A hypoglycemic and hypolipidemic traditional Chinese medicine composition, a preparation method and application thereof

Abstract

The application relates to the technical field of traditional Chinese medicine preparation, and discloses a preparation method of a traditional Chinese medicine composition for reducing blood sugar and blood fat, which comprises the following steps: raw material preparation, selecting dried mature zanthoxylum bungeanum pericarps as main medicine, ensuring that the raw material is fresh and free of impurities, and preparing auxiliary material sodium carboxymethyl cellulose; zanthoxylum bungeanum crushing and sieving, crushing the dried zanthoxylum bungeanum pericarps by using a crusher to obtain fine and uniform zanthoxylum bungeanum powder; auxiliary material preparation, preparing a 0.3% sodium carboxymethyl cellulose solution as an excipient for mixing with the zanthoxylum bungeanum powder, so as to improve the formability and stability of the medicine; mixing and homogenization, mixing the sieved zanthoxylum bungeanum powder with the 0.3% sodium carboxymethyl cellulose solution under the condition of heating to 50-100 DEG C. The preparation method of the traditional Chinese medicine composition for reducing blood sugar and blood fat aims to prepare a traditional Chinese medicine composition containing zanthoxylum bungeanum, and the composition has a new use of reducing blood sugar and blood fat.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine preparation technology, specifically to a traditional Chinese medicine composition for lowering blood sugar and lipids, its preparation method, and its application. Background Technology

[0002] Non-alcoholic fatty liver disease (NAFLD) is a chronic liver disease caused by excessive fat deposition in the liver. It is a major cause of liver transplantation and hepatocellular carcinoma and has become one of the world's serious public health problems, with a high prevalence (25% to 45%) that is constantly increasing. However, there are currently no FDA-approved specific drugs for the clinical treatment of NAFLD. At the same time, there is a close relationship between non-alcoholic fatty liver and abnormal blood sugar. Severe non-alcoholic fatty liver may cause hyperglycemia because liver damage affects its normal uptake, storage and utilization of glucose. Meanwhile, abnormal blood sugar may also lead to increased fat deposition in the liver, thereby aggravating the condition of non-alcoholic fatty liver.

[0003] Although the applications of Sichuan pepper in daily life, clinical practice, and experiments have been widely reported, the effects and mechanisms by which Sichuan pepper improves NAFLD induced by a high-fat diet remain unclear. This severely limits the application of Sichuan pepper in improving NAFLD and related chronic metabolic diseases. Numerous preclinical models and clinical studies have confirmed that the occurrence of NAFLD is often accompanied by dysbiosis in the structure, composition, and metabolic function of the gut microbiota, which can induce abnormal lipid metabolism. For example, compared with healthy individuals, NAFLD patients have a higher proportion of Bacteroidetes and Firmicutes, lower gut microbiota diversity, and altered gut microbiota metabolites. Thus, regulating the structure of the gut microbiota can improve microbial and metabolic disorders associated with pathological states. Notably, appropriate dietary intake of Sichuan pepper can improve gut microbiota homeostasis. For instance, Sichuan pepper volatile oil can alleviate chronic unpredictable stress-induced anxiety in rats by restoring stress-induced gut microbiota imbalance. Furthermore, Sichuan pepper can reduce colitis in mice by increasing the levels of Lactobacillus and Bifidobacterium. Therefore, by studying the structure of the gut microbiota and its metabolite levels, the potential mechanisms by which Sichuan pepper improves NAFLD can be further revealed. Summary of the Invention

[0004] The purpose of this invention is to prepare a traditional Chinese medicine composition containing Sichuan pepper, which has new uses for lowering blood sugar and lipids. The invention proposes a traditional Chinese medicine composition for lowering blood sugar and lipids, its preparation method and application.

[0005] The technical solution of the present invention to solve the above-mentioned technical problems is as follows:

[0006] A method for preparing a traditional Chinese medicine composition for lowering blood sugar and lipids includes the following steps:

[0007] S10. Raw material preparation: Select dried and mature pepper peel as the main ingredient, ensuring that the raw materials are fresh and free of impurities, and prepare excipient sodium carboxymethyl cellulose.

