Use of a traditional Chinese medicine composition in preparation of a preparation for promoting growth of butyrate-producing bacteria and increasing butyrate yield

Abstract

The application belongs to the technical field of biology, and discloses application of a traditional Chinese medicine composition in preparation of a preparation for promoting growth of butyrate-producing bacteria and increasing butyrate yield, wherein the traditional Chinese medicine composition is composed of the following raw medicinal materials in parts by weight: 18-25 parts of angelica sinensis, 12-18 parts of persicae semen, 12-18 parts of safflower, 25-32 parts of rehmanniae radix praeparata, 20-25 parts of paeoniae rubra radix, 12-18 parts of chuanxiong rhizoma, 32-48 parts of astragali radix, 25-32 parts of epimedii herba and 8-12 parts of cinnamomi cortex. The experiment proves that the culture medium prepared by using the traditional Chinese medicine composition can directly promote growth of butyrate-producing bacteria and increase butyrate yield.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, specifically relating to the application of a traditional Chinese medicine composition in the preparation of formulations that promote the growth of butyrate-producing bacteria and increase butyrate production. Background Technology

[0002] Traditional Chinese medicine formulas are composed of various bioactive phytochemicals, which often exert systemic therapeutic effects through interactions with the gut microbiota. Increasing research indicates that regulating the gut microbiota is a key mechanism by which herbal preparations exert their therapeutic efficacy. Modified Taohong Siwu Decoction (MTHSWD) is developed based on Taohong Siwu Decoction, which first appeared in the Qing Dynasty medical classic *Jinkui Yaolue*. Taohong Siwu Decoction is a classic Chinese medicine formula traditionally used in TCM to invigorate blood circulation and remove blood stasis, thus having a therapeutic effect on coronary heart disease. Clinical practice and previous studies have confirmed that the modified Taohong Siwu Decoction formula, obtained by adding or subtracting specific herbs, significantly enhances efficacy compared to using Taohong Siwu Decoction alone.

[0003] This invention has shown through experiments that MTHSWD can selectively enrich bacteria that produce short-chain fatty acids (SCFAs), restore the microbial butyrate synthesis capacity, and increase the level of circulating butyrate. Therefore, this invention proposes the application of a traditional Chinese medicine composition in the preparation of formulations that promote the growth of butyrate-producing bacteria and increase butyrate production. Summary of the Invention

[0004] This invention aims to provide an application of a traditional Chinese medicine composition in the preparation of formulations that promote the growth of butyrate-producing bacteria and increase butyrate production. In vitro experiments have confirmed that butyrate can significantly inhibit the expression of inflammatory factors and reduce inflammatory responses. It can be used to prepare pharmaceutical compositions, foods, health products or cosmetics for the treatment or prevention of inflammation-related diseases.

[0005] To achieve the above objectives, the technical solution of the present invention is as follows:

[0006] In a first aspect, the present invention provides an application of a traditional Chinese medicine composition in the preparation of an agent that promotes the growth of butyrate-producing bacteria and increases butyrate production. The traditional Chinese medicine composition is characterized by comprising the following raw medicinal materials in parts by weight: Angelica sinensis 18-25 parts, Prunus persica 12-18 parts, Carthamus tinctorius 12-18 parts, Rehmannia glutinosa 25-32 parts, Paeonia lactiflora 20-25 parts, Ligusticum chuanxiong 12-18 parts, Astragalus membranaceus 32-48 parts, Epimedium brevicornu 25-32 parts, and Cinnamomum cassia 8-12 parts.

[0007] Preferably, the traditional Chinese medicine composition consists of the following raw medicinal materials in parts by weight: 22 parts Angelica sinensis, 16 parts Prunus persica, 16 parts Carthamus tinctorius, 27 parts Rehmannia glutinosa, 22 parts Paeonia lactiflora, 16 parts Ligusticum chuanxiong, 40 parts Astragalus membranaceus, 27 parts Epimedium brevicornu, and 10 parts Cinnamomum cassia.

