CN115463172A - Traditional Chinese medicine composition for treating gastrointestinal mucosal injury - Google Patents

Traditional Chinese medicine composition for treating gastrointestinal mucosal injury Download PDF

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CN115463172A
CN115463172A CN202211156848.4A CN202211156848A CN115463172A CN 115463172 A CN115463172 A CN 115463172A CN 202211156848 A CN202211156848 A CN 202211156848A CN 115463172 A CN115463172 A CN 115463172A
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时小红
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Abstract

The invention discloses a traditional Chinese medicine composition for treating gastrointestinal mucosa injury, which comprises the following raw materials in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment. The invention is safe and effective, treats both principal and secondary aspects of diseases, and has obvious curative effect on alimentary canal mucosal erosion and ulcer.

Description

Traditional Chinese medicine composition for treating gastrointestinal mucosal injury
Technical Field
The invention relates to the technical field of traditional Chinese medicine production, and particularly relates to a traditional Chinese medicine composition for treating gastrointestinal mucosal injury.
Background
The gastrointestinal mucosa injury refers to tissue injury including oral mucosa, esophagus, stomach, duodenum, large and small intestinal erosion, ulcer mucosa lesion and the like. Clinical medicine has studied the pathological mechanism from many aspects deeply, and developed membrane protective agent, proton pump inhibitor, anti-infection and immunosuppressant etc. medicine, but mucosa healing quality, long-term recurrence rate, autoimmune function improvement and pylorus helicobacterium reinfection etc. are still difficult to be done to the best, some medicines have long-term carcinogenic risk, can't be used for a long time, have not specificity and curative, the state of an illness is repeated.
By integrating the descriptions in the documents of ' observation of the effect of traditional Chinese medicine decoction piece decoction on treating gastric ulcer ' (' clinical reasonable medication ' 13 volume 2A (pages 67-68) ' research on the treatment effect of traditional Chinese medicine Xiangsha Liujunzi decoction on gastric ulcer ' (' Chinese modern medicine application ' 2018 volume 12 volume 21 phases 205-206) ' Chinese special disease TCM clinical diagnosis and treatment for digestive department ' (' Beijing people health Press ' 2005 pages 27-72) ', the Chinese herbal medicines developed at present for mucosal injury research mainly comprise: the traditional Chinese medicine composition comprises (1) single traditional Chinese medicines including liquorice, rheum officinale, elecampane, cinnamon, mangnolia officinalis, inula flower, poria cocos, bighead atractylodes rhizome, immature bitter orange, codonopsis pilosula, cuttle bone, thunberg fritillary bulb, bletilla striata, dandelion and the like, (2) compound traditional Chinese medicines including radix bupleuri liver soothing powder, dajianzhong decoction, monkshood middle regulating decoction, three-yellow heart-purging decoction, astragalus root middle-purging decoction, rehabilitation new liquor, qi regulating and stomach harmonizing decoction, clear decoction, five-flavor disinfection decoction, fructus amomi and six-monarch decoction, stomach nourishing granules, radix bupleuri and stomach tonifying decoction, liver soothing and spleen strengthening decoction, stomach nourishing decoction, zuojin pills, yang harmonizing decoction, stomach invigorating and ulcer healing tablets and the like.
Aiming at recurrent oral ulcer, the commonly used western medicine is thalidomide which has quick response, but can only cure symptoms, but can not cure the root cause, relapse is easy after stopping the medicine, and has very large side effect, especially has serious teratogenicity to fetuses, and can cause infertility, and common adverse reactions comprise dry oral and nasal mucosa, burnout, somnolence, dizziness, rash, constipation, nausea, abdominal pain, facial edema, and the like, which can possibly cause polyneuritis, anaphylactic reaction and the like.
Existing treatments, which are not curative, provide only temporary relief from symptoms and may be associated with serious side effects. The health problem is still lack of effective, safe and long-term adverse reaction free medicines. There is an urgent need to find new natural compounds with strong activity and acceptable safety, promoting the rapid healing of erosions or ulcers, preventing the recurrence of canceration. After thousands of years of practice and development, the traditional Chinese medicine can be subjected to syndrome differentiation and formula under the guidance of the theory of traditional Chinese medicine, is strictly processed and prepared, and can play the comprehensive role of multiple targets, multiple ways and multiple links. Has specific advantages in the aspects of improving clinical symptoms of pathological changes, improving the maturity of regenerated mucous membranes, improving healing quality, reducing long-term recurrence rate and the like, and is an effective and feasible treatment idea.
Disclosure of Invention
The invention aims to provide the traditional Chinese medicine composition for treating the gastrointestinal mucosal injury, which is safe and effective, treats both principal and secondary aspects of diseases, and particularly has obvious curative effects on the gastrointestinal erosion and ulcer.
The technical scheme adopted by the invention for solving the technical problem is as follows:
a traditional Chinese medicine composition for treating gastrointestinal mucosal injury comprises the following raw materials in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment.
Preferably, the feed comprises the following raw materials in parts by weight: 30 parts of astragalus membranaceus, 20 parts of cotton rose hibiscus leaves, 15 parts of glossy privet fruits, 10 parts of rhizoma rhei chinensis, 10 parts of phoenix membrane and 6 parts of rhizoma homalomenae.