[0008] S20. Grinding and sieving Sichuan peppercorns: The dried Sichuan peppercorn peels are ground using a grinder to obtain fine and uniform Sichuan peppercorn powder.

[0009] S30. Preparation of excipients: Prepare a 0.3% sodium carboxymethyl cellulose solution as an excipient to mix with Sichuan pepper powder to improve the shapeability and stability of the drug.

[0010] S40. Mixing and homogenization: The sieved pepper powder is mixed with a 0.3% sodium carboxymethyl cellulose solution under heating conditions of 50-100℃. Ultrasonic treatment is used to promote the thorough mixing of powder and solution by utilizing the cavitation effect and stirring action of ultrasound, ensuring the uniform distribution of drug components.

[0011] S50. Formulation: As needed, the uniformly mixed traditional Chinese medicine composition is formulated into drugs of different dosage forms.

[0012] Based on the above technical solution, the present invention can be further improved as follows.

[0013] Furthermore, in step S20, the pulverized pepper powder is sieved through a 200-mesh sieve to ensure uniform powder particle size.

[0014] Furthermore, the drug dosage form is one or more of the following: decoction, tea, granules, pills, powder, tablets, capsules, or oral liquid.

[0015] A traditional Chinese medicine composition for lowering blood sugar and lipids, which has the effects of lowering blood sugar and lipids.

[0016] The application of a traditional Chinese medicine composition for lowering blood sugar and lipids, used as a drug or health product for the prevention or treatment of diseases related to hyperglycemia and hyperlipidemia.

[0017] Furthermore, it is applicable to diseases related to high blood sugar and high blood lipids, including but not limited to coronary heart disease, metabolic syndrome, and diabetes.

[0018] Furthermore, it can be used in drugs, health products, foods, or food additives that lower blood sugar and blood lipids.

[0019] Compared with the prior art, the technical solution of this application has the following beneficial technical effects:

[0020] (1) The Chinese herbal composition containing Sichuan pepper can effectively alleviate NAFLD, such as: reducing mouse body weight and epididymal fat weight; improving serum and liver TC, TG, LDL-C and blood glucose levels; and reducing hepatocyte lipid droplet area.

[0021] (2) The biochemical indicators and pathological section results, compared with body weight and organ weight, further confirmed that the Chinese medicine composition containing Sichuan pepper can effectively alleviate fat accumulation.

[0022] (3) Liver transcriptome analysis showed that the Chinese herbal composition containing Sichuan pepper played a key role in improving NAFLD by inhibiting SREBP-1 and activating the PPARα / CPT-1α signaling pathway. In addition, the Chinese herbal composition containing Sichuan pepper could inhibit the expression of HMGCR (encoded by the Hmgcr gene) in NAFLD mice and regulate cholesterol synthesis in hepatocytes.

[0023] (4) After treatment with the Chinese herbal composition containing Sichuan pepper, the relative abundance of gut microbiota closely related to metabolic diseases was significantly alleviated. Correlation analysis between gut microbiota and biochemical pathological indicators and differential metabolic biomarkers revealed that the Chinese herbal composition containing Sichuan pepper could alleviate NAFLD by improving the gut microbiota and metabolic biomarkers in mice. Attached Figure Description

[0024] Figure 1 This invention provides for the identification of the chemical components of the traditional Chinese medicine composition containing Sichuan pepper.

[0025] Figure 2 The changes in body weight, liver fat, and epididymal fat in mice fed a high-fat diet after administration of the traditional Chinese medicine composition containing Sichuan pepper of the present invention were studied.

[0026] Figure 3 This invention relates to the role of a traditional Chinese medicine composition containing Sichuan pepper in improving hepatic steatosis induced by a high-fat diet.

[0027] Figure 4 The AUC of the traditional Chinese medicine composition containing Sichuan pepper of the present invention was obtained in the oral glucose tolerance test (OGTT) and calculated based on the glucose concentration after glucose loading.

[0028] Figure 5 For liver transcriptome RNA sequencing analysis.

[0029] Figure 6 This invention relates to the regulatory effect of the Sichuan pepper-containing traditional Chinese medicine composition on fatty acid decomposition and biosynthesis.

