CN116327830A - Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof - Google Patents

Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof Download PDF

Info

Publication number
CN116327830A
CN116327830A CN202310493293.0A CN202310493293A CN116327830A CN 116327830 A CN116327830 A CN 116327830A CN 202310493293 A CN202310493293 A CN 202310493293A CN 116327830 A CN116327830 A CN 116327830A
Authority
CN
China
Prior art keywords
ethanol solution
intestinal flora
chinese medicine
traditional chinese
medicine composition
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CN202310493293.0A
Other languages
Chinese (zh)
Other versions
CN116327830B (en
Inventor
任守忠
牛海艳
谢意强
朱菲菲
支芸芸
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Hainan Medical College
Original Assignee
Hainan Medical College
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Hainan Medical College filed Critical Hainan Medical College
Priority to CN202310493293.0A priority Critical patent/CN116327830B/en
Publication of CN116327830A publication Critical patent/CN116327830A/en
Application granted granted Critical
Publication of CN116327830B publication Critical patent/CN116327830B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Classifications

    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/70Polygonaceae (Buckwheat family), e.g. spineflower or dock
    • A61K36/704Polygonum, e.g. knotweed
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/71Ranunculaceae (Buttercup family), e.g. larkspur, hepatica, hydrastis, columbine or goldenseal
    • A61K36/718Coptis (goldthread)
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00Drugs for disorders of the alimentary tract or the digestive system
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P39/00General protective or antinoxious agents
    • A61P39/06Free radical scavengers or antioxidants
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/30Extraction of the material
    • A61K2236/31Extraction of the material involving untreated material, e.g. fruit juice or sap obtained from fresh plants
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/30Extraction of the material
    • A61K2236/33Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
    • A61K2236/333Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/50Methods involving additional extraction steps
    • A61K2236/55Liquid-liquid separation; Phase separation
    • YGENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02ATECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Landscapes

  • Health & Medical Sciences (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Epidemiology (AREA)
  • Microbiology (AREA)
  • Medical Informatics (AREA)
  • Botany (AREA)
  • Alternative & Traditional Medicine (AREA)
  • Organic Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Mycology (AREA)
  • Toxicology (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Medicines Containing Plant Substances (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)

Abstract

The invention discloses a preparation method of a traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora, which comprises polygonum hydropiper and coptis chinensis in a mass ratio of 1.8-2.2:1, wherein effective parts of the polygonum hydropiper and the coptis chinensis are extracted by a specific preparation method, the structures and compositions of the intestinal microbiota are obviously changed after the polygonum hydropiper and the coptis chinensis are extracted, the abundance of probiotics such as lactobacillus and Acermanni are obviously up-regulated, the abundance of harmful microbiota such as bacillus prismatic, molluscum and the like is obviously down-regulated, and the metabolite levels including tyrosine and steroid hormone are regulated.

