JP7287937B2 - Composition used for regulation of intestinal permeability and/or treatment and/or prevention of intestinal wall leakage-related diseases - Google Patents

Description

The present disclosure relates to a Chinese herbal medicine complex or a Chinese herbal medicine complex extract, especially for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases, comprising the Chinese medicine complex medicine or the Chinese medicine complex extract. It relates to the composition used and the use of the herbal compound or the herbal compound extract.

The intestinal barrier regulates intestinal homeostasis, and its major functions include: (1) water and ion transport: the intestinal tract carries approximately 9 liters of fluid per day; It can be processed and is mainly absorbed by the small intestine and can handle the process of fluid absorption and secretion through transcellular or intercellular pathways. (2) Antigen regulation: regulation of antigens in the intestinal lumen is the key to maintaining adequate immunity, and specific dendritic cells can be activated by altering intercellular permeability. . (3) Immune defense: The intestinal lumen can assist in immune defense in the process of washing away microorganisms and toxins (the outer mucus layer has a large number of bacteria and the inner layer is kept sterile).

The intestinal barrier consists primarily of several layers of defense mechanisms that limit the transfer of antigens within the intestinal lumen. The intestinal barrier comprises a single layer of semipermeable epithelial cells whose apical junctions adhesion and tight junctions proteins connect the epithelial cells and Paracellular antigens and molecules can be controlled to cross the epithelium. Intestinal epithelial cells transport antigens and molecules from the intestinal lumen to the mucosa by a transcellular pathway. Certain epithelial cells, such as M cells (Microfold cells, M cells), can transport antigens in the intestinal lumen to phagocytes and lymphocytes in the epithelium to initiate an immune response. Goblet cells, Paneth cells and enterocytes can secrete mucins and antimicrobial peptides (AMPs), which aggregate into the mucus layer. can be. Plasma cells of the lamina propria of intestinal epithelial cells can also secrete IgA. Intestinal epithelial cells also have many microbial recognition receptors (MRRs), such as Toll-like receptors (TLRs) and NOD-like receptors, and are capable of specific Microbial associated molecular patterns (MAMPs) can be identified. Intestinal microbes identified by intestinal epithelial cells trigger the secretion of cytokines and other immunomodulatory factors that help elicit immunomodulatory responses to counteract intestinal microbes and maintain intestinal homeostasis. Become.

The intestinal barrier functions to prevent the entry of toxic antigens and to maintain the health of the intestinal tract, and enteric bacteria are important constituents of the intestinal mucosal barrier. When the intestinal flora is in balance, normal functioning of the body can be maintained. On the other hand, the gut flora is out of balance with chronic stress, chronic constipation, and exposure to environmental toxins (e.g., taking antibiotics, unhealthy eating, etc.) killing more bacteria. As a result, the intestinal mucosa is destroyed by bad bacteria, and food debris and toxins enter the bloodstream. This phenomenon is called leaky gut, or increased intestinal permeability. is. Intestinal wall leakage or increased intestinal permeability also causes an inflammatory response.

Intestinal wall seepage can be caused by a number of factors and can manifest in many different clinical manifestations or illnesses. Currently known diseases associated with abnormal intestinal barrier function and imbalanced intestinal flora include Inflammatory Bowel Disease (IBD), coeliac disease (or celiac disease), hypersensitivity irritable bowel syndrome, acute pancreatitis, non-alcoholic fatty liver disease (NAFLD), alcoholic cirrhosis, type 1 and type 2 diabetes mellitus, obesity, chronic kidney disease, cardiovascular disease, multiple organ failure syndrome (shock, burns and trauma), AIDS, asthma ), eczema, psoriasis, autism, depression, anxiety, schizophrenia, bipolar disorder, Alzheimer's disease ( Alzheimer's disease, Parkinson's disease, multiple sclerosis, Amyotrophic lateral sclerosis (ALS), ankylosing spondylitis, fibromyalgia , chronic sleep fragmentation, insomnia, and the like.

However, there are currently no herbal medicines that have been proven to be useful for treating diseases associated with leaky gut, so there is currently a need for new herbal medicines that can be used to treat diseases associated with leaky gut. .

The present disclosure provides a composition for use in regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal leakage, the composition comprising a Chinese herbal medicine complex or a Chinese herbal medicine complex extract. The Chinese herbal compound includes reishi mushroom, jujube, longan flesh and lotus seed. In addition, the herbal complex extract includes ganoderma lucidum extract, jujube extract, longan extract and lotus seed extract.

In order to make the above and other objects, features and advantages of the present invention more clear and comprehensible, preferred embodiments are listed below and described in detail as follows in conjunction with the accompanying drawings.
[Invention 1001]
A composition for use in modulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage, comprising:
containing a herbal compound or a herbal compound extract,
Said Chinese herbal medicine compound is:
Ganoderma lucidum;
jujube;
longan meat; and
lotus seeds, including
The herbal medicine complex extract is:
reishi extract;
Jujube extract;
longan meat extract; and
A composition comprising a lotus seed extract.
[Invention 1002]
The composition of the present invention 1001 for use in regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage, wherein the ganoderma lucidum includes red turf, purple turf, or antler turf.
[Invention 1003]
The jujube includes ash jujube, chicken heart jujube, winter jujube, large jujube, small jujube or golden jujube, for the regulation of intestinal permeability and/or treatment and/or prevention of diseases associated with intestinal wall leakage according to the present invention 1001. Composition used.
[Invention 1004]
The composition for use in regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage according to the invention 1001, wherein the longan meat includes powdered shell longan meat, red shell longan meat, or green shell longan meat.
[Invention 1005]
The composition used for controlling intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage according to the present invention 1001, wherein the lotus seeds include red lotus seeds or white lotus seeds.
[Invention 1006]
The intestinal tract of the present invention 1001, wherein the composition used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage contains 0.2 to 20 mg of reishi acid A per gram of composition. A composition for use in modulating permeability and/or treating and/or preventing diseases associated with intestinal wall leakage.
[Invention 1007]
The present invention 1001, wherein the weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed in the herbal medicine compound is 0.1-15:0.6-2:0.6-2:0.6-5. and/or for the treatment and/or prevention of intestinal wall leakage-related diseases.
[Invention 1008]
The weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed in the traditional Chinese medicine complex drug is 1:1:1:1, according to the present invention 1001, for controlling intestinal permeability and/or treating intestinal wall leakage-related diseases. Compositions for therapeutic and/or prophylactic use.
[Invention 1009]
The weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed in the Chinese herbal medicine complex is 3:1:1:1, according to the present invention 1001 for controlling intestinal permeability and/or treating intestinal wall leakage-related diseases. Compositions for therapeutic and/or prophylactic use.
[Invention 1010]
Ganoderma lucidum, jujube, longan flesh and lotus seed as raw materials necessary for obtaining the ganoderma lucidum extract, jujube extract, longan flesh extract and lotus seed extract contained in the aforementioned herbal medicine complex extract, respectively. Modulation of intestinal permeability and/or treatment of intestinal leakage-related diseases and /or a composition for prophylaxis.
[Invention 1011]
Ganoderma lucidum, jujube, longan flesh and lotus seed as raw materials necessary for obtaining the ganoderma lucidum extract, jujube extract, longan flesh extract and lotus seed extract contained in the aforementioned herbal medicine complex extract, respectively. The composition of the present invention 1001 for use in regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage, wherein the weight ratio is 1:1:1:1.
[Invention 1012]
Ganoderma lucidum, jujube, longan flesh and lotus seed as raw materials necessary for obtaining the ganoderma lucidum extract, jujube extract, longan flesh extract and lotus seed extract contained in the aforementioned herbal medicine complex extract, respectively. The composition of the present invention 1001 used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage, wherein the weight ratio is 3:1:1:1.
[Invention 1013]
The intestinal wall seepage-related diseases include inflammatory bowel disease, celiac disease, irritable bowel syndrome, acute pancreatitis, non-alcoholic fatty liver disease, alcoholic cirrhosis, type 1 diabetes, type 2 diabetes, obesity, and chronic kidney disease. disease, cardiovascular disease, multiple organ disorders, AIDS, asthma, eczema, psoriasis, autism, depression, anxiety, schizophrenia, bipolar disorder, Alzheimer's disease, Parkinson's disease, multiple sclerosis, muscle atrophy Modulation of intestinal permeability and/or treatment and/or prevention of intestinal wall seepage-related diseases of the invention 1001, including lateral sclerosis, ankylosing spondylitis, fibromyalgia, chronic sleep fragmentation or insomnia composition for use in
[Invention 1014]
A composition of the invention 1001 for use in modulating intestinal permeability and/or treating and/or preventing intestinal wall seepage-related diseases, further comprising a pharmaceutically acceptable carrier or salt.
[Invention 1015]
In the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of diseases associated with intestinal wall leakage, the content of the herbal medicine composite or the herbal medicine complex extract may be 10-99.5 wt%, Compositions of the present invention 1014 used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage.
[Invention 1016]
The regulation of intestinal permeability and/or the intestinal tract of the present invention 1001, wherein the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of diseases associated with intestinal wall leakage is a pharmaceutical composition or a healthcare composition. A composition for the treatment and/or prevention of a leak-related disease.
[Invention 1017]
Oral forms of said pharmaceutical or health care compositions include tablets, granules, powders, pellet-in-capsules, capsules, coated tablets, solutions or instant powders of the enteric permeable composition of the invention 1016. A composition for use in the regulation and/or treatment and/or prevention of leaky gut-related diseases.

