Traditional Chinese medicine composition for treating metabolism-related fatty liver disease (MASLD) and application thereof

Abstract

The invention relates to a traditional Chinese medicine composition for treating metabolism-related fatty liver disease (MASLD) and application thereof. The traditional Chinese medicine composition is prepared from seven traditional Chinese medicines including pericarpium citri reticulatae viride, hawthorn, lotus leaf, rhizoma polygonati, salvia miltiorrhiza, polygonum cuspidatum and oriental wormwood. According to the traditional Chinese medicine composition, through the synergistic effect of multiple components, multiple targets and multiple pathways, glucose and lipid metabolism disorder, inflammation change, mitochondrial homeostasis imbalance and oxidative stress of MASLD patients are remarkably improved. Proved by preclinical pharmacodynamic verification, the seven-ingredient lipid-lowering formula can be used for remarkably reducing the body weight, liver weight and fat content of MASLD model animals and lipid deposition in liver tissues, improving the blood lipid level and insulin resistance phenomena and reducing the release of inflammatory factors. The seven-component lipid-lowering formula replaces natural traditional Chinese medicines, is high in safety, small in side effect and suitable for being taken for a long time, and provides a new treatment choice for MASLD patients.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of pharmaceutical compositions, in particular to a traditional Chinese medicine composition for treating metabolic associated steatohepatitis (MASLD) and application thereof. BACKGROUND

[0002] Metabolic associated steatohepatitis (MASLD), also known as non-alcoholic fatty liver disease (NAFLD), is a chronic liver disease that is widely prevalent worldwide and has become one of the most common chronic liver diseases in the world and China, and the disease burden continues to increase.

[0003] For the treatment of MASLD, the current main method is to combine lifestyle intervention and drug therapy. Lifestyle intervention includes diet control and weight management, exercise therapy, behavior intervention and weight loss surgery, among which diet control and exercise therapy are the cornerstone of the prevention and treatment of MASLD, and low-calorie diet combined with moderate-intensity aerobic exercise is recommended as a first-line measure. Drug therapy mainly includes hypoglycemic / weight loss drugs (such as metformin, SGLT-2 inhibitors, GLP-1 receptor agonists, etc.), antioxidants (such as vitamin E, silymarin), lipid-lowering drugs (such as statins), etc. These drugs are mainly used to improve insulin resistance, reduce liver fat deposition, and regulate blood lipid levels. However, despite the availability of multiple treatment options, the treatment of MASLD is still not ideal, and there are many problems to be solved. The pathogenesis of MASLD is complex and involves multiple pathological mechanisms, and single-target drugs are difficult to intervene comprehensively. It is of practical significance to develop multi-component, multi-target, multi-pathway and safe and effective compound traditional Chinese medicines.

[0004] In order to solve the above problems, the applicant proposes a traditional Chinese medicine composition for treating metabolic associated steatohepatitis (MASLD) and application thereof. SUMMARY

[0005] The purpose of the present application is to provide a traditional Chinese medicine composition for treating metabolic associated steatohepatitis (MASLD) and application thereof to solve the problems in the prior art.

[0006] To achieve the above-mentioned purpose, the present application provides the following technical solution: a traditional Chinese medicine composition for treating metabolic associated steatohepatitis (MASLD), which is made of the following raw materials by weight:

[0007] Pericarpium Citri Reticulatae Viride: 3-15 parts; Polygonum cuspidatum: 2-10 parts; Crataegus pinnatifida: 3-15 parts; Polygonatum sibiricum: 3-15 parts; Herba Artemisiae Scopariae: 1-8 parts; Folium Nelumbinis: 3-15 parts; Salvia miltiorrhiza: 2-10 parts.

[0008] Optionally, the preparation method comprises the following steps:

[0009] Medicine preparation: According to the above proportion, each flavor of traditional Chinese medicine is weighed to ensure that the quality of the medicine meets the standard of the pharmacopoeia.

[0010] Decoction: Add the medicine to the appropriate amount of water, soak for 30 minutes, then boil with a strong fire, and then simmer with a weak fire for 30-60 minutes. Filter out the medicinal liquid.

[0011] Concentration: Concentrate the filtered medicinal liquid to the appropriate volume, or further prepare into pills, powders, granules, etc. according to needs.

[0012] Preparation: According to clinical needs, the concentrated liquid or dried extract is prepared into appropriate dosage forms such as capsules, tablets, oral liquids, etc.

[0013] Optionally, in the preparation of a drug for treating metabolic-related fatty liver disease.