[0008] Preferably, the preparation is a culture medium composed of an extract of traditional Chinese medicine composition, phosphate buffer, and enhanced Clostridium culture medium.

[0009] Preferably, the amount of the herbal composition extract is as follows: Angelica sinensis 18-25g, Prunus persica 12-18g, Carthamus tinctorius 12-18g, Rehmannia glutinosa 25-32g, Paeonia lactiflora 20-25g, Ligusticum chuanxiong 12-18g, Astragalus membranaceus 32-48g, Epimedium brevicornu 25-32g, and Cinnamomum cassia 8-12g. The herbal composition extract is diluted with phosphate buffer as needed, and the volume ratio of the diluted herbal composition extract to the enhanced Clostridium difficile culture medium is 1:7 to 1:9.

[0010] Preferably, the amount of the herbal composition extract is 22g of Angelica sinensis, 16g of peach kernel, 16g of safflower, 27g of Rehmannia glutinosa, 22g of Paeonia lactiflora, 16g of Ligusticum chuanxiong, 40g of Astragalus membranaceus, 27g of Epimedium brevicornu, and 10g of cinnamon. The herbal composition extract is diluted with phosphate buffer as needed, and the volume ratio of the diluted herbal composition extract to the enhanced Clostridium difficile culture medium is 1:9.

[0011] Preferably, the method for preparing the extract of the traditional Chinese medicine composition includes: mixing all the medicinal materials in the traditional Chinese medicine composition, soaking them in water, extracting twice, filtering, combining the two filtrates, and concentrating to obtain the extract of the traditional Chinese medicine composition.

[0012] Preferably, the amount of water added is 1L.

[0013] Preferably, the soaking time is 25-45 minutes.

[0014] Preferably, the extraction time is 20-45 minutes.

[0015] Preferably, the filtration method is to filter through four layers of gauze.

[0016] Preferably, the concentration is achieved by evaporation and concentration in a rotary evaporator at 60°C to 70°C.

[0017] In a first aspect, the present invention provides a method for promoting the growth of butyrate-producing bacteria and increasing butyrate production. The method comprises: culturing the bacteria in a culture medium containing an extract of a traditional Chinese medicine composition, wherein the extract is prepared from the following raw medicinal materials in parts by weight: Angelica sinensis 18-25 parts, Prunus persica 12-18 parts, Carthamus tinctorius 12-18 parts, Rehmannia glutinosa 25-32 parts, Paeonia lactiflora 20-25 parts, Ligusticum chuanxiong 12-18 parts, Astragalus membranaceus 32-48 parts, Epimedium brevicornu 25-32 parts, and Cinnamomum cassia 8-12 parts.

[0018] Preferably, the method is as follows: culturing with a culture medium containing an extract of a traditional Chinese medicine composition, wherein the extract of the traditional Chinese medicine composition is prepared from the following raw medicinal materials in parts by weight: 22 parts Angelica sinensis, 16 parts Prunus persica, 16 parts Carthamus tinctorius, 27 parts Rehmannia glutinosa, 22 parts Paeonia lactiflora, 16 parts Ligusticum chuanxiong, 40 parts Astragalus membranaceus, 27 parts Epimedium brevicornu, and 10 parts Cinnamomum cassia.

[0019] Compared with existing technologies, the beneficial effects of this solution are:

[0020] 1. This study reveals for the first time the targeted regulatory effect of MTHSWD (i.e., the traditional Chinese medicine composition described in this invention) on intestinal butyrate-producing bacteria.

[0021] Taohong Siwu Decoction is known to be used for promoting blood circulation and removing blood stasis, but this invention is the first to confirm through metagenomic sequencing that its modified formula (with the addition of Astragalus membranaceus, Epimedium, and Cinnamomum cassia) can selectively enrich SCFA-producing gut microbiota (such as Trichophytonceae and Bacteroides), establishing a new association between the traditional "promoting blood circulation and removing blood stasis" effect and the modern "gut-heart axis" microecological mechanism.