1A to 1P show the results of the cell-killing concentration test of each test sample prepared in Example 1, respectively. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. See description of FIG. 1A. Figure 2 shows the results of an intestinal epithelial cell permeability test of single component extracts. ***: p<0.001 compared with negative control group. Data are presented as mean±standard deviation (Mean±SD) (n=3). The statistical method is one-way ANOVA. Figure 2 shows the results of an intestinal epithelial cell permeability test of a multi-component extract. Compared with negative control group: *: p<0.05; ***: p<0.001. Compared to sample 15A: #: p<0.05; ##: p<0.01; ###: p<0.001. Compared to sample 1: φ: p<0.05, φφ: p<0.01, φφφ: p<0.001. Data are presented as mean±s.d. (n=3). The statistical method is one-way analysis of variance. Figures 4A to 4D show the results of the intestinal epithelial cell permeability test of the four multi-component extracts with different formulation ratios. Compared with negative control group: *: p<0.05; **: p<0.01; ***: p<0.001. Compared to sample 15A: #: p<0.05; ##: p<0.01; ###: p<0.001. Compared to sample 1: φ: p<0.05, φφ: p<0.01, φφφ: p<0.001. Data are presented as mean±s.d. (n=3). The statistical method is one-way analysis of variance. See description of FIG. 4A. See description of FIG. 4A. See description of FIG. 4A. Figure 2 shows the results of an intestinal epithelial cell permeability test for different concentrations of Sample 15B. Compared with rhamnose-treated negative control group: *: p<0.05; in positive control group and each experimental group, rhamnose-treated group compared with rhamnose-untreated: #: p<0.05. Data are presented as mean±s.d. (n=3). The statistical method is one-way analysis of variance. Figures 6A to 6C show the effect of sample 15B on gene expression of CLDN3, OCLN and TJP1, respectively. The negative control group is untreated cells and the positive control group is cells treated with berberine chloride (BBC) at 50 μM (18.6 μg/mL). Compared with negative control group: *: p<0.05; **: p<0.01; ns: not significant. Data are presented as mean±s.d. (n=3). The statistical method is one-way analysis of variance. See description of FIG. 6A. See description of FIG. 6A. 15B shows the effect of sample 15B on DSS-induced mouse intestinal permeability. Compared with untreated group: #: p<0.05. Compared with negative control group: *: p<0.05. Data are presented as mean±standard deviation (n=3-4). The statistical method is t test. 15B shows the effect of sample 15B on circadian rhythm disrupters. Compared with untreated group: ###: p<0.001. Compared with negative control group: ***: p<0.001. Data are presented as mean±s.d. (n=6). Statistical methods for data are one-way analysis of variance and Dunnett's test. Figures 9A, 9B and 9C show the effect of Sample 15B on time spent in the hole, number of hole-pokes and exploratory ability of rats in tests assessing motor activity and exploratory behavior. ing. Compared with Sham group: #: p<0.05, ##: p<0.01. Compared with control group: *: p<0.05, **: p<0.01. Data are presented as mean±s.d. (n=6). Statistical methods for data are one-way analysis of variance and Dunnett's test. See description of FIG. 9A. See description of FIG. 9A. Figure 2 shows the effect of Sample 15B on rats' time to find a safe platform in the pool in the spatial performance in Morris water maze test. Compared with Sham group: #: p<0.05. Compared with control group: **: p<0.01; ***: p<0.001. Data are presented as mean±s.d. (n=6). Statistical methods for data are one-way analysis of variance and Dunnett's test. Figure 2 shows the effect of Sample 15B on rats' time to reach the pool safety platform in the non-spatial performance in Morris water maze test. Compared with Sham group: ###: p<0.001. Compared with control group: *: p<0.05; ns: not significant. Data are presented as mean±s.d. (n=6). Statistical methods for data are one-way analysis of variance and Dunnett's test. Figures 12A and 12B show the effect of sample 15B on AChE activity in rat hippocampal and frontal cortex regions in a β-amyloid-induced learning disability animal model. Compared with Sham group: #: p<0.05, ###: p<0.001. Compared with control group: **: p<0.01; ***: p<0.001; ns: not significant. Data are statistical methods. Data are one-way analysis of variance and Dunnett's test. See description of FIG. 12A. 2 shows the results of high performance liquid chromatography (HPLC) analysis of sample 15B.

The present disclosure is a composition for use in regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage, comprising, but not limited to, a Chinese herbal compound or a Chinese herbal compound extract. A good composition can be provided, but is not limited to this.

The “intestinal wall seepage-related disease” described in the present disclosure is not particularly limited as long as it is a disease associated with intestinal wall seepage. For example, a leaky-associated disease may be alleviated and/or treated and/or its symptoms or per se by modulating intestinal permeability and/or by reducing and/or treating and/or preventing leaky gut. or a disease that can be prevented, or a symptom or itself that is alleviated and/or associated with the expression of a leaky-associated gene and/or protein and by modulating the expression of the leaky-associated gene and/or protein It may include, but is not limited to, diseases that can be treated and/or prevented. Examples of leaky gut related diseases include, but are not limited to, inflammatory bowel disease (such as colitis), celiac disease, irritable bowel syndrome, acute pancreatitis, non-alcoholic fatty liver disease, alcoholic cirrhosis, Type 1 diabetes, type 2 diabetes, obesity, chronic kidney disease, cardiovascular disease, multiple organ disorders, AIDS, asthma, eczema, psoriasis, psychiatric disorders (e.g., autism, depression, anxiety, schizophrenia, bipolar disorder, etc.), neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, or a combination thereof), ankylosing spondylitis, fibromyalgia, sleep disorders (e.g., chronic sleep fragmentation, insomnia or a combination thereof) or any combination of the above.

In the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure, the above-mentioned Chinese herbal compound drugs include, but are not limited to, ganoderma lucidum, jujube, longan pulp and lotus seed, and the herbal complex extracts may include, but are not limited to, ganoderma lucidum extract, jujube extract, longan extract and lotus seed extract. The Chinese herbal medicine complex or the Chinese herbal complex extract has a modulating effect on intestinal permeability and/or has a therapeutic and/or preventive effect on intestinal wall seepage-related diseases.

In the composition used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage of the present disclosure, ganoderma lucidum in the Chinese herbal medicine complex, or one of the raw materials for preparing the Chinese herbal medicine complex extract The source reishi may include, but is not limited to, red turf, purple turf, antler reishi, or any combination thereof. In one embodiment, the ganoderma lucidum in the Chinese herbal medicine complex or one of the raw materials for the preparation of the Chinese herbal medicine complex extract may be Ganoderma lucidum.

In the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure, jujube in the Chinese herbal medicine composite drug, or one of the raw materials for producing the Chinese herbal medicine composite extract Different jujubes can include, but are not limited to, gray jujubes, chicken heart jujubes, winter jujubes, large jujubes, small jujubes, golden jujubes, or any combination thereof. In one embodiment, the jujube in the Chinese herbal medicine complex or the jujube as one of raw materials for the preparation of the Chinese medicine complex extract may be ash jujube.

In the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure, it is one of the raw materials for preparing the longan meat in the Chinese herbal medicine complex or the Chinese herbal medicine complex extract. Longan meat may include, but is not limited to, powdered shell longan meat, red shell longan meat or green shell longan meat or any combination thereof. In one embodiment, the longan meat in the Chinese herbal medicine complex or one of the raw materials for the preparation of the traditional Chinese medicine complex extract may be powdered longan meat.

In addition, in the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure, the lotus seed in the above-mentioned Chinese herbal medicine compound or raw materials for producing the above-mentioned Chinese herbal compound extract lotus seeds may include, but are not limited to, red lotus seeds, white lotus seeds, or combinations thereof. In one embodiment, the lotus seed in the Chinese herbal medicine complex or the lotus seed that is one of the raw materials for preparing the Chinese herbal medicine complex extract may be the red lotus seed.

In a specific embodiment, ganoderma lucidum, jujube, longan flesh and lotus seed in the above-mentioned Chinese herbal medicine composite drug, or ganoderma lucidum, jujube, longan flesh and lotus seed as raw materials for preparing the above-mentioned traditional Chinese medicine complex extract, respectively, It can be ash jujube, powdered longan meat and red lotus fruit.

Compositions for use in modulating intestinal permeability and/or treating and/or preventing intestinal leakage-related diseases of the present disclosure contain at least about 0.2 mg of reishi acid A per gram of composition. Well, for example, at least about 0.2 to 20 mg of reishi acid A per gram of composition, such as about 0.2 mg, about 0.25 mg, about 0.3 mg, about 0.4 mg, about 0.5 mg, about 0 Including, but not limited to, .8 mg, about 1 mg, about 2 mg, about 5 mg, about 10 mg, about 15 mg, about 20 mg, and the like. In one embodiment, the minimum content of reishi acid A is used to validate the composition used for modulating intestinal permeability and/or treating and/or preventing intestinal wall seepage-related diseases of the present disclosure. be able to.

In one embodiment, in the composition for use in regulating intestinal permeability and/or treating and/or preventing intestinal wall leakage-related diseases of the present disclosure, the Chinese herbal medicine complex contains ganoderma lucidum, jujube, longan flesh and lotus root. The actual weight ratio is about 0.1-15:0.6-2:0.6-2:0.6-5, for example about 0.3-12:0.8-1.5:0.8- 1.5:0.8-4, about 0.5-10:1:1:1-3, about 1:1:1:1, about 0.5:1:1:1, about 3:1: 1:1, about 6:1:1:1, about 10:1:1:1, about 1:1:1:3, etc., but not limited thereto. In certain embodiments, in the compositions used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal permeation of the present disclosure, ganoderma lucidum, jujube, longan pulp and lotus are used in the above herbal medicine complexes. can be 1:1:1:1. In another specific embodiment, in the composition used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal permeation of the present disclosure, in the above-mentioned Chinese herbal medicine complex, ganoderma lucidum, jujube, longan meat and lotus seed weight ratio can be 3:1:1:1.

