Application of tetrastigma hemsleyanum vine and leaf or extract thereof in preparation of medicine for treating irritable bowel syndrome

Abstract

The invention provides application of tetrastigma hemsleyanum vine leaves or an extract thereof in preparation of a medicine for treating irritable bowel syndrome. Clinical and pharmacodynamic tests prove that the radix tetrastigme vine and leaf extract has a remarkable curative effect on the diarrhea-type irritable bowel syndrome, and clinical medication is safe. The tetrastigma hemsleyanum vine and leaf extract disclosed by the invention not only has a remarkable curative effect on treating diarrhea type irritable bowel syndrome and clinical medication safety, but also is stable in property and controllable in quality after being subjected to processes such as concentration or drying and the like, and can ensure that the drug effect is consistent. The pharmaceutical composition is comprehensive in action mechanism, can play a role in multiple targets of regulating an intestinal nervous system, improving dysbacteriosis, reducing inflammation and the like, also can be in synergistic interaction with other medicines, is high in safety, wide in applicable crowd and convenient in dosage form, and is beneficial to improving the compliance of a patient. In addition, the tetrastigma hemsleyanum vine and leaf resources are rich, the extraction process is simple and environmentally friendly, the raw materials are easy to obtain and low in cost, large-scale production and sustainable development are facilitated, and good economic and social benefits are achieved.

Description

Technical Field

[0001] The present invention relates to the use of Illigera trifoliata leaves or extracts thereof in the preparation of a medicament for treating irritable bowel syndrome. Background Art

[0002] Irritable bowel syndrome is a common chronic disease, affecting approximately 11.2% of the world's population, characterized by recurrent abdominal pain related to defecation and changes in stool frequency or form. Diarrhea-predominant irritable bowel syndrome (IBS-D) is a subtype of irritable bowel syndrome. Epidemiological survey results show that the proportion of IBS-D in functional gastrointestinal diseases in China is 35.6%.

[0003] The leaves of Illigera trifoliata are pungent, slightly sour, sweet, and cool in nature and flavor. They belong to the lung and liver meridians and have the effects of clearing heat and detoxifying, and promoting blood circulation and relaxing tendons. Clinically, they are used for treating swollen and painful throat due to heat toxin, sores and ulcers, snake and insect bites; traumatic injuries, stasis pain, and rheumatic arthralgia.

[0004] The leaves of Illigera trifoliata have the reputation of being a plant antibiotic, which can effectively improve the immune function of the human body and have good therapeutic effects on various primary cancers and metastatic cancers such as esophageal cancer, gastric cancer, lung cancer, liver cancer, kidney cancer, pancreatic cancer, gallbladder cancer, breast cancer, cervical cancer, leukemia, lymphoma, ovarian cancer, bladder cancer, and prostate cancer. Zhong Xiaoming, et al., Effects of Illigera trifoliata extract on local and systemic immune functions of the intestine in scalded rats, Chinese Journal of Medicinal Materials, Vol. 29, No. 9, September 2006, disclosed that Illigera trifoliata extract has a corrective effect on local and systemic immune disorders in the intestine caused by scald. Zhang Xiaoxiong, et al., Research progress on chemical constituents and pharmacological effects of Illigera trifoliata, Fujian Light Industry and Textile, No. 2, February 2024, Illigera trifoliata contains various compounds such as flavonoids, phenolic acids, terpenoids, and polysaccharides. Pharmacological studies have found that Illigera trifoliata has various effects such as anti-cancer, anti-inflammatory, antibacterial, antiviral, and regulating the body's immunity, but the research on the activity and biological mechanism of its single compound is not deep enough. Currently, there is no report on the use of Illigera trifoliata or its extracts for treating irritable bowel syndrome. Summary of the Invention

[0005] The present invention provides a new use of Illigera trifoliata leaves or extracts thereof.

[0006] The present invention provides the use of Illigera trifoliata leaves or extracts thereof in the preparation of a medicament for treating irritable bowel syndrome.