[0014] Optionally, the traditional Chinese medicine composition can significantly improve the following pathological aspects of MASLD patients:

[0015] Disorder of glycolipid metabolism;

[0016] Inflammatory changes;

[0017] Mitochondrial homeostasis imbalance;

[0018] Oxidative stress.

[0019] Optionally, the traditional Chinese medicine composition exerts therapeutic effects on MASLD through the following mechanisms:

[0020] Up-regulating the expression of PPARα, CYP3A4, etc.

[0021] Regulating lipid metabolism and inflammation-related pathways

[0022] Optionally, when preparing a drug, the traditional Chinese medicine composition can be mixed with other drugs or excipients to prepare a dosage form suitable for clinical use.

[0023] Optionally, when treating MASLD, the traditional Chinese medicine composition can be used alone or in combination with other treatment methods.

[0024] Beneficial effects: Significant therapeutic advantages:

[0025] Multi-component, multi-target, and multi-pathway synergistic effect: Seven-flavor lipid-lowering formula is a replacement of seven traditional Chinese medicines carefully matched, containing multiple active ingredients such as luteolin, quercetin, and baicalin. These components can comprehensively regulate the metabolism and immune function of the body through multi-target and multi-pathway synergistic effect, thereby exerting a comprehensive therapeutic effect on MASLD. Compared with single-component or single-target Western medicine, seven-flavor lipid-lowering formula can more comprehensively intervene in the complex pathological mechanism of MASLD, reducing the side effects and drug resistance that may be caused by single-target treatment.

[0026] Overall regulation, treating both symptoms and root causes: Based on the holistic concept of traditional Chinese medicine, Qiwijiangshi Decoction can regulate the balance of qi and blood, yin and yang in the body as a whole by replacing various functions such as soothing the liver, regulating qi, eliminating stagnation, promoting blood circulation, and strengthening the body. It not only treats the symptoms of MASLD, but also fundamentally improves the pathological state of MASLD, achieving both symptoms and root causes.

[0027] High safety and fewer side effects: As a traditional Chinese medicine compound, Qiwijiangshi Decoction is composed of natural plant medicines, which have higher safety and fewer side effects compared to chemically synthesized Western medicines. Both clinical trials and animal experiments have shown that Qiwijiangshi Decoction effectively treats MASLD while having no significant toxic and side effects on other organs and systems of the body, and patients have good tolerance.

[0028] Improved specific symptoms:

[0029] Significant improvement in glucose and lipid metabolism disorders: Qiwijiangshi Decoction can significantly reduce the body weight, liver weight, spleen weight, and liver index of MASLD patients, and reduce the deposition of fat in the liver and adipose tissue. At the same time, it can also regulate blood lipid levels, reduce the content of total cholesterol (TC), triglycerides (TG), and low-density lipoprotein cholesterol (LDL-C) in serum, and increase the content of high-density lipoprotein cholesterol (HDL-C), thereby effectively improving lipid metabolism disorders.

[0030] Effective reduction of insulin resistance: MASLD patients often have insulin resistance, which is one of the important mechanisms of disease progression. Qiwijiangshi Decoction can significantly reduce insulin resistance by promoting glucose uptake and utilization and improving insulin sensitivity, thereby improving abnormal glucose metabolism.

[0031] Significant relief of inflammatory response: Inflammatory response plays an important role in the pathogenesis of MASLD. Qiwijiangshi Decoction can significantly reduce the levels of inflammatory factors in the serum of MASLD patients, such as interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β), thereby reducing the inflammatory response in the liver and whole body and protecting hepatocytes from further damage.

[0032] Effective improvement of mitochondrial function: Mitochondrial dysfunction is one of the important pathological mechanisms of MASLD. Qiwijiangshi Decoction can regulate the expression of mitochondrial-related functional genes, improve mitochondrial ultrastructure, increase mitochondrial DNA relative copy number, and restore mitochondrial membrane potential, thereby improving the energy metabolism capacity of hepatocytes, reducing oxidative stress and lipid peroxidation damage.

[0033] Improvement of quality of life:

[0034] Significant relief of clinical symptoms: MASLD patients often have symptoms such as fatigue, loss of appetite, abdominal distension, etc., which seriously affect the quality of life. Qiwijiangzhi Decoction can significantly improve these symptoms and improve the quality of life of patients. At the same time, by regulating the overall functional state of the body, Qiwijiangzhi Decoction can also improve the sleep quality and mental state of patients, making patients feel overall physical and mental improvement.

[0035] Effective prevention of complications: If MASLD is not treated in time, it may progress to serious complications such as cirrhosis and liver cancer, which seriously threaten the life and health of patients. Qiwijiangzhi Decoction can significantly reduce the risk of these complications by effectively treating MASLD. In addition, long-term use of Qiwijiangzhi Decoction can also reduce the incidence of metabolic diseases such as cardiovascular disease and diabetes in MASLD patients, further improving the quality of life of patients.

