample stock solution (concentration 1 mg / mL). Dilute with complete culture medium to prepare the sample working solution. Take a 96-well plate, wait for the cells to adhere and grow for 24 h, discard the culture medium in each well, wash twice with PBS, and add the drug at various concentrations. Set up 5 groups of 3T3-L1 cells: Control, HXD-50-1 (1.25, 2.5, 5, 10 μg / mL), and then incubate for 48 h. In this example, HXD-50-1 is compound CG.

[0089] 1.3 Remove the culture medium containing the sample (CG). Wash each well three times with PBS, add 100 μL of culture medium containing 0.5 mg / mL LMT in each well, and incubate in a 5% CO2 incubator at 37°C for 4 hours. Discard the supernatant and add 100 μL of DMSO to each well. Gently shake for 10 min and then measure the absorbance at 570 nm.

[0090] Data analysis: Cell viability was calculated (vs. control group), and concentrations with inhibition rate <10% were selected for subsequent experiments.

[0091] The results are as follows Figure 6 As shown, Figure 6 HXD-50-1, namely (+)-catechin 7-O-β-D-glucopyranoside (CG), showed in MTT assays that the mean cell viability was 95.4% and 99.8% at CG concentrations of 1.25 μg / mL and 2.5 μg / mL, respectively. This indicates that CG concentrations at these experimental concentrations had almost no inhibitory effect on 3T3-L1 preadipocytes. Therefore, concentrations of 1.2 and 2.5 μg / mL were selected for subsequent experiments.

[0092] 2. Cell culture and differentiation induction (mainly used to evaluate the inhibitory effect of compound CG on differentiation and lipid droplet accumulation): 2.1 3T3-L1 cells (3×10⁻⁶) were added. 4 (Numbers / well) were inoculated into 96-well culture plates and cultured in DMEM medium containing 10% heat-inactivated fetal bovine serum (FBS) and 1% penicillin-streptomycin for 2 days at 37°C in a 5% CO2 incubator.

[0093] 2.2 Differentiation Induction: Two days after complete cell confluence (designated as day 0), the treatment group was switched to induction medium I (DMEM medium containing 0.5 mmol / L IBMX, 1 μmol / L dexamethasone, and 10 μg / mL insulin), while the control group was switched to fresh DMEM medium containing 1% antibiotics and 10% heat-inactivated fetal bovine serum. Three days after induction (day 3), the treatment group was switched to induction medium II (DMEM medium containing 10 μg / mL insulin), while the control group was switched to fresh DMEM medium containing 1% antibiotics and 10% heat-inactivated fetal bovine serum. From day 5 onwards, the treatment group was switched to induction medium II every two days until day 8, when over 80% of the cells had differentiated and matured, and the cells were filled with lipid droplets.

[0094] 3. Drug treatment and Oil Red O staining (the effect of CG on lipid accumulation during the differentiation of 3T3-L1 preadipocytes was determined using Oil Red O staining). After induction and differentiation, 3T3-L1 preadipocytes were treated with different concentrations of CG (1.25 μg / mL and 2.5 μg / mL) for 8 days. The intracellular lipid droplet content was then detected using Oil Red O staining. The specific steps are as follows: 3.1 Starting on day 0 of induced differentiation, different concentrations of active ingredient I (i.e., compound CG, at concentrations of 1.25 μg / mL and 2.5 μg / mL, respectively) were added to the induction medium. A non-induced group (Preadipocyte) and an induced differentiation control group (Adipocyte) were established.

[0095] 3.2 On day 8 of differentiation, the culture medium was aspirated and the cells were washed twice with PBS.

[0096] 3.3 Fix cells with 10% formalin at room temperature for 30 minutes.

[0097] 3.4 Soak the culture plate twice with 60% isopropanol for 5 minutes each time, then blow dry the plate until the wells are completely dry.

[0098] 3.5 Add freshly prepared Oil Red O working solution and stain at room temperature for 30 minutes.