[0007] Among them, the Illigera trifoliata leaf extract is an aqueous extract of Illigera trifoliata leaves.

[0008] Among them, the medicament is a medicament for treating diarrhea-predominant irritable bowel syndrome.

[0009] Among them, the Tetrastigma hemsleyanum Diels et Gilg. vine leaves are the dried aerial vine leaves of Tetrastigma hemsleyanum Diels et Gilg., a plant of the genus Tetrastigma in the Vitaceae family.

[0010] Among them, the preparation method of the water extract of Tetrastigma hemsleyanum Diels et Gilg. vine leaves is as follows:

[0011] a. Take the dried or fresh Tetrastigma hemsleyanum Diels et Gilg. vine leaves.

[0012] b. Soak in water, decoct, concentrate, and dry to obtain the product.

[0013] The drug uses the original Tetrastigma hemsleyanum Diels et Gilg. vine leaves or the water extract as the active ingredient, and is prepared into a pharmaceutically common preparation by adding pharmaceutically acceptable excipients or auxiliary ingredients.

[0014] The preparation is an oral preparation.

[0015] The oral preparation is tablets, capsules, pills, oral liquids, or granules.

[0016] Clinical and pharmacodynamic experiments have proved that the Tetrastigma hemsleyanum Diels et Gilg. vine leaf extract has a significant curative effect on diarrhea-predominant irritable bowel syndrome, and the clinical medication is safe. In addition to having a significant curative effect on diarrhea-predominant irritable bowel syndrome and clinical medication safety, the Tetrastigma hemsleyanum Diels et Gilg. vine leaf extract of the present invention also has many advantages. Its extract is processed by processes such as concentration or drying, with stable properties and controllable quality, which can ensure consistent drug effects. Its action mechanism is comprehensive and may play a role through multiple targets such as regulating the intestinal nervous system, improving dysbacteriosis, and reducing inflammation, and can also synergistically enhance the effects with other drugs. In terms of clinical application, due to its high safety, wide range of applicable populations, and convenient dosage forms, it helps to improve patient compliance. In addition, the resources of Tetrastigma hemsleyanum Diels et Gilg. vine leaves are abundant, the extraction process is simple and environmentally friendly, the raw materials are easily available and the cost is low, which is conducive to large-scale production and sustainable development, and has good economic and social benefits. Description of the Drawings

[0017] Figure 1 Effects on the manifestations of diarrhea-predominant irritable bowel syndrome mice (n = 10) (Note: Compared with the normal group, ##P < 0.01; compared with the model group, *P < 0.05, **P < 0.01);

[0018] Figure 2 Effects on the number of activities in the open field test of diarrhea-predominant irritable bowel syndrome mice (n = 10) (Note: Compared with the normal group, ##P < 0.01; compared with the model group, *P < 0.05, **P < 0.01);

[0019] Figure 3Effect of Tetrastigma hemsleyanum Diels et Gilg Leaf Extract on Intestinal Barrier in Mice with Diarrhea-Predominant Irritable Bowel Syndrome (n = 10) (Note: Compared with the normal group, ##P < 0.01; compared with the model group, *P < 0.05, **P < 0.01). Detailed implementation manners

[0020] Example 1 Preparation of the water extract of Tetrastigma hemsleyanum Diels et Gilg leaf of the present invention

[0021] Take 5 kg of fresh Tetrastigma hemsleyanum Diels et Gilg leaves or 1 kg of dried Tetrastigma hemsleyanum Diels et Gilg leaves, wash, chop, soak in 12 times the amount of water for 1 hour, heat to boiling and boil for 90 min, decoct twice, filter, remove the medicinal residues, and concentrate to obtain an extract of 0.5 g / mL; or a powder of Tetrastigma hemsleyanum Diels et Gilg leaf extract obtained by spray drying or freeze drying.

[0022] The medicinal material of Tetrastigma hemsleyanum Diels et Gilg is from the Tetrastigma hemsleyanum Diels et Gilg planting base in Ninghua, Sanming. It was identified by Senior Experimentalist Fan Shiming of the Medicinal Plant Laboratory of the School of Pharmacy, Fujian University of Traditional Chinese Medicine as the dried vine leaves of Tetrastigma hemsleyanum Diels et Gilg. The sample is stored in the specimen room of the School of Pharmacy, Fujian University of Traditional Chinese Medicine.