No research report of the traditional Chinese medicine composition consisting of the 6 medicines is reported in the literature. In human body experiments, better clinical effects can be seen on clinical symptoms and mucosal tissue healing, and meanwhile, the ulcer healing soup can promote proliferation of human gastric mucosal cells in vitro basic experiments. Experiments show that the expression of 312 genes has significant difference compared with a control group, wherein 233 genes are significantly up-regulated, 79 genes are significantly down-regulated (the difference multiple is more than or equal to 2, and P is less than 0.05), and the 312 different genes are enriched on 210 signal paths in the KEGG database. These pathways can be attributed to three major aspects, namely cellular, metabolic and cancer. Wherein the metabolism-related pathway comprises Cell adhesion molecules such as Cell Adhesion Molecules (CAMs), rap1 Signaling pathway, JAK-STAT, mitogen-activated protein kinase Signaling pathway (MAPK Signaling pathway), cytoplasmic Signaling pathway (Cytosalic DNA-sensing pathway), signaling pathway regulating stem Cell pluripotency (Signaling pathway regulation of stem cells), epithelial Cell Signaling in Hp infection (Epithelial Cell Signaling in Helicobacter pylori infection), and the like. Metabolic pathways (glycerophosphopeptide metabolism), galactose metabolism (Galactose metabolism), PPAR (Pyrimidine metabolism), fructose and mannose metabolism (Fructose and mannose metabolism), and the like. The cancer-related pathway includes a PI3K-Akt signaling pathway (PI 3K-Akt signaling pathway), a p53 signaling pathway (p 53 signaling pathway), and the like. The channel enrichment analysis shows that the ulcer healing soup influences the epithelial function of the gastric mucosa by regulating and controlling the signal transmission process of the channels. Therefore, the invention focuses on regulating immunity, mobilizing self-repairing ability, and simultaneously performing local repair and promoting mucosa healing. The gene which is shown to be differentially expressed with cell proliferation participates in the growth and development of cells, has interaction with the invention, and promotes and accelerates the occurrence of clinical erosion and ulcer healing through a specific path. In addition, a total extract containing 823 active ingredients, such as the ingredient of the active ingredient, namely the xanthorrhizone, is obtained through extraction and mass spectrometry of compound compounds, and according to the literature on the influence of the xanthorrhizone with different doses on severe acute pancreatitis of rats (J. Microcirculatory journal, 2010, volume 20, pages 3-19), the total extract can inhibit inflammation and tumor occurrence and signal paths in the development process, and plays an anti-inflammatory and anti-tumor role, and chemical ingredients of the total extract, such as expansion of smooth muscle and blood vessels, improvement of microcirculation disturbance, increase of local blood flow and the like. According to the record of Chinese medicine dictionary, the pharmacological research of single medicine shows that the Chinese rhubarb is the processed product of licorice and excrement juice and mainly contains liquiritin, liquiritigenin, isoliquiritigenin, glycyrrhizic acid, glycyrrhetinic acid and licochalcone A. Human urine is a sediment generated by the change of pH value in urine after human urine is left for a long time. Uric acid, urate, calcium sulfate, calcium hydrogen phosphate and sometimes a plurality of amino acids are precipitated in the acidic environment; precipitated in alkaline environment are calcium carbonate, magnesium ammonium phosphate, calcium phosphate, magnesium phosphate, ammonium urate, calcium oxalate, etc. Folium Hibisci Mutabilis contains flavonoid glycoside, phenols, amino acids, tannin, and reducing sugar. The main component of the eggshell membrane is keratin, and a small amount of viscous protein fiber is clamped in the keratin. Radix astragali contains sucrose, glucuronic acid, mucus, several amino acids, bitter monosodium glutamate, choline, betaine, folic acid, 2',4' -dihydroxy-5, 6-dimethoxy isoflavan and androgenetic bamboo. Linoleic acid and linolenic acid are separated from saponified product of inner Mongolia radix astragali lipid, and non-saponified part contains beta-sitosterol. The glossy privet fruit contains oleanolic acid, mannitol, glucose, palmitic acid, stearic acid, oleic acid and linoleic acid, the peel contains oleanolic acid, acetyl oleanolic acid and androgenic acid, the seed contains 14.9% of fatty oil, the content of palmitic acid and stearic acid in the oil is 19.5%, and the content of oleic acid and linolenic acid is 80.5%.
In the invention, astragalus root, sweet and warm in taste, toxin expelling and tissue regeneration promoting, diuresis inducing and swelling diminishing, yang lifting and sinking promoting, and middle warmer tonifying are monarch drugs and mainly used for treating ulcer without ulceration or ulceration for a long time; zhonghuang for treating sores and ulcers through clearing heat, cooling blood and removing toxicity; the Chinese medicinal materials are white in the middle, clear heat, reduce pathogenic fire and remove blood stasis, are mainly used for treating mouth and tongue sores and are used as ministerial medicaments; the medicinal composition is matched with the cotton rose hibiscus leaves to cool blood, detoxify, detumescence and relieve pain, and is mainly used for treating carbuncle, cellulitis and swelling; fenghuang membrane can nourish yin and treat unhealed ulcer; fructus Ligustri Lucidi has effects of nourishing liver and kidney, and regulating immunity, and is used as adjuvant drug. Mild medicine property, treats both principal and secondary aspects of diseases, promotes the power of mucosa repair, accelerates mucosa healing, and eliminates the root of relapse.