In one embodiment, in the composition used for regulating intestinal permeability and/or treating and/or preventing intestinal wall leakage-related diseases of the present disclosure, the form of each drug contained in the above-mentioned Chinese herbal medicine composite drug includes: It may include, but is not limited to, whole herbal medicines, small pieces/small pieces obtained by cutting herbal medicines, powders obtained by pulverizing herbal medicines, etc., or any combination thereof.

Also, in one embodiment, the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure comprises the above-described herbal drug combination, and the intestinal permeability of the present disclosure Compositions for sex regulation and/or treatment and/or prevention of leaky gut related disorders may be packaged in filter bags. In this embodiment, a filter bag packaged with a composition used for regulating intestinal permeability and/or treating and/or preventing intestinal wall leakage-related diseases of the present disclosure is percolated with a solvent, and then an exudate is obtained. In certain embodiments, filter bags packaged with compositions for use in modulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage of the present disclosure can be infused with water and then taken directly. A leachate can be obtained.

Further, in one embodiment, in the composition used for regulating intestinal permeability and/or treating and/or preventing intestinal wall leakage-related diseases of the present disclosure, ganoderma lucidum extract contained in the above-mentioned Chinese herbal medicine complex extract, The weight ratio of ganoderma lucidum, jujube, longan flesh, and lotus seed, which are raw materials necessary for obtaining jujube extract, longan flesh extract, and lotus seed extract, respectively, is about 0.1-15:0.6-2. : 0.6-2: 0.6-5, for example about 0.3-12: 0.8-1.5: 0.8-1.5: 0.8-4, about 0.5-10: 1:1:1 to 3, about 1:1:1:1, about 0.5:1:1:1, about 3:1:1:1, about 6:1:1:1, about 10:1 : 1:1, about 1:1:1:3, etc., but not limited thereto. In a specific embodiment, in the composition used for regulating intestinal permeability and/or treating and/or preventing intestinal wall leakage-related diseases of the present disclosure, ganoderma lucidum extract, jujube extract contained in the above-mentioned herbal medicine complex extract The weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed, which are the raw materials required to obtain the plant, longan flesh extract and lotus seed extract, respectively, can be 1:1:1:1. In another specific embodiment, in the composition used for regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage of the present disclosure, ganoderma lucidum extract contained in the above-mentioned Chinese herbal medicine complex extract, The weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed, which are raw materials required to obtain jujube extract, longan flesh extract and lotus seed extract, respectively, can be 3:1:1:1.

Further, in one embodiment, in the composition for use in regulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage of the present disclosure, in the Chinese herbal complex extract, the ganoderma lucidum extract, the The weight ratio of jujube extract, longan flesh extract and lotus seed extract is about 1-40:20-200:20-180:5-130, such as about 5-35:40-180:40-170: 10-120, about 9.3:150.1:136.8:56.2, about 5.3:172.28:156.37:64.25, about 6.18:100.5:91.21 : 112.45, but is not limited to this.

There is no particular limitation on the method of obtaining the Chinese herbal medicine complex extract in the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure. It is only necessary that the method be such that a mixture of extracts can be obtained.

In one embodiment, the herbal medicine complex extract in the compositions used for modulating intestinal permeability and/or treating and/or preventing leaky gut-related diseases of the present disclosure can be obtained by a method. The method may include, but is not limited to, the following steps.

First, each drug contained in the raw materials used to form the above-mentioned Chinese herbal medicine complex extract is subjected to an extraction process to obtain an extract of each drug. For example, the ganoderma lucidum, jujube, longan flesh and lotus seed contained in the raw materials used to form the above-mentioned herbal medicine complex extract are subjected to an extraction process with a solvent, respectively, to obtain the respective ganoderma lucidum extract, jujube extract, Longan flesh extract and lotus seed extract are obtained.

The resulting extracts of each drug substance are then mixed to form a mixed extract. For example, the above-obtained ganoderma lucidum extract, jujube extract, longan extract and lotus seed extract are mixed to form a mixed extract, and thus the above-mentioned Chinese herbal complex extract is obtained.

The solvents used in each extraction process for each drug substance in the production feedstock may be the same or different, and the solvents used in each extraction process interact with each other in the subsequent step of mixing each extraction liquid. Any solvent may be used as long as it does not affect the components in the extract to be mixed. In certain embodiments, both solvents used in each extraction process for each drug substance in the production feedstock may be water.

The temperature employed for each extraction process for each drug substance in the raw material may be the same or different, is not subject to any particular limitations, and can be adjusted as needed. For example, the temperature to be adopted for each extraction process of each drug substance in the production raw material depends on the environment during the extraction process (e.g. environmental temperature, humidity, pressure, etc.), the type of drug to be extracted, and the type of drug to be extracted. It can be adjusted according to the condition of the drug (eg, moisture content, weight, etc.), the time of extraction to proceed and/or the type of extraction solvent used, etc., but is not limited thereto. For example, the temperature employed in each extraction process for each drug substance in the production raw material is a temperature above the freezing point and below the boiling point of the solvent used in each extraction process. In one embodiment, the solvent used in each extraction process of each drug substance in the make-up feedstock is all water, and the temperature employed in each extraction process of each drug substance in the make-up feedstock is about 0-100° C., such as about 4°C, about 5°C, about 10°C, about 15°C, about 20°C, about 25°C, about 30°C, about 35°C, about 37°C, about 40°C, about 50°C, about 60°C, about 70°C , about 80° C., about 90° C., about 100° C., etc., but are not limited thereto.

Similar to the above, the progress time of each extraction process for each drug substance in the raw material can be the same or different, there is no particular limitation, and can be adjusted as needed. . For example, the progress time of each extraction process for each drug substance in the raw material is determined by the environment during the extraction process (e.g., environmental temperature, humidity, pressure, etc.), the type of drug to be extracted, and the extraction process. It can be adjusted according to the state of the drug (for example, moisture content, weight, etc.), the extraction temperature to be employed and/or the type of extraction solvent used, etc., but is not limited to this. For example, the progress time of each extraction process for each drug substance in the raw material is about 0.1 to 10 hours, such as about 0.1 hours, about 0.5 hours, about 1 hour, about 1.5 hours, about It can be, but is not limited to, 2 hours, about 2.5 hours, about 5 hours, about 8 hours, about 10 hours, and the like. In one embodiment, the solvent used for each extraction process of each drug substance in the make-up feedstock is all water, and the duration of each extraction process of each drug substance in the make-up feedstock is about 0.1 to 10 hours, such as about 0.1 hours, about 0.5 hours, about 1 hour, about 1.5 hours, about 2 hours, about 2.5 hours, about 5 hours, about 8 hours, about 10 hours, etc. It is not limited to this.

Furthermore, the above method of obtaining the Chinese herbal medicine complex extract in the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure, optionally each obtained After the step of mixing the extracts of the ingredients to form a mixed extract, the step of drying the mixed extract may be further included. The drying method is not particularly limited as long as it is a method for drying the extract, and examples thereof include oven drying and freeze drying. In one embodiment, the drying of the mixed extract is performed by freeze-drying.

Also, in another embodiment, the herbal medicine complex extracts in the compositions used for modulating intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage of the present disclosure can be obtained by other methods. can. The method may include, but is not limited to, the following steps.

First, each medicinal material contained in the preparation raw materials used to form the Chinese herbal medicine complex extract is mixed to form a raw material mixture. For example, ganoderma lucidum, jujube, longan pulp and lotus seed, which are included in the raw materials used to form the aforementioned herbal medicine complex extract, are mixed to form a raw material mixture.

The raw material mixture is then subjected to an extraction process to obtain a mixed extract. For example, a raw material mixture containing ganoderma lucidum, jujube, longan flesh and lotus seed is subjected to an extraction process to obtain a mixed extract, and the above-mentioned traditional Chinese medicine complex extract.

The solvent used in the extraction process for the raw material mixture is not particularly limited as long as it does not adversely affect the components in the extract. In certain embodiments, the solvent used in the extraction process for the feed mixture may be water.

The temperature employed in the extraction process for the raw material mixture is also not particularly limited and can be adjusted as required. For example, the temperature employed in the extraction process for the raw material mixture depends on the environment in which the extraction process is carried out (e.g. environmental temperature, humidity, pressure, etc.), the type of drug contained in the raw drug to be extracted, and the type of drug to be extracted. It can be adjusted according to the state of the raw material mixture to be used (eg, moisture content, weight, etc.), the time of extraction to proceed and/or the type of extraction solvent used, etc., but is not limited thereto. For example, the temperature employed in the extraction process for the raw material mixture is above the freezing point and below the boiling point of the solvent used in the extraction process. In one embodiment, the solvent used in the extraction process for the feed mixture is water and the temperature employed in the extraction process for the feed mixture is about 0-100°C, such as about 4°C, about 5°C, about 10°C, about 15°C. , about 20°C, about 25°C, about 30°C, about 35°C, about 37°C, about 40°C, about 50°C, about 60°C, about 70°C, about 80°C, about 90°C, about 100°C, etc. can be, but is not limited to.