[0099] 3.6 Discard the staining solution and wash repeatedly with distilled water until no red dye is washed off.

[0100] 3.7 Wash twice with 60% isopropanol (5 minutes each time), dry, add 100% isopropanol to extract the bound dye, and gently shake for 10 minutes; during differentiation, treat cells with specific concentrations of compound CG, and perform Oil Red O staining on days 1, 3, 5, and 8 of differentiation as described above.

[0101] 3.8 Take the extract and measure the absorbance at 510 nm using an ELISA reader. The results are expressed as the relative percentage of fully differentiated cells.

[0102] 4. Cell grouping and modeling After 3T3-L1 cells were induced to differentiate to day 10 (forming mature adipocytes), they were divided into two groups and intervention groups: a blank control group and a drug intervention group. The blank control group was replaced with fresh complete culture medium containing an equal volume of solvent (such as 0.1% DMSO). The intervention groups were cultured in fresh complete culture medium containing compound CG at concentrations of (2.5 μg / mL and 1.25 μg / mL), respectively. After 24 hours of culture, the cells were thawed with PBS buffer for 1 hour.

[0103] The results showed that active ingredient I (i.e. compound CG) could significantly inhibit the accumulation of lipid droplets during the differentiation of 3T3-L1 preadipocytes in a concentration-dependent manner. Example 4

[0104] Example 4 investigates the weight loss efficacy of compound CG in animals.

[0105] 1. Animal experimental methods 1.1 Grouping and Administration Forty-eight healthy male SPF-grade C57BL / 6J mice (mean weight: 19.3 g) aged 6-8 weeks were selected and divided into 6 groups (N=8). The positive control group (orlistat) received a dose of 10 mg / kg. The CG administration groups received low, medium, and high doses of 30 mg / kg, 60 mg / kg, and 120 mg / kg, respectively, prepared as a suspension in 2 mL of water. The suspension was freshly prepared and administered by gavage at a volume of 10 mL / kg once daily for 12 weeks, with weekly recording of body weight and food intake. The normal control group and the high-fat control group received 2 mL of 0.9% saline by gavage. Specific drug administration and grouping details are as follows: 1) Normal control group (NC): Normal feed + physiological saline by gavage.

[0106] 2) High-fat model group (HFD): 45% high-fat diet + physiological saline by gavage.

[0107] 3) Positive control group (OT): High-fat diet + orlistat (10 mg / kg) by gavage.

[0108] 4) Low-dose CG group (CG-L): High-fat diet + oral gavage of active ingredient I (30 mg / kg).

[0109] 5) CG medium-dose group (CG-M): high-fat diet + oral gavage of active ingredient I (60 mg / kg).

[0110] 6) High-dose CG group (CG-H): High-fat diet + oral gavage of active ingredient I (120 mg / kg).

[0111] 1.2 Sample collection: Fasting for 12 hours after the last administration (free access to water), blood was collected from the eyeballs after anesthesia, serum was separated, and the tissue was dissected and separated. White adipose tissue (perrenal and epididymal) and brown adipose tissue (posterior scapular region and liver) were collected, weighed, and the fat coefficient and liver coefficient were calculated.

[0112] 1.3 Analysis and Testing: Weight, height, food intake, and other indicators were measured weekly; two days before the end of the experiment, fasting for 12 hours was required, and an oral glucose tolerance test was conducted. 1.4 The serum biochemical analyzer measures four lipid parameters in the serum: triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C); and two liver function parameters: aspartate aminotransferase (AST) and alanine aminotransferase (ALT).

[0113] 1.5 Histopathological examination: Fresh liver, white adipose tissue and brown adipose tissue were fixed in 10% neutral formaldehyde, embedded in paraffin, sectioned, stained with H&E and Oil Red O, and observed under an optical microscope.

[0114] 2. Animal experiment results: 2.1 Effects on mouse body weight Body weight is a key indicator for assessing obesity. The dynamic changes in body weight of the six groups of mice after 12 weeks of continuous feeding are shown below. Figure 9 As shown, all groups exhibited an overall upward trend. Compared to the HFD group, the weight gain of mice in all drug treatment groups was reduced, with a decrease of 17.43% in the positive control group (p < 0.05), 11.39% in the low-dose CG group (p < 0.05), 13.43% in the medium-dose CG group (p < 0.05), and 20.77% in the high-dose CG group (p < 0.05). Notably, the average weight gain in the high-dose CG group was not statistically different from that in the normal control group (p > 0.05). The body fat coefficient of the CG-H group was significantly lower than that of the HFD group, and even slightly lower than that of the NC group.

[0115] 2.2 Effects of CG on fat coefficient and liver coefficient in obese mice The results of the effect on the fat coefficient in mice are as follows: Figure 10 As shown in the figure: The difference between the HFD group and the NC group was significant. Compared with the HFD group, the fat coefficient of all treatment groups was reduced. The CG-H group showed the smallest reduction, with the greatest difference from the HFD group. Notably, the fat coefficient of the CG-H group was even lower than that of the NC group. The effect on the liver coefficient is shown in the figure: the NC group had the smallest reduction, showing a significant difference compared to the other groups. Compared with the HFD group, the fat coefficient of all treatment groups was reduced. With increasing CG dosage, the fat coefficient showed a gradually decreasing trend.

[0116] 2.3 Effects of CG on blood lipid levels in obese mice Obesity is considered a disease caused by a variety of factors. Figure 11 The figure shows the changes in blood lipids in each group of rats. Figure 11As shown, the HFD group had the highest levels of TG (2.39 ± 0.10) and TC (10.90 ± 0.45), significantly higher than the NC group (p<0.05), being 4.89 times and 3.02 times higher, respectively. Compared with the HFD group, the TG levels in the OT group, CG-L group, CG-M group, and CG-H group decreased by 81.92%, 56.41%, 40.48%, and 77.36%, respectively, with significant differences (p<0.05).

[0117] Compared to the HFD group, the TC content in the OT group, CG-L group, CG-M group, and CG-H group also decreased by 59.77%, 41.26%, 57.10%, and 69.83%, respectively, showing significant differences. The TC content decreased in all groups (p<0.05).

[0118] Figure 11 The results in C showed that the HFD group (5.93 ± 0.26) had the highest LDL-C content, which was 6.17 times that of the NC group. Meanwhile, the LDL-C content in each treatment group gradually decreased with increasing dose (CG-L 6.24 ± 0.28, CG-M 2.42 ± 0.16, CG-H 1.66 ± 0.22). However, there was no significant difference between the CG-L group and the HFD group (p>0.05), and the CG-H group was slightly lower than the OT group (1.24 ± 0.10).

[0119] Figure 11 In group D, compared with the NC group, the high-fat diet (HFD) group showed a significant decrease in high-density lipoprotein cholesterol (HDL-C) levels (p<0.05). Meanwhile, in all CG intervention groups, HDL-C levels showed a numerical dose-dependent increasing trend (CG-L 1.12 ± 0.10, CG-M 1.47 ± 0.04, CG-H 1.79 ± 0.05). Compared with the HFD group, there was no significant difference in HDL-C levels in the CG-L group (p>0.05), while the HDL-C levels in the CG-H group were significantly increased (p<0.01). These results indicate that CG significantly improves lipid dyslipidemia induced by a high-fat diet.

[0120] 2.4 Effects of CG on alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in obese mice (i.e., effects on liver function) Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) are markers of liver damage and are commonly used to indicate the state of liver health. Figure 12 As shown, ALT and AST activities in mice fed a high-fat diet (HFD) were significantly higher than those in the normal control group (NC). pThe levels of ALT and AST were <0.05, which were 3.64 times and 5.40 times higher than those in the NC group, respectively, indicating that HFD successfully induced liver injury. Compared with the HFD group, both the positive control group (OT group) and the CG intervention group significantly reduced ALT and AST levels (<0.05), which were 3.64 times and 5.40 times higher than those in the NC group, respectively, indicating that HFD successfully induced liver injury. p (<0.05), and moreover, in each dose group of CG, the reduction in ALT and AST was more significant with increasing dosage. This indicates that CG has a significant protective effect on the liver in a dose-dependent manner.