Description

Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof
Technical Field
The invention relates to the field of traditional Chinese medicine compositions for resisting oxidation and regulating intestinal flora, in particular to a preparation method of a traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora.
Background
The dynamic balance of intestinal microorganisms is closely related to human health, and intestinal microbiota can regulate gastrointestinal homeostasis, host metabolism and immune system, and development of various diseases. Intestinal microbiota dysregulation and altered metabolic phenotype are closely related to the pathophysiology of various diseases, such as GUs and Inflammatory Bowel Disease (IBD), among others. The traditional Chinese medicine or medicated diet has beneficial effects in maintaining intestinal function integrity and relieving mucosa injury by regulating intestinal dysbacteriosis. Various compounds contained in the traditional Chinese medicine inevitably interact with intestinal microbiota after oral administration, and the stable state of the intestinal microbiota is restored by regulating the community structure and metabolism of the intestinal microbiota, and pathological and clinical symptoms are relieved, so that the treatment effect is achieved. Meanwhile, the intestinal microbiota can metabolize or convert active substances of the traditional Chinese medicine, so that the curative effect of the traditional Chinese medicine is changed, the damage of toxic components to the body is weakened, adverse reactions of the medicine to the body are avoided, and the traditional Chinese medicine plays a better role in preventing and treating diseases.
Different intestinal microorganisms have different effects on host health. Lactobacillus and Acremonium belong to the beneficial bacteria. The lactobacillus has effects in relieving inflammation, preventing pathogen invasion and clostridium difficile infection, activating phagocytosis of macrophage, and enhancing immunity. Pharmacological actions of Ackermana include alleviating intestinal inflammation, delaying aging, reducing blood lipid and blood glucose, enhancing immunity, regulating nervous system function, and anti-inflammatory and anticancer effects. The mechanism of action is to improve the thickness of intestinal mucus, maintain the integrity of the intestinal barrier, and prevent invasion of foreign pathogens or toxic substances. Traditional Chinese medicine plays a key role in a variety of diseases by targeting intestinal flora. Lactobacillus and AKK bacteria may be one of the main targets of traditional Chinese medicine, and significant increase in AKK bacteria abundance after treatment with traditional Chinese medicine is a strong evidence that may suggest that excessive lactobacillus and akmann bacteria may have beneficial effects on physical health.
Disclosure of Invention
Accordingly, the present invention provides a preparation method for antioxidant and intestinal flora regulation, which solves the above problems.
The technical scheme of the invention is realized as follows:
a Chinese medicinal composition for resisting oxidation and regulating intestinal flora comprises herba Polygoni Hydropiperis and Coptidis rhizoma at a mass ratio of 1.8-2.2:1.
Further, the preparation method of the traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora comprises the following steps:
(1) Drying and pulverizing herba Polygoni Hydropiperis, processing herba Polygoni Capitati powder by distillation, soaking distilled herba Polygoni Capitati powder in ethanol solution to obtain extractive solution, and concentrating to obtain herba Polygoni Capitati extract;
(2) Extracting the polygonum hydropiper extract by petroleum ether and ethyl acetate in sequence to obtain an ethyl acetate part;
(3) Adsorbing the ethyl acetate part by macroporous resin, and eluting with ethanol solvent to obtain the effective part of herba Polygoni Hydropiperis;
(4) Pulverizing Coptidis rhizoma to obtain Coptidis rhizoma powder, and extracting Coptidis rhizoma powder with subcritical water to obtain Coptidis rhizoma extractive solution;
(5) Adsorbing Coptidis rhizoma extractive solution with macroporous resin, sequentially eluting with water, 80-90wt% ethanol solution, 50-60deg.C ethanol solution and 10-20wt% ethanol solution, collecting eluate, and drying to obtain Coptidis rhizoma effective component.
Further, in the step (1), the distillation temperature of the distillation method is 55-65 ℃ and the distillation time is 30-40min.
Further, in the step (1), the mass ratio of the polygonum hydropiper powder to the ethanol solution is 1:8-10, the soaking time is 15-18h, the soaking times are 2-3 times, the concentration of the ethanol solution is 75-85wt%, and the concentration is carried out until the water content is 30-32%.
Further, in the step (2), the mass ratio of the polygonum hydropiper extract to the petroleum ether to the ethyl acetate is 1:3-5:8-10.
Further, in the step (3), the weight ratio of the coptis extract to the macroporous resin is 1:5-10, and the coptis extract and the macroporous resin are eluted by ethanol solution with the volume of 3-5 times of the column volume, wherein the concentration of the ethanol solution is 30-90wt%.