[0030] Figure 7 This invention relates to the effect of the traditional Chinese medicine composition containing Sichuan pepper on the regulation of intestinal microbial structure.

[0031] Figure 8 For fecal metabolomics analysis.

[0032] Figure 9 This study analyzes the correlation between gut microbiota and differential metabolic markers after treatment with the traditional Chinese medicine composition containing Sichuan pepper of this invention. Detailed Implementation

[0033] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0034] A method for preparing a traditional Chinese medicine composition for lowering blood sugar and lipids includes the following steps:

[0035] S10. Raw material preparation: Select dried and mature pepper peel as the main ingredient, ensuring that the raw materials are fresh and free of impurities, and prepare excipient sodium carboxymethyl cellulose.

[0036] S20. Grinding and sieving Sichuan peppercorns: The dried Sichuan peppercorn peels are ground using a grinder to obtain fine and uniform Sichuan peppercorn powder. In step S20, the ground Sichuan peppercorn powder is sieved through a 200-mesh sieve to ensure uniform powder particle size.

[0037] S30. Preparation of excipients: Prepare a 0.3% sodium carboxymethyl cellulose solution as an excipient to mix with Sichuan pepper powder to improve the shapeability and stability of the drug.

[0038] S40. Mixing and homogenization: The sieved pepper powder is mixed with a 0.3% sodium carboxymethyl cellulose solution under heating conditions of 50-100℃. Ultrasonic treatment is used to promote the thorough mixing of powder and solution by utilizing the cavitation effect and stirring action of ultrasound, ensuring the uniform distribution of drug components.

[0039] S50. Formulation: As needed, the uniformly mixed Chinese herbal medicine composition is formulated into different dosage forms of medicine, such as decoction, tea, granules, pills, powder, tablets, capsules, or oral liquid, or one or more of these dosage forms.

[0040] A traditional Chinese medicine composition for lowering blood sugar and lipids, which has the effects of lowering blood sugar and lipids.

[0041] The main chemical components of the traditional Chinese medicine composition containing Sichuan pepper also need to be identified using UHPLC-Q-Orbitrap HRMS.

[0042] This invention provides the application of the aforementioned traditional Chinese medicine composition in lowering blood sugar and lipids. The main research steps are as follows:

[0043] The lipid-lowering effect was evaluated by monitoring changes in body weight, liver weight, and epididymal fat weight in mice fed a high-fat diet after administration.

[0044] The lipid-lowering and hypoglycemic effects were evaluated by monitoring changes in serum cholesterol, triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), blood glucose, and hepatic triglyceride levels in mice fed a high-fat diet after drug administration, combined with pathological sections of liver and epididymal adipose tissue.

[0045] Liver transcriptome sequencing was analyzed to explore the potential molecular mechanism by which a traditional Chinese medicine composition containing Sichuan pepper improves NAFLD. Real-time PCR and immunofluorescence techniques were used to investigate the regulatory effects of the Sichuan pepper-containing traditional Chinese medicine composition on genes and corresponding proteins enriched in these pathways.

[0046] The effects of a traditional Chinese medicine composition containing Sichuan pepper on the gut microbiota composition of NAFLD mice were investigated using 16S rRNA sequencing of colon contents.

[0047] The mechanism by which the Chinese herbal composition containing Sichuan pepper improves NAFLD was further explored using non-targeted metabolomics methods to examine the microbial metabolomics profile.

[0048] Chemical composition identification of traditional Chinese medicine compositions containing Sichuan pepper

[0049] The Ultimate 3000™ system and the Q Exactive™ Plus system were analyzed using an ACQUITY UPLC HSS T3 column (2.1 mm × 100 mm, 1.8 μm). The column temperature was 35 °C. The autosampler temperature was 4 °C, and the injection volume was 5 μL. The mobile phase was acetonitrile (A) and 0.1% formic acid in water (B), with the following elution gradient: 0–12 min, 30%–85% A; 12–14 min, 85% A; 14–15 min, 85%–30% A; 15–17 min, 30% A.

[0050] like Figure 1 As shown, five chemical components were determined in the traditional Chinese medicine composition containing Sichuan pepper: Zanthoamide A, Hyperoside, Zanthoamides B, 3-Carene, and Hydroxy-α-sanshool.

[0051] Changes in body weight, liver fat, and epididymal fat in mice fed a high-fat diet after administration of a traditional Chinese medicine composition containing Sichuan pepper.

[0052] The methods used in animal research are as follows:

[0053] All male C57BL / 6J mice (6 weeks old) were purchased from Beijing Huafukang Biotechnology Co., Ltd. Mice were housed in a constant temperature (22±1℃) environment with a 12h / 12h light / dark cycle, and were fed standard food and water. All animal experiments were approved by the Ethics Committee of Traditional Chinese Medicine, Peking University.

[0054] After one week of acclimatization, mice were randomly divided into a control group (Group C, standard diet, n=6) and a high-fat diet (HFD) group. Five weeks later, the HFD group mice were randomly re-divided into a model group (Group M, n=6), a positive control berberine group (BBR group, n=6), and a group containing a traditional Chinese medicine composition of Sichuan pepper (ZBM group, n=6). The mice were then administered the medicine by gavage. Groups C and M were administered PBS (10 mL / kg bw / d), the BBR group was administered BBR (120 mg / kg bw / d), and the ZBM group was administered ZBM (1.5 g / kg bw / d; the raw medicinal materials were of equal weight). The dosage of BBR and ZBM was determined according to previous literature. Before the end of the experiment, mice in Group C were given a standard diet daily, while mice in Group M and the drug groups (BBR and ZBM groups) were given HFD daily. Mouse weight was recorded every other day. After 12 weeks, all mice were sacrificed, and the corresponding tissues were collected for subsequent experiments. A schematic diagram of the animal experimental design is shown below. Figure 2 As shown in Figure A.

[0055] The specific research results are shown below:

[0056] To demonstrate that ZBM has an ameliorative effect on NAFLD, we induced an NAFLD model in C57BL / 6J mice by feeding them with HFD. ZBM, a phytochemical, was used as a positive control because it is believed to differ from traditional statins in its cholesterol-lowering effects. Figure 2 B shows the weight gain curves for all mice, with group C mice showing a slight increase in weight throughout the experiment. However, the weight of group M mice was 1.22 times that of group C (p<0.05), and increased significantly starting from week 3. From week 5, mice were administered ZBM and BBR, respectively. As expected, the intervention with ZBM and BBR significantly alleviated HFD-induced weight gain by week 9 (p<0.05), indicating that ZBM and BBR have a good ameliorative effect on NAFLD. To further verify this claim, at week 12, representative experimental samples, including body weight, liver tissue, and epididymal fat, were collected, photographed, and weighed, as shown below. Figure 2As shown in C, compared with group C mice, group M mice showed significantly increased body weight, liver weight, and epididymal fat weight by 91.55%, 55.86%, and 191%, respectively (p<0.05). The body weight (31.10±0.61g vs. 35.16±0.32g, p<0.05), liver weight (1.15±0.12g vs. 1.44±0.16g, p<0.05), and epididymal fat weight (0.74±0.11g vs. 1.10±0.15g, p<0.05) of mice in group M were significantly lower than those in group M. The results indicate that ZBM can effectively inhibit HFD-induced increases in body weight, liver tissue, and epididymal fat in NAFLD mice.

[0057] The effects of traditional Chinese medicine compositions containing Sichuan pepper on improving hepatic steatosis and hyperglycemia induced by a high-fat diet.

[0058] The effect of a traditional Chinese medicine composition containing Sichuan pepper on improving lipid levels in mice fed a high-fat diet was verified using the following method:

[0059] Biochemical assays: Blood samples and liver tissue were analyzed using a biochemical analyzer and a biochemical reagent kit, respectively. Blood samples were centrifuged. Serum TC, TG, and LDL-C levels were measured using a 7180 fully automated biochemical analyzer (Hitachi, Tokyo, Japan). Frozen liver tissue was accurately weighed and recorded, and ethanol was added at a ratio of weight (g):volume (mL) of 1:9. The liver tissue was thoroughly homogenized in an ice bath using a cell homogenizer and an ultrasonic cell disruptor. These samples were centrifuged (2500 rpm, 10 min). The supernatant was separated and analyzed using a biochemical reagent kit.

[0060] Histological analysis: Mouse liver tissue was fixed, sectioned, and blocked, then stained with Oil Red O and HE. Epididymal adipose tissue was stained with HE after routine treatment. Histopathological changes were observed under a fluorescence inverted microscope.

[0061] For details, please see [link / details]. Figure 3 and Figure 4 .

[0062] The occurrence of NAFLD is often associated with abnormalities in blood lipids and other indicators. Mouse serum biochemical parameters include... Figure 3As shown in A, compared with group C, mice in group M exhibited abnormal lipid metabolism, including TC (4.79±0.57 mmol·L⁻¹ vs. 2.92±0.21 mmol·L⁻¹, p<0.05), triglycerides (TG, 1.56±0.14 mmol·L⁻¹ vs. 0.59±0.12 mmol·L⁻¹, p<0.05), and LDL-C (0.76±0.08 mmol·L⁻¹ vs. 0.43±0.06 mmol·L⁻¹, p<0.05). Interestingly, treatment with ZBM and BBR significantly improved hyperlipidemia in mice. In the ZBM group, TC, TG, and LDL-C decreased to 16.70%, 23.59%, and 40.19%, respectively; while in the BBR group, these figures were 44.05%, 17.11%, and 35.53%, respectively. Meanwhile, liver cholesterol and TG levels in untreated HFD mice were 1.11-fold and 1.18-fold higher than those in group C, respectively. Correspondingly, blood glucose levels in group M mice were higher than those in group C (p<0.05), and the ZBM and BBR intervention groups significantly regulated blood glucose levels. Figure 3 Images B show HE-stained and Oil Red O-stained liver tissue images, respectively. Compared to group C, group M mice showed more fat vacuoles and red lipid droplets in their livers, indicating that HFD successfully induced hepatic steatosis. Interestingly, no obvious lipid droplets were observed in liver pathological sections after ZBM and BBR intervention. These results clearly demonstrate that ZBM has a very significant effect on improving lipid deposition in mice. HE-stained images of epididymal adipose tissue showed that the epididymal fat in group C was round and uniform in size. Compared to group C, the adipocytes in group M were significantly hypertrophic, specifically manifested as increased cell diameter. ZBM and BBR significantly inhibited epididymal adipocyte hypertrophy. These biochemical indicators and pathological section results, compared with body weight and organ weight, further confirm that ZBM can effectively alleviate fat accumulation.

[0063] Oral glucose tolerance test (OGTT) results as follows Figure 4 As shown in Figure A, blood glucose levels in all groups rapidly increased within 30 minutes after administration, followed by a slow decline over 120 minutes. Notably, blood glucose levels in group M were higher than those in groups C and ZBM. Furthermore, compared to group M, the AUC (Average Value of Blood Glucose) calculated based on OGTT in mice in groups C and ZBM were significantly lower. Figure 4 B) Significantly reduced. The results indicate that ZBM can improve the elevated blood glucose levels caused by a high-sugar, high-fat diet.

[0064] Liver transcriptome analysis

[0065] Liver transcriptome sequencing analysis was performed to explore the potential molecular mechanisms by which ZBM improves NAFLD. The methods are as follows:

[0066] Total RNA was extracted from liver tissue; RNA purity was assessed using a NanoRhatometer@spectrophotometer (IMPLEN, Santa Clara, CA, USA); RNA integrity was evaluated using a Bioanalyzer 2100 system (Agilent Technologies, Santa Clara, CA, USA); sequencing libraries were prepared using the Illumina NEBNext Ultra™ RNA Library Preparation Kit (NEB) and further sequenced on an Illumina Novaseq 6000 platform (Illumina, San Diego, CA, USA). Each group underwent three biological replicates. Raw data were screened, sequenced, and differentially expressed genes were subjected to GO and KEGG functional annotation and pathway enrichment analysis.

[0067] For details, please see [link / details]. Figure 5 .

[0068] The hierarchical clustering trend of DEGs associated with the NAFLD phenotype was also consistent with previous biochemical and pathological indicators. Figure 5 A). Analysis results of GO and KEGG functional annotations ( Figure 5 (BC) indicates that HFD affects multiple molecular functions, cellular components, and biological processes in mice, such as metabolic and cellular processes. The identified shared DEGs mainly involve biological functions related to the endocrine system, transport and catabolism, signal transduction, lipid metabolism, energy metabolism, and carbohydrate metabolism. Bubble plots of GO and KEGG pathway enrichment show that DEGs are mainly enriched in pathways related to lipid metabolism disorders, such as those regulating lipid storage, fatty acid degradation, and the PPAR signaling pathway. Figure 5 DE). Simultaneously, through gene enrichment chord maps and gene expression-related network maps (… Figure 5 FG) can visually show the relationship between DEGs and lipid metabolism disorder-related pathways.

[0069] Verification of the regulatory effects of a traditional Chinese medicine composition containing Sichuan pepper on fatty acid decomposition and biosynthesis.

[0070] The regulatory role of ZBM in the enriched genes and corresponding proteins of these pathways was studied using real-time PCR and immunofluorescence techniques. The specific methods are as follows:

[0071] Real-time PCR analysis: After liver tissue was extracted, RNA was extracted, reverse transcribed, and amplified according to the kit instructions. The reverse transcription program was 50℃ for 15 min; the PCR amplification program was 95℃ for 5 min; 35×(95℃, 30 s; 55℃, 30 s; 72℃, 1 min). The relative expression levels of corresponding mRNAs were quantified using a 2-ΔΔCT method. The primer sequences used are listed in Table 1.

[0072] Immunofluorescence: Paraffin sections were dewaxed and rehydrated, antigen was recovered, serum was used for blocking, and the sections were incubated with primary antibody (PPARα, GB13148; HMGCR, GB13132, Servicebio), then with secondary antibody (CY3 conjugated with goat anti-rabbit; GB21303; Servicebio), and restained with DAPI (G1012; Servicebio). Fluorescence images were examined under a fluorescence microscope (Morrell, USA).

[0073] Table 1 Primer sequences used in the study

[0074]

[0075] For details, please see [link / details]. Figure 6 .

[0076] The regulatory role of ZBM in the enriched genes and corresponding proteins of these pathways was investigated using real-time PCR and immunofluorescence techniques. Figure 6 As shown in Figure A, a total of 6 DEGs involved in these pathways were identified, of which 4 genes were significantly upregulated by ZBM and 2 genes were significantly downregulated by ZBM. Specifically, the mRNA levels of Srebf1 (an important gene involved in de novo lipogenesis) and Hmgcr (a key enzyme in de novo cholesterol synthesis) were significantly induced by HFD and then significantly downregulated by ZBM. Figure 6 B and Figure 6 C). Immunofluorescence results consistently showed that the expression level of HMGCR in the liver of group M was significantly higher than that of group C. ZBM intervention significantly decreased HMGCR expression level, while BBR intervention did not significantly decrease it. Furthermore, real-time PCR results confirmed that the mRNA levels of Ppara and Cpt1a (regulator of fatty acid oxidation) were significantly decreased in mice fed HFD, while the mRNA levels of Ppara and Cpt1a were significantly increased after BBR and ZBM treatment. Figure 6 D and Figure 6 E). Immunofluorescence results also confirmed that ZBM significantly induced the expression of PPARα in NAFLD mice (E). Figure 6 F).

[0077] The effects of traditional Chinese medicine compositions containing Sichuan pepper on the regulation of gut microbiota structure

[0078] The effects of ZBM on the gut microbiota composition of NAFLD mice were investigated using 16S rRNA sequencing of colonic contents. The specific methods are as follows:

[0079] Bacterial genomes were extracted from mouse intestinal contents, and DNA concentration was determined. PCR amplification, purification, and quantification were performed. Pairing and sequencing were conducted using Illumina MiSeq. Raw fastq files were quality filtered using QIIME (version 1.17). Sequences with 97% similarity were classified as OTUs (Operational Taxonomic Units) using USEARCH software. Alpha diversity (Shannon and Simpson indices) and β diversity (principal coordinate analysis based on binary Jacques distance) were analyzed using QIIME.

[0080] For details, please see [link / details]. Figure 7 ;

[0081] α-diversity results ( Figure 7 AC analysis showed that the community richness and diversity in group M were significantly lower than those in group C (Shannon index: p = 0.00087; Simpson index: p = 0.0017), while the community richness and diversity in ZBM-treated mice were significantly higher than those in group M (Shannon index: p = 0.04594; Simpson index: p = 0.07144), indicating that ZBM can restore the gut microbiota diversity of HFD mice. Next, we investigated the β-diversity of the microbial communities in each group using principal coordinate analysis (PCoA). Figure 7 As shown in Figure C, the microbial communities of each group exhibited clear separation and spatial clustering. The ZBM and C groups were significantly farther from the M group, indicating substantial differences between the M group's microbial community and the ZBM and C groups. This also suggests that the composition of the gut microbiome underwent significant changes during HFD feeding and ZBM intervention. Correspondingly, the community composition and species abundance of each sample were analyzed at different taxonomic levels. At the phylum level, compared to the C group, the relative abundance of Bacteroidetes was significantly decreased and the relative abundance of Actinobacteria was significantly increased in the M group. After ZBM intervention, the relative abundance of Bacteroidetes was slightly increased compared to the M group, while the relative abundance of Actinobacteria was significantly decreased. Furthermore, the genus-level microbial community structure was further analyzed. Figure 7 D). As Figure 7 As shown in Figure E, compared with group C, the relative abundance of harmful bacteria Coriobacteriaceae_UCG-002, Ileibacterium, and Romboutsia in group M showed an increasing trend, while the relative abundance of Lachnospiraceae_NK4A136_group and Desulfovibrio was lower. After ZBM treatment, the relative abundance of these bacteria closely related to metabolic diseases caused by HFD was significantly reduced, indicating that the relative abundance of bacteria in the ZBM group showed a trend of gradually recovering towards that of group C.

[0082] Including fecal metabolomics and related analysis

[0083] Mouse fecal samples were removed from the freezer (-80℃) and thawed at 4℃. The feces were weighed in 2mL centrifuge tubes; 200μL of ultrapure water was added; the samples were vortexed and sonicated for 3 min each; 400μL of dichloromethane was added; and the supernatant was collected by centrifugation. After removing the supernatant, 400μL of methanol-water (V:V = 80:20) was added to the precipitate and centrifuged. The combined supernatants were placed in centrifuge tubes, dried under nitrogen, and then dissolved in methanol. The supernatant was collected and analyzed using an Ultimate 3000™ system and a q Exactive™ plus system. The column temperature was 35℃. The autosampler temperature was 4℃, and the injection volume was 5μL. The mobile phase was acetonitrile (A)-0.1% formic acid water (B), and the elution gradient was: 0-12 min, 30%-85% A; 12-14 min, 85% A; 14-15 min, 85%-30% A; 15-17 min, 30% A.

[0084] Data preprocessing and statistical analysis: Raw LC-MS data were preprocessed using MarkerLynx (Waters, MA, USA). MetaboAnalyst 5.0 was used for data filtering, missing value estimation, trade-offs, and data normalization. Data matrices were imported into SIMCA-P 14.1 (Umetrics, ume, Sweden) software for multivariate statistical analysis (PCA and OPLS-DA). Potential biomarkers were identified using p-values ​​(p<0.05), VIP (VIP>1), and trade-offs (FC, FC>1 or <0.5). Based on the identified biomarkers, pathway analysis was performed using MetaboAnalyst 5.0.

[0085] For details, please see [link / details]. Figure 8-9 .

[0086] Fecal metabolomics experiments further revealed the metabolism of the gut microbiota. It has been reported that dyssynergistic biosynthesis of valine, leucine, and isoleucine may lead to oxidative stress in NAFLD; coenzyme A (CoA) overexpression can cause insulin resistance. The results showed that ZBM affects the biosynthesis of valine, leucine, and isoleucine, as well as pantothenic acid and coenzyme A in NAFLD mice. Compared with the M group, ZBM reduced L-isoleucine and coenzyme A levels, which may help prevent oxidative stress damage and insulin resistance. Oxidative stress and insulin resistance are also important causes of NAFLD. Furthermore, ZBM regulates niacin and nicotinamide metabolism, as well as pyruvate metabolism. Nicotinamide and niacin are involved in the synthesis and catabolism of carbohydrates, lipids, and proteins. Compared with HFD-induced metabolic diseases, niacin abundance increased after ZBM intervention. Pyruvate is involved in multiple metabolic pathways (protein, carbohydrate, and lipid metabolism) for bioenergy production. Notably, ZBM increased pyruvate levels in HFD-fed mice, which may promote lipid metabolism. Furthermore, HFD-induced mice exhibited higher blood glucose levels, indicating increased gluconeogenesis and decreased glycolysis in HFD mice. The study found that ZBM can improve blood glucose levels in an HFD-induced NAFLD model and affect glycolysis / gluconeogenesis.

[0087] Statistical analysis

[0088] Data are expressed as mean ± standard deviation and statistical analysis was performed using Prism 7.0 (GraphPad, La Jolla, CA, USA). Independent samples t-tests were used for comparisons between two groups. One-way ANOVA with Tukey's post-hoc test was used for comparisons among multiple groups. A p-value < 0.05 was considered statistically significant.

[0089] The application of a traditional Chinese medicine composition for lowering blood sugar and lipids, used in drugs or health products for the prevention or treatment of diseases related to hyperglycemia and hyperlipidemia, suitable for diseases related to hyperglycemia and hyperlipidemia including but not limited to coronary heart disease, metabolic syndrome, diabetes, etc., and used in drugs, health products, foods or food additives for lowering blood sugar and lipids.

[0090] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0091] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A method for preparing a traditional Chinese medicine composition for lowering blood sugar and lipids, characterized in that, Includes the following steps: S10. Raw material preparation: Select dried and mature pepper peel as the main ingredient, ensuring that the raw materials are fresh and free of impurities, and prepare excipient sodium carboxymethyl cellulose. S20. Grinding and sieving Sichuan peppercorns: The dried Sichuan peppercorn peels are ground using a grinder to obtain fine and uniform Sichuan peppercorn powder. S30. Preparation of excipients: Prepare a 0.3% sodium carboxymethyl cellulose solution as an excipient to mix with Sichuan pepper powder to improve the shapeability and stability of the drug. S40. Mixing and homogenization: The sieved pepper powder is mixed with a 0.3% sodium carboxymethyl cellulose solution under heating conditions of 50-100℃. Ultrasonic treatment is used to promote the thorough mixing of powder and solution by utilizing the cavitation effect and stirring action of ultrasound, ensuring the uniform distribution of drug components. S50. Formulation: As needed, the uniformly mixed traditional Chinese medicine composition is formulated into drugs of different dosage forms.

2. The method for preparing a traditional Chinese medicine composition for lowering blood sugar and lipids according to claim 1, characterized in that, In step S20, the pulverized pepper powder is sieved through a 200-mesh sieve to ensure uniform particle size.

3. The method for preparing a traditional Chinese medicine composition for lowering blood sugar and lipids according to claim 1, characterized in that, The drug dosage form is one or more of the following: decoction, tea, granules, pills, powder, tablets, capsules, or oral liquid.

4. The traditional Chinese medicine composition prepared by the method according to claim 1, characterized in that, This composition has the effects of lowering blood sugar and lowering blood lipids.

5. The application of a traditional Chinese medicine composition as described in any one of claims 1 to 3, characterized in that, Drugs or health products used to prevent or treat diseases related to high blood sugar and high blood lipids.

6. The application of the traditional Chinese medicine composition for lowering blood sugar and lipids as described in claim 5, characterized in that, It is suitable for diseases related to high blood sugar and high blood lipids, including but not limited to coronary heart disease, metabolic syndrome, and diabetes.

7. The application of the traditional Chinese medicine composition for lowering blood sugar and lipids as described in claim 5, characterized in that, Used in drugs, health products, foods, or food additives that lower blood sugar and blood lipids.

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