[0023] The beneficial effects of the present invention are demonstrated by the following clinical trials and pharmacodynamic experiments.

[0024] Experimental Example 1 Clinical trial of the water extract of Tetrastigma hemsleyanum Diels et Gilg leaf of the present invention

[0025] The present invention selects 66 patients with diarrhea-predominant irritable bowel syndrome as the observation objects, aiming to clearly analyze the clinical efficacy of the water extract of Tetrastigma hemsleyanum Diels et Gilg leaf in the treatment of diarrhea-predominant irritable bowel syndrome. The report is as follows.

[0026] 1. Materials and methods

[0027] General information 66 patients with diarrhea-predominant irritable bowel syndrome who visited the People's Hospital Affiliated to Fujian University of Traditional Chinese Medicine from May 2022 to August 2023 were randomly divided into a control group and a research group, with 33 patients in each group. In the control group, there were 16 males and 17 females; the course of disease was 0.5 - 3.5 years, and the average course of disease was (1.77 ± 0.32) years. In the research group, there were 15 males and 18 females; the course of disease was 1 - 4 years, and the average course of disease was (1.89 ± 0.56) years. Inclusion and exclusion criteria Inclusion criteria: 1) Western medicine diagnosis criteria: This study strictly followed the standardized process in the "Expert Consensus on Irritable Bowel Syndrome in China 2020" to conduct a comprehensive and systematic Western medicine evaluation of the included patients to ensure that the patients met the medical diagnosis criteria of irritable bowel syndrome (IBS). 2) Age limit: The age range of the patients included in this study was 18 - 75 years old. This age range was selected to ensure the extensiveness of the research sample and the practicality of the clinical research. 3) Informed consent: At the level of the patients and their families, we emphasized that before participating in the study, they must fully understand the medical background, purpose, methods, potential risks, and expected benefits of the study and sign an informed consent form on the basis of full knowledge to comply with the norms of medical ethics. 4) Treatment history and compliance: The patients included in the study needed to ensure that they had not received any form of treatment for irritable bowel syndrome (IBS) before the start of the study, including but not limited to drug treatment, traditional Chinese medicine treatment, etc. At the same time, the patients needed to show a high degree of treatment compliance and be able to strictly follow the doctor's medical guidance to ensure the scientificity and reliability of the study.

[0028] Exclusion criteria: 1) Patients with allergic reactions to the drugs or treatment methods involved in this study will be excluded to ensure the safety of the patients and avoid unnecessary allergic reactions; 2) Patients with combined intestinal organic lesions will not be included in the study; 3) Patients with a history of mental illness or current consciousness disorders, resulting in the inability to communicate normally with doctors or researchers, will be excluded to ensure effective communication during the study process and the accuracy of the data; 4) Patients with severe liver and kidney insufficiency are not suitable to participate in this study to avoid possible abnormal drug metabolism or toxic reactions during the study process and ensure the safety of the patients' lives; 5) If a patient chooses to withdraw midway during the study process, their participation will also be terminated.

[0029] In the treatment plan for the patients in the control group, montmorillonite powder was selected as the drug for the control group. In the treatment plan for the research group, in addition to the conventional therapy, the patients received the treatment with the water extract of Tetrastigma planicaule leaves. In the dosing plan, it was recommended that the patients take the medicine in a warm manner, that is, keep the liquid medicine at an appropriate temperature for taking to promote the absorption of the drug and the exertion of the drug effect. One dose of the decoction of Tetrastigma planicaule leaves (30 grams of dried Tetrastigma planicaule leaves) should be taken daily and divided into two equal amounts for taking in the morning and evening.