Similar to the above, the duration of each extraction process for each drug substance in the raw material is not particularly limited and can be adjusted as needed. For example, the progress time of each extraction process for each drug substance in the raw material to be produced depends on the environment during the extraction process (e.g. environmental temperature, humidity, pressure, etc.), and the drug substances contained in the raw drug substance to be extracted. , the state of the raw material mixture to be extracted (e.g., water content, weight, etc.), the extraction temperature employed and/or the type of extraction solvent used, etc., but is not limited thereto. For example, the duration of each extraction process for each drug substance in the raw material to be made is about 0.1 to 10 hours, such as about 0.1 hours, about 0.5 hours, about 1 hour, about 1.5 hours, It may be about 2 hours, about 2.5 hours, about 5 hours, about 8 hours, about 10 hours, etc., but is not so limited. In one embodiment, the solvent used for each extraction process of each drug substance in the raw material is water, and the duration of each extraction process of each drug substance in the raw material is about 0.1 to 10 hours, For example, about 0.1 hours, about 0.5 hours, about 1 hour, about 1.5 hours, about 2 hours, about 2.5 hours, about 5 hours, about 8 hours, about 10 hours. , but not limited to.

Furthermore, the above method of obtaining the Chinese herbal complex extract in the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure optionally includes each of the obtained After the step of mixing the extracts of the raw materials to form a mixed extract, the step of drying the mixed extract may be further included. The drying method is not particularly limited as long as it can dry the extract, and examples thereof include oven drying and freeze drying. In one embodiment, the drying of the mixed extract is performed by freeze-drying.

In one embodiment, the composition used for the regulation of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases of the present disclosure described above includes the above-mentioned Chinese herbal medicine complex drug or Chinese herbal medicine complex extract, as well as pharmaceutical In this embodiment, the above-described modulating intestinal permeability and/or treatment of leaky-related diseases and/or Alternatively, in the composition used for prevention, the content of the herbal compound drug or herbal compound extract may be about 10 to 99.5 wt%, such as about 10 to 50 wt%, about 50 to 99.5 wt%, etc. is not limited to In this embodiment, the compositions used for regulating intestinal permeability and/or treating and/or preventing intestinal wall seepage-related diseases of the present disclosure described above may be pharmaceutical compositions or healthcare compositions, It is not limited to this.

Pharmaceutically acceptable carriers include, but are not limited to, pharmaceutical agents such as solvents, dispersion media, coatings, antibacterial and antifungal agents, and isotonic and absorption delaying agents. compatible with therapeutic administration. Pharmaceutical compositions can be formulated into dosage forms for different modes of administration using conventional methods.

Also, the pharmaceutically acceptable salts include, but are not limited to, inorganic cations, inorganic protons, alkali metal salts such as sodium salts, potassium salts or amine salts, such as magnesium salts, calcium salts. Salts containing divalent or tetravalent cations such as alkaline earth metal salts, zinc salts, aluminum salts or zirconium salts may be included. It may also be an organic salt such as dicyclohexylamine salt, methyl-D-glucamine, arginine, lysine, histidine, amino acid salt such as glutamine.

Also, examples of compositions for use in modulating intestinal permeability and/or treating and/or preventing intestinal leakage-related diseases of the present disclosure include, but are not limited to, pharmaceutical compositions or healthcare compositions. obtain.

The pharmaceutical or healthcare compositions described in this disclosure may be administered parenterally, orally, by inhalation spray, or via an implanted reservoir. Parenterally includes application to any part of the skin or as needed, subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, Intrasynovial, intrasternal intrathecal, intralesional injection, and infusion techniques may be included.

Oral forms of the pharmaceutical or healthcare compositions described in this disclosure include tablets, granules, powders, pellet-in-capsules, capsules, coated tablets, emulsions, solutions, aqueous suspensions. may include, but are not limited to, aqueous suspensions, dispersions, instant powders, and the like.

In certain embodiments, the compositions of the present disclosure for use in modulating intestinal permeability and/or treating and/or preventing leaky gut-related diseases can be pharmaceutical compositions. In this particular embodiment, the pharmaceutical composition may be in an oral dosage form, which includes tablets, granules, powders, pellets-in-capsules, capsules, coated tablets, emulsions, solutions, It may include, but is not limited to, aqueous suspensions, dispersions, and the like. Also, in this particular embodiment, the pharmaceutical compositions include, but are not limited to, pharmaceutical compositions for use in reducing and/or treating and/or preventing inflammatory bowel disease, neurodegenerative diseases and/or sleep disorders. can be

In another specific embodiment, the composition of the present disclosure for use in modulating intestinal permeability and/or treating and/or preventing leaky gut-related diseases can be a healthcare composition. In this particular embodiment, the health care composition may be a health care food, and health care food formats include tablets, granules, powders, pellets-in-capsules, capsules, coated tablets, emulsions, solutions. can include, but are not limited to, formulations, aqueous suspensions, dispersions, instant powders, and the like. Also, in this particular embodiment, the health care compositions may include, but are not limited to, health care compositions that prevent and/or ameliorate inflammatory bowel disease, neurodegenerative diseases and/or sleep disorders. However, it is not limited to this.

Based on the above, the present disclosure is used to prepare compositions of Chinese herbal medicine complexes or Chinese herbal complex extracts for use in modulating intestinal permeability and/or treating and/or preventing intestinal wall seepage-related diseases. Applications can also be provided.

The present disclosure can also provide use of the herbal medicinal complexes or herbal medicinal complex extracts to make compositions for use in reducing and/or treating and/or preventing inflammatory bowel disease. Inflammatory bowel diseases as referred to herein may include, but are not limited to, colitis and the like. In one embodiment, the inflammatory bowel disease referred to herein is colitis.

The present disclosure can also provide uses of the herbal medicinal complexes or herbal medicinal complex extracts for use in making compositions for reducing and/or treating and/or preventing neurodegenerative diseases. Neurodegenerative diseases referred to herein may include, but are not limited to, Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis or a combination thereof, and the like. In one embodiment, the inflammatory bowel disease described herein is Alzheimer's disease.

The present disclosure can also provide uses of the herbal medicinal complexes or herbal medicinal complex extracts for use in making compositions for reducing and/or treating and/or preventing sleep disorders. Sleep disorders as referred to herein may include, but are not limited to, chronic sleep fragmentation, insomnia or a combination thereof. In one embodiment, the sleep disorder described herein is insomnia.

In each of the uses of the Chinese herbal medicine complex or the Chinese herbal complex extract of the present disclosure described above, the Chinese herbal medicine complex or the Chinese herbal complex extract has a modulating effect on intestinal permeability and/or intestinal wall leakage-related It has a therapeutic and/or prophylactic effect on diseases.

In addition, all descriptions of the Chinese herbal medicine complex or the Chinese herbal complex extract according to each of the above-mentioned uses of the present disclosure refer to the above-mentioned regulation of intestinal permeability and/or treatment of intestinal wall leakage-related diseases and/or The description of the Chinese herbal compound or the Chinese herbal compound extract in the paragraph describing the composition for prevention can be referred to and will not be repeated here.

In one embodiment, in each of the above uses of the herbal medicinal complex or herbal medicinal complex extract of the present disclosure, the pharmaceutically acceptable carrier or salt is also used to control intestinal permeability regulation and/or intestinal wall leakage-related diseases. can be used to make a composition for the treatment and/or prevention of

Also for the description of pharmaceutically acceptable carriers or salts, in the paragraph describing compositions used for modulating intestinal permeability and/or treating and/or preventing intestinal wall seepage-related diseases of the present disclosure, as well. Relevant descriptions can be referred to and will not be repeated here.

In addition, all relevant descriptions of the compositions made in each of the applications of the present disclosure set forth above are directed to the modulation of intestinal permeability and/or treatment and/or prevention of intestinal wall seepage-related disorders of the present disclosure set forth above. Reference may be made to, but not limited to, all descriptions in the paragraphs describing the compositions used.

Based on the foregoing, the present disclosure can also provide methods of modulating intestinal permeability and/or treating and/or preventing leaky gut-related disorders. The method includes administering, but not limited to, any of the compositions of the present disclosure for use in modulating intestinal permeability and/or treating and/or preventing a leaky gut-related disease to an individual in need thereof. can include doing

Based on the foregoing, the present disclosure can also provide methods of treating inflammatory bowel disease. The method includes administering, but not limited to, any of the compositions of the present disclosure for use in modulating intestinal permeability and/or treating and/or preventing a leaky gut-related disease to an individual in need thereof. can include doing Inflammatory bowel diseases as referred to herein may include, but are not limited to, colitis and the like.

Based on the foregoing, the present disclosure can also provide methods of treating neurodegenerative diseases. The method includes administering, but not limited to, any of the compositions of the present disclosure for use in modulating intestinal permeability and/or treating and/or preventing a leaky gut-related disease to an individual in need thereof. can include doing Neurodegenerative diseases referred to herein may include, but are not limited to, Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis or a combination thereof, and the like.

Based on the foregoing, the present disclosure can also provide methods of treating sleep disorders. The method includes, but is not limited to, administering any of the compositions of the present disclosure for use in modulating intestinal permeability and/or treating and/or preventing a leaky gut-related disease to an individual in need thereof. administering. Sleep disorders as referred to herein may include, but are not limited to, chronic sleep fragmentation, insomnia or a combination thereof.