[0022] 2. Restoring the butyrate metabolism pathway at the gene function level

[0023] This invention not only observes changes in gut microbiota abundance, but also demonstrates through KEGG pathway analysis that Jiawei Taohong Siwu Decoction can significantly restore the butyrate metabolism pathway damaged after myocardial infarction and upregulate the gene expression of key enzymes in butyrate biosynthesis, thus confirming its potential to restore butyrate synthesis in gut microbiota at the molecular functional level.

[0024] 3. Targeted metabolomics confirmed that butyrate was the most significant response metabolite.

[0025] Through quantitative analysis of targeted metabolomics by LC-MS, this invention clearly points out for the first time that after intervention with Jiawei Taohong Siwu Decoction, the recovery of butyrate levels in feces and serum is far greater than that of other short-chain fatty acids such as acetic acid and propionic acid, thus accurately identifying butyrate as its core pharmacodynamic biomarker.

[0026] 4. In vitro experiments provide evidence of causality.

[0027] Through anaerobic fecal culture and pure culture experiments of representative butyric acid-producing bacteria such as *F. prausnitzii*, this invention eliminates in vivo interference and directly proves that the extract of this formula can promote the proliferation of butyric acid-producing bacteria in a dose-dependent manner and directly increase the butyric acid yield in the culture supernatant, achieving an upgrade of evidence from "correlation" to "causation".

[0028] 5. Provide clear and industrially feasible preparation and application solutions.

[0029] This invention provides a precise weight ratio of modified Taohong Siwu Decoction (MTHSWD) (Angelica sinensis 22 parts, Prunus persica 16 parts, Carthamus tinctorius 16 parts, Rehmannia glutinosa 27 parts, Paeonia lactiflora 22 parts, Ligusticum chuanxiong 16 parts, Astragalus membranaceus 40 parts, Epimedium brevicornu 27 parts, Cinnamomum cassia 10 parts), and specifies the volume ratio (1:9) of the extract to the enhanced Clostridium difficile culture medium and the specific extraction process parameters, providing a clear industrial implementation path for its application in the preparation of prebiotic preparations, functional foods or pharmaceuticals that promote butyrate production. Attached Figure Description

[0030] Figure 1 Alpha diversity analysis based on Ace, Chao, Shannon and Simpson indices (n=4);

[0031] Figure 2 Results of principal coordinate analysis (PCoA) based on intergroup species-level Bray-Curtis distance (n=4);

[0032] Figure 3 LEfSe analysis showed differentially enriched microbial taxa among the groups (n=4).

[0033] Figure 4 Linear discriminant analysis (LDA) scores for key microbial taxa (n=4);

[0034] Figure 5 The results of intergroup comparisons of the abundance of major genera and species highlight the significant enrichment of SCFA-producing bacteria after MTHSWD treatment (n=4).

[0035] Figure 6 KEGG pathway analysis for butyrate and pyruvate metabolism during butyrate and propionic acid formation (n=4)

[0036] Figure 7 For the comparison of butyrate metabolism among the three groups (n=4);

[0037] Figure 8 Comparison of the relative expression levels of key enzyme genes in the butyrate biosynthesis pathway (n=4).

[0038] Figure 9 Heatmaps and bar charts showing the content of major short-chain fatty acids (acetic acid, propionic acid, butyric acid, and others) in feces of each group (n=4);

[0039] Figure 10 Heatmaps and bar charts showing the content of major short-chain fatty acids in serum of each group (n=4).

[0040] Figure 11MTHSD can directly promote the growth of fecal bacteria in mice (A is the asepticity of the culture medium verified by the OD600 growth curve; B is the OD600 growth curve of total fecal bacteria after MTHSD treatment (n=3)).

[0041] Figure 12 The results of qPCR analysis of representative SCFA-producing bacteria (Clostridium plasminogen var. spp., Lactobacillus spp., Lactobacillus murineis, Bifidobacterium) (n=3);

[0042] Figure 13 To quantify butyric acid production in fecal bacterial culture supernatant using ELISA (n=3);

[0043] Figure 14 OD600 growth curves of Clostridium plasminogen lysate after MTHSWD treatment (n=3). Detailed Implementation

[0044] To facilitate understanding of the present invention by those skilled in the art, the technical solution of the present invention will be further described in detail below with reference to embodiments and accompanying drawings. Obviously, the described embodiments are merely some, not all, of the embodiments of the present invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort should fall within the scope of protection of the present invention.

[0045] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0046] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0047] The materials and methods involved in the following embodiments are as follows:

[0048] 1. Animals

[0049] All male C57BL / 6J mice (7 weeks old, weighing 20±2 g) were purchased from Shanghai Jihui Laboratory Animal Breeding Co., Ltd. (Shanghai, China) and housed at the Laboratory Animal Center of Shanghai University of Traditional Chinese Medicine (Shanghai, China). Mice were housed in a controlled environment (temperature 23±2°C, humidity 50%-70%), maintaining a 12-hour light / dark cycle, and had free access to standard feed and water. All animal experiments were approved by the Animal Ethics Committee of Shanghai University of Traditional Chinese Medicine (Approval No.: PZSHUTCM2308010001).

[0050] 2. Mouse model of myocardial infarction (MI)

[0051] After a one-week acclimatization period, 8-week-old mice underwent left anterior descending (LAD) coronary artery ligation to induce myocardial infarction. Mice were anesthetized using mechanical ventilation with a 2% isoflurane and oxygen mixture at a flow rate of 0.4 L / min. After iodine disinfection, the third to fourth intercostal spaces of each mouse were opened via blunt thoracotomy to expose the heart. The LAD coronary artery was then ligated approximately 2 mm below the pulmonary conus using a 6-0 suture needle, and the heart was repositioned. After repositioning the heart and compressing the pleural cavity to expel air, the pleural cavity and skin were sutured using a 4-0 suture needle. Mice in the sham-operated group underwent the same procedure but without arterial ligation.

[0052] 3. Preparation of MTHSWD extract

[0053] MTHSWD is composed of the following raw medicinal materials in parts by weight: Angelica sinensis 22 parts, peach kernel 16 parts, safflower 16 parts, prepared rehmannia root 27 parts, red peony root 22 parts, chuanxiong rhizome 16 parts, astragalus root 40 parts, epimedium 27 parts, and cinnamon 10 parts.

[0054] The crude extract of MTHSWD was provided by Shuguang Hospital affiliated to Shanghai University of Traditional Chinese Medicine (Shanghai, China). The preparation method of MTHSWD is as follows: The raw medicinal materials were mixed with distilled water (mass-volume ratio of 196g:1L, 1 part of MTHSWD raw medicinal materials = 1g) and soaked for 30 minutes to obtain a mixture. The mixture was extracted with boiling water for 30 minutes and then filtered through 4 layers of gauze. The filtrate was collected. The residue was extracted with boiling water for 20 minutes and then filtered through 4 layers of gauze. The two filtrates were combined and evaporated using a rotary evaporator at 60°C. The concentrated volume was 360mL.

[0055] The daily dosage in mice was calculated based on the clinically used human equivalent dose, with the ratio of MTHSWD-L, MTHSWD-M, and MTHSWD-H being 1:2:4 (455 mg / kg, 910 mg / kg, and 1820 mg / kg daily, respectively, the dosage corresponding to the weight of the raw materials). The captopril group received 5 mg / kg captopril (Shanghai Pharmaceutical, China) daily. Mice were administered the drug via gavage daily starting on the third day after LAD ligation for 4 weeks.