[0036] Significant improvement in patient compliance: Qiwijiangzhi Decoction, as a traditional Chinese medicine compound, is relatively easy for patients to accept in terms of taste and method of taking, and its safety and effectiveness have been clinically verified, so patients have increased confidence in treatment, and treatment compliance has also improved accordingly. This helps to ensure that patients take medicine on time and in the right amount, thereby improving treatment effectiveness. BRIEF DESCRIPTION OF DRAWINGS

[0037] Figure 1 is a flowchart of an embodiment of the present application;

[0038] Figure 2 is a general appearance of each group of mice in the embodiment of the present application

[0039] Figure 3 is the body weight growth change of each group of mice in the embodiment (A) of the present application, and (B) the body weight at the end of the 18th week

[0040] Figure 4 is the change in food intake of each group of mice during the administration period (A), and the change in water intake of each group of mice during the administration period (B)

[0041] Figure 5 is the liver weight (A), spleen weight (B), and liver index (C) in the embodiment of the present application

[0042] Figure 6 is the iBAT weight (A) and eWAT weight (B) in the embodiment of the present application

[0043] Figure 7 is the general appearance of the liver of each group of mice in the embodiment of the present application

[0044] Figure 8 is the HE staining of mouse liver tissue (x200) in the embodiment of the present application

[0045] Figure 9 Liver tissue oil red O staining of the embodiment of the present application: (A) staining photo (x400), (B) quantitative result

[0046] Figure 10 The first row (A) liver weight (g) (B) spleen weight (g) (C) liver index (g) of the embodiment of the present application;

[0047] The second row (A) brown fat (g), (B) white fat (g);

[0048] The third row (A) liver TG (mg / g), (B) liver tissue TG (mg / g);

[0049] Figure 11 The (A) serum TC level, (B) serum TG level, (C) serum HDL-C level, (D) serum LDL-C level of the embodiment of the present application;

[0050] Figure 12 The first row (A) serum TC level, (B) serum TG level, the second row (C) serum HDL-C level, (D) serum LDL-C level of the embodiment of the present application; The third row (A) fasting 8h blood glucose, (B) serum insulin, (C) HOMA-IR index;

[0051] Figure 13 The (A) serum ATL, (B) serum ASL of the embodiment of the present application;

[0052] Figure 14 The first row (A) serum ATL level, (B) serum ASL level of the embodiment of the present application; The second row (A) serum IL-6 level, (B) serum TNF-α level, (C) serum IL-1β level DETAILED DESCRIPTION

[0053] The preferred embodiments of the present application are described below with reference to the accompanying drawings, so that the technical content of the present application is more clear and easy to understand. The present application can be embodied in many different forms, and the protection scope of the present application is not limited to the embodiments mentioned in the text.

[0054] The present application provides a traditional Chinese medicine composition for treating MASLD, named Qiwjiangzhi recipe. The traditional Chinese medicine composition is carefully prepared from the following raw medicinal materials in a certain proportion by weight: pericarpium citri reticulatae viride 15g, fructus crataegi 15g, folium loti 15g, rhizoma polygonati 15g, salvia miltiorrhiza 12g, giant knotweed 9g, and herba artemisiae 6g.

[0055] Preparation process of Qiwjiangzhi recipe

[0056] The preparation process of Qiwjiangzhi recipe includes the following fine steps:

[0057] Medicinal material selection and pretreatment: Strictly select high-quality traditional Chinese medicinal materials that meet the standards of the Chinese Pharmacopoeia, remove impurities, and perform necessary cleaning and slicing to ensure the purity and effectiveness of the medicinal materials.

[0058] Scientific decoction and concentration: Mix the pretreated traditional Chinese medicinal materials in proportion, add an appropriate amount of pure water, and use advanced decoction equipment for three times of decoction. After each decoction, remove the dregs through a precision filtration system and collect the filtrate. Combine the filtrates obtained from the three decoctions and use vacuum concentration technology for concentration treatment to obtain a 13.8g / ml concentration of the Seven-Ingredient Lipid-Lowering Formula substitute concentrated liquid. This step aims to fully extract the effective components from the medicinal materials while removing ineffective impurities, thereby improving the purity and efficacy of the liquid.

[0059] Quality detection and control: Strict quality detection is performed on the concentrated liquid, including appearance, identification, inspection, and content determination to ensure that the liquid meets the relevant quality standards.