Individuals described in this disclosure may include, but are not limited to, vertebrates. The vertebrates may include, but are not limited to, fish, amphibians, reptiles, birds or mammals. Examples of mammals can include, but are not limited to, humans, orangutans, monkeys, cows, horses, donkeys, dogs, cats, rabbits, guinea pigs, rats, mice, and the like. In one embodiment, the individual is human.

Example

Example 1: Preparation of test samples

40 g of Ganoderma lucidum, 40 g of ash jujube, 40 g of powdered husk longan flesh and 40 g of red lotus seeds were refluxed and extracted with 400 g of water for 2 hours, respectively. ), longan flesh extract (iii) (286 g) and lotus seed extract (iv) (313 g), respectively.

The total weight of the various medicinal materials in the raw materials to be used is 1 g, and the amount of the extract of the various medicinal materials is mixed according to the weight corresponding to the content (ratio) in the raw materials. 16 test samples were prepared as shown in .

(Table 1)

For example, 1/40 of the weight (241 g x (1/40) = 6.025 g) of a single-component rouge reishi sample, that is, the rouge reishi extract (i) obtained above (241 g x (1/40) = 6.025 g) (corresponding to 1 g of Kazuno Reishi medicinal material), and freeze-dried.

For example, a combination sample of two kinds of composite components, Reishi mushroom and Jujube, that is, the extract (i) of Reishi mushroom obtained above is added to 1/80 of its weight (241 g x (1/80) = 3 013 g), and 1/80 of the weight of the jujube extract (ii) obtained above (338 g x (1/80) = 4.225 g) was taken (0.5 g of Kazuno Reishi medicinal material). and 0.5 g of jujube medicinal materials, the total weight of the medicinal materials is 1 g), both were well mixed and then freeze-dried.

For example, a combination sample of three kinds of composite components, Reishi mushroom, jujube and longan meat, that is, the Reishi mushroom extract (i) obtained above is added in an amount of 1/120 of its weight (241 g × (1/120) = 2.008 g), the jujube extract (ii) obtained above was taken in an amount of 1/120 of its weight (338 g x (1/120) = 2.817 g), and the above obtained 1/120 of its weight (286 g x (1/120) = 2.383 g) of longan extract (iii) was taken (0.33 g of reishi mushroom extract, 0.33 g of jujube extract, and 0.33 g of longan extract). Corresponding to 0.33 g, the total weight of the drug is 1 g), the three were thoroughly mixed and then lyophilized to obtain the desired sample.

A sample of a combination of the four composite components, rouge reishi, jujube, longan flesh and lotus seed (1:1:1:1), that is, the rouge reishi extract (i) obtained above was added to 1 of its weight. /160 (241 g x (1/160) = 1.506 g), and the jujube extract (ii) obtained above is added in an amount of 1/160 of its weight (338 g x (1/160) = 2 .113 g), 1/160 of its weight (286 g x (1/160) = 1.788 g) of the longan flesh extract (iii) obtained above, and the lotus root obtained above 1/160 of its weight (313 g x (1/160) = 1.956 g) of fruit extract (iv) was taken (0.25 g of reishi medicinal agent, 0.25 g of jujube medicinal agent, longan meat medicinal agent corresponding to 0.25g and 0.25g lotus seed medicinal, the total weight of the medicinal is 1g).

Sixteen test samples, shown in Table 1, were made according to the rules set forth above.

The dry weight of each medicinal extract contained in each test sample can be estimated, for example, by the estimation methods shown in Tables 2 to 5 below. Also, based on the dry weight of each extract obtained corresponding to 1 g of each drug substance listed in Tables 2 to 5 below, the extraction rate of each extract can be calculated by the following formula.
Extraction rate (%) = dry weight of extract/weight of corresponding drug × 100

Extraction rates for each extract are as follows:

Extraction rate of Reishi extract: 3.71% (0.0371g/1g x 100)

Extraction rate of jujube extract: 60.3% (0.603 g/1 g x 100)

Extraction rate of longan meat extract: 54.73% (0.5473g/1g×100)

Extraction rate of lotus seed extract: 22.49% (0.225 g/1 g x 100)

注：試験サンプル６の乾燥重量２９２．２ｍｇ≒それに含まれる霊芝抽出物の推定乾燥重量１８．６ｍｇ＋それに含まれる竜眼肉抽出物の推定乾燥重量２７３．７ｍｇ (Table 2) Exemplary method for estimating the dry weight of each medicinal extract contained in a two-component test sample

Note: 292.2 mg dry weight of test sample 6 ≈ 18.6 mg estimated dry weight of ganoderma lucidum extract it contains + 273.7 mg estimated dry weight of longan extract it contains

注：試験サンプル１１の乾燥重量３９５．８ｍｇ≒それに含まれる霊芝抽出物の推定乾燥重量１２．４ｍｇ＋それに含まれる棗抽出物の推定乾燥重量２０１ｍｇ＋それに含まれる竜眼肉抽出物の推定乾燥重量１８２．４ｍｇ (Table 3) Exemplary method for estimating the dry weight of each medicinal extract contained in a three-component test sample

Note: 395.8 mg dry weight of test sample 11 ≈ estimated dry weight of ganoderma lucidum extract it contains 12.4 mg + estimated dry weight of jujube extract it contains 201 mg + estimated dry weight of longan extract it contains 182.4 mg

注：試験サンプル１５Ａの乾燥重量３５２．４ｍｇ≒それに含まれる霊芝抽出物の推定乾燥重量９．３ｍｇ＋それに含まれる棗抽出物の推定乾燥重量１５０．１ｍｇ＋それに含まれる竜眼肉抽出物の推定乾燥重量１３６．８ｍｇ＋それに含まれる蓮の実抽出物の推定乾燥重量５６．２ｍｇ (Table 4) Exemplary method for estimating the dry weight of each medicinal extract contained in a four-component test sample

Note: 352.4 mg dry weight of test sample 15A ≈ 9.3 mg estimated dry weight of ganoderma lucidum extract it contains + 150.1 mg estimated dry weight of jujube extract it contains + 136 estimated dry weight of longan extract it contains .8mg + estimated dry weight of lotus seed extract it contains 56.2mg

注：試験サンプル１５Ｂの乾燥重量２４７．８ｍｇ≒それに含まれる霊芝抽出物の推定乾燥重量１８．６ｍｇ＋それに含まれる棗抽出物の推定乾燥重量１００．５ｍｇ＋それに含まれる竜眼肉抽出物の推定乾燥重量９１．２ｍｇ＋それに含まれる蓮の実抽出物の推定乾燥重量３７．５ｍｇ (Table 5) Exemplary method for estimating the dry weight of each medicinal extract contained in the four-component test sample

Note: 247.8 mg dry weight of test sample 15B ≈ estimated dry weight of ganoderma lucidum extract it contains 18.6 mg + estimated dry weight of jujube extract it contains 100.5 mg + estimated dry weight of longan extract it contains 91 .2mg + estimated dry weight of lotus seed extract it contains 37.5mg

Example 2

A. Method

A cytotoxic concentration test was performed on the 16 types of test samples prepared above.

Cytotoxic concentration testing of test samples was performed with human colon adenocarcinoma cells Caco-2.

Cells were divided into a negative control group (NC) and 16 sets of experimental groups each using a different test sample. The negative control group was untreated cells, and for each experimental group, the test sample was diluted from the highest concentration of 200 μg/mL to each test concentration by two-fold dilution (the test sample was first diluted with ddH ₂ O). 100 mg/mL, then diluted with cell culture medium to a concentration of 200 μg/mL, then further diluted to each test concentration), and different concentrations of test samples were added to human colon adenocarcinoma cells Caco-2 and 96, respectively. Placed in a well culture plate and co-cultured for 48 hours.

Then, 0.5 mg/mL of MTT was added to each of the negative control group and each experimental group and cultured for 4 hours, after which the medium was removed, the blue-violet Formazan crystals were dissolved in DMSO, and the absorbance value at 570 nm was measured.

Cell viability was evaluated according to the formula shown below.
Cell viability = (570 nm absorbance value of experimental group/570 nm absorbance value of negative control group (NC)) x 100%

B. result

The results are as shown in Figures 1A to 1P.

From Figures 1A to 1P, it is possible to know the maximum concentration that can be used in the absence of cell death for each sample shown in Table 6.

(Table 6) Maximum concentration of each sample that can be used without cell death

Example 3: Effect of single component extract on intestinal epithelial cell permeability

A. Method

Using an intestinal permeability system constructed with human intestinal epithelial cells, the effect of each extract obtained in Example 1 on intestinal permeability was evaluated (Pham VT, Seifert N, Richard N, et al. The effects of fermentation products of prebiotic fibers on gut barrier and immune functions in vitro [published correction appears in PeerJ. 2018 Aug 17;6:. Steinert, Robert [corrected to Steinert, Robert E]]. PeerJ. 2018;6:e5288. Published 2018 Aug 10 doi:10.7717/peerj.5288). Detailed implementation steps are as follows.

After culturing human colon adenocarcinoma cells Caco-2 in a transwell culture plate for 21 days, the cells were divided into a negative control group (NC), a positive control group (PC), and four experimental groups. We performed an in vitro intestinal wall leakage platform test. Negative controls are untreated cells. The positive control group was cells treated with 50 μM berberine chloride (Valenzano MC, DiGuilio K, Mercado J, Teter M, To J, Ferraro B, et al. (2015) Remodeling of Tight Junctions and Enhancement of Barrier Integrity of the CACO-2 Intestinal Epithelial Cell Layer by Micronutrients. PLoS ONE 10(7): e0133926.).