Further, in the step (4), the subcritical water temperature is 150-170 ℃, the extraction pressure is 10-13MPa, the flow rate is 15-18L/h, and the extraction time is 140-160min.
Further, in the step (5), the mass ratio of the coptis extract to the macroporous resin is 1:7-9, the water consumption is 3-5 times of the column volume, the ethanol solution with the concentration of 80-90wt% is 7-9 times of the column volume, the ethanol solution with the concentration of 50-60wt% is 1-2 times of the column volume, and the ethanol solution with the concentration of 10-20wt% is 3-4 times of the column volume.
Further, in the step (3), the type of the macroporous resin is AB-8, and in the step (5), the type of the macroporous resin is polystyrene type macroporous resin.
The invention has the beneficial effects that:
the Chinese medicine extract consists of the polygonum hydropiper and the coptis chinensis, the specific extraction method is adopted to extract the effective components in the polygonum hydropiper and the coptis chinensis, the structure and the composition of intestinal microbiota are obviously changed after the polygonum hydropiper and the coptis chinensis extract are used, the abundance of probiotics such as lactobacillus and Acermannin are obviously up-regulated, the abundance of harmful microbiota such as bacillus prismatic, mollis and the like is obviously down-regulated, and the metabolite level including tyrosine and steroid hormone is regulated.
Pharmacodynamics research shows that the composition has obvious effect of improving intestinal flora disorder, and can raise the expression of antioxidant enzyme and strengthen antioxidant capacity.
Drawings
FIG. 1A comparison of SOD activities
FIG. 2 influence of example 1 composition on intestinal microbiologic diversity
FIG. 3 influence of example 1 composition on intestinal microbiota composition
FIGS. 4-6 Effect of example 1 compositions the present compositions on intestinal microflora
FIG. 7 Effect of example 1 composition on Lactobacillus entericus composition and abundance
FIG. 8 influence of important metabolic pathways and differential metabolites of the composition of example 1
FIG. 9 comparison of the effects of example 1 compositions on normal GES-1 cell proliferation
FIG. 10 example 1 composition pair H 2 O 2 Induction of GES-1 cell damage effects
Detailed Description
In order to better understand the technical content of the present invention, the following provides specific examples to further illustrate the present invention.
Example 1
Weighing the polygonum hydropiper and the coptis chinensis with the mass ratio of 2:1.
(1) Drying and pulverizing herba Polygoni Hydropiperis whole herb to obtain herba Polygoni Capitati powder, distilling herba Polygoni Capitati powder at 55-65deg.C for 30-40min, soaking in 75-85wt% ethanol solution for 15-18 hr for 2-3 times to obtain extractive solution, and concentrating until the water content is 30-32% to obtain herba Polygoni Capitati extract;
(2) Extracting the polygonum hydropiper extract by petroleum ether and ethyl acetate in sequence, wherein the mass ratio of the polygonum hydropiper extract to the petroleum ether to the ethyl acetate is 1:3-5:8-10, and obtaining an ethyl acetate part;
(3) Adsorbing the ethyl acetate part by using AB-8 macroporous resin, eluting with 3-5 times of 60wt% ethanol solution with the concentration of column volume to obtain the effective part of herba Polygoni Hydropiperis with the weight ratio of Coptidis rhizoma extract to HPD-100 macroporous resin of 1:5-10;
(4) Pulverizing Coptidis rhizoma to obtain Coptidis rhizoma powder, extracting Coptidis rhizoma powder with subcritical water as solvent at 150-170deg.C under 10-13MPa and flow rate of 15-18L/h for 140-160min to obtain Coptidis rhizoma extractive solution;
(5) Adsorbing the coptis chinensis extract by using polystyrene macroporous resin, wherein the mass ratio of the coptis chinensis extract to the polystyrene macroporous resin is 1:8, sequentially adopting 4 times of column volume of water, 8 times of column volume of ethanol solution with the concentration of 85wt%, 1.5 times of column volume of ethanol solution with the concentration of 55wt%, and 3.5 times of column volume of ethanol solution with the concentration of 15wt% to perform elution, collecting eluent, and drying to obtain the coptis chinensis effective part.
(6) Combining the effective parts of herba Polygoni Hydropiperis and Coptidis rhizoma to obtain the final product.
Test example 1
The invention tests that the intestinal flora disorder is induced by constraint water immersion stress
1. Test materials:
reagent:
Figure BDA0004211038860000051
pro Kit, (QIAGEN, USA); fastpfu DNA Polymerase (TransGen, china); axyPrepDNA gel recovery kit (Axygen, USA); NEXTFLEX. Rapid DNA-Seq Kit (Bioo Scientific, USA); miSeq Reagent Kit v3 (Illumina, U.S.A.)
Instrument: an analytical electronic balance (Sartorius cerdolis, germany); cryo-mill (Retsch leichi, germany); cryogenic centrifuges (Eppendorf Ai Bende, germany); vortex mixer (Crystal essence Qi in U.S.); PCR instrument (ABI)
Figure BDA0004211038860000052
Model 9700); sequencer (Illumina Miseq, usa).
2. The test method comprises the following steps:
SD rats were randomly divided into 3 groups: normal group, model group, present composition group, 10 per group. Distilled water is given to the normal group and the model group, and the administration group is administered for 1 time a day for 6 days continuously; before molding, the rats are fasted without water inhibition for 24 hours, after the last administration, the rats except the normal group are placed into a self-made rat cage, the head faces upwards, limbs are fixed, then the rat cage is vertically immersed into a constant-temperature water tank (19+/-1 ℃), the water level reaches the xiphoid process position of the rats, and the molding is finished after the last 7 hours. After the molding, the abdominal cavity is anesthetized with 10% chloral hydrate, the abdominal aorta is collected with blood sample, and the blood serum is collected by centrifugation. Then, 3 faeces are taken from the colon, put into liquid nitrogen and then transferred to-80 ℃ for preservation.
Serum SOD activity assay: firstly, carrying out a small sample pre-experiment, diluting serum stock solution to 5-10 times by using normal saline, completing all experimental operations according to the instruction of a SOD assay (WST-1 method) kit, detecting the OD value of a micro-pore plate, calculating the SOD inhibition rate, analyzing the optimal dilution ratio of samples, controlling the SOD inhibition rate of each group of samples to be between 40 and 60 percent as much as possible, and then carrying out a large amount of formal experiments, wherein the calculation method refers to the instruction.
Sequencing experiment procedure: total DNA extraction and detection, PCR amplification, PCR product identification, purification and quantification, PE library construction and Illumina sequencing.
Bioinformatics analysis: PE reads obtained by Miseq sequencing are spliced according to an overlap relation, quality control and filtering are carried out on sequence quality, and OTU cluster analysis and species taxonomy analysis are carried out after samples are distinguished. Based on the OTU cluster analysis result, various diversity index analyses can be performed, and the sequencing depth is detected; based on the taxonomic information, statistics of the community structure can be performed at various classification levels. Based on the analysis, a series of deep statistical and visual analysis of community structure, phylogenetic development and the like can be performed.
3. Experimental results
1. Referring to fig. 1, the SOD results show that the SOD activity of the model group is significantly reduced compared with that of the normal group, and the SOD activity (P < 0.01) can be significantly improved after pretreatment of the composition of the invention compared with the model group.
2. The effective components in the coptis chinensis and polygonum hydropiper extracts in the invention have influence on the diversity of intestinal microorganisms: alpha diversity analysis was performed on the 16S rRNA sequencing results based on a 97% similarity level. Statistical analysis of the alpha diversity index shows that the diversity of the model group is not different from that of the control group, and shows that the influence of stress stimulation on the diversity of intestinal microorganisms is small, and the composition treatment obviously increases the diversity of intestinal microbial communities. As the number of sampling sequences increases, the sparse curve tends to flatten, indicating enough information for sequencing depth to cover most microbial species, and the results are shown in fig. 2.
3. In the invention, the effective components in the coptis chinensis and polygonum hydropiper extracts have influence on the intestinal microbial composition: the PLS-DA is used for analyzing the composition profile of intestinal microbiota among the model group, the control group and the composition group, and the effect of the effective components in the coptis chinensis and polygonum hydropiper extracts on the intestinal microbiota structure. As shown in fig. 3, the model group and the control group are completely separated and distributed in different quadrants, and there is a significant difference, which indicates that stress causes a significant change in the structure of the intestinal microbiota of the rat, while the treatment group of the composition is completely separated from the model group, which indicates that the composition shows a significant effect on the structure of the intestinal microbiota.
4. In the invention, the effective components in the coptis chinensis and polygonum hydropiper extracts have influence on intestinal microflora: the composition treatment changes at the genus level of intestinal microorganisms more significantly than the portal level. At the phylum level, the composition of dominant bacteria in each group is substantially similar, with the two dominant phylum being the firmicutes phylum and the bacteroides phylum. Verrucomicrobiota abundance was significantly reduced in the model group compared to the normal group, whereas Verrucomicrobiota abundance was reversed after treatment with the present composition, and firmicutes abundance was significantly increased compared to the model group. At the genus level, the highest relative abundance of 5 genera in the three groups were lactobacillus, romidepsia, norank_f __ Muribaculaceae, alloprevotella and Akkermansia, respectively. The abundance of lactobacillus and ackermanni in the model group was significantly reduced compared to the normal group, while the abundance of clostridium_ucg-014 and eubacterium_co-pro-stances was increased. The abundance of lactobacillus and ackermannia is significantly increased after the treatment of the present composition, while the abundance of clostridium_ucg-014 and eubacterium_co-pro stanols is significantly decreased. These reactions show that the effective components in the coptis chinensis and polygonum hydropiper extracts have obvious regulation effect on intestinal microbiota in the invention, and are shown in figures 4-6.