[0060] Flexible preparation and packaging: According to actual needs, the concentrated liquid can be diluted to an appropriate concentration to prepare various dosage forms such as oral liquids, capsules, and tablets for convenient patient use. Advanced packaging technology is used to ensure the stability and safety of the liquid.

[0061] Mechanism of action of the Seven-Ingredient Lipid-Lowering Formula

[0062] Through in-depth scientific research, the multi-component, multi-target, and multi-pathway synergistic mechanism of the Seven-Ingredient Lipid-Lowering Formula in treating MASLD has been revealed:

[0063] Multi-component synergistic effect: Each traditional Chinese medicine in the Seven-Ingredient Lipid-Lowering Formula contains multiple active components that synergize with each other in the body to jointly regulate metabolic and immune functions. For example, hesperidin in pericarpium citri reticulatae has antioxidant and anti-inflammatory effects; quercetin in hawthorn can regulate lipid metabolism; and polysaccharides in polygonatum can improve insulin resistance.

[0064] Key target regulation: The Seven-Ingredient Lipid-Lowering Formula may exert therapeutic effects by regulating multiple key targets closely related to the pathogenesis of MASLD. These targets include:

[0065] PPARα and PPARγ: As important nuclear receptors regulating glucose and lipid metabolism and immune processes, PPARα and PPARγ play a crucial role in the pathogenesis of MASLD. The Seven-Ingredient Lipid-Lowering Formula may activate these receptors to improve glucose and lipid metabolism disorders and insulin resistance in MASLD patients.

[0066] CYP3A4: As a key enzyme in drug metabolism, CYP3A4 may play an important role in the treatment of MASLD. Some components in Qishijiangzhi Decoction may affect drug metabolism and efficacy by regulating the activity of CYP3A4.

[0067] AKT1, STAT3, SREBF1: Key molecules in these signaling pathways also play an important role in the pathogenesis of MASLD. Qishijiangzhi Decoction may affect the pathological process of MASLD by regulating the expression or activity of these molecules.

[0068] Pathway network regulation: The mechanism of action of Qishijiangzhi Decoction involves the regulation of multiple signaling pathways, forming a complex pathway network. These pathways include the PPAR signaling pathway, adipokine signaling pathway, and insulin signaling pathway, which interweave and influence each other, collectively regulating the pathological process of MASLD. Qishijiangzhi Decoction regulates these pathways to exert a comprehensive therapeutic effect on MASLD.

[0069] To verify the effectiveness and safety of Qishijiangzhi Decoction, a large number of experimental studies were conducted, including in vivo experiments, in vitro experiments, transcriptomic analysis, and gut microbiota analysis.

[0070] In vivo experiments (C57BL / 6J mouse model):

[0071] Experimental design: A high-fat high-sugar diet was used to induce MASLD mouse models, which were fed for 18 weeks. From the 14th week, mice were randomly divided into model group, Qishijiangzhi Decoction replacement low-dose group, Qishijiangzhi Decoction replacement medium-dose group, Qishijiangzhi Decoction replacement high-dose group, and positive control group (simvastatin), with daily gavage administration for 5 consecutive weeks.

[0072] Experimental results: Qishijiangzhi Decoction replacement at each dose significantly reduced the body weight, liver weight, spleen weight, and liver index of MASLD mice; significantly reduced lipid deposition in liver tissue, reduced TC, TG, and LDL-C content in serum, and increased HDL-C content; significantly reduced serum inflammatory factor levels (such as IL-6, TNF-α, and IL-1β); improved insulin resistance, and reduced HOMA-IR index.

[0073] In vitro experiments (HepG2 cell model):

[0074] Experimental design: A mixture of palmitic acid and oleic acid was used to induce HepG2 cells to establish a MASLD cell model. Different concentrations of Qishijiangzhi Decoction-containing drug serum groups were set up, as well as a positive control group (simvastatin) for intervention treatment.

[0075] Experimental results: QWZL can significantly reduce lipid accumulation and TC, TG content in cells; promote glucose uptake, improve insulin sensitivity; improve mitochondrial structure and function, increase mitochondrial DNA relative copy number, up-regulate mitochondrial-related gene expression; reduce ROS and MDA levels, increase SOD and GSH-Px activity, thereby reducing oxidative stress damage.

[0076] Transcriptomic analysis:

[0077] Experimental method: High-throughput sequencing technology was used to analyze the transcriptome of liver tissue of MASLD mice before and after QWZL intervention.

[0078] Experimental results: After QWZL intervention, there were 595 differentially expressed genes in the liver tissue of mice, of which 242 were up-regulated and 353 were down-regulated. These differentially expressed genes were significantly enriched in PPAR signaling pathway, adipocytokine signaling pathway, insulin signaling pathway and other pathways closely related to MASLD.