The 6 components are reasonably compatible and act synergistically, can effectively repair gastrointestinal mucosal injury, can regulate immunity, mobilize self-repair capacity, simultaneously carry out local repair and promote mucosal healing, is safe and effective, treats both principal and secondary aspects of diseases, and simultaneously has the effects of changing microbial flora at pathological changes, promoting the propagation of beneficial bacteria and inhibiting the growth of harmful bacteria, thereby repairing from a micro-ecological environment.
The invention is used for clinically treating erosion of oral mucosa, esophagus, stomach, duodenum, large intestine, small intestine, rectum and the like, ulcer and other mucosal lesions, has obvious curative effect, and can also be used for cervical erosion lesions. And can be used for preventing and treating chronic atrophic gastritis. The invention is summarized in long-term clinical practice according to the guidance of the traditional Chinese medicine theory by the inventor and aims at clinical digestive tract mucous membrane injury lesion design. Different from the prior single medicine or a certain active ingredient, the medicine can reduce the occurrence of drug resistance in the clinic, is more suitable for the clinical practice of the traditional Chinese medicine, and has higher application value. Because the traditional treatment method has no specific medicine for modern diseases such as digestive tract mucosa injury, the accurate treatment is slightly insufficient. The invention is drawn to mucosal injury, and the recent and long-term effects of clinical observation are satisfactory.
Preferably, the traditional Chinese medicine composition is prepared into tablets, capsules, water aqua, suppository or powder.
The ulcer healing oral spray comprises 2-3g/mL of crude drugs, and the crude drugs comprise the following components in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment.
Preferably, the preparation method comprises the following steps:
(1) Mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix et rhizoma Rhei and membrana Follicularis ovi according to formula amount, adding water according to the proportion of 1g medicinal materials and 10-15mL water, soaking for 1-2 hours, decocting for 3 times to obtain decoction, filtering, concentrating under reduced pressure to obtain primary concentrated solution containing crude drug 1-1.5g/mL, cooling, adding 3 times volume of 95% ethanol into the primary concentrated solution, standing overnight, sucking supernatant, recovering ethanol, and concentrating to obtain secondary concentrated solution containing crude drug 2-3g/m L; adding 95% ethanol 3 times the volume of the secondary concentrated solution, standing overnight, sucking supernatant, recovering ethanol, and concentrating to obtain tertiary concentrated solution containing crude drug 4-6 g/mL;
weighing homo according to formula amount, mixing homo according to the proportion of adding 8-10mL70-80% ethanol into 1g medicinal material, extracting at 50-60 deg.C for 2-3 times, each time for 1-2 hr, filtering, and mixing extractive solutions;
(2) Filtering the third concentrated solution, mixing with the human urine sediment extractive solution to obtain the final product with crude drug content of 2-3g/mL, and packaging.
The ulcer healing suppository is prepared by mixing 10-20 wt% of medicinal powder and 80-90 wt% of polyoxyethylene stearate, wherein the medicinal powder comprises the following components in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment.
Preferably, the medicinal powder is prepared by mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, calculus bovis, membrana Follicularis ovi and homo, micronizing, and sieving.
The invention has the beneficial effects that: clinical tests, cytology and genomics experiments prove that the traditional Chinese medicine compound has obvious curative effect on preventing and treating the gastrointestinal mucosal injury, and a mechanism of the component for preventing and treating the gastrointestinal mucosal injury is preliminarily discussed.
Drawings
FIG. 1: the patients with chronic oral ulcer treated by the ulcer healing decoction are observed by naked eyes, and the mucous membrane (lower lip ulcer) shows before and after treatment: before 1C intervention, 1D prognosis, chronic erosive gastritis under a microscope and pathological manifestations: 1E stomach body has multiple erosive foci, and 1F pathology (stomach body erosion HP positive). Shows that: the ulcer healing decoction has good repairing effect on the clinical digestive tract mucosa injury.
FIG. 2 is a graph of the concentration of Yuyang decoction administered as a function of cell proliferation activity; when the administration concentration of the ulcer healing decoction is 80 mu g/mL, the promotion effect on the proliferation activity of gastric mucosal epithelial cells is strongest; the abscissa represents the administration concentration of the Yuyang decoction, and the ordinate represents the cell proliferation activity.
FIG. 3 is a graph showing the relationship between the administration time of the ulcer healing decoction and the cell proliferation activity, wherein the concentration of the ulcer healing decoction is 80 μ g/mL, and the effect of promoting the proliferation activity of gastric mucosal epithelial cells is optimal after the ulcer healing decoction is administered for 24 hours; the abscissa represents the administration time of the Yuyang decoction, and the ordinate represents the cell proliferation activity.