[0030] 2. Observation indicators

[0031] Quantitative assessment of gastrointestinal symptoms: Referring to the "Expert Consensus on Quantitative Criteria for Gastrointestinal Symptoms 2017", the gastrointestinal symptoms of patients were accurately evaluated, and a systematic quantitative assessment of the clinical symptoms of the two groups of patients before and after treatment was carried out. This assessment process comprehensively covered four key dimensions: stool condition, degree of abdominal distension, frequency of defecation, and degree of abdominal pain, aiming to reflect the changes in the patient's condition through objective data. During the assessment process, the internationally common Likert four-level scoring method was used to quantitatively evaluate the patient's symptoms, which were divided into four grades, representing asymptomatic (normal), mild symptoms, moderate symptoms, and severe symptoms in sequence from 0 to 3.

[0032] Tracking of adverse reactions: During the entire treatment period, the possible adverse reactions of the two groups of patients were tracked and recorded. These adverse reactions included but were not limited to symptoms such as local skin discomfort (such as itching), abnormal gastrointestinal function (such as dyspepsia), and nervous system reactions (such as dizziness).

[0033] Detection of laboratory indicators: Before treatment and 28 days after treatment, in order to accurately quantitatively evaluate the dynamic changes in the concentration of neurotransmitters in the serum of the two groups of patients, strictly following the medical experiment standards, the enzyme-linked immunosorbent assay (ELISA) was used to highly sensitively detect 5-hydroxytryptamine (5-HT) and neuropeptide Y (NPY) in the serum samples.

[0034] 3. Results

[0035] 3.1. Comparison of the clinical efficacy results of the two groups: See Table 1.

[0036] Table 1 Comparison of the clinical efficacy results of the two groups

[0037]

[0038] Note: Compared with the control group, #P < 0.05

[0039] 3.2 Comparison of the gastrointestinal symptom scores of the two groups before and after treatment is shown in Table 2.

[0040]

[0041] Note: Compared with before treatment, #P < 0.05; compared with the control group, △P < 0.05

[0042] 3.3. Comparison of the serum 5-HT and NPY levels of the two groups before and after treatment is shown in Table 3.

[0043] Table 3 Comparison of the serum 5-HT and NPY levels of the two groups before and after treatment

[0044]

[0045] Note: Compared with before treatment, #P < 0.05; compared with the control group, △P < 0.05

[0046] 3.4. Comparison of adverse reactions between the two groups: The total incidence rate in the study group was 3.75%, and that in the control group was 7.19%.

[0047] Summary: The clinical treatment effect of the study group was significantly better than that of the control group in terms of the total effective rate, and the difference was statistically significant (P < 0.05). The improvement in abdominal distension, defecation frequency, stool characteristics, and abdominal pain degree in the study group was better than that in the control group (P < 0.05). After the treatment cycle, the serum 5-hydroxytryptamine (5-HT) and neuropeptide Y (NPY) indexes of the two groups showed a downward trend, and the difference was statistically significant (P < 0.05). The treatment of diarrhea-predominant irritable bowel syndrome with Illigera trifoliata leaves has significant curative effects and obvious advantages in improving clinical symptoms and laboratory indexes.

[0048] Experimental Example 2 Experiment on the effect of the extract of Illigera trifoliata leaves of the present invention on diarrhea-predominant irritable bowel syndrome mice

[0049] 1 Materials and methods

[0050] 1.1 Animal grouping and model establishment: Forty healthy male SPF-grade ICR mice, 6 weeks old, with a body weight of (200 ± 20) g. After 7 days of adaptive feeding, the mice were randomly divided into a normal group, a model group, a montmorillonite powder group, and an aqueous extract group of Illigera trifoliata leaves, with 10 mice in each group. There was no significant difference in body weight among the groups (P > 0.05). Except for the normal group, the mice in the other groups were intragastrically administered with senna leaf extract at 4°C (10 mL / kg) twice a day for 10 consecutive days. During this period, the mice were fed irregularly and had an abnormal diet to establish a diarrhea-predominant irritable bowel syndrome model.