[0079] Intestinal flora analysis:

[0080] Experimental method: 16S rRNA sequencing technology was used to analyze the intestinal flora of MASLD mice before and after QWZL intervention.

[0081] Experimental results: After QWZL intervention, the richness and diversity of the intestinal flora of mice increased significantly, the relative abundance of beneficial bacteria (such as Bifidobacterium, Bacteroides) increased, and the relative abundance of potential pathogenic bacteria (such as Faecalibacterium, Erysipelothrix) decreased. Meanwhile, functional prediction analysis showed that QWZL may play a role in anti-MASLD by up-regulating anti-MASLD-related pathways (such as PPAR signaling pathway, peroxisome pathway), while inhibiting pro-MASLD-related pathways (such as fatty acid biosynthesis, glycerolipid and glycerophospholipid metabolism pathway).

[0082] The following provides three patent embodiments of QWZL (QWZL replacement), which respectively describe the preparation methods, experimental steps and experimental results of its oral liquid preparation, capsule preparation and tablet preparation. For ease of reading, the contents of the embodiments are appropriately integrated.

[0083] The oral liquid preparation of the Seven-Ingredient Lipid-Lowering Formula is prepared by the following steps: First, select the standard Chinese medicinal materials such as pericarpium citri reticulatae, hawthorn, lotus leaf, polygonatum, salvia miltiorrhiza, giant knotweed, and caper, mix them in proportion, and then soak them in a suitable amount of purified water. Next, use a decoction device to decoct three times, filter the dregs after each decoction, combine the filtrates, and concentrate them to obtain a concentrated medicinal liquid with a crude drug concentration of 13.8 g / ml. Then, dilute the concentrated medicinal liquid according to actual needs, add an appropriate amount of flavoring agents and preservatives, stir uniformly, and fill into sterile bottles to prepare the oral liquid preparation. Finally, the oral liquid is subjected to quality testing to ensure compliance with relevant quality standards. In order to verify the effectiveness and safety of the oral liquid preparation, in vivo and in vitro experiments are conducted. The in vivo experiment selects a MASLD mouse model induced by high-fat and high-sugar diet, observes and records the mouse body weight, liver weight, and other indicators, detects serum biochemical indicators and liver pathological changes. The experimental results show that the oral liquid preparation can significantly reduce the body weight, liver weight, and liver index of MASLD mice, improve lipid deposition and blood lipid levels, reduce inflammation factor levels, and improve insulin sensitivity. The in vitro experiment selects a HepG2 cell model and detects intracellular lipid deposition, TC and TG content, and other indicators. The experimental results show that the oral liquid preparation can significantly reduce intracellular lipid deposition, reduce TC and TG content, enhance glucose uptake capacity, reduce oxidative stress levels, and improve mitochondrial function.

[0084] The preparation method of the Seven-Ingredient Lipid-Lowering Formula capsule is similar to that of the oral liquid preparation, but there are differences in the preparation and deployment steps. Specifically, the dry extract obtained by decoction and concentration is pulverized into fine powder, mixed uniformly with an appropriate amount of auxiliary materials to form granules, and then filled into capsule shells to prepare the capsule preparation. Each capsule contains 0.5 g of dry extract, and the capsule is subjected to quality testing to ensure compliance with relevant quality standards. In order to verify the effectiveness and safety of the capsule preparation, in vivo and in vitro experiments are also conducted. The in vivo experiment results show that the capsule preparation can significantly reduce the body weight, liver weight, and liver index of MASLD mice, improve lipid deposition and blood lipid levels, reduce inflammation factor levels, and improve insulin sensitivity. The in vitro experiment results show that the capsule preparation can significantly reduce intracellular lipid deposition, reduce TC and TG content, enhance glucose uptake capacity, reduce oxidative stress levels, and improve mitochondrial function.

[0085] The preparation method of the tablet preparation of Qiwijiangzhi Decoction is similar to that of the oral liquid and capsule preparations, but the formulation and molding steps are different. Specifically, the dry extract obtained by decoction and concentration is vacuum dried and pulverized into fine powder, which is then mixed with an appropriate amount of excipients to form granules, which are then compressed into tablets. Each tablet contains 0.5 g of dry extract, and the tablets are subjected to quality testing to ensure compliance with relevant quality standards. To verify the effectiveness and safety of the tablet preparation, in vivo and in vitro experiments were also conducted. The in vivo experiment results showed that the tablet preparation significantly reduced the body weight, liver weight, and liver index of MASLD mice, improved lipid deposition and blood lipid levels, reduced inflammation factor levels, and improved insulin sensitivity. The in vitro experiment results showed that the tablet preparation significantly reduced intracellular lipid deposition, reduced TC and TG content, enhanced glucose uptake capacity, reduced oxidative stress levels, and improved mitochondrial function.