FIG. 4 is an experiment of the difference in gene expression of human gastric mucosal epithelial cells (GES-1) in vitro: 312 genes are differentially expressed in the cell proliferation, wherein the up-regulation 233 and the down-regulation 79 are enriched to 210 signal paths by 312 differential genes. Group1: control, group2: a medicine group for experiments. Description of the drawings: the ulcer healing soup has a large effect on GES-1 cells, and influences the epithelial function of the gastric mucosa by regulating and controlling the signal transmission process of various channels.
Detailed Description
The technical solution of the present invention will be further specifically described below by way of specific examples.
In the present invention, the raw materials and equipment used are commercially available or commonly used in the art, unless otherwise specified. The methods in the following examples are conventional in the art unless otherwise specified.
Example 1:
a traditional Chinese medicine composition for treating gastrointestinal mucosa injury comprises the following raw materials in parts by weight: 25 parts of astragalus membranaceus, 23 parts of cotton rose leaves, 18 parts of glossy privet fruits, 8 parts of rheum officinale, 8 parts of phoenix membrane and 8 parts of homalomena leucocephala.
Example 2:
a traditional Chinese medicine composition for treating gastrointestinal mucosa injury comprises the following raw materials in parts by weight: 35 parts of astragalus membranaceus, 16 parts of cotton rose hibiscus leaves, 12 parts of glossy privet fruits, 12 parts of rhizoma rhei chinensis, 12 parts of membrana Follicularis ovi and 5 parts of rhizoma homalomenae.
Example 3:
a traditional Chinese medicine composition for treating gastrointestinal mucosal injury comprises the following raw materials in parts by weight: 30 parts of astragalus membranaceus, 20 parts of cotton rose hibiscus leaves, 15 parts of glossy privet fruit, 10 parts of rheum officinale, 10 parts of phoenix-egg membrane and 6 parts of human urine sediment.
Example 4:
the ulcer healing oral spray comprises 2g/mL of crude drugs, and the crude drugs comprise the following components in parts by weight: 25 parts of astragalus membranaceus, 23 parts of cotton rose hibiscus leaves, 18 parts of glossy privet fruit, 8 parts of rheum officinale, 8 parts of phoenix-egg membrane and 8 parts of human urine sediment.
The preparation method comprises the following steps:
(1) Mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei and membrana Follicularis ovi according to formula amount, soaking in water for 1 hr according to the proportion of 1g medicinal material and 10mL water, decocting for 3 times (the first time is 1.5 hr, the second time is 1 hr, the third time is 40min, the water addition amount of the second and third times is 1g medicinal material and 5 mL) to obtain decoction, filtering, concentrating under reduced pressure to obtain primary concentrated solution containing crude drug 1g/mL, cooling, adding 95% ethanol 3 times volume of the primary concentrated solution, standing overnight, absorbing supernatant, recovering ethanol and concentrating to obtain secondary concentrated solution containing crude drug 2g/m L; adding 95% ethanol with 3 times volume of the secondary concentrated solution, standing overnight, sucking supernatant, recovering ethanol, and concentrating to obtain tertiary concentrated solution containing crude drug 4 g/mL;
weighing human urine sediment according to the formula ratio, mixing and extracting the human urine sediment according to the proportion of 1g of medicinal material added with 8mL of 80% ethanol, extracting for 3 times at 50 ℃ for 1 hour each time, filtering, and combining the extracting solutions;
(2) Filtering the third concentrated solution, mixing with the human urine sediment extractive solution to obtain the final product with crude drug content of 2g/mL, and packaging.
Example 5:
the ulcer healing oral spray comprises 3g/mL of crude drugs, and the crude drugs comprise the following components in parts by weight: 35 parts of astragalus membranaceus, 16 parts of cotton rose hibiscus leaves, 12 parts of glossy privet fruit, 12 parts of rheum officinale, 12 parts of phoenix-egg membrane and 5 parts of human urine sediment.
The preparation method comprises the following steps:
(1) Mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei and membrana Follicularis ovi according to formula amount, soaking in water for 2 hr according to the proportion of 1g medicinal material and 5mL water, decocting for 3 times (the first time is 1.5 hr, the second time is 1 hr, the third time is 40min, the water addition amount of the second and third times is 1g medicinal material and 5 mL) to obtain decoction, filtering, concentrating under reduced pressure to obtain primary concentrated solution containing crude drug 1.5g/mL, cooling, adding 95% ethanol 3 times volume of the primary concentrated solution, standing overnight, absorbing supernatant, recovering ethanol, and concentrating to obtain secondary concentrated solution containing crude drug 3g/m L; adding 95% ethanol with 3 times volume of the secondary concentrated solution, standing overnight, sucking supernatant, recovering ethanol, and concentrating to 6g/mL crude drug to obtain tertiary concentrated solution;
weighing the human urine sediment according to the formula ratio, mixing and extracting the human urine sediment according to the proportion of 1g of medicinal materials and 10mL of 75% ethanol, extracting for 2 times at 60 ℃ for 2 hours each time, filtering, and combining the extracting solutions;
(2) Filtering the third concentrated solution, mixing with the human urine sediment extractive solution to obtain the final product with crude drug content of 3g/mL, and packaging.