5. Effect of the present composition on the composition and abundance of lactobacillus in the intestinal tract: further analysis of the composition and variation of Lactobacillus in intestinal microorganisms showed that the composition had a significant effect on Lactobacillus and that the abundance of Lactobacillus after treatment with the composition increased significantly, mainly including Lactobacillus murinus, lacillus intestinalis, lacobacillus johnsonii, lactobacillus reuteri, gun_metal_g __ Lactobacillus, uncultured _bacterium_g __ Lactobacillus and unclassified_g __ Lactobacillus. The effective components in the coptis chinensis and polygonum hydropiper extracts in the invention can obviously reverse stress-induced intestinal microbial disorder of rats, and are shown in figure 7.
6. The effect of active ingredients in coptis chinensis and polygonum hydropiper extracts on important metabolic pathways and differential metabolites in the invention: there are two important metabolic pathways, tyrosine metabolism and steroid hormone biosynthesis. The metabolic products of these two metabolic pathways are significantly different from each other, including tyrosine metabolites such as L-tyrosine and L-dopa, and steroid hormone biosynthesis substances such as 21-deoxycortisol, dihydrocortisol, and tetrahydrocortisone. The composition can reduce L-tyrosine and L-dopa levels and 21-deoxycortisol, dihydrocortisol and tetrahydrocortisol levels, thereby improving tyrosine metabolism and HPA axis disorder, as shown in figure 8.
7. Analysis of the correlation of intestinal microbiota with fecal metabolites: at the genus level, the abundance changes of Acremonium and Lactobacillus are inversely related to the changes of tyrosine, 21-hydroxy pregnenolone, D-galactose and cholic acid. These differential metabolites are key fecal metabolites that regulate ackermanni and lactobacillus. The effective components in the coptis chinensis and polygonum hydropiper extracts regulate the changes of fecal metabolites by improving the intestinal microflora composition.
Experimental example 2 cell experiment
1. Effect on normal GES-1 cell proliferation
Taking GES-1 cells in logarithmic growth phase, adding 0.25% trypsin 1mL, digesting at room temperature for 1min, adding 1640 culture solution to terminate digestion, and preparing into concentration of 2×10 5 Inoculating single cell suspension of each/mL on 48-well plate according to cell solution of 100 μL/well, respectively adding composition of example 1 (low concentration 0.5mg/mL, medium concentration 1mg/mL, high concentration 5 mg/mL) of different concentrations after cell adhesion, arranging 6 parallel compound wells for each group of samples, arranging model control group and blank control group, and setting 37 ℃ and 5% CO 2 After 24h of incubation, the stock solution was discarded, 10ul of CC-K8 reagent and 90ul of medium were added to each well, and after 3h of incubation, the absorbance was measured at 450nm using an ELISA reader. Cell viability was determined by CCK-8 method.
2. For H 2 O 2 Induction of GES-1 cell damage effects
Taking GES-1 cells in logarithmic growth phase, adding 0.25% trypsin 1mL, digesting at room temperature for 1min, adding 1640 culture solution to terminate digestion, and preparing into concentration of 2×10 5 individual/mL monolithsInoculating cell suspension into 96-well plate at cell solution of 100 μl/well, adding extract after cell adhesion, arranging 6 parallel multiple wells for each group of samples, arranging negative control group and blank group without cells, and adding 5% CO at 37deg.C 2 After culturing for 24H, the stock solution is discarded, and 200uM H is added 2 O 2 After 4h of injury, the culture broth was aspirated, 10ul of CC-K8 reagent and 90ul of medium were added to each well, and after 3h of incubation, the absorbance was measured at 450nm using an ELISA reader.
Referring to FIG. 9, the traditional Chinese medicine composition prepared in example 1 has different concentrations capable of obviously promoting normal GES-1 cell proliferation and has obvious dose-dependent relationship.
Referring to FIG. 10, GES-1 cells were subjected to 200uM H 2 O 2 After 4 hours of induced injury, the cell survival rate of the model group is obviously reduced compared with that of the normal group; compared with the model group, the compositions of the invention can obviously improve the cell survival rate (p after pretreatment for 12 hours at low and high doses<0.05)。
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.

