A traditional Chinese medicine composition for protecting gastric mucosa and application thereof

Abstract

The application belongs to the technical field of traditional Chinese medicine, and particularly relates to a traditional Chinese medicine composition for protecting gastric mucosa and application. The raw material of the traditional Chinese medicine composition is Corydalis and Fructus Meliae, and the preparation method comprises the following steps: firstly, Corydalis is extracted with water, and filtered to obtain Corydalis extract and Corydalis residue; then, Fructus Meliae and the Corydalis residue are mixed with water, and enzyme is added for enzymolysis to obtain an enzymolysis solution; finally, the Corydalis extract and the enzymolysis solution are mixed, sterilized, fermented with probiotic bacteria liquid, and filtered to obtain the traditional Chinese medicine composition. The composition prepared by the preparation method has a significantly better protection effect on gastric mucosa than other methods due to the reasonable combination of Fructus Meliae and Corydalis, can significantly reduce the gastric mucosa lesion degree of experimental gastric mucosa injury model animals, and improve the repair ability of gastric mucosa.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine technology, specifically relating to a traditional Chinese medicine composition for protecting the gastric mucosa and its application. Background Technology

[0002] The gastric mucosa is an important component of the stomach wall and plays a vital protective role in the stomach. However, due to various factors such as irregular eating habits, high stress levels, Helicobacter pylori infection, and long-term use of certain medications, the incidence of gastric mucosal damage is increasing, leading to a series of gastric diseases such as gastritis and gastric ulcers, which seriously affect quality of life.

[0003] Currently, there are many types of drugs on the market for protecting the gastric mucosa, including chemical drugs such as bismuth preparations and aluminum preparations, as well as some traditional Chinese medicine preparations. However, chemical drugs often have varying degrees of side effects. For example, long-term use of bismuth preparations may lead to bismuth accumulation in the body, causing adverse reactions in the nervous system; aluminum preparations may affect the absorption of phosphorus in the intestines, etc.

[0004] Traditional Chinese medicine has many advantages, but some products also have problems such as toxic components, multiple ingredients and complex composition making quality control difficult, and unclear efficacy advantages.

[0005] Sichuan pepper (Chuanlianzi) is the dried, mature fruit of the plant *Melia toosendan* (family Meliaceae), and it has the effects of soothing the liver and clearing heat, promoting qi circulation and relieving pain, and killing parasites. Corydalis rhizome (Yuanhusuo) is the dried tuber of the plant *Corydalis yanhusuo* (family Papaveraceae), and it has the effects of promoting blood circulation, promoting qi circulation, and relieving pain. In traditional Chinese medicine theory, both are often used to treat liver qi stagnation, stomach pain, and other ailments. However, there is currently limited in-depth research on the protective effect of the combination of Sichuan pepper and Corydalis rhizome on the gastric mucosa. Furthermore, Sichuan pepper contains toxic components such as azadirachtin, and there are no reports on how to obtain a stable composition that significantly protects the gastric mucosa and removes toxic byproducts through specific preparation processes. Summary of the Invention

[0006] In view of the problems existing in the prior art, the present invention provides a traditional Chinese medicine composition for protecting the gastric mucosa and its application.

[0007] To achieve the purpose of the invention, the following technical solution is adopted:

[0008] A traditional Chinese medicine composition for protecting the gastric mucosa, characterized in that the preparation method of the traditional Chinese medicine composition includes the following steps:

[0009] (1) Add water to extract Corydalis rhizome, filter, and obtain Corydalis rhizome extract and Corydalis rhizome residue;

[0010] (2) Mix the residue of Sichuan pepper and corydalis with water, add enzymes for enzymatic hydrolysis, and obtain the enzymatic hydrolysate;

[0011] (3) Mix the Corydalis extract and the enzymatic hydrolysate, sterilize, add probiotic liquid for fermentation, and filter to obtain the final product.

[0012] Preferably, the mass ratio of Corydalis yanhusuo to Melia toosendan is 1:0.5-2, the mass ratio of Corydalis yanhusuo to water in step (1) is 1:5-12, the extraction is a heating reflux extraction, the number of extractions is 1-3 times, and the extraction time for each extraction is 0.5-1h.

[0013] Preferably, the enzymes in step (2) are pectinase, xylanase and tanninase.

[0014] Preferably, the mass ratio of pectinase, xylanase and tanninase is 1:0.2-0.5:0.8-1.5.

[0015] Preferably, the pH of the enzymatic hydrolysis in step (2) is 3.5-6.5, the hydrolysis time is 3-5 hours, and the hydrolysis temperature is 40-60°C.

[0016] Preferably, in step (2), the mass of water added is 1-5 times the total mass of the residue of Sichuan pepper and Corydalis rhizome, and the amount of enzyme added is 0.1-0.5% of the mass of water added.

[0017] Preferably, the probiotic liquid in step (3) includes inoculating Lactobacillus casei LC-12 into a seed culture medium and fermenting it at 30-38°C for 12-24 hours to obtain the probiotic liquid.

[0018] Preferably, the inoculation amount of the probiotic solution in step (3) is 0.5-1.5% of the total volume of the Corydalis extract and the enzymatic hydrolysate.

[0019] Preferably, the fermentation temperature in step (3) is 26-30℃ and the fermentation time is 12-72h.

[0020] Another objective of this invention is to provide the application of the above-mentioned traditional Chinese medicine composition in the preparation of products that protect the gastric mucosa.

[0021] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0022] (1) The composition prepared by the method of the present invention has a significantly better protective effect on the gastric mucosa than other processes. Experimental results show that the composition can significantly reduce the degree of gastric mucosal lesions in experimental gastric mucosal injury model animals, improve the antioxidant capacity and repair capacity of the gastric mucosa, and its efficacy is better than that of commercially available Sanjiu Weitai and first-line drugs such as cimetidine and omeprazole.

[0023] (2) The preparation method of this invention can effectively enrich the active ingredients and remove impurities. Through strict screening of raw materials and optimization of extraction and fermentation processes, the content of azadirachtin has been reduced, good safety data has been accumulated, and the potential hepatotoxicity of the prescription has been reduced.

[0024] (3) The compositions of the present invention have high safety after preparation, providing a reliable guarantee for clinical application and have broad application prospects: Due to the high incidence of diseases related to gastric mucosal injury, the market demand for drugs that protect the gastric mucosa is huge. The compositions of the present invention have significant gastric mucosal protective activity, good quality stability and safety, and can be used to prepare drugs in various dosage forms. New pH-dependent formulations can be given priority for targeted release in the stomach to improve their bioavailability. Attached Figure Description

[0025] Figure 1 Example images showing the gross morphological observation results of stomach tissue in each group of rats.

[0026] Figure 2 The graph shows the changes in the levels of serum inflammatory factors IL-6, TNF-α, PGE2, and CRP in rats. Detailed Implementation

[0027] All raw materials used below are commercially available conventional materials. Lactobacillus casei LC-12 was purchased from Shandong Zhongke Jiayi Bioengineering Co., Ltd., and the suppliers of pectinase, cellulase, xylanase, saccharifying enzyme and tanninase are Nanning Dongheng Huadao Biotechnology Co., Ltd.

[0028] Example 1

[0029] A traditional Chinese medicine composition for protecting the gastric mucosa, comprising Corydalis yanhusuo and Melia toosendan in a 1:1 mass ratio, and prepared by the following steps:

[0030] (1) Add 8 times the amount of water to Corydalis rhizome and heat under reflux for extraction. The extraction is performed twice, with each extraction lasting 0.5 hours. Filter the mixture and combine the filtrates to obtain Corydalis rhizome extract and Corydalis rhizome residue.

[0031] (2) Mix the residue of Sichuan pepper and corydalis with 5 times the amount of water, and add 0.5% of the water mass of enzyme (mass ratio 1:0.2:1).

[0032] The pectinase, xylanase and tanninase of 1 were enzymatically hydrolyzed at pH 4.5 and 50℃ for 4 hours to obtain the enzymatic hydrolysate.

[0033] (3) Mix the Corydalis extract and the enzymatic hydrolysate, sterilize at 120°C for 20 min, add 1% of the total volume of the Corydalis extract and the enzymatic hydrolysate as probiotic liquid, ferment at 30°C for 24 h, filter, concentrate under reduced pressure at 65°C, and dry at 105°C to obtain the final product.

[0034] The probiotic culture was prepared by inoculating Lactobacillus casei LC-12 into a seed culture medium (composed of 1.0g glucose, 1.0g peptone, 0.5g yeast powder, 0.5g sodium chloride and 100mL pure water) and fermenting at 35℃ for 24h.

[0035] Example 2

[0036] A traditional Chinese medicine composition for protecting the gastric mucosa, comprising Corydalis yanhusuo and Melia toosendan in a 1:1 mass ratio, and prepared by the following steps:

[0037] (1) Add 5 times the amount of water to Corydalis rhizome and heat and reflux to extract. The extraction is performed 3 times, and the extraction time is 0.5h each time. Filter and combine the filtrates to obtain Corydalis rhizome extract and Corydalis rhizome residue.

[0038] (2) Mix the residue of Sichuan pepper and Corydalis rhizome with 1 times the amount of water, add 0.5% of the water mass of a mixture of pectinase, xylanase and tanninase in a mass ratio of 1:0.2:0.8, and enzymatically hydrolyze at pH 3.5 and 60℃ for 3 hours to obtain the enzymatic hydrolysate.

[0039] (3) Mix the Corydalis extract and the enzymatic hydrolysate, sterilize at 120°C for 20 min, add 0.5% probiotic liquid of the total volume of Corydalis extract and enzymatic hydrolysate, ferment at 26°C for 12 h, filter, concentrate under reduced pressure at 65°C, and dry at 105°C to obtain the final product.

[0040] The probiotic culture was prepared by inoculating Lactobacillus casei LC-12 into a seed culture medium (composed of 1.0g glucose, 1.0g peptone, 0.5g yeast powder, 0.5g sodium chloride and 100mL pure water) and fermenting at 38℃ for 12h.

[0041] Example 3

[0042] A traditional Chinese medicine composition for protecting the gastric mucosa, comprising Corydalis yanhusuo and Melia toosendan in a 1:1 mass ratio, and prepared by the following steps:

[0043] (1) Add 12 times the amount of water to Corydalis rhizome, heat and reflux to extract for 1 hour, filter, and obtain Corydalis rhizome extract and Corydalis rhizome residue;

[0044] (2) Mix the residue of Sichuan pepper and Corydalis rhizome with twice the amount of water, add 0.5% of the water mass of a mixture of pectinase, xylanase and tanninase in a mass ratio of 1:0.5:1.5, and enzymatically hydrolyze at pH 6.5 and 40℃ for 5 hours to obtain the enzymatic hydrolysate.

[0045] (3) Mix the Corydalis extract and the enzymatic hydrolysate, sterilize at 120°C for 20 min, add 1.5% probiotic liquid of the total volume of Corydalis extract and enzymatic hydrolysate, ferment at 26°C for 12 h, filter, concentrate under reduced pressure at 65°C, and dry at 105°C to obtain the final product.

[0046] The probiotic culture was prepared by inoculating Lactobacillus casei LC-12 into a seed culture medium (composed of 1.0g glucose, 1.0g peptone, 0.5g yeast powder, 0.5g sodium chloride, and 100mL pure water) and fermenting it at 30℃ for 12 hours.

[0047] Example 4

[0048] The difference between this embodiment and Example 1 is the probiotic used, specifically Lactobacillus casei CGMCC 1.29. The bacterial culture was prepared by inoculating Lactobacillus casei CGMCC 1.29 into a seed culture medium (composed of 1.0g glucose, 1.0g peptone, 0.5g yeast extract, 0.5g sodium chloride, and 100mL pure water) and fermenting at 28°C for 24 hours to obtain the probiotic culture.

[0049] Example 5

[0050] The difference between this embodiment and Example 1 is that the enzymes are different; specifically, a mixture of pectinase, cellulase, and saccharifying enzyme in a 1:0.2:1 ratio is used.

[0051] Example 6

[0052] The difference between this embodiment and Embodiment 1 is that fermentation is not performed; specifically:

[0053] A traditional Chinese medicine composition for protecting the gastric mucosa, comprising Corydalis yanhusuo and Melia toosendan in a 1:1 mass ratio, and prepared by the following steps:

[0054] (1) Add 8 times the amount of water to Corydalis rhizome and heat under reflux for extraction. The extraction is performed twice, with each extraction lasting 0.5 hours. Filter the mixture and combine the filtrates to obtain Corydalis rhizome extract and Corydalis rhizome residue.

[0055] (2) Mix the residue of Sichuan pepper and corydalis with 5 times the amount of water, and add 0.5% of the water mass of enzyme (mass ratio 1:0.2:1).

[0056] 1. Pectinase, xylanase and tanninase were enzymatically hydrolyzed at pH 4.5 and 50℃ for 4 hours to obtain the enzymatic hydrolysate;

[0057] (3) Mix the Corydalis extract and the enzymatic hydrolysate, concentrate under reduced pressure at 65°C, and dry at 105°C to obtain the final product.

[0058] Example 7

[0059] The difference between this embodiment and Embodiment 1 lies in the different probiotics, specifically Lactobacillus plantarum JYLP-002 (purchased from Shandong Zhongke Jiayi). The culture of its bacterial liquid is to inoculate Lactobacillus plantarum JYLP-002 into a seed culture medium (the composition is 1.0 g of glucose, 1.0 g of peptone, 0.5 g of yeast powder, 0.5 g of sodium chloride, and 100 mL of pure water), and ferment at 37 °C for 24 h to obtain the probiotic bacterial liquid.

[0060] Pharmacodynamic experiment, animal pharmacodynamic experiment for treating gastric mucosal injury

[0061] 1.1 Experimental animals

[0062] Male SD rats (200±20 g), 36 - 56 days old. Purchased from Zhuhai Besttone Biotechnology Co., Ltd., the license number for the experimental unit is SYXK (Guangdong) 2022 - 0125, and the license number is SCXK (Guangdong) 2020 - 0051. Ethics number: gdpulacspf2022552. The SD rats are housed in the Experimental Animal Center of Guangdong Pharmaceutical University, with natural light, a temperature of 25 °C, and a relative humidity of 65 - 70%. Free access to water. All operations during the experiment comply with the experimental ethical principles of the Experimental Animal Center of Guangdong Pharmaceutical University.

[0063] 1.2 Drugs and reagents

[0064] Samples prepared in Embodiments 1 - 7, cimetidine (batch number: C13095070, Shanghai Macklin Biochemical Co., Ltd.), Sanjiuweitai Granules (batch number 2304031F, China Resources Sanjiu Medical & Pharmaceutical Co., Ltd.), ultrapure water (prepared in the laboratory), rat interleukin-6 (IL-6) ELISA detection kit, rat tumor necrosis factor α (TNF-α) ELISA detection kit, rat prostaglandin E2 (PGE2) ELISA detection kit, rat C-reactive protein (CRP) ELISA detection kit, rat superoxide dismutase (SOD) ELISA detection kit, rat malondialdehyde (MDA) ELISA detection kit.

[0065] 1.3 Animal grouping and drug administration

[0066] Healthy SPF-grade male SD rats, aged 36-56 days (200±20g), were given free access to food and acclimatized for three days. They were then randomly divided into 11 groups based on body weight: control group (CON), model group (MOD), Sanjiuweitai group (SJWT), cimetidine group (CTD), and groups 1-7 (Examples 1-7), with 10 rats in each group. The control and model groups were administered distilled water by gavage. The Sanjiuweitai group received 4.2g / kg / day of Sanjiuweitai solution by gavage; the cimetidine group received 42mg / kg / day of cimetidine solution by gavage; and groups 1-7 received their respective drug solutions prepared at 0.375g / kg / day of raw drug. Administered the drugs at fixed times daily for one week. For 24 hours prior to the last administration, the rats were fasted but allowed free access to water. 1.5 hours after the last administration, all rats except the control group were given 1mL of anhydrous ethanol, while the control group received distilled water by gavage. One hour later, SD rats were anesthetized with inhaled isoflurane and samples were collected.

[0067] 1.4 Detection of biochemical indicators in gastric tissue

[0068] A 1cm x 1cm section of gastric tissue was excised from the visibly damaged area and stored in a cryovial at -80°C. Biochemical markers of the gastric tissue included malondialdehyde (MDA) and superoxide dismutase (SOD). The procedure was performed according to the kit instructions.

[0069] 1.5 Gross morphological observation and scoring of gastric tissue

[0070] After anesthetizing experimental rats with isoflurane, the entire stomach was removed, the pylorus was ligated, and an appropriate amount of 10% paraformaldehyde solution was instilled. After fixation for 20 minutes, the stomach was cut open at the greater curvature, rinsed with PBS, and the gastric mucosa was observed and photographed under a stereoscope to record the damage. Simultaneously, the damage to the gastric mucosa was observed, bleeding points and bleeding bands were counted, and the length and width of the bleeding bands were measured using a ruler. The degree of gastric damage in each group of rats was scored according to the criteria in Tables 1 and 2.

[0071] Table 1. Score Table for Severity of Gastric Injury

[0072]

[0073] Table 2 Scoring criteria for bleeding points and bleeding streaks in gastric injuries.

[0074]

[0075] Observation indicators: The degree of gastric mucosal damage in rats in each experimental group was expressed as the damage incidence rate, damage score index, and damage inhibition rate.

[0076] Total score = Bleeding point score + Length score + Width score × 2;

[0077] Incidence of injury (%) = Number of rats with bleeding or ulceration / Number of rats in the group × 100;

[0078] Injury Score Index = Sum of group injury scores / Number of animals in the group;

[0079] Damage inhibition rate (%) = (AB) / A × 100;

[0080] In the formula, A represents the damage integral of the model group; B represents the damage integral of the dose group.

[0081] 2. Statistical methods

[0082] Experimental data were statistically analyzed using Graph Pad Prism 9.0.0, and results are expressed as mean ± standard deviation (x ± s). The t-test was used to compare data between two groups, and one-way ANOVA was used to compare means among multiple groups. P < 0.05 indicated statistical significance, and P < 0.01 indicated significant statistical significance.

[0083] 3. Experimental Results and Analysis

[0084] 3.1 Changes in the activity of SOD and the content of MDA, antioxidant-related factors, in the gastric tissue of rats in each group.

[0085] The results showed that compared with the control group, the SOD level in the gastric tissue of rats in the model group was significantly decreased (P<0.01), the MDA level was significantly increased (P<0.01), and the SOD / MDA ratio was significantly decreased (P<0.01). Compared with the model group, the SOD level in each treatment group was significantly increased (P<0.01), and the MDA level was significantly decreased (P<0.01), showing a dose-related relationship. Only the SOD and MDA levels in groups 1-3 were close to those of the positive control group, especially group 1 in Example 1, with no statistically significant differences. The results are shown in Table 3.

[0086] Table 3

[0087]

[0088]

[0089] Note: Compared with the control group, *P<0.05, **P<0.01; compared with the model group, #P<0.05, ##P<0.01; compared with Example 1 group, &P<0.05, &&P<0.01.

[0090] 3.2 Gross observation results of gastric tissue

[0091] The results showed that the gastric mucosa of rats in the control group was smooth, without congestion, edema, hemorrhage, or erosion; the gastric mucosa of the model group showed punctate and linear hemorrhages, and large areas of erosion. Compared with the model group, punctate or linear hemorrhages were still observed in all treatment groups, but the degree of gastric damage was improved. The Sanjiu Weitai group and the cimetidine group had only a few hemorrhages, and the gastric surface was smoother than that of the model group, with no erosion observed; the 6th example group showed improvement; the 4th-5th and 7th examples showed less bleeding, reduced inflammation, and significant improvement; the 1st-3th examples showed a significant increase in the protective effect on the gastric mucosa, with only small hemorrhage spots and no diffuse inflammation of the whole stomach. The results are as follows. Figure 1 .

[0092] 3.3 Results of the investigation on the incidence of gastric injury, injury score index, and injury inhibition rate

[0093] The results are shown in Table 4 below.

[0094] Table 4

[0095]

[0096]

[0097] Note: Compared with the control group, *P<0.05, **P<0.01; compared with the model group, #P<0.05, ##P<0.01; compared with Example 1 group, &P<0.05, &&P<0.01.

[0098] The results showed that, compared with the control group, the gastric mucosa of all rats in the model group was damaged; compared with the model group, the incidence of gastritis in the positive drug group (CTD, SJWT) and Examples 1-7 was reduced. Among them, Examples 1-3 showed the best effect. Compared with the model group, the total gastric injury score of the positive drug group (CTD, SJWT) and the total gastric injury score of Examples 1-7 were significantly reduced.

[0099] This invention further investigated and verified the levels of IL-6, TNF-α, PGE2, and CRP in the serum of rats in preferred embodiments 1-3. The results showed that, compared with the control group, the corresponding indicators in the model group rats were significantly increased (P < 0.01). Compared with the model group, the levels of IL-6, TNF-α, PGE2, and CRP in the serum of rats in embodiments 1-3 were significantly decreased (P < 0.01). Specific results are detailed below. Figure 2 .

[0100] Summary: In the preliminary stage of this study, the content of azadirachtin was determined using the detection method for *Melia toosendan* fruit in the Chinese Pharmacopoeia. The results showed that the composition prepared using the method of this invention had a lower azadirachtin content than compositions prepared using other processes. This indicates that the extraction and fermentation process of this invention converts azadirachtin, reducing its content. Combined with the aforementioned pharmacodynamic experiments of this invention, the obtained extract effectively improved the protective effect on the gastric mucosa, indicating that it enriched more effective active components or reduced the irritation to the gastric mucosa, effectively inhibiting gastric damage. The specific pharmacodynamic material basis needs further confirmation.

Claims

1. A traditional Chinese medicine composition for protecting the gastric mucosa, characterized in that, The raw materials for the traditional Chinese medicine composition are Corydalis yanhusuo and Melia toosendan, and the preparation method includes the following steps: (1) Corydalis extract was extracted with water and filtered to obtain Corydalis extract and Corydalis residue; (2) Mix the residue of Sichuan pepper and corydalis with water, add enzymes for enzymatic hydrolysis, and obtain the enzymatic hydrolysate; (3) Mix the Corydalis extract and the enzymatic hydrolysate, sterilize, add probiotic liquid for fermentation, and filter to obtain the final product; The enzymes mentioned in step (2) are pectinase, xylanase, and tanninase; The probiotic liquid in step (3) involves inoculating Lactobacillus casei LC-12 into a seed culture medium and fermenting it at 30-38℃ for 12-24 hours to obtain the probiotic liquid.

2. The traditional Chinese medicine composition according to claim 1, characterized in that, The mass ratio of Corydalis rhizome to Melia toosendan is 1:0.5-2, the mass ratio of Corydalis rhizome to water in step (1) is 1:5-12, the extraction is a heating reflux extraction, the number of extractions is 1-3 times, and the extraction time for each extraction is 0.5-1h.

3. The traditional Chinese medicine composition according to claim 1, characterized in that, The mass ratio of pectinase, xylanase, and tanninase is 1:0.2-0.5:0.8-1.

5.

4. The traditional Chinese medicine composition according to claim 1, characterized in that, The pH of the enzymatic hydrolysis in step (2) is 3.5-6.5, the hydrolysis time is 3-5 hours, and the hydrolysis temperature is 40-60℃.

5. The traditional Chinese medicine composition according to claim 1, characterized in that, In step (2), the mass of water added is 1-5 times the total mass of the residue of Sichuan pepper and Corydalis rhizome, and the amount of enzyme added is 0.1-0.5% of the mass of water added.

6. The traditional Chinese medicine composition according to claim 1, characterized in that, The amount of probiotic culture inoculated in step (3) is 0.5-1.5% of the total volume of Corydalis extract and enzymatic hydrolysate.

7. The traditional Chinese medicine composition according to claim 1, characterized in that, The fermentation temperature in step (3) is 26-30℃, and the fermentation time is 12-72h.

8. The use of the traditional Chinese medicine composition according to any one of claims 1-7 in the preparation of a gastric mucosa protectant.

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