Example 6:
the ulcer healing oral spray comprises 2g/mL of crude drug, and the crude drug comprises the following components in parts by weight: 30 parts of astragalus membranaceus, 20 parts of cotton rose hibiscus leaves, 15 parts of glossy privet fruits, 10 parts of rhizoma rhei chinensis, 10 parts of phoenix membrane and 6 parts of rhizoma homalomenae.
The preparation method comprises the following steps:
(1) Mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei and membrana Follicularis ovi according to formula amount, soaking in water for 1 hr according to the proportion of 1g medicinal material and 10mL water, decocting for 3 times (the first time is 1.5 hr, the second time is 1 hr, the third time is 40min, the water addition amount of the second and third times is 1g medicinal material and 5 mL) to obtain decoction, filtering, concentrating under reduced pressure to obtain primary concentrated solution containing crude drug 1g/mL, cooling, adding 95% ethanol 3 times volume of the primary concentrated solution, standing overnight, absorbing supernatant, recovering ethanol and concentrating to obtain secondary concentrated solution containing crude drug 2g/m L; adding 95% ethanol with 3 times volume of the secondary concentrated solution, standing overnight, sucking supernatant, recovering ethanol, and concentrating to obtain a tertiary concentrated solution containing 4g/mL crude drug;
weighing the human urine sediment according to the formula ratio, mixing and extracting the human urine sediment according to the proportion of adding 8mL70% ethanol into 1g of medicinal materials, extracting for 3 times at 50 ℃, extracting for 1 hour each time, filtering, and combining the extracting solutions;
(2) Filtering the third concentrated solution, mixing with the human urine sediment extractive solution to obtain the final product with crude drug content of 2g/mL, and packaging.
Example 7:
the ulcer healing suppository is prepared by mixing 10% of medicinal powder and 90% of polyoxyethylene stearate in percentage by weight, wherein the medicinal powder consists of the following components in parts by weight: 25 parts of astragalus membranaceus, 23 parts of cotton rose hibiscus leaves, 18 parts of glossy privet fruit, 8 parts of rheum officinale, 8 parts of phoenix-egg membrane and 8 parts of human urine sediment. The medicinal powder is prepared by mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei, membrana Follicularis ovi and homo, micronizing, and sieving.
Example 8:
the ulcer healing suppository is prepared by mixing 20% of medicinal powder and 80% of polyoxyethylene stearate in percentage by weight, wherein the medicinal powder consists of the following components in parts by weight: 35 parts of astragalus membranaceus, 16 parts of cotton rose hibiscus leaves, 12 parts of glossy privet fruit, 12 parts of rheum officinale, 12 parts of phoenix-egg membrane and 5 parts of human urine sediment. The medicinal powder is prepared by mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei, membrana Follicularis ovi and homo, micronizing, and sieving.
Example 9:
the ulcer healing suppository is prepared by mixing 15 wt% of medicinal powder and 85 wt% of polyoxyethylene stearate, wherein the medicinal powder comprises the following components in parts by weight: 30 parts of astragalus membranaceus, 20 parts of cotton rose hibiscus leaves, 15 parts of glossy privet fruit, 10 parts of rheum officinale, 10 parts of phoenix-egg membrane and 6 parts of human urine sediment. The medicinal powder is prepared by mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei, membrana Follicularis ovi and homo, micronizing, and sieving.
Test 1:
the subjects adopted in the study specifically include 30 recurrent oral ulcers, 30 gastric ulcers (including duodenal ulcers) and 100 chronic erosive gastritis, which are included in the population from 2016 (04) months to 2017 (02) months, and are treated on the basis of the traditional Chinese medicine for different periods of 1-6 months.
The prescription is composed of 30g of astragalus mongholicus, 20g of cotton rose hibiscus leaves, 15g of glossy privet fruits, 10g of human bezoar (decocted by coating), 10g of membrana Follicularis ovi and 6g of human urine sediment (dried and crushed in a baking oven at 50 ℃ and sieved by a 20-mesh sieve to be prepared into powder, and the powder is swallowed by 2 times 6g per day). Soaking Chinese medicinal decoction pieces (without human urine) for 2 hr, heating to 100 deg.C, decocting to obtain decoction for 2 times, pulverizing human urine into powder, and administering the powder with the decoction for 2 times a day. The invention intervenes in the clinical symptoms of canker sore pathological changes, can be cured, and improves the main and secondary symptoms. Follow-up shows that the ulcer healing quality is good, and the long-term effect is obvious (figure 1). The effective rate reaches 93 percent.
Test 2: the invention relates to the experiment of proliferation and gene expression of human gastric mucosal epithelial cells (GES-1)
2.1 methods of investigation
2.1.1 preparation of Compound Chinese medicine
The medicines used in the research are prepared from decoction piece preparations (produced by Zhejiang tung Juntang Chinese medicinal decoction pieces Co., ltd.), 10g of Renzhonghuang, 6g of Renzhong Bai, 20g of Hibiscus mutabilis leaves, 10g of Fenghuang, 30g of radix astragali and 15g of glossy privet fruit; decocting for 2 times, metering to 1000ml, packaging into 50ml centrifuge tubes, 20ml per tube, freezing overnight at-80 deg.C, cooling to-98 deg.C with Labogen cold trap half an hour in advance, drying the sample in the freeze drying system for 48h to obtain dry powder (number of freeze dryer 2016714306, SP Scientific company in USA), packaging, and storing at-20 deg.C.