Claims (9)

1. A traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora is characterized by comprising polygonum hydropiper and coptis chinensis in a mass ratio of 1.8-2.2:1.
2. The method for preparing the traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora according to claim 1, comprising the following steps:
(1) Drying and pulverizing herba Polygoni Hydropiperis, processing herba Polygoni Capitati powder by distillation, soaking distilled herba Polygoni Capitati powder in ethanol solution to obtain extractive solution, and concentrating to obtain herba Polygoni Capitati extract;
(2) Extracting the polygonum hydropiper extract by petroleum ether and ethyl acetate in sequence to obtain an ethyl acetate part;
(3) Adsorbing the ethyl acetate part by macroporous resin, and eluting with ethanol solvent to obtain the effective part of herba Polygoni Hydropiperis;
(4) Pulverizing Coptidis rhizoma to obtain Coptidis rhizoma powder, and extracting Coptidis rhizoma powder with subcritical water to obtain Coptidis rhizoma extractive solution;
(5) Adsorbing Coptidis rhizoma extractive solution with macroporous resin, sequentially eluting with water, 80-90wt% ethanol solution, 50-60deg.C ethanol solution and 10-20wt% ethanol solution, collecting eluate, and drying to obtain Coptidis rhizoma effective component.
3. The method for preparing an antioxidant and intestinal flora-regulating Chinese medicinal composition according to claim 2, wherein in the step (1), the distillation temperature is 55-65deg.C, and the distillation time is 30-40min.
4. The method for preparing the traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora according to claim 2, wherein in the step (1), the mass ratio of the polygonum hydropiper powder to the ethanol solution is 1:8-10, the soaking time is 15-18h, the soaking times are 2-3 times, the concentration of the ethanol solution is 75-85wt%, and the concentration is 30-32%.
5. The method for preparing the traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora according to claim 2, wherein in the step (2), the mass ratio of the polygonum hydropiper extract to the petroleum ether to the ethyl acetate is 1:3-5:8-10.
6. The method for preparing the traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora according to claim 2, wherein in the step (3), the weight ratio of the coptis extract to the macroporous resin is 1:5-10, the coptis extract and the macroporous resin are eluted by ethanol solution with the volume of 3-5 times of the column volume, and the concentration of the ethanol solution is 30-90wt%.
7. The method for preparing an antioxidant and intestinal flora-regulating Chinese medicinal composition according to claim 2, wherein in the step (4), the subcritical water temperature is 150-170 ℃, the extraction pressure is 10-13MPa, the flow rate is 15-18L/h, and the extraction time is 140-160min.
8. The method for preparing the traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora according to claim 2, wherein in the step (5), the mass ratio of the coptis chinensis extracting solution to the macroporous resin is 1:7-9, the water consumption is 3-5 times of column volume, the ethanol solution with the concentration of 80-90wt% is 7-9 times of column volume, the ethanol solution with the concentration of 50-60wt% is 1-2 times of column volume, and the ethanol solution with the concentration of 10-20wt% is 3-4 times of column volume.
9. The method for preparing the traditional Chinese medicine composition for resisting oxidation and regulating intestinal flora according to claim 2, wherein in the step (3), the type of the macroporous resin is AB-8, and in the step (5), the type of the macroporous resin is polystyrene type macroporous resin.
CN202310493293.0A 2023-05-04 2023-05-04 Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof Active CN116327830B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202310493293.0A CN116327830B (en) 2023-05-04 2023-05-04 Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202310493293.0A CN116327830B (en) 2023-05-04 2023-05-04 Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof

Publications (2)

Publication Number Publication Date
CN116327830A true CN116327830A (en) 2023-06-27
CN116327830B CN116327830B (en) 2024-04-12

Family

ID=86878945

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202310493293.0A Active CN116327830B (en) 2023-05-04 2023-05-04 Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof

Country Status (1)

Country Link
CN (1) CN116327830B (en)

Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001022934A2 (en) * 1999-09-24 2001-04-05 Yng Wong Quing Non Delivery of small doses of ingestible treatments
WO2012094834A1 (en) * 2011-01-14 2012-07-19 广东药学院 Compound traditional chinese medicine extract for prevention and treatment of glycometabolism disorders and preparation method thereof
CN104138442A (en) * 2014-07-19 2014-11-12 扬中牧乐药业有限公司 Compound Chinese herbal medicine extracting solution preventing and treating sea eel bacterial enteritis and preparing method of compound Chinese herbal medicine extracting solution
CN108969584A (en) * 2018-10-24 2018-12-11 海南医学院 A kind of composition and its preparation method and application
CN110279764A (en) * 2019-07-16 2019-09-27 广西富凤农牧集团有限公司 A kind of fermented tcm additive and preparation method thereof improving poultry intestinal tract health
CN111821311A (en) * 2019-04-18 2020-10-27 中国医学科学院药物研究所 Application of berberine hydrochloride and stachyose composition in regulating intestinal flora
CN113975317A (en) * 2021-11-22 2022-01-28 美益添生物医药(武汉)有限公司 Application of coptis chinensis in preparation of product for promoting proliferation of beneficial bacteria in intestinal tract

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001022934A2 (en) * 1999-09-24 2001-04-05 Yng Wong Quing Non Delivery of small doses of ingestible treatments
WO2012094834A1 (en) * 2011-01-14 2012-07-19 广东药学院 Compound traditional chinese medicine extract for prevention and treatment of glycometabolism disorders and preparation method thereof
CN104138442A (en) * 2014-07-19 2014-11-12 扬中牧乐药业有限公司 Compound Chinese herbal medicine extracting solution preventing and treating sea eel bacterial enteritis and preparing method of compound Chinese herbal medicine extracting solution
CN108969584A (en) * 2018-10-24 2018-12-11 海南医学院 A kind of composition and its preparation method and application
CN111821311A (en) * 2019-04-18 2020-10-27 中国医学科学院药物研究所 Application of berberine hydrochloride and stachyose composition in regulating intestinal flora
CN110279764A (en) * 2019-07-16 2019-09-27 广西富凤农牧集团有限公司 A kind of fermented tcm additive and preparation method thereof improving poultry intestinal tract health
CN113975317A (en) * 2021-11-22 2022-01-28 美益添生物医药(武汉)有限公司 Application of coptis chinensis in preparation of product for promoting proliferation of beneficial bacteria in intestinal tract