[0056] 4. Metagenomic sequencing

[0057] Four weeks after oral administration of the drug to mice, fresh fecal samples were collected from the sham-operated group, MI group, and MTHSWD group (n=4 mice per group). Total genomic DNA was extracted using the FastPure fecal DNA extraction kit (magnetic bead method) (Majorbio Biotech, Shanghai, China). The concentration and purity of the extracted DNA were determined using Synergy HTX and NanoDrop2000, respectively. The DNA was fragmented to an average size of approximately 350 bp using a Covaris M220 (Gene Company Limited, China), and paired-end sequencing libraries were constructed using the nextflex Rapid DNA-Seq system (Bioo Scientific, Austin, USA). Paired-end sequencing was performed by Majorbio Biotech, and data analysis was conducted on the Majorbio cloud platform (https: / / cloud.majorbio.com / ). The metagenomic sequencing data has been stored in the NCBI Short Read Database.

[0058] 5. Targeted metabolomics analysis

[0059] Short-chain fatty acids (SCFAs) in serum and fecal samples were quantified using targeted metabolomics analysis. Samples were processed according to standard procedures and analyzed by liquid chromatography-mass spectrometry (LC-MS) according to established protocols. LC / MS analysis of samples was performed on an ExionLC AD system (equipped with a QTRAP® 6500+ mass spectrometer, Sciex Corporation, USA) used by Majorbio Biotech. Data were analyzed using Sciex software. All ion fragments were automatically identified and integrated using default parameters, and all integration results were manually verified. Metabolite concentrations were calculated based on linear regression standard curves.

[0060] Example 1: MTHSWD exerts its effects by regulating the gut microbiota.

[0061] Given the inherent interaction between traditional Chinese medicine formulas and the gut microbiota, we further conducted metagenomic sequencing research to explore whether MTHSWD exerts its function by regulating the gut microbiota.

[0062] In this embodiment, MI model mice were first randomly divided into two groups: the MTHSWD group (denoted as MTHSWD-MI) and the model group (denoted as MI). Mice in the MTHSWD group received MTHSWD (1820 mg / kg) intervention, while mice in the model group and the sham-operated group received no intervention. Four weeks after the mice were administered the medication via gastric administration, fresh fecal samples were collected from the sham-operated group, the MI group, and the MTHSWD group (n=4 mice in each group) for metagenomic sequencing.

[0063] After intervention, compared with the sham-operated group, the gut microbiota α-diversity indices (Ace and Chao indices representing community richness, and Shannon and Simpson indices representing community diversity) in the MTHSWD group mice all showed a decreasing trend in community richness. Although there were no significant differences in gut microbiota α-diversity indices among the sham-operated group, the model group, and the MTHSWD group, the α-diversity level in the MTHSWD treatment group showed a trend of recovering to the level of the sham-operated group compared with the model group. Figure 1 Notably, principal coordinate analysis (PCoA) revealed significant clustering differences among the three groups, indicating substantial differences in microbial community structure and species abundance. Figure 2 ).

[0064] Linear discriminant analysis of effect size (LEfSe) and LDA analysis confirmed that MTHSWD can increase the abundance of Lachnospiraceae, Bacteroides, Muribactae, and Lawsonibacter, which are known to produce short-chain fatty acids (SCFAs). Figure 3 and Figure 4 The fold changes in abundance of selected genera and species among the groups are shown in the figure. Figure 5 In summary, these results indicate that MTHSWD can regulate the gut microbiota of MI mice and increase the abundance of SCFA-producing bacteria.

[0065] Example 2: MTHSWD can enhance butyrate metabolism and butyrate production after MIG.

[0066] Given that microbiome analysis showed a significant enrichment of SCFA enzyme-producing bacteria after MTHSWD treatment, this example further analyzed the KEGG pathway for SCFA metabolism (including butyrate metabolism and pyruvate metabolism).

[0067] The results showed that only butyrate metabolism differed significantly among the three groups: the MI group had lower butyrate metabolism levels, while the MTHSWD group returned to normal levels. Figure 6 and Figure 7In-depth analysis of the butyrate metabolic pathway revealed that MTHSWD significantly enhanced the activity of key enzymes in the butyrate biosynthesis pathway. Figure 8 Therefore, we hypothesize that MTHSWD may promote microbial butyrate production.

[0068] To verify this hypothesis, this embodiment used targeted metabolomics technology to detect the levels of short-chain fatty acids (SCFAs) in feces and serum after MTHSWD treatment. Among the various SCFAs, butyrate showed the most significant change after MTHSWD treatment: compared with the sham-operated group, the butyrate level in MI mice was significantly reduced, indicating that butyrate production function was impaired after MI; notably, MTHSWD administration could significantly restore butyrate levels, with a recovery magnitude exceeding that of other SCFAs ( Figure 9 and Figure 10 ).

[0069] In summary, these results indicate that butyrate is the most significant microbial metabolite responding to MTHSWD intervention.

[0070] Example 3: MTHSWD directly promotes the growth of SCFA-producing bacteria and enhances in vitro butyrate production.

[0071] To further verify the causal relationship between MTHSWD and microbial metabolic function, this example further investigated whether MTHSWD directly promotes the production of SCFA-producing bacteria and butyrate in vitro. Five g of fresh feces from conventional mice were collected and thoroughly homogenized with 40 mL of PBS. The homogenate was centrifuged at 800 g for 3 min to collect the supernatant, then centrifuged at 12,000 g for 15 min to obtain the precipitate, which was resuspended in PBS. This step was repeated three times. The final precipitate was mixed with brain and heart infusion medium (Solepro, Beijing, China) and diluted to a ΔOD600 value less than 0.1 (ΔOD600 = OD600 of bacterial solution - OD600 of sterile solution). MTHSWD (50, 100, and 200 mg / mL, corresponding to the weight of the drug) or PBS, filtered through 0.22 μm, or added to the fecal bacteria, and cultured anaerobically at 37°C for 24 h. OD600 values ​​were recorded at 0, 8, 12, and 24 h to assess bacterial load. After centrifugation, total bacterial DNA was extracted from the precipitate for further real-time quantitative polymerase chain reaction analysis, and the supernatant was collected for quantification of butyric acid content.

[0072] The OD600 growth curve was measured in a culture medium composed of extracts of traditional Chinese medicine, phosphate buffer, and enhanced Clostridium culture medium to ensure its sterility. Figure 11 (A) MTHSWD treatment (50, 100, and 200 mg / mL) significantly increased the total bacterial load within 24 hours of incubation. Figure 11The growth of B) and representative SCFA-producing bacteria (all of which can produce butyrate in the culture system), including *F. prausnitzii*, *Lactobacillus*, *L. murinus*, and *Bifidobacterium*. Figure 12 As expected, butyrate levels were also significantly increased in cultures treated with MTHSWD. Figure 13 Consistently, OD600 growth curves showed enhanced proliferation of *Plannocarya platyspori* (a representative butyrate-producing bacterium) after exposure to MTHSWD. Figure 14 This was accompanied by an increase in butyrate production. These results indicate that MTHSWD can directly promote the growth of butyrate-producing bacteria and increase butyrate production.

[0073] The above specific embodiments are merely explanations of the present invention and are not intended to limit the present invention. After reading this specification, those skilled in the art can make modifications to these embodiments without contributing any inventive step, but as long as they are within the scope of the claims of the present invention, they are protected by patent law.

Claims

1. The application of a traditional Chinese medicine composition in the preparation of a formulation that promotes the growth of butyrate-producing bacteria and increases butyrate production, characterized in that, The traditional Chinese medicine composition consists of the following raw medicinal materials in parts by weight: Angelica sinensis 18-25 parts, peach kernel 12-18 parts, safflower 12-18 parts, prepared rehmannia root 25-32 parts, red peony root 20-25 parts, chuanxiong rhizome 12-18 parts, astragalus root 32-48 parts, epimedium root 25-32 parts, and cinnamon bark 8-12 parts.

2. According to claim 1, the traditional Chinese medicine composition comprises the following raw medicinal materials in parts by weight: Angelica sinensis 22 parts, peach kernel 16 parts, safflower 16 parts, prepared rehmannia root 27 parts, red peony root 22 parts, chuanxiong rhizome 16 parts, astragalus root 40 parts, epimedium 27 parts, cinnamon 10 parts.