2.1.2 determination of reasonable drug concentration and action time of GES-1
2.1.2.1 MTT colorimetric method for detecting influence of different concentrations of Yuyang decoction (the invention) on GES-1 cell proliferation activity, yuyang decoction (obtained by adding water to dry powder of 2.1.1) is added into GES-1 cell culture solution, and the drug concentrations are respectively 0 mug/mL, 10 mug/mL, 20 mug/mL, 40 mug/mL, 80 mug/mL, 160 mug/mL, 320 mug/mL, 640 mug/mL, 1280 mug/mL and 2560 mug/mL. The influence of different concentrations of the ulcer healing decoction on the proliferation activity of the GES-1 cells is detected by an MTT method, and the reasonable drug concentration of the ulcer healing decoction in the GES-1 cell culture solution is determined according to the influence. The method comprises the following steps: cells were added at 100. Mu.l/well (about 1X 10) to 96-well plates 4 ) Each group of 5 duplicate wells. The above ulcer healing decoction with different concentrations is added respectively, and the blank control group is added with culture solution only and is not added with ulcer healing decoction. 37 ℃ C. 5% CO 2 After culturing for 48h in the incubator, adding MTT, incubating for 4h, adding 150 μ l DMSO, shaking for 10min, and detecting absorbance at wavelength of 570 nm.
2.1.2.2 Effect of Yuyang Tang on the proliferative Activity of GES-1 cells with different action time
The GES-1 cells were subcultured in medium, and the cells were seeded into 5 96-well plates at 100. Mu.l/well (about 1X 10) 4 ) 6 per group of 6 double wells, 37 ℃ 5% 2 After the cells are cultured in an incubator overnight, the cells are acted on at the concentration of the ulcer healing decoction of 80 mu g/mL, and the cells are continuously cultured according to the action time of the ulcer healing decoction: 12h,24h,36h and 48h, all 4 groups. The blank control group was not supplemented with Yuyang decoction. After the drug action time is reached, the drug action is stopped, MTT is added for incubation for 4h, then DMSO (150 mu l) is added, oscillation is carried out for 10min, and the absorbance value is detected at the position of 570 nm.
2.1.3 cell genome experiment protocol:
2.1.3.1 genomic RNA extraction
Sample RNA was extracted using the TriZol method.
2.1.3.2RNA sequencing and analysis method.
Detection of RNA in the Normal group, sequencing of the intervention group of the invention, and use of DEGUST: (http://victorian- bioinformatics-consortium.github.io/degust/) And screening out genes with obviously changed expression.
Analysis of RNA sequencing results described how significantly altered expressed genes affected the proliferation of GES-1 cells. Meanwhile, the therapeutic mechanism of the compound traditional Chinese medicine for promoting cell proliferation is described from the perspective of molecular biology.
As a result:
(I) influence of Yuyang Tang on the proliferative Activity of GES-1 cells (FIG. 2)
TABLE 1 Effect of Yuyang Tang at different concentrations on the proliferative Activity of GES-1 cells
Figure BDA0003856673480000081
Figure BDA0003856673480000091
Note: (1), (2) and (3) P < 0.001vs.0.00. Mu.g/mL
Based on the above experiments, it was confirmed that the optimal concentration of Yuyang decoction acting on GES-1 cells was 80. Mu.g/mL. Therefore, the concentration condition of 80 mug/mL can be used when selecting the optimal action time of the ulcer healing decoction.
(II) Effect of Yuyang Tang on GES-1 cell proliferation Activity with different action time (FIG. 3)
TABLE 2 Effect of different incubation times of Yuyang decoction on the proliferative Activity of GES-1 cells
80.00(μg/mL)
1h 1.015±0.031
2h 1.016±0.020
4h 1.054±0.037
6h 1.074±0.065
12h 1.160±0.059
24h 1.325±0.034(1)
36h 1.306±0.026(2)
48h 1.147±0.038
0.00(μg/mL) 1.000±0.023
Note: (1) (2) P < 0.05vs.0.00. Mu.g/mL
The ulcer healing soup in the tables 1 and 2 has no toxicity to GES-1 cells within the range of 0-24 h, and the cell activity begins to decline when the ulcer healing soup has the action time of 36-48 h. Therefore, the influence of Yuyang decoction on the gene expression of the GES-1 cells can be studied within the range of the concentration of 80.00 mu g/mL and the action time of 24 h.
GES-1 cell differential gene expression analysis
1. Sample collection and preparation
1. RNA extraction and detection: agilent 2100bioanalyzer: the integrity and total amount of RNA were accurately determined.