[0086] Pharmacodynamic evaluation of the composition for intervention of MASLD

[0087] 36 SPF male C57BL / 6J mice aged 6 weeks, weighing between 18-20 g, were provided by Chengdu Dashuo Experimental Animal Co., Ltd. (Animal Use License No. SYXK (Chuan) 2024-049). The mice were randomly divided into 6 cages, with 6 mice in each cage, and were raised at a temperature of 22-24°C, a relative humidity of 50%-60%, alternating light and dark illumination, and free access to food and water. Before the formal start of the experiment, the mice were adapted to the ordinary feed for one week. All procedures involving experimental animals in the study were carried out in accordance with the Guidelines for the Care and Use of Laboratory Animals (8th edition), and all animal experiments were approved by the Experimental Animal Ethics Committee of Chengdu University of Traditional Chinese Medicine, with approval number 2024174.

[0088] Pericarpium Citri Reticulatae (Batch No. 202108002, produced in Sichuan), Crataegus Pinnatifida (Batch No. 231101, produced in Shandong), Polygonatum sibiricum (Batch No. 23062701, produced in Guizhou), Salvia miltiorrhiza (Batch No. 202406005, produced in Shandong), Polygonum cuspidatum (Batch No. 230305, produced in Hubei), Artemisia capillaris (Batch No. 24022001, produced in Gansu), and Nelumbo nucifera (Batch No. 23031301, produced in Zhejiang) were all purchased from the Traditional Chinese Medicine Pharmacy of Gansu University of Traditional Chinese Medicine. The above-mentioned medicinal materials were decocted and concentrated according to the original prescription dosage to prepare the experimental drug.

[0089] The decoction method includes the following five steps: (1) Soak the six kinds of medicinal materials, namely green tangerine peel, hawthorn, polygonatum, salvia miltiorrhiza, polygonum cuspidatum and artemisia capillaris, in cold water for 60 minutes, and soak the lotus leaf separately for 5 minutes; (2) Boil the soaked medicinal materials over high heat until boiling, then reduce to low heat and keep it simmering for 30 minutes. After that, add the soaked lotus leaf and decoct for another 10 minutes; (3) Pour out the first decoction, add an appropriate amount of hot water and decoct for another 20 minutes; (4) Pour out the second decoction, add an appropriate amount of hot water and decoct for a third time for about 10 minutes; (5) Mix the above three decoctions and concentrate them using a rotary evaporator to finally obtain a concentrated decoction of the composition with a raw drug concentration of 13.8 g / ml.

[0090] Table 1 Main experimental consumables and reagents

[0091]

[0092] Table 2 Main Experimental Instruments

[0093]

[0094] Experimental methods

[0095] One cage of mice was randomly selected as the blank control group and fed with normal diet (product number: DS001, Chengdu Dashuo Experimental Animal Co., Ltd.) and distilled water. The other five cages of mice were fed with high-fat diet (product number: XTCD60, Jiangsu Xietong Pharmaceutical Biotechnology Co., Ltd.; energy ratio of fat:carbohydrate:protein = 60%:20%:20%) combined with high-sugar drinking water (concentration: 42 g / L; preparation method: dissolve 46.2 g fructose and 37.8 g sucrose in 1 L of distilled water and bring the volume to 2 L) for 18 weeks to induce the MASLD model.

[0096] The clinical equivalent dose of mice per day is 13 mg / g. In the experiment, 1 times (13 mg / g), 2 times (26 mg / g), and 4 times (52 mg / g) of the clinical equivalent dose of the composition were set as the low dose (Qiyi Jiangzhi Fang Substitution-L), the medium dose (Qiyi Jiangzhi Fang Substitution-M), and the high dose (Qiyi Jiangzhi Fang Substitution-H) of the composition, respectively. After the end of the 13th week, 5 cages of mice fed with high-fat feed combined with high-sugar drinking water were randomly divided into the Model group, the Qiyi Jiangzhi Fang Substitution-L group, the Qiyi Jiangzhi Fang Substitution-M group, the Qiyi Jiangzhi Fang Substitution-H group, and the XFTT group (positive control drug: simvastatin tablets, purchased from the pharmacy of the Affiliated Hospital of Gansu University of Chinese Medicine, batch number: H20084486). The daily clinical equivalent dose of simvastatin is 0.2 μg / g. From the 14th week, the mice in the Qiyi Jiangzhi Fang Substitution-L group, the Qiyi Jiangzhi Fang Substitution-M group, the Qiyi Jiangzhi Fang Substitution-H group, and the XFTT group were continuously administered by gavage according to the corresponding doses of each group for 5 days, while the Control group and the Model group were administered with the same volume of distilled water by gavage. During the entire experiment, the behavior states of food intake, water intake, activity, spirit, and hair of the mice in each group were observed and recorded daily, and the weights were recorded.