[0051] After the model was established, the normal group and the model group were given distilled water, the montmorillonite powder group and the aqueous extract group of Illigera trifoliata leaves were dissolved in warm water and intragastrically administered (equivalent to 30 grams of the original medicinal materials used by humans) once a day for 14 consecutive days.

[0052] 1.2 Index detection

[0053] 1.3.1 Stool characteristics and body weight

[0054] Record the body weight of the mice, observe their defecation conditions and conduct Bristol scoring. The scoring criteria are shown in Table 4.

[0055] Table 4 Bristol scoring criteria

[0056]

[0057] 1.3.2 Behavioral testing On the last day of the experiment, the mice were placed in a special transparent plastic cage and fixed. The catheter and its tail were tied together with tape. The lubricated latex balloon (length 45 cm) was assembled into the thrombectomy catheter and placed in the colon. The other end of the catheter was connected to a syringe filled with water for colon dilation. After the mice adapted to the environment, water was gradually injected into the balloon to dilate the intestine, and each time lasted for 20 seconds. The capacity thresholds of abdominal lifting and back arching were observed respectively. After the mice rested for 1 hour, the open field test was performed. The mice were placed in a 40cm×40cm×30cm box. After 2 minutes, the number of horizontal and vertical activities was recorded, and the activity within 5 minutes was observed. After each mouse was tested, the area and cage were cleaned with 75% ethanol before the next test. After completing all behavioral tests, the mice were anesthetized by intraperitoneal injection of 3% sodium pentobarbital, blood was collected from the abdominal aorta, and the mice were killed to collect colon tissue and contents.

[0058] 1.2.3 HE staining of colon tissue

[0059] The colon tissue was fixed with 4% paraformaldehyde, embedded in paraffin, dehydrated, sliced, stained with hematoxylin for 10-20 minutes, eosin for 3-5 minutes, and the pathological morphological changes were observed under a microscope after sealing.

[0060] 1.2.4 TUNEL staining of colon tissue was performed according to the instructions of the TUNEL kit. The tissue sections were dewaxed into water, microwave-fixed with citric acid, and incubated with the TUNEL reaction solution at 37°C in the dark for 60 min. The sections were washed with PBS three times for 5 min each time, and the nuclei were stained with DAPI for 15 min. The sections were scanned and images were collected. The nuclei of apoptotic cells were displayed in green, and the nuclei of normal cells were displayed in blue.

[0061] 1.2.5 Serum or colon tissue LPS, LDH, D-LA, 5-HT, NDY, SP, NK-1 level detection

[0062] ELISA kits were used to detect the levels of LPS, LDH, D-LA, 5-HT, NDY, SP, and NK-1 in serum or colon tissue in strict accordance with the instructions, and the absorbance at a wavelength of 450 nm was detected by an enzyme reader.

[0063] 2 Results

[0064] 2.1 Effects of water extract of Ilex trifoliata leaves on symptoms of diarrhea-predominant irritable bowel syndrome in mice

[0065] Compared with the normal group, the body weight of the mice in the model group decreased (P<0.01), and the Bristol score of feces increased (P<0.01); compared with the model group, there were no significant changes in the body weight and Bristol score of feces in the montmorillonite powder group (P>0.05), while the body weight of the mice in the aqueous extract group of Tetrastigma planicaule (Hook.) Merr. leaves increased (P<0.01), and the Bristol score of feces decreased (P<0.05), as shown in Figure 1 A - 1B. Compared with the normal group, the abdominal lift value and back arch value of the mice in the model group decreased (P<0.01), indicating an increase in visceral sensitivity; compared with the model group, there were no significant changes in the abdominal lift value and back arch value of the mice in the montmorillonite powder group (P>0.05), while the abdominal lift value and back arch value of the mice in the aqueous extract group of Tetrastigma planicaule (Hook.) Merr. leaves increased (P<0.05), indicating a decrease in visceral sensitivity, as shown in Figure 1 C.