2. Library construction and quality inspection: the starting RNA for the library was total RNA, mRNA with a polyA tail was enriched by Oligo (dT) magnetic beads, and the resulting mRNA was randomly fragmented with divalent cations in a Fragmentation Buffer. Using the fragmented mRNA as a template and random oligonucleotide as a primer, synthesizing a first strand of cDNA in an M-MuLV reverse transcriptase system, then degrading the RNA strand by RNaseH, and synthesizing a second strand of cDNA by using dNTPs as a raw material under a DNA polymerase I system. And (3) carrying out end repair on the purified double-stranded cDNA, adding an A tail and connecting with a sequencing joint, screening cDNA about 370-420 bp by using AMPure XP beads, carrying out PCR amplification, purifying a PCR product by using the AMPure XP beads again, and finally obtaining a library. After the library is constructed, firstly, a Qubit2.0 Fluorometer is used for preliminary quantification, the library is diluted to 1.5ng/ul, then, an insert size of the library is detected by using an Agilent 2100bioanalyzer, and after the insert size meets the expectation, qRT-PCR is used for accurately quantifying the effective concentration of the library (the effective concentration of the library is higher than 2 nM) so as to ensure the quality of the library.
3. Sequencing on a computer: after the library was qualified, different libraries were polung as required for effective concentration and amount of off-machine data of interest, sequenced with Illumina NovaSeq 6000 and generated 150bp paired end reads. The basic principle of Sequencing is Sequencing by Synthesis (Sequencing by Synthesis). Adding four kinds of fluorescence-labeled dNTPs, DNA polymerase and a joint primer into a sequenced flow cell for amplification, releasing corresponding fluorescence every time one fluorescently-labeled dNTP is added when each sequencing cluster extends a complementary chain, and acquiring sequence information of a fragment to be detected by a sequencer through capturing a fluorescence signal and converting an optical signal into a sequencing peak through computer software.
2. Data analysis
1. Data quality control
2. Sequence alignment to reference genome
3. Quantification of gene expression levels
4. Differential expression analysis
5. Differential Gene enrichment analysis
Results of Gene analysis
Compared with the control group, 312 expression difference genes were obtained in the experimental group, of which 233 genes were up-regulated and 79 were down-regulated (compared with the existing database, fig. 4). The expression up-regulated in relation to cell proliferation is: DP-1,2, smad2,3,4, SCF, kip1,2, rb, P300, DNA-PK, ATMATR, PCNA, smc1, stag1,2, rad21, mps1, mad2, bubR1, bub3, APC/C, 14-3-3, cycH, CDK7, CDK1, cycB, CDK1, SCF, P107, 130, rb, ab1, HDAC, DP-1,2, ORC, dbf4, CDK1, rb, cycB; genes with down-regulated expression include E 2 F 4,5 、C-Myc、Miz1、Skp2、Cip1、P53、14-3-3σ、CDK4,6、CycA、Skp2、MCM、Wee、Cdh1、Cdc14。
Test 3:
the Chinese medicinal composition is prepared by clinically intervening on 10g of Renzhonghuang (bag decoction), 6g of Renzhong (powder grinding, swallowing, 2 times a day), 20g of Hibiscus mutabilis leaves, 10g of Fenghuang, 30g of Astragalus membranaceus and 15g of glossy privet fruit for 7 days, and decocting with water for 2 times per day or decocting according to the proportion.
14 tongue coating samples and 6 healthy control groups are collected before and after the application of the medicine, and the tongue coating samples are collected according to the specified requirements. The tongue coating sample collection method comprises the following steps: the oral professional photographed early morning before collection. Scraping with a swab, starting from the middle of the back of the tongue, 3 times from the back to the front, scraping the center of the tongue 1cm 2 About 5s. Vortex immediately in a MoBio buffer containing 750 ul. The sponge on the swab should be pressed several more times, about 20s, against the centrifuge wall to ensure transfer of bacteria into the buffer. The buffer was then cryopreserved at-80 ℃ until the experiment was started. The results are shown in the following table:
TABLE 3 comparison of flora abundance before and after administration of pathogenic flora in tongue coating of patients with recurrent oral ulcer
Class of flora Normal control group Oral ulcer group Yuyang decoction treatment group
Proteobacteria 177043.17±55296 201595.60±133919 156596.90±119899.47
Bacteroides sp 148.6±65.85 255.67±104.47 139.05±173.16
Ciliate chrysosporium 79895±33241.5 90409±57941.70 83744.00±78047.89
Neisseria 83108.25±24717.32 91880.29±88431.34 82826.33±61546.11
Note: p <0.05vs. Proteobacteria, bacteroides, trichizae and Neisseria associated with normal control group oral mucosa erosion tongue coating bacterial colony. It is verified that the main pathogenic factor of pathogenic microorganisms is due to the increased proportion of pathogenic bacteria.
TABLE 4 comparison of abundance of the tongue coating-beneficial bacteria and short-chain fatty acid-producing bacteria for recurrent oral ulcer
Class of flora Normal control group Oral ulcer group Yuyang decoction treatment group
Pleurotus ostreatus 321221.6±49074.20 303034.14±139909.70 325030.95±81641.60
Enterococcus faecalis 179±0 45.5±13.44 113.05±153.96
Eubacterium (producing SCFAs bacteria) 399.10±564.59 164.46±111.30 -
Clostridium (producing SCFAs bacteria) 2360.96±2581.75 1711.16±1633.10 -
Butyric acid vibrio (SCFAs producing bacterium) 1172.32±952.24 749.99±619.93 -
Note: p <0.05vs. normal control.