[0097] At the end of the 18th week (8 h of fasting without water after the last administration), the sampling operation was performed according to the following steps:

[0098] (1) After weighing and recording the mice, the mice were anesthetized by intraperitoneal injection of 0.3% sodium pentobarbital solution 0.1 ml / 10 g. (2) After blood sampling in the box, the mice were sacrificed by cervical dislocation, and the whole blood was centrifuged at 3000 r / min for 10 min after standing for 2 h, and the supernatant was taken and stored at -80°C for later use. (3) Epididymal white adipose tissue (eWAT) was taken, weighed, and recorded; the cecum was taken out, the intestinal contents were separated, and placed in a sterile centrifuge tube and stored at -80°C; the spleen was taken, weighed, and recorded; the liver was taken, weighed, and recorded; after taking a photo of the liver, about 1 cm * 1 cm tissue in the middle of the large lobe was taken and immersed in 4% paraformaldehyde solution, and the rest of the tissue was stored at -80°C; interscapular brown adipose tissue (iBAT) was taken, weighed, and recorded.

[0099] Liver histopathology and lipid content detection

[0100] HE and oil red O staining were used to observe liver histopathology and lipid deposition, Image J was used for quantitative analysis, and the TG and TC contents were determined by using a kit.

[0101] Serum biochemical index detection

[0102] The automatic biochemical analyzer was used to detect the levels of TC, TG, HDL-C, LDL-C, ALT and AST in the serum of mice. The blood glucose level of mice after fasting for 8 hours was measured by a blood glucose meter, and the levels of IL-6, TNF-α, IL-1β and insulin in the serum of mice were detected by ELISA kit; at the same time, the insulin resistance index (HOMA-IR) was calculated, and the formula was HOMA-IR = (fasting blood glucose x fasting insulin) / 22.5.

[0103] Statistical analysis

[0104] The online software BrightStat was used for statistical analysis, and the graphs were drawn by GraphPad Prism 10.0 software. Kruskal-Wallis H test was used to analyze the overall difference between multiple groups, and when the test results showed that there were differences between multiple groups, Conover test was further used for pairwise comparison analysis. P < 0.05 was considered to have statistical difference.

[0105] Results and conclusions

[0106] General situation of mouse model

[0107] During the experiment, the Control group mice were in good overall condition, with good hair color, free movement, very flexible reaction, normal feed and water consumption, and relatively stable feeding rhythm. The Model group mice had relatively greasy fur, significantly reduced daily activity, and even showed a tendency to lie down, with significantly increased body size. The XFTT group, Qishen Jiangzhi Decoction replacement-H group, Qishen Jiangzhi Decoction replacement-M group and Qishen Jiangzhi Decoction replacement-L group mice showed a state between the Model group and the Control group, i.e. good hair luster, free movement, relatively agile reaction, and significantly lower body size growth rate than the Model group. The general situation of typical mice in each group is shown in Figure 2 .

[0108] Effect of the composition on the body weight of mice

[0109] As shown in Figure 3 , high-fat feed combined with high-sugar drinking water can significantly increase the body weight of mice, while the composition and XFTT can significantly inhibit the increase of body weight of mice (see Figure 2 ).

[0110] Effect of the composition on the food and water intake of mice

[0111] Compared to the Control group, the Model group had a significantly reduced food intake; however, there was no significant difference in food intake among the Model group, the Seven-Ingredient Lipid-Lowering Formula Replacement-H group, the Seven-Ingredient Lipid-Lowering Formula Replacement-M group, the Seven-Ingredient Lipid-Lowering Formula Replacement-L group, and the XFTT group, as shown in the figure. Figure 4 As shown.

[0112] The Model group and the Seven-Ingredient Lipid-Lowering Formula Replacement-M group drank more water than the Control group. Meanwhile, there was no significant difference in water intake among the XFTT group, the Seven-Ingredient Lipid-Lowering Formula Replacement-H group, and the Seven-Ingredient Lipid-Lowering Formula Replacement-L group (see below). Figure 4 As shown.

[0113] This composition can reduce liver, spleen, and fat weight and liver index in model mice.

[0114] (1) Liver and spleen weight and liver index: This composition can reduce liver weight, reduce spleen weight, and lower liver index, such as Figure 5 As shown.