Four sets of experimental groups, respectively, sample 1, sample 2, sample 3 and sample 4 at concentrations that do not cause cell death (sample 1: 3 μg/mL; sample 2: 100 μg/mL; sample 3: 100 μg/mL; Sample 4: 100 μg/mL) (samples were first prepared with ddH ₂ O to a concentration of 100 mg/mL, then diluted with cell culture medium to a concentration of 200 μg/mL, then further diluted to each test concentration). bottom).

After the above treatment, the cells of each group were cultured for 48 hours, then the cells were induced with 350 mg/mL of Rhamnolipids to induce cell permeabilization.

Next, FITC-dextran 4 (FD4) fluorescent dye was added to the inner plate of the transwell culture plate and allowed to react for 4 hours, after which the liquid in the lower well of the culture plate was sucked and the fluorescence intensity was detected. (excitation light wavelength: 485 nm; emission light wavelength: 538 nm).

Intestinal cell permeability was assessed by the fluorescence intensity of the liquid obtained from the lower wells, based on the formula shown below.
FD4 leakage rate = (FD4 fluorescence value of test sample/FD4 fluorescence value of negative control group (NC)) x 100%

B. result

The results are as shown in FIG.

Figure 2 shows that sample 1 (reishi extract), sample 2 (jujube extract), sample 3 (longan extract) and sample 4 (lotus seed extract) all significantly reduced the FD4 leakage rate. This indicates that all of Samples 1 to 4 have the activity of suppressing intestinal permeation.

Example 4: Effect of multicomponent extract on intestinal epithelial cell permeability

A. Method

Using an intestinal permeability system constructed with human intestinal epithelial cells, the effect of each extract obtained in Example 1 on intestinal permeability was evaluated. Detailed implementation steps are as follows.

After culturing human colon adenocarcinoma cells Caco-2 in transwell culture plates for 21 days, the cells were divided into a negative control group (NC), a positive control group (PC), and 9 sets of experimental groups, and subjected to in vitro intestinal wall A seepage platform test was performed. Negative controls are untreated cells. The positive control group is cells treated with 50 μM berberine chloride. The nine experimental groups are cells treated with 1.5 μg/mL Sample 1, Sample 2, Sample 3, Sample 4, Sample 6, Sample 9, Sample 11, Sample 12 and Sample 15A, respectively.

After the above treatment, the cells in each group were cultured for 48 hours, and then the cells were induced to permeabilize with rhamnolipid 350 mg/mL.

Next, FITC-dextran 4 (FD4) fluorescent dye was added to the inner plate of the transwell culture plate and allowed to react for 4 hours, after which the liquid in the lower wells of the culture plate was aspirated and the fluorescence intensity was detected. (excitation light wavelength: 485 nm; emission light wavelength: 538 nm).

Intestinal cell permeability was assessed by the fluorescence intensity of the liquid obtained from the lower wells, based on the formula shown below.
FD4 leakage rate = (FD4 fluorescence value of test sample/FD4 fluorescence value of negative control group (NC)) x 100%

B. result

The results are as shown in FIG.

FIG. 3 shows the concentration of sample 2 (jujube extract), sample 3 (longan flesh extract) and sample 4 (lotus seed extract) lowered from the concentration of 100 μg/mL employed in Example 3 to 1.5 μg/mL. Samples 2, 3 and 4 did not significantly reduce the FD4 leakage rate after 20 minutes, indicating that only Sample 1 was able to significantly reduce the FD4 leakage rate.

In addition, FIG. 3 shows sample 12 (reishi mushroom extract + jujube extract + lotus seed extract (the weight ratio of each medicinal raw material to obtain each extract is 1: 1:1)) and sample 15A (reishi mushroom extract + jujube extract + longan flesh extract + lotus seed extract (the weight ratio of each medicinal raw material to obtain each extract is 1:1:1:1). )) was able to significantly reduce the FD4 leakage rate. At the same working concentration (1.5 μg/mL), Sample 15A was able to inhibit the FD4 leakage rate to a greater extent than Samples 1 and 12, indicating that Sample 15A was superior to intestinal penetration. It shows that it has a high activity.

Example 5: Effect of different ratios of four complexes on intestinal epithelial cell permeability

A. Method

Using an intestinal permeability system constructed with human intestinal epithelial cells, the effect of each extract obtained in Example 1 on intestinal permeability was evaluated. Detailed implementation steps are as follows.

After culturing human colon adenocarcinoma cells Caco-2 in transwell culture plates for 21 days, the cells were divided into a negative control group (NC), a positive control group (PC), and 6 sets of experimental groups, and were subjected to ex vivo intestinal wall A seepage platform test was performed.

In this experiment, four batches were run. In each batch, the negative controls were untreated cells and the positive controls were cells treated with 50 μM berberine chloride.

In the first batch, six experimental groups are cells treated with 1.5 μg/mL of Sample 1, Sample 15A, Sample 15B, Sample 15C, Sample 15D and Sample 15E, respectively.

In the second batch, six experimental groups are cells treated with 1.5 μg/mL of Sample 1, Sample 15A, Sample 15F, Sample 15G, Sample 15H and Sample 15I, respectively.

In a third batch, six experimental groups are cells treated with 1.5 μg/mL Sample 1, Sample 15A, Sample 15J, Sample 15K, Sample 15L and Sample 15M, respectively.

In the fourth batch, six experimental groups are cells treated with 1.5 μg/mL Sample 1, Sample 15A, Sample 15N, Sample 15O, Sample 15P and Sample 15Q, respectively.

After the above treatment, the cells in each group were cultured for 48 hours, and then the cells were induced to permeabilize with 350 mg/mL of rhamnolipid.

Next, FITC-dextran 4 (FD4) fluorescent dye was added to the inner plate of the transwell culture plate and allowed to react for 4 hours, after which the liquid in the lower wells of the culture plate was aspirated and the fluorescence intensity was detected. (excitation light wavelength: 485 nm; emission light wavelength: 538 nm).

Intestinal cell permeability was assessed by the fluorescence intensity of the liquid obtained from the lower wells, based on the formula shown below.
FD4 leakage rate = (FD4 fluorescence value of test sample/FD4 fluorescence value of negative control group (NC)) x 100%

B. result

The results of the first batch experiment, the second batch experiment, the third batch experiment, and the fourth batch experiment are shown in FIGS. 4A, 4B, 4C, and 4D, respectively. .

As can be seen from FIGS. 4A, 4B, 4C and 4D, sample 15A (reishi mushroom extract + jujube extract + longan extract + lotus seed extract (each drug used to obtain each extract) The weight ratio of raw materials is 1:1:1:1)) and sample 15B (reishi mushroom extract + jujube extract + longan extract + lotus seed extract (each medicinal raw material used to obtain each extract). The weight ratio of 3:1:1:1)) has excellent activity of suppressing intestinal permeation, and sample 15A was able to suppress intestinal permeation more remarkably than sample 15B.

Example 6: Effect of four complex component extracts on intestinal epithelial cell permeability

Sample 15B (reishi mushroom extract + jujube extract + longan extract + lotus seed extract (used for obtaining each extract) obtained in Example 1 was confirmed by an intestinal permeability system constructed with human intestinal epithelial cells. The weight ratio of each medicinal raw material used was 3:1:1:1)), and the effect on intestinal permeability was evaluated. Detailed implementation steps are as follows.

After culturing human colon adenocarcinoma cells Caco-2 in transwell culture plates for 21 days, the cells were divided into 2 negative control groups (NC), 2 positive control groups (PC) and 6 experimental groups. We performed an in vitro intestinal wall leakage platform test. Two sets of negative controls are untreated cells. Two sets of positive controls are cells treated with 50 μM berberine chloride. Six experimental groups were treated with Sample 15B at concentrations of 0.3 μg/mL, 0.3 μg/mL, 0.6 μg/mL, 0.6 μg/mL, 1.5 μg/mL and 1.5 μg/mL, respectively. are cells.

After the above treatment, the cells of each group were cultured for 48 hours, and then, for half of the groups in the experiment (including the negative control group, the positive control group and the experimental group treated with three concentrations of Sample 15B), Cell permeabilization was induced with rhamnolipid at 350 mg/mL, the cells in the other half of the group in the experiment were not treated with rhamnolipid.

Next, FITC-dextran 4 (FD4) fluorescent dye was added to the inner plate of the transwell culture plate and allowed to react for 4 hours, after which the liquid in the lower wells of the culture plate was aspirated and the fluorescence intensity was detected. (excitation light wavelength: 485 nm; emission light wavelength: 538 nm).

Intestinal cell permeability was assessed by the fluorescence intensity of the liquid obtained from the lower wells, based on the formula shown below.

FD4 leakage rate=(FD4 fluorescence value of test sample/FD4 fluorescence value of negative control group (NC))×100%.

B. result

The results are as shown in FIG. None of the cells in the group not treated with rhamnolipid showed any distinct fluorescence intensity. Among the negative control groups, the fluorescence intensity of the rhamnolipid-induced group was significantly enhanced compared to the group not induced with rhamnolipids. Concentration sample 15B (Ganoderma lucidum extract + jujube extract + longan flesh extract + lotus seed extract (the weight ratio of each medicinal raw material used to obtain each extract is 3:1:1:1)) All of them were able to significantly reduce the fluorescence intensity, which indicates that they have activity to suppress intestinal permeation.

Example 7: Effect of four complex component extracts on intestinal epithelial cell tight junction protein gene expression level

A. Method

The cells of the negative control group, positive control group and experimental group (treated with sample 15B at a concentration of 1.5 μg/mL) in the rhamnolipid-induced group of Example 6 were collected respectively.

Total RNA of cells was isolated by Total RNA Purification Kit (GeneMark, Taichung, Taiwan). The resulting total RNA was subjected to reverse transcription-polymerase chain reaction to reverse transcribe messenger RNA (mRNA) into cDNA using the Maxima First Strand cDNA Synthesis Kit (Thermo Fisher Scientific) according to the manufacturer's protocol.

Real-time quantitative polymerase chain reaction was then performed on the cDNA samples to measure the expression levels of CLDN3, OCLN and TJP1 mRNA. Differences in gene expression were calculated by the relative quantitative data analysis 2- ^(ΔΔCT) method, and (Glyceraldehyde 3-phosphate dehydrogenase, GAPDH) was used as a reference gene. The primers used for each gene are shown in Table 7.

(Table 7) Primers used for each gene

B. result

The results are shown in Figures 6A, 6B and 6C. As shown in the results, both the positive control group and the experimental group were able to significantly increase the expression levels of the genes CLDN3, OCLN and TJP1 compared to the negative control group.

Example 8: Effect of four complex component extracts on Dextran Sulfate Sodium Salt (DSS)-induced abnormal intestinal permeability in animals

A. Method

Animals used in this experiment are 5-6 week old C57BL/6 male mice (purchased from the National Laboratory Animal Center).

Dextran Sulfate Sodium Salt (DSS)-induced intestinal permeability in an animal model (Laroui H, Ingersoll SA, Liu HC, Baker MT, Ayyadurai S, et al. (2012) Dextran Sodium Sulfate (DSS) Induces Colitis in Mice by Forming Nano-Lipocomplexes with Medium-Chain-Length Fatty Acids in the Colon. PLoS ONE 7(3): e32084. doi:10.1371/journal.pone.0032084).

Mice were divided into an untreated group (Naive), a negative control group (vehicle: water), a positive control group (5-aminosalicylic acid, 5-ASA) and three sets of experimental groups (Sample 15B (reishi extract)). (The weight ratio of each medicinal raw material used to obtain each extract is 3:1:1:1))). Mice in the untreated group received no treatment. Mice in the negative control group were given water containing 0.5% (v/v) dextran sulfate sodium (MP Biomedicals) to induce the development of intestinal permeability defects; 0.5% (v/v) sodium dextran sulfate and 200 μM 5-aminosalicylic acid. Three experimental groups of mice received 0.5% (v/v) dextran sulfate sodium and 100 mg/kg sample 15B, 0.5% (v/v) dextran sulfate sodium and 200 mg/kg sample 15B, and 0.5% (v/v) dextran sulfate sodium and 400 mg/kg of sample 15B, respectively.

The experimental period was 14 days. On the day of termination of the experiment, mice were fasted for 3-4 hours and then given 500 mg/kg FITC-dextran (FD4, Sigma). Two hours later, mice were sacrificed, blood was collected and serum was separated.

FITC-dextran content in serum of mice was determined by a multifunctional microplate spectrophotometer (Molecular Device, FlexStation® 3) and served as an index of intestinal permeability.

B. result

As shown in FIG. 7, the serum FITC-dextran concentration in the negative control group treated with 0.5% (v/v) DSS was significantly increased compared to the untreated group, indicating that It shows that 0.5% (v/v) dextran sulfate sodium could induce an increase in mouse intestinal permeability. In addition, compared with the negative control group, both the positive control group and the experimental group were able to reduce the serum FITC-dextran concentration, which indicated that sample 15B (reishi extract + jujube extract + longan extract + lotus seed extract (the weight ratio of each drug substance used to obtain each extract is 3:1:1:1)) can improve the intestinal permeability increase induced by sodium dextran sulfate. showing.

Example 9: Effect of four complex component extracts on circadian rhythm disruption animal models

A. Method

Animals used in this experiment are 8-week-old C57BL/6 male mice (purchased from the National Laboratory Animal Center).

Mice were operated on to attach electrodes to the skull and record electroencephalograms. After a recovery period of 7 days, the mice were maintained for 2 weeks under a normal light/dark cycle of 12 hours:12 hours, and the changes in the brain waves of the mice for 24 hours were recorded as a baseline.

Mice were then divided into an untreated group (Naive), a negative control group (Vehicle), a positive control group (Diazepam) and an experimental group (Sample 15B (Ganoderma lucidum extract + jujube extract + longan flesh extract + lotus seed extract). (The weight ratio of each drug substance used to obtain each extract is 3:1:1:1))) Mice in the untreated group continued to have a normal light-dark cycle (12 hours light-dark cycle). The mice in the negative control group, the positive control group, and the experimental group were raised for 7 days under conditions of a light/dark cycle of 7 hours: 7 hours to induce circadian disruption. During the breeding period, mice in the negative control group were given the same amount of sterilized water, mice in the positive control group were given diazepam (orally 2.5 mg/kg), Mice were given 200 mg/kg of sample 15B.

On the 4th day after the circadian rhythm was disturbed, changes in the brain waves of the mice were recorded for 24 hours, and the rapid eye movement period (REM) and the non-rapid eye movement period (NREM) were recorded. ) were analyzed for sleep cycle parameters, such as the proportion (%) of time.

B. result

The results are as shown in FIG. As shown in the results, compared with the untreated group, both the ratio of the rapid eye movement period and the ratio of the non-rapid eye movement period of the negative control group were significantly decreased, indicating that , indicating that circadian rhythm disruption could induce insomnia in mice. In addition, compared to the negative control group, both the positive control group and the experimental group were able to significantly increase the ratio of the time during the non-rapid eye movement period, and had the effect of alleviating insomnia in mice. rice field.

Example 10: Effect of four complex component extracts on amyloid β-induced animal learning and manipulative deficits

A. Method

An animal model in which senile plaque-like deposits were formed in the brain of animals induced by β-amyloid (Kim, H.Y., Lee, D.K., Chung, B.R., Kim, H.V., Kim, Y. Intracerebroventricular Injection of Amyloid- β Peptides in Normal Mice to Acutely Induce Alzheimer-like Cognitive Deficits. J. Vis. Exp. (109), e53308, doi:10.3791/53308 (2016). . The experimental steps are as follows:

The animals used in this experiment were male Sprague-Dawley rats (weighing 300-330 g, purchased from Lesco).

Rats were divided into Sham group, control group and experimental group. Rats in the Sham group were injected with artificial cerebrospinal fluid, rats in the control group were injected with 2.5 μL of β-amyloid β peptide 1-40, and rats in the experimental group were injected with 2.5 μL of β-amyloid peptide 1-40. Amyloid peptide 1-40 was injected and sample 15B was orally administered (127 mg/kg/day) for 30 days.

On days 22-23 of the experiment, tests were performed to assess motor activity and exploratory behavior.

The assessment of motor activity and exploratory behavior is described below:

An evaluation test of motor activity and exploratory behavior of rats was carried out in an experimental box. The experiment box has a size of 40 cm in length, width and height. Also, the experiment box has a bottom plate made of stainless steel, and the bottom plate has 16 holes with a diameter of 3 cm, which are arranged in a 4×4 matrix, and the distance between two adjacent holes is 4 cm. , the distance between each side of the matrix and the hole is 7 cm. Rat movements were detected and recorded by sensors TruScan Line E63-01HS, TruScan Sensor E63-22 (Coulbourn Instruments International Corporation) and analyzed with analysis software Coulbourn Instruments'The Habitest System. The measurement time for each rat was 15 minutes. Measures include time spent on the hole, number of punches, and search ability.

Water maze experiments were performed on days 24-28 of the experiment.

Describe the water maze experiment as follows:

The pool was divided into 4 quadrants and a safety platform was fixed in the 4th quadrant. Rats were placed in a pool (each rat was placed in a different quadrant for each experiment) and trained twice daily for 2 minutes each time. If the rat found the safe platform within 2 minutes, it was allowed to rest for 30 seconds, removed from the pool and rested for 30 seconds before the next training session. If the rat did not find the safety platform within 2 minutes, it was placed on the safety platform, allowed to rest for 30 seconds, then removed from the pool and allowed to rest for 30 seconds before the next training; I trained for 4 days.

A water maze spatial performance in Morris water maze test and a water maze non-spatial performance in Morris water maze test were then performed, respectively. In the water maze spatial performance test, a reference point was established at the relative position of the safety platform platform, then the rat was placed in the first quadrant and the time it took for the rat to reach the original safety platform in the pool was recorded. bottom. For the water maze non-spatial performance test, after removing the reference point, the rat was placed in the first quadrant and the time it took the rat to reach the original safety platform in the pool was recorded.

The day after behavioral testing of all animals was completed, hippocampus and frontal cortex regions of the brain were removed for AChE activity analysis and protein content measurements.

B. result

Figures 9A, 9B and 9C show the results of evaluation tests of motor activity and exploratory behavior in rats. The results showed that the control group rats given β-amyloid peptide 1-40 had reduced time spent in the hole, number of hole pokes, and exploration ability compared to the Sham group rats. In contrast, in the experimental group, sample 15B ameliorated the time spent in the hole, the number of punches in the hole, and the decline in exploration ability induced by β-amyloid peptide 1-40 in rats, and improved the time spent in the hole in rats. It has been shown that the number of punches in the hole and the searching ability could be improved.

Figures 10 and 11 show the results of the rat water maze test.

Figure 10 shows the time rats found the safe platform in the water maze spatial performance test. As shown in FIG. 10, rats in the control group given β-amyloid peptide 1-40 had a significantly longer time to reach the safety platform of the pool compared to rats in the Sham group. In contrast, the experimental group showed that sample 15B was able to ameliorate the spatial performance impairment that appeared on days 1 and 2 caused by rat β-amyloid peptide 1-40 (i.e. , reducing the time it takes to reach the safety platform).

Figure 11 shows the time the rats reached the safety platform in the water maze non-spatial performance test.

Compared to rats in the Sham group, rats in the control group given β-amyloid peptide 1-40 had a significantly longer time to find the area where the safety platform was placed. In contrast, the experimental group showed that sample 15B was able to ameliorate the reference memory deficit caused by β-amyloid peptide 1-40 in rats (i.e., a safety platform was placed in the pool). time to find the cracked area).

FIGS. 12A and 12B show rat hippocampal and rat frontal cortex AChE activity, respectively, in an animal model of β-amyloid-induced learning impairment.

As shown in FIGS. 12A and 12B, AChE activity in the hippocampal and frontal cortex regions of the control group rats fed β-amyloid peptide 1-40 was clearly increased compared to the Sham group rats. . In contrast, the experimental group showed that sample 15B was able to ameliorate the phenomenon of increased AChE activity in hippocampal and frontal cortex regions of rats induced by β-amyloid peptide 1-40.

Example 11: Confirmation of Reishi Acid A Content of Four Composite Components

A. Method

Content measurement of Ganoderic acid A:

1. Preparation of Control Standard Stock Solution:

Accurately weighed 10 mg of the reference standard Ganoderic acid A (Ganoderic acid A) was placed in a 10 mL volumetric flask, added with a methanol solution, and ultrasonically shaken to completely dissolve the Ganodermic acid A. The solution was aliquoted to 10 mL to contain 1 mg of reishi acid A per mL of solution to obtain the control standard solution (1 mg/mL).

2. Repeatability test:

1 mL of the control standard solution (1 mg/mL) was taken and placed in each 10 mL volumetric flask and then made up to volume with methanol. The solution (0.1 mg/mL) was subjected to five replicate injection analyzes by HPLC (repeatability test) and the relative standard deviation (RSD ) was calculated.

3. Preparation of calibration curve (reishi acid A standard):

A control standard stock solution was taken and diluted with methanol to prepare five concentrations of reishi acid A standard based on the sample concentration. The concentration range must include 80-120% concentration of reishi acid A in the sample, and each standard solution at five concentrations was subjected to high performance liquid chromatography (HPLC). Injections were analyzed and linear regression plotted with peak area versus concentration. R2 must be greater than or equal to 0.995.

4. Preparation of test material:

Put 1.0 g of the test substance (Ganoderma lucidum extract (Sample 1) or Sample 15B) into a 20 mL volumetric flask, then add about 10 mL of pure water, then place the volumetric flask in a water bath and subject it to ultrasonic shaking for 10 minutes. bottom. After standing still and cooling, pure water was added to the volumetric flask and quantified to 20 mL to prepare a test substance solution. An appropriate amount of the test substance solution was centrifuged (10000 rpm, 5 minutes) and then filtered through a 0.45 μm filtration membrane, followed by high-performance liquid chromatography analysis to analyze the content of reishi acid A in the test substance. .

5. High-performance liquid chromatography analysis conditions:

Chromatography column type: Inertsil 5 ODS-2 4.6 x 250 mm or equivalent

Detection wavelength: 257 nm

Pump flow rate: 1.0 mL/min

Analysis time for each sample: 120 minutes

Total injection volume: 20 μL

Preparation of mobile phase:
Mobile phase A = 10% MeOH in _CH3CN (v/v)
Mobile phase B = 0.075% aqueous phosphoric acid (522 μL of 85% phosphoric acid (density = 1.685 g/mL) in a 1 L volumetric flask and diluted to the mark with pure water).

6. Calculation of reishi acid A content

Reishi acid A content (mg/g) =
Reishi acid A concentration in test substance (mg/mL) x 20 (mL)

B. result

The reishi acid A standard, reishi extract (Sample 1), and sample 15B, each prepared as described above, were subjected to the above high-performance liquid chromatography analysis to determine the reishi acid A content in sample 15B. was calculated. The results are shown in FIG.

Please refer to FIG. From the calibration curve of the standards described above, the content of reishi acid A in the reishi extract and sample 15B can be calculated as 66.3 mg/g and 4.96 mg/g, respectively.

By determining the reishi acid A content in the sample, the quality of the sample can be confirmed.

Although the present invention has been disclosed above in terms of preferred embodiments, this is not intended to limit the invention, and a person skilled in the art can make certain changes and modifications without departing from the spirit and scope of the invention. may be added, and the protection scope of the present invention shall therefore be as defined in the appended claims.

Claims (14)

A composition for treating and/or preventing intestinal permeability inhibition and/or intestinal wall leakage-related diseases, comprising:
Containing only Chinese herbal medicine compound or only Chinese herbal compound extract as an active ingredient ,
Said Chinese herbal medicine compound is:
Ganoderma lucidum;
jujube;
Consisting of longan meat; and lotus seeds,
The herbal medicine complex extract is:
reishi extract;
Jujube extract;
longan flesh extract; and lotus seed extract,
The weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed in the traditional Chinese medicine compound is 0.3-12:0.8-1.5:0.8-1.5:0.8-4. and
Ganoderma lucidum, jujube, longan flesh, and lotus seed as raw materials necessary for obtaining the ganoderma lucidum extract, jujube extract, longan flesh extract, and lotus seed extract contained in the aforementioned herbal medicine complex extract, respectively is 0.3-12:0.8-1.5:0.8-1.5:0.8-4,
The intestinal wall seepage-related diseases include inflammatory bowel disease, celiac disease, irritable bowel syndrome, acute pancreatitis, non-alcoholic fatty liver disease, alcoholic cirrhosis, type 1 diabetes, type 2 diabetes, obesity, and chronic kidney disease. disease, multiple organ disorders, AIDS, asthma, eczema, psoriasis, autism, depression, anxiety, schizophrenia, bipolar disorder, Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis Ankylosing spondylitis, or fibromyalgia . 2. The composition for use in inhibiting intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall seepage according to claim 1, wherein said reishi mushrooms include red turf, purple turf, or antler reishi. The jujube includes ash jujube, chicken heart jujube, winter jujube, large jujube, small jujube or golden jujube, according to claim 1, wherein inhibition of intestinal permeability and/or treatment and/or intestinal wall leakage-related diseases Compositions for prophylaxis. The composition for use in inhibiting intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage according to claim 1, wherein the longan meat includes powdered shell longan meat, red shell longan meat, or green shell longan meat. The composition for use in inhibiting intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage according to claim 1, wherein the lotus seeds include red lotus seeds or white lotus seeds. 2. The composition according to claim 1, wherein 0.2 to 20 mg of reishi acid A is contained per 1 g of the composition used for the inhibition of intestinal permeability and/or the treatment and/or prevention of diseases associated with intestinal wall leakage. and/or treatment and/or prevention of intestinal wall leakage-related diseases. Inhibition of intestinal permeability and/or intestinal wall leakage according to claim 1, wherein the weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed in the traditional Chinese medicine compound is 1:1:1:1. A composition for use in treating and/or preventing disease. Inhibition of intestinal permeability and/or intestinal wall leakage according to claim 1, wherein the weight ratio of ganoderma lucidum, jujube, longan flesh and lotus seed in the traditional Chinese medicine compound is 3:1:1:1. A composition for use in treating and/or preventing disease. Ganoderma lucidum, jujube, longan flesh and lotus seed as raw materials necessary for obtaining the ganoderma lucidum extract, jujube extract, longan flesh extract and lotus seed extract contained in the aforementioned herbal medicine complex extract, respectively. 2. The composition according to claim 1, wherein the weight ratio is 1:1:1:1, for the treatment and/or prevention of diseases associated with inhibition of intestinal permeability and/or intestinal wall seepage. Ganoderma lucidum, jujube, longan flesh and lotus seed as raw materials necessary for obtaining the ganoderma lucidum extract, jujube extract, longan flesh extract and lotus seed extract contained in the aforementioned herbal medicine complex extract, respectively. 2. The composition according to claim 1, wherein the weight ratio is 3:1:1:1, for the treatment and/or prevention of intestinal permeability inhibition and/or intestinal wall seepage-related diseases. 2. The composition for treating and/or preventing intestinal permeability inhibition and/or intestinal wall seepage-related diseases according to claim 1, further comprising a pharmaceutically acceptable carrier or salt. In the composition used for the inhibition of intestinal permeability and/or the treatment and/or prevention of intestinal wall leakage-related diseases, the content of the herbal medicine composite or the herbal medicine complex extract may be 10 to 99.5 wt%. 12. A composition for use in inhibiting intestinal permeability and/or treating and/or preventing diseases associated with intestinal wall leakage according to claim 11 . Inhibition of intestinal permeability and/or according to claim 1, wherein the composition used for inhibition of intestinal permeability and/or treatment and/or prevention of intestinal wall leakage-related diseases is a pharmaceutical composition or a healthcare composition. Or a composition used for the treatment and/or prevention of diseases associated with intestinal wall leakage. 14. The intestinal permeation according to claim 13 , wherein oral forms of said pharmaceutical or healthcare composition include tablets, granules, powders, pellet-in-capsules, capsules, coated tablets, solutions or instant powders. A composition for use in treating and/or preventing diseases associated with sexual suppression and/or intestinal wall leakage.

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