[0121] 2.5 Effects of CG on the morphological and pathological analysis of liver tissue in obese rats, the results are as follows: Figure 13 , 14 As shown in Figure 15.

[0122] 2.5.1 Results of liver histopathological examination as follows Figure 13 As shown, hepatocytes in the NC group exhibited normal morphology, with no cytoplasmic vacuolation (i.e., fatty degeneration) or inflammatory response. In contrast, the HFD group showed significant hepatocyte degeneration and inflammatory cell infiltration. Compared to the HFD group, the lesion severity in the CG-L group was similar to that in the HFD group, the lesion severity in the CG-M group was slightly reduced, and the liver lesion severity in the CG-H and OT groups was the mildest with no significant difference, having essentially returned to a normal state. In conclusion, CG has an ameliorative and repairing effect on hepatocyte fatty degeneration.

[0123] 2.5.2 As Figure 14 As shown, compared to the LFD group, the HFD group showed a greater number of unilateral lipid droplets in the cells, indicating a high concentration of lipid droplets and reduced activity in the brown adipose tissue. In contrast, the NC and CG groups showed a significant decrease in unilateral lipid droplets and a substantial increase in multi-ocular lipid droplets, suggesting that the positive control drugs orlistat and CG significantly improved the lipid content in brown adipose tissue. These results indicate that CG has a certain protective effect on the normal morphology of brown adipose tissue.

[0124] 2.5.3 The external characteristic of obesity is increased body weight, while the internal characteristic is an increase in the number and size of fat cells. For example... Figure 15 As shown, within the observation field, the adipocytes in the NC group were smaller in outline, more compactly arranged, and more numerous. Compared to the NC group, the adipocytes in the HFD group showed significantly larger cell outlines and looser arrangement, indicating a significant increase in the volume of white adipocytes. In all drug-treated groups, the adipocytes were smaller to varying degrees, and the cell arrangement was more compact.

[0125] Overall, the CG-H group showed a significant reduction in hepatic steatosis, with white adipocytes decreasing in size and becoming more densely packed. In brown adipose tissue, there were fewer uniocular lipid droplets and more multiocular lipid droplets, with a morphology similar to the NC group. Example 5

[0126] Example 5 provides the application of compound CG in formulation (tablet preparation), including formulation composition and preparation method. Details are as follows: 1. Prepare 1000 tablets according to the following prescription: 1.1 Active ingredient I ((+)-catechin 7-O-β-D-glucopyranoside): 50 g; 1.2 Microcrystalline cellulose: 80 g; (Model MCC PH101, conforming to the 2020 edition of the Chinese Pharmacopoeia, Anhui Shanhe Pharmaceutical Excipients Co., Ltd.) 1.3 Pregelatinized starch: 50 g; (Model Starch 1500, conforming to the Chinese Pharmacopoeia, Shanghai Changwei Pharmaceutical Excipients Technology Co., Ltd.) 1.4 Crospovidone: 10 g; (Model: Kollidon® CL, conforms to the Chinese Pharmacopoeia, Shanghai Linchen Pharmaceutical Technology Co., Ltd.) 1.5 Magnesium stearate: 3 g (Pharmaceutical excipient grade, conforming to the Chinese Pharmacopoeia, Anhui Hongyuan Pharmaceutical Excipients Co., Ltd.).

[0127] 2.1 The active ingredient I (i.e., compound CG in this application), microcrystalline cellulose, pregelatinized starch and cross-linked polyvinylpyrrolidone were first passed through a 100-mesh sieve, and then placed in a V-shaped mixing tank and mixed for 30 minutes until homogeneous.

[0128] 2.2 Add magnesium stearate and continue mixing for 5 minutes.

[0129] 2.3 Use a single-punch tablet press and adjust the tablet weight to 0.243 g / tablet for tableting.