[0066] 2.2 Effects of the extract of Tetrastigma planicaule (Hook.) Merr. leaves on the emotional disorder in mice with diarrhea - predominant irritable bowel syndrome

[0067] As Figure 2 shown, compared with the normal group, the number of horizontal activities and vertical activities of the mice in the model group decreased (P<0.01); compared with the model group, the number of horizontal activities of the mice in the montmorillonite powder group increased (P<0.05), and there was no significant change in the number of vertical activities (P>0.05), while the number of horizontal activities and vertical activities of the mice in the aqueous extract group of Tetrastigma planicaule (Hook.) Merr. leaves increased (P<0.01).

[0068] 2.3 Effects of the extract of Tetrastigma planicaule (Hook.) Merr. leaves on the colon tissue injury in mice with diarrhea - predominant irritable bowel syndrome

[0069] It can be seen from the pathological sections that the colon tissue structure of the mice in the normal group was intact, the mucosal layer, submucosal layer, muscular layer and adventitia were clearly stratified, and no obvious pathological changes and cell apoptosis were observed; compared with the normal group, a large number of inflammatory cells aggregated in the lamina propria of the colon mucosa of the mice in the model group, mainly lymphocytes and neutrophils, and apoptosis occurred in the colon tissue cells, and the proportion of apoptotic cells increased (P<0.01); compared with the model group, the infiltration of inflammatory cells in the colon mucosa tissue of the mice in the montmorillonite powder group and the aqueous extract group of Tetrastigma planicaule (Hook.) Merr. leaves was reduced, and the proportion of apoptotic cells decreased (P<0.01), and the effect of the aqueous extract group of Tetrastigma planicaule (Hook.) Merr. leaves was better (P<0.01).

[0070] 2.4 Effects of the extract of Tetrastigma planicaule (Hook.) Merr. leaves on the intestinal barrier in mice with diarrhea - predominant irritable bowel syndrome are as Figure 3As shown in Figures A to 3C, compared with the normal group, the levels of serum LPS, LDH, and D-LA in the model group increased (P < 0.01); compared with the model group, there were no significant changes in the levels of serum LPS, LDH, and D-LA in the montmorillonite powder group and the probiotic group (P > 0.05), while the levels of serum LPS, LDH, and D-LA in the aqueous extract group of Tetrastigma hemsleyanum Diels et Gilg leaves decreased (P < 0.05, P < 0.01). As Figure 3 As shown in Figures D to 3E, compared with the normal group, the Occludin and ZO-1 mRNA in the colon tissue of the model group decreased (P < 0.01); compared with the model group, there were no significant changes in the expression of Occludin and ZO-1 mRNA in the colon tissue of the montmorillonite powder group and the probiotic group (P > 0.05), while the expression of Occludin and ZO-1 mRNA in the colon tissue of the aqueous extract group of Tetrastigma hemsleyanum Diels et Gilg leaves increased (P < 0.05).

Claims

1. Use of the leaves of Ivy Leaf or its extract in the preparation of a medicine for treating irritable bowel syndrome.

2. The use according to claim 1, characterized in that: The extract of Semen Triptera leaf is a water extract of Semen Triptera leaf.

3. The use according to claim 1 or 2, characterized in that: The medicine is a medicine for treating diarrhea-type irritable bowel syndrome.

4. The use according to any one of claims 1 to 3, characterized in that: The three-leaf vine leaf is derived from the above-ground vine leaf part of Tetrastigma hemsleyanum Diels et Gilg., a plant of the genus Tetrastigma in the family Vitaceae.

5. The use according to any one of claims 1 to 3, characterized in that: The preparation method of the water extract of Ilex trifoliata leaves is as follows: a. Take dried or fresh Ivy leaves; b. Soak in water, boil, concentrate and dry.

6. The use according to any one of claims 1 to 3, characterized in that: The medicine is prepared from the original medicinal material or water extract of the leaves of Sinensis trifoliata as the active ingredient, and pharmaceutically acceptable excipients or auxiliary ingredients are added to form a commonly used pharmaceutical preparation.

7. The use according to claim 6, characterized in that: The preparation is an oral preparation.

8. The use according to claim 7, characterized in that: The oral preparations are tablets, capsules, pills, oral liquids and granules.

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