[0115] (2) Weight of brown and white fat: This composition can effectively reduce the iBAT and eWAT weights in MASLD model mice, such as Figure 6 As shown.

[0116] This composition improves the histopathological features of liver tissue in model mice.

[0117] (1) Pathological changes in liver tissue: The livers of mice in the Control group were red and uniform in texture, while the livers of mice in the Model group were significantly larger, with blunt edges and a more noticeable greasy feel. After intervention with this combination, the livers of mice were reduced to varying degrees, the sharpness of the edges was restored, and the greasy feel was reduced.

[0118] like Figure 7 As shown, HE staining indicates that the liver pathological changes in the MASLD mouse model were improved after intervention with this composition.

[0119] like Figure 8 As shown, Oil Red O staining results indicated that lipid droplets in hepatocytes of the MASLD mouse model were reduced to varying degrees after intervention with this composition; quantitative results showed that this composition could significantly reduce the Oil Red lipid content in the liver tissue of the Model group mice.

[0120] like Figure 9 As shown, (2) Lipid content in liver tissue: This composition can reduce the TC and TG content in liver tissue and shows an obvious dose-response relationship, that is, the higher the dose of this composition, the more obvious the effect.

[0121] like Figure 10As shown, (1) lipid metabolism indicators: serum TC, TG, HDL-C and LDL-C levels were measured. The results showed that the composition could improve the blood lipid levels of MASLD model mice.

[0122] like Figure 11 As shown, (2) glucose metabolism indicators: 8h fasting blood glucose, serum insulin level and HOMA-IR value were detected, indicating that the composition can improve the abnormal glucose metabolism level of MASLD model mice.

[0123] like Figure 12 As shown, (3) Liver enzyme indicators: ALT and AST levels were measured. After administration, the serum ALT and AST levels of mice decreased, but the difference in this combination was not statistically significant.

[0124] like Figure 13 As shown, (4) Inflammatory markers: Serum IL-6, TNF-α and IL-1β were detected, and the results showed that the composition could significantly improve the inflammatory response of the MASLD mouse model.

[0125] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. It will be apparent to those skilled in the art that the invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from its spirit or essential characteristics. Therefore, the embodiments should be considered illustrative and non-limiting in all respects, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, all variations falling within the meaning and scope of equivalents of the claims are intended to be included within the scope of the invention. No reference numerals in the claims should be construed as limiting the scope of the claims.

[0126] Furthermore, it should be understood that although this specification describes embodiments, not every embodiment contains only one independent technical solution. This narrative style is merely for clarity. Those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A traditional Chinese medicine composition for treating metabolic associated steatohepatitis (MASLD), characterized in that, The traditional Chinese medicine composition is made of raw medicinal materials in the following weight parts: Green tangerine peel: 3-15 parts; Giant knotweed: 2-10 parts; Hawthorn: 3-15 parts; Yellow ginseng: 3-15 parts; Chinese wormwood: 1-8 parts; Lotus leaf: 3-15 parts; Salvia miltiorrhiza: 2-10 parts.

2. The traditional Chinese medicine composition according to claim 1, wherein the preparation method comprises the following steps: Medicinal material preparation: weigh each kind of traditional Chinese medicine according to the above proportion, and ensure that the medicinal material quality meets the pharmacopoeia standard. Decoction: add medicinal materials into appropriate amount of water, soak for 30 minutes, then boil with strong fire, and then decoct with medium fire for 30-60 minutes, filter out medicinal liquid. Concentration: concentrate the filtered medicinal liquid to appropriate volume, or further make into pills, powders, granules, etc. according to needs. Preparation: according to clinical needs, concentrate the liquid or dry extract into appropriate dosage form, such as capsule, tablet, oral liquid, etc.

3. The traditional Chinese medicine composition according to claim 1 or 2, for use in the preparation of a drug for treating metabolic associated fatty liver disease.

4. Use according to claim 3, characterized in that, The traditional Chinese medicine composition can significantly improve the following pathological links of MASLD patients: Disorder of glycolipid metabolism; Inflammatory changes; Mitochondrial homeostasis imbalance; Oxidative stress.

5. Use according to claim 4, characterized in that, The traditional Chinese medicine composition plays a role in treating MASLD through the following mechanisms: Up-regulating the expression of PPARα, CYP3A4 and other genes Regulating lipid metabolism and inflammation-related pathways.

6. Use according to claim 3, characterized in that, When the traditional Chinese medicine composition is used in the preparation of a drug, it can be mixed with other drugs or excipients to make dosage forms suitable for clinical use.

7. Use according to claim 3, characterized in that, When the traditional Chinese medicine composition is used in the treatment of MASLD, it can be used alone or in combination with other treatment methods.

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