3. The application according to claim 2, characterized in that, The preparation is a culture medium composed of extracts of traditional Chinese medicine composition, phosphate buffer, and enhanced Clostridium culture medium.

4. The application according to claim 3, characterized in that, The dosage of the herbal composition extract is as follows: Angelica sinensis 18-25g, Prunus persica 12-18g, Carthamus tinctorius 12-18g, Rehmannia glutinosa 25-32g, Paeonia lactiflora 20-25g, Ligusticum chuanxiong 12-18g, Astragalus membranaceus 32-48g, Epimedium brevicornu 25-32g, and Cinnamomum cassia 8-12g. The herbal composition extract is diluted with phosphate buffer as needed, and the volume ratio of the diluted herbal composition extract to the enhanced Clostridium difficile culture medium is 1:7 to 1:

9.

5. The application according to claim 3, characterized in that, The amount of the herbal composition extract used is as follows: Angelica sinensis 22g, Prunus persica 16g, Carthamus tinctorius 16g, Rehmannia glutinosa 27g, Paeonia lactiflora 22g, Ligusticum chuanxiong 16g, Astragalus membranaceus 40g, Epimedium brevicornu 27g, and Cinnamomum cassia 10g. The herbal composition extract is diluted with phosphate buffer as needed, and the volume ratio of the diluted herbal composition extract to the enhanced Clostridium difficile culture medium is 1:

9.

6. The application according to claim 3, characterized in that, The method for preparing the extract of the traditional Chinese medicine composition includes: mixing all the medicinal materials in the traditional Chinese medicine composition, soaking them in water, extracting twice, filtering, combining the two filtrates, and concentrating to obtain the extract of the traditional Chinese medicine composition.

7. The application according to claim 6, characterized in that, The soaking time is 25-45 minutes.

8. The application according to claim 6, characterized in that, The extraction time is 20-45 minutes.

9. The application according to claim 6, characterized in that, The filtration method involves filtering through four layers of gauze.

10. The application according to claim 6, characterized in that, The concentration is achieved by evaporation and concentration in a rotary evaporator at 60°C to 70°C.

11. The application according to claim 6, characterized in that, The amount of water added is 1L. The traditional Chinese medicine composition is mixed with distilled water and soaked for 30 minutes to obtain a mixture. The mixture is extracted with boiling water for 30 minutes and filtered with 4 layers of gauze. The filter residue is then extracted with boiling water for 20 minutes and filtered with 4 layers of gauze again. The two filtrates are combined and evaporated in a rotary evaporator at 60°C.

12. A method for promoting the growth of butyrate-producing bacteria and increasing butyrate production, characterized in that, The method is as follows: culturing with a culture medium containing an extract of a traditional Chinese medicine composition, wherein the extract is prepared from the following raw medicinal materials in parts by weight: Angelica sinensis 18-25 parts, Prunus persica 12-18 parts, Carthamus tinctorius 12-18 parts, Rehmannia glutinosa 25-32 parts, Paeonia lactiflora 20-25 parts, Ligusticum chuanxiong 12-18 parts, Astragalus membranaceus 32-48 parts, Epimedium brevicornu 25-32 parts, and Cinnamomum cassia 8-12 parts.

13. A method for promoting the growth of butyrate-producing bacteria and increasing butyrate production according to claim 12, wherein the method comprises: culturing the bacteria in a culture medium containing an extract of a traditional Chinese medicine composition, wherein the extract of the traditional Chinese medicine composition is prepared from the following raw medicinal materials in parts by weight: 22 parts Angelica sinensis, 16 parts Prunus persica, 16 parts Carthamus tinctorius, 27 parts Rehmannia glutinosa, 22 parts Paeonia lactiflora, 16 parts Ligusticum chuanxiong, 40 parts Astragalus membranaceus, 27 parts Epimedium brevicornu, and 10 parts Cinnamomum cassia.

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