[0130] 2.4 The resulting tablets have a smooth surface, uniform color, and their weight variation and hardness meet the requirements of the Chinese Pharmacopoeia. Each tablet contains 100 mg of active ingredient.

Claims

1. An active ingredient from red beans, characterized in that, Identified by NMR and mass spectrometry, the active ingredient of the red bean is (+)-catechin 7-O-β-D-glucopyranoside, with the following structural formula: ； The method for preparing the active ingredients of red beans includes the following steps: 1) Crude extract extraction: Weigh red bean seeds, crush and sieve them, and then extract them 2-3 times with an ethanol aqueous solution at room temperature; combine the extracts, concentrate them under reduced pressure, and then extract them sequentially with petroleum ether, ethyl acetate and n-butanol to obtain the crude extract; 2) Macroporous adsorption resin enrichment: The crude extract obtained in step 1) was subjected to macroporous resin chromatography to obtain five components: A, B, C, D, and E. 3) Silica gel column chromatography separation: Take the C component and E component obtained in step 2), and separate the C component and E component by silica gel column chromatography. The C component is separated by silica gel column chromatography to obtain four sub-components: C1, C2, C3 and C4. 4) Recrystallization purification: The C1 sub-fraction of the C component obtained in step 2) is recrystallized and purified to obtain the active ingredients of red beans.

2. According to the active ingredient of red bean as described in claim 1, in step 2), the macroporous resin chromatography step is as follows: the n-butanol extract obtained in step 1) is placed on a macroporous resin and eluted sequentially with pure water and 20%, 50%, 70%, and 95% ethanol aqueous solutions, and then concentrated under reduced pressure. The ethanol aqueous solutions are all volume fractions.

3. According to claim 1, the active ingredient of red bean, in step 3), the separation of component C by silica gel column chromatography is as follows: component C is thoroughly ground and mixed with 80-100 mesh silica gel, and a wet column is packed using 100-200 mesh silica gel as the stationary phase; during the elution stage, mobile phase I is used for stepwise gradient elution, with the gradient ratios being the following volume ratios: 9:2:0.1 → 8:2:0.2 → 7:3:0.

5. After each ratio is stabilized and eluted until the target component elutes, the next ratio is switched. During the elution process, each fraction was qualitatively analyzed by thin-layer chromatography, similar fractions were combined, and the combined fractions were concentrated under reduced pressure. Mobile phase I is a dichloromethane-methanol-water system.

4. According to claim 1, the active ingredient of red bean, in step 4), the recrystallization purification step is to dissolve the C1 sub-component in component C by heating with methanol at 40~50℃ for 1~2h, then let it stand for 0.5~2h to crystallize, filter, and repeat the same dissolution-crystallization-filtration procedure as above 3~5 times to obtain a white solid, that is, the active ingredient of red bean.

5. The use of the active ingredient of red bean according to any one of claims 1 to 4 in the preparation of a drug for treating metabolic syndrome.

6. The use of the red bean active ingredient according to any one of claims 1 to 4 in the preparation of pancreatic lipase and / or α-glucosidase inhibitors.

7. The use of the active ingredient of red bean according to any one of claims 1 to 4 in the preparation of drugs or functional foods for the prevention or treatment of obesity.

8. The use of the active ingredient of red bean according to any one of claims 1 to 4 in the preparation of weight loss drugs or weight loss foods.

9. The use of the active ingredient of red bean according to any one of claims 1 to 4 in the preparation of a weight-loss formulation, characterized in that, The preparation is any one of tablets, capsules, oral preparations, or injections.

10. A formulation comprising the active ingredient of red bean as described in claim 1, characterized in that, The preparation is any one of tablets, capsules, oral preparations, and injections. When the preparation is a tablet, the mass of each component in the tablet is: 40-60g of the red bean active ingredient, 70-90g of microcrystalline cellulose, 40-60g of pregelatinized starch, 5-15g of crospovidone, and 1-5g of magnesium stearate.

Images