And (3) analyzing the components of the ulcer healing decoction composition metabonomics: estradiol, butyric acid, berberine, acetoacetate, toluic acid, aminopentanoic acid, pyruvic acid, phloroglucinol, oxycodone, norepinephrine, ornithine, lysine, methionine, glutamic acid, mirtazapine, tryptophan, lysine, epinephrine, isovaleric acid, glutathione, aminobutyric acid, glucoside, and the like.
Researches find that after the ulcer healing decoction intervenes in oral mucosal erosion, the tongue coating of a patient has the phenomenon of quick recovery of beneficial bacteria. Butyric acid is essential for maintaining the integrity of tight junctions between epithelial cells of the intestinal tract. Literature research shows that butyric acid as an energy metabolism substrate of cell mitochondria can promote gene expression and cell proliferation and differentiation, is a main nutrient substance of epithelial cells, provides 70% of total energy for the epithelial cells, and promotes the formation of normal cells by stabilizing DNA and repairing damage. Animal models of butyrate intragastric administration also showed significant increases in parietal cell number, density, and mucosal thickness. The metabonomics of the invention contain butyric acid component, which is important for repairing digestive tract mucosal erosion. From the observation in table 4, it was found that the number of species producing SCFAs was insufficient in the patients with erosive oral mucosa, indicating an increase in the microbiota consuming SCFAs in the patients. The obvious metabolic disorder of the SCFAs is suggested, and the metabolic species of the SCFAs are related to mucosal erosion. SCFAs play a key role in mucosal injury, repair, and regeneration by maintaining mucosal epithelial cell stability. The species reduction unbalance condition of SCFAs in oral ulcer erosion patients is shown, and the pathological mechanism of mucosal erosion and the curative effect mechanism of curing mucosal erosion by the compound traditional Chinese medicine are also shown.
The above-described embodiments are only preferred embodiments of the present invention, and are not intended to limit the present invention in any way, and other variations and modifications may be made without departing from the spirit of the invention as set forth in the claims.

Claims (7)

1. A traditional Chinese medicine composition for treating gastrointestinal mucosa injury is characterized by comprising the following raw materials in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment.
2. The traditional Chinese medicine composition for treating the gastrointestinal mucosal injury according to claim 1, which is characterized by comprising the following raw materials in parts by weight: 30 parts of astragalus membranaceus, 20 parts of cotton rose hibiscus leaves, 15 parts of glossy privet fruit, 10 parts of rheum officinale, 10 parts of phoenix-egg membrane and 6 parts of human urine sediment.
3. The traditional Chinese medicine composition for treating the gastrointestinal mucosal injury according to claim 1 or 2, wherein the traditional Chinese medicine composition is prepared into tablets, capsules, water aqua, suppositories or powder.
4. The ulcer healing oral spray is characterized in that the content of crude drugs in the ulcer healing oral spray is 2-3g/mL, and the crude drugs comprise the following components in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment.
5. The ulcer oral spray according to claim 4, which is prepared by the following steps:
(1) Mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, radix Et rhizoma Rhei and membrana Follicularis ovi according to formula amount, soaking in water for 1-2 hr according to the proportion of 1g medicinal materials and 10-15mL water, decocting for 3 times to obtain decoction, filtering, concentrating under reduced pressure to obtain primary concentrate containing crude drug 1-1.5g/mL, cooling, adding 3 times volume of 95% ethanol into the primary concentrate, standing overnight, sucking supernatant, recovering ethanol, and concentrating to obtain secondary concentrate containing crude drug 2-3 g/mL; adding 95% ethanol with 3 times volume of the secondary concentrated solution, standing overnight, sucking supernatant, recovering ethanol, and concentrating to obtain a third concentrated solution containing crude drug 4-6 g/mL;
weighing homo according to the formula ratio, mixing 1g of Chinese medicinal materials with 8-10mL of 70-80% ethanol, extracting at 50-60 deg.C for 2-3 times, each time for 1-2 hr, filtering, and mixing extractive solutions;
(2) Filtering the third concentrated solution, mixing with the human urine sediment extractive solution to obtain the final product with crude drug content of 2-3g/mL, and packaging.
6. The ulcer healing suppository is characterized by being prepared by mixing 10-20% of medicinal powder and 80-90% of polyoxyethylene stearate in percentage by weight, wherein the medicinal powder comprises the following components in parts by weight: 25-35 parts of astragalus membranaceus, 16-23 parts of cotton rose hibiscus leaves, 12-18 parts of glossy privet fruits, 8-12 parts of rheum officinale, 8-12 parts of membrana Follicularis ovi and 5-8 parts of human urine sediment.
7. The ulcer suppository as claimed in claim 6, wherein the powder is prepared by mixing radix astragali, folium Hibisci Mutabilis, fructus Ligustri Lucidi, homo, membrana Follicularis ovi and homo, micronizing, and sieving.
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