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
SHOUZHONG REN;等: "Polygonum hydropiper extract attenuates ethanolinduced gastric damage through antioxidant and antiinflammatory pathways", BRAZILIAN JOURNAL OF MEDICAL AND BIOLOGICAL RESEARCH, vol. 54, no. 08, pages 1 - 9 *
任守忠;等: "枫蓼提取物抗炎、止泻作用的物质基础研究", 时珍国医国药, vol. 27, no. 01, 20 January 2016 (2016-01-20), pages 32 - 34 *
杨卫丽;曾祥周;张俊清;任守忠;陈峰;刘明生;: "牛耳枫与辣蓼提取物药效学研究", 时珍国医国药, vol. 19, no. 09, pages 2229 - 2231 *
聂敏仁, 等: "中西医结合治疗观赏犬细小病毒病", 河南畜牧兽医, vol. 15, no. 01, pages 1 *

Also Published As

Publication number Publication date
CN116327830B (en) 2024-04-12

Similar Documents

Publication Publication Date Title
CN114507621B (en) Lactobacillus plantarum and application thereof in reducing uric acid, weight and inflammation
CN113209177B (en) Method for extracting flavonoid compounds in citrus peels with assistance of ultrasonic waves
Yi et al. Lactobacillus plantarum CQPC02-fermented soybean milk improves loperamide-induced constipation in mice
AU2020101537A4 (en) A kind of probiotic and application thereof in gastric injury
CN113549582B (en) Licorice fermentation liquor with effects of resisting oxidation, relieving acute alcoholic liver injury and regulating intestinal flora and application thereof
CN116445346B (en) Lactobacillus reuteri for improving polycystic ovary syndrome and application thereof
CN111569007A (en) Probiotic traditional Chinese medicine co-culture extracting solution, preparation method and application
CN114634899B (en) Lactobacillus fermentum and application thereof
CN115287239A (en) Lactobacillus plantarum capable of degrading nucleosides and purines in vitro and reducing uric acid and application thereof
Wang et al. Effects of Scutellaria baicalensis Georgi. on intestinal flora in rats with spleen deficiency and damp-heat
CN110771875A (en) Method for fermenting ginseng by using lactobacillus
CN116327830B (en) Traditional Chinese medicine composition for resisting oxidization and regulating intestinal flora and preparation method thereof
WO2021109879A1 (en) Composition having wholesome personalized intestinal flora diversity function and application
CN111560447A (en) Method for evaluating regulation capacity of hericium erinaceus beta-glucan on intestinal flora
CN116355804A (en) Lactobacillus reuteri for treating polycystic ovary syndrome and application thereof
WO2008064521A1 (en) Lactobacillus fermentum cms-h002 and use thereof
Liu et al. Intestinal bacteria are involved in radix glycyrrhizae and radix euphorbiae pekinensis incompatibility
CN114869916A (en) Lactobacillus plantarum metazoan and application thereof in intestinal microecology modification and cholesterol reduction products
CN114717161A (en) Lactobacillus fermentum and application thereof
CN118028190B (en) Lactobacillus mucilaginosus BAS313 and application thereof in preparation of hypoglycemic products
CN116836872B (en) Bacillus subtilis polysaccharide alpha-amylase inhibitor and application thereof
CN117866850B (en) Lactobacillus plantarum and application thereof in platycodon grandiflorum fermentation
CN113546117B (en) Preparation method for improving content of geniposide
CN117965401B (en) Lactobacillus rhamnosus AFY01 and application of product thereof in inflammatory colon cancer
CN110123824A (en) Ilexsaponin A1New application

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant