CN113908224A

CN113908224A - A Chinese medicinal composition with immunoregulatory function, and its preparation method

Info

Publication number: CN113908224A
Application number: CN202111365315.2A
Authority: CN
Inventors: 刘雅杰
Original assignee: Changchun Shoudao Biotechnology Co ltd
Current assignee: Changchun Shoudao Biotechnology Co ltd
Priority date: 2021-11-18
Filing date: 2021-11-18
Publication date: 2022-01-11

Abstract

The invention relates to a traditional Chinese medicine composition with an immunoregulation function and a preparation method thereof, belonging to the field of traditional Chinese medicines. The feed is prepared from the following raw materials in parts by weight: 1-6 parts of rehmannia, 2-5 parts of tree peony bark, 2-5 parts of salvia miltiorrhiza, 1-6 parts of astragalus membranaceus, 2-5 parts of fingered citron, 2-5 parts of thunberg fritillary bulb, 2-5 parts of wild buckwheat rhizome and 1-6 parts of radix glehniae. The Chinese medicinal composition is used for immunoregulation and cancer prevention, has the advantages of delicate and concise medicinal composition and clear main components, regulates the spleen and stomach through various medicaments, promotes blood circulation to remove blood stasis, nourishes yin, clears heat, cools blood and activates blood to achieve the immunoregulation effect and the cancer prevention and treatment effect, adopts a water decoction technology extraction technology, not only keeps the medication characteristics of the traditional Chinese medicine, but also can maximally extract and retain the effective components of the medicaments in the prescription, exerts the comprehensive effect of the prescription, and has very wide prospect in the future.

Description

A Chinese medicinal composition with immunoregulatory function, and its preparation method

Technical Field

The invention relates to the technical field of traditional Chinese medicines, and in particular relates to a traditional Chinese medicine composition with an immunoregulation function and a preparation method thereof.

Background

In the modern times, cancer patients are increasing, and the most commonly occurring cancers are lung cancer, liver cancer, stomach cancer, esophagus cancer, colon cancer and rectal cancer. The incidence of breast cancer is also on a growing trend. Data published by the international anticancer consortium indicate that more deaths occur worldwide due to cancer than AIDS, malaria and tuberculosis add together. Cancer has become a major health hazard, and if the cancer cannot be effectively treated and prevented, the cancer will have great influence on our productive life.

At present, in order to effectively prevent cancer, the improvement of the immunoregulation capability is one of effective ways, and a plurality of medicinal materials have the immunoregulation function, which is counted as follows:

immunoregulation function of rehmannia: the rehmannia glutinosa libosch glycoside A monomer in rehmannia glutinosa libosch has the effects of enhancing humoral immunity and cellular immunity, and rehmannia glutinosa libosch polysaccharide has a remarkable immunity promoting effect and can enhance the proportion of T lymphocytes in peripheral blood to enhance the effect of immunocytes; the prepared rehmannia root decoction can increase the number of white blood cells around the mouse; the rehmanniae radix decoction can accelerate DNA biosynthesis in spleen lymphocyte and enhance interleukin-II production. The decoction of fresh rehmannia root can enhance the nonspecific immunity and T, B lymphocyte function of yin-deficiency mice. The polysaccharide component of rehmannia glutinosa Libosch has been found to have an effect of promoting the proliferation of immune cells in studies by Cabernet Sauvignon et al. Compared with western medicine antitumor drugs, the traditional Chinese medicine rehmannia has no cytotoxic effect and has no direct killing effect on tumor cells. Modern researches show that stachyose and catalpol in rehmannia have anticancer effect. The rehmanniae radix polysaccharide is hydrolyzed due to thermal instability during processing, so that rehmanniae radix has good anticancer effect and can inhibit antigen cell growth. Control experiments show that the Liuwei Dihuang Wan and the Sijunzi Tang improve the life quality of patients with tumor chemotherapy. The LIUWEIDIHUANG pill has effects of regulating immunity, inhibiting protooncogene, enhancing expression of anticancer gene, reducing toxicity, enhancing efficacy, and preventing and treating tumor by inhibiting generation of inflammatory factor. Rehmannia also has a certain effect on the cardiovascular system, and many researches prove that the rehmannia has the effect of strengthening the heart, and the chemical component of the rehmannia for playing the role of positive inotropic action is mainly catalpol in the rehmannia. Lihongwei and the like are modeled by adult male rats, a myocardial ischemia model is established, catalpol is injected before an operation and is compared with a control group to find that the catalpol can obviously improve the cardiac function and reduce myocardial infarction. The sunrise is tested by using myocardial cells of suckling mice, and researches on hypoxia and reoxygenation injuries find that the rehmannia polysaccharide can repair the morphology of the myocardial cells and increase the survival number of the cells. Zhao Li Juan finds that the rehmannia root can improve the clinical symptoms and cardiac insufficiency of patients with coronary heart disease.

The immunoregulation function of the tree peony bark: the Wangyun and the like find that in the research on the influence of the moutan bark on the immunity, a rat inhales paeonol in an atomizing inhalation mode, immune organ index data before and after inhalation are measured, the immune function of immune cells in the rat inhaling the paeonol is obviously enhanced compared with that of a control group of rats, and the immune functions of immune organs such as spleen, thymus and the like in an experimental group are enhanced, and experiments show that the paeonol component in the moutan bark has better curative effect on enhancing the immunity of the organism. Jiangli research finds that the moutan bark has the effect of resisting tumors by inhibiting the proliferation of tumor cells, inhibiting newly generated tumor vessels, inhibiting the metastasis of cancer cells, inhibiting the transcription of cancer genes, inducing the apoptosis of cancer cells and acting on cancer cells together with chemical drugs. Paeonol can inhibit proliferation and induce apoptosis of colon cancer, hepatocarcinoma and gastric cancer, and cortex moutan has tumor inhibiting rate over 50%, so cortex moutan has certain anti-tumor effect. The activity mechanism of the moutan bark is discussed from the cellular level by Tianfuzhong and the like, and the paeonol can inhibit the proliferation of tumor cells, induce the apoptosis of the tumor cells, inhibit the migration and the invasion of the tumor cells and inhibit the formation of tumor neovascularization, thereby triggering the immune regulation reaction and inhibiting the chronic inflammatory reaction of the tumor. Cortex moutan can be used as Chinese medicinal material and chemotherapy medicine for improving microcirculation disturbance. Zhengyanyanyanyan decoction component has been found to inhibit the activation and proliferation of lymphocyte and has better immunosuppressive action on ulcerative colitis mouse. In the research of extracting effective components of the cortex moutan radicis of the Ouleira, the synergistic effect of the cortex moutan radicis through the regulation of cellular immunity and humoral immunity is found, the activity of Th1 cells is enhanced, the activity of Th2 cells is reduced, the proliferation and differentiation of islet B cells are promoted, and the effects of reducing blood sugar and treating diabetes are achieved. Zulixia et al found that polysaccharides in cortex moutan have immunoregulatory effect on immunodeficient mice in response surface method optimized extraction process of polysaccharides in cortex moutan. In addition, it also has certain bacteriostatic action.

The immunoregulation function of the thunberg fritillary bulb is as follows: peimine in Fritillaria thunbergii achieves the anti-tumor effect by inhibiting the proliferation of tumor cells and promoting the apoptosis of the tumor cells. The peimine inhibits cell proliferation by blocking a cell cycle at a G1 stage, and the inhibition effect of the cell proliferation is dependent on time and concentration, so that the research finds that the peimine has a significant inhibition effect on 80 percent of the cell inhibition rate of breast cancer cells. The peimine can remarkably inhibit the proliferation of cancer cells and has the effect of resisting tumors.

The immune regulation function of wild buckwheat rhizome: the wild buckwheat rhizome extract can inhibit the proliferation of human esophageal cancer cell strains CaEs-17 cells and promote the apoptosis of the cells, and has the function of resisting cancers. The polyphenol component Fr4 in rhizoma Fagopyri Dibotryis can significantly inhibit the growth of mouse Lewis lung cancer by down-regulating the expression of metalloproteinase-9 (MMP-9), thereby achieving the anti-tumor effect. The flavone (RCFGB) of herba Trifolii Pratentis in rhizoma Fagopyri Dibotryis has anticancer effect by inhibiting IL-6 protein expression and inhibiting human gastric cancer SGC7901 cell migration ability. The wild buckwheat rhizome extract can inhibit the growth of liver cancer cells HepG2 by down-regulating the expression of the Tima-1 gene and up-regulating the expression of the nm23-H1 gene, thereby playing a role in resisting tumor invasion. The wild buckwheat rhizome extract has an inhibiting effect on the cancer cells, has an obvious inhibiting effect on cancer cells such as HCT116 (colon), U2OS (bone) and the like, has a slight inhibiting effect on Hela (cervix) and OVCAR-3 (ovary) cancer cells, and also has an inhibiting effect on the growth of prostate cancer cells DU145 and brain cancer cells T98G when the concentration reaches a certain value.

The immunoregulation function of the salvia miltiorrhiza: the function of the salvia miltiorrhiza plays a role in bidirectional internal regulation of immune response. The danshensu has obvious inhibition effect on the secretion of various cytokines by rat kupffer cells under the stimulation of LPS, and the tanshinone IIA has obvious inhibition effect on migration and chemotaxis of human leucocytes. The salvianolic acid B can obviously improve phagocytosis, liver and spleen index and clearance index of mouse macrophage, thereby enhancing non-specific immune function. In addition, researches show that the salvia miltiorrhiza polysaccharide also has remarkable immunoregulation activity, the salvia miltiorrhiza polysaccharide can remarkably enhance the phagocytic function of mouse macrophages, stimulate lymphocyte proliferation, improve mouse thymus index caused by dinitrofluorobenzene, inhibit increase of vascular permeability and ear swelling, and down-regulate the expression of cell factors IL-1 beta, human leukocyte interferon alpha and inducible nitric oxide synthase mRNA, thereby indicating that the salvia miltiorrhiza has stronger immunoregulation function.

The immunoregulation function of fingered citron: a large number of pharmacological and clinical researches show that the polysaccharide compound is an immunomodulator which can activate immune cells and improve the immune function of an organism and has no toxic or side effect on normal cells. In recent years, studies have reported that some natural polysaccharides enhance the immunomodulatory activity of macrophages by increasing NO levels and cytokine production. Polysaccharides can bind to specific membrane receptors on the surface of macrophages, such as Toll-like receptors, complement receptor 3, dendritic cell-associated C-type lectin 1, and mannose receptor, and then trigger several different intracellular signal transduction pathways. The influence of the fructus citri sarcodactylis polysaccharide on the immunologic function of mice with low immunologic function caused by cyclophosphamide is researched by the Digitalis. The fructus Citri Sarcodactylis polysaccharide can promote the increase of the level of interleukin 6(IL-6) outside macrophages of mice with low immune function, and has no influence on the concentration of IL-6 in macrophages. IL-6 can affect the proliferative differentiation of B cells and directly or indirectly enhance the tumoricidal activity of natural killer cells and cytotoxic T cells. He and the like are used for extracting, separating and purifying the crude polysaccharide of the fingered citron to obtain the polysaccharide with good uniformity, which is named as FCp-3, and the influence of the FCp-3 on the proliferation of splenocytes and thymocytes in vitro is measured. The results show that: FCp-3 shows obvious splenocyte proliferation promoting activity when the concentration reaches 25 mu g/ml, and can promote the proliferation of thymocytes, and the bergamot polysaccharide FCp-3 can be used as a novel immunomodulator.

The immunoregulation function of radix glehniae is as follows: influence of polysaccharides in Adenophora stricta on immunoregulation. Researches show that the radix glehniae crude polysaccharide (GLP) can remarkably enhance the killing rate of NK cells and the T lymphocyte transformation function of hyperthyroidism yin-deficiency mice spleen, and improve the content of serum immunoglobulin IgM and IgG. The purified components of the radix glehniae polysaccharide have a good promotion effect on the proliferation of spleen lymphocytes in vitro and have good in vitro immune activity. The radix glehniae extract is administrated by taking cyclophosphamide as a positive control and gavage to BALB/c mice, so that the radix glehniae can increase the thymus and spleen quality of the mice, enhance the phagocytic capability of macrophages in abdominal cavities of the mice on neutrophils, and improve the tumor killing rate of lymphocytes of the mice and the killing capability of natural killer cells. The radix Glehniae supercritical fluid extract can also increase the number of T lymphocyte sub-population and T lymphocyte, obviously increase the ratio of helper T cell to suppressive T cell (Th/Ts), and has effect of enhancing cellular immunity. The alcohol extract of the aqueous extract of the stem and leaf of the Laiyang radix glehniae has the function of inhibiting the reduction of the white blood cell number and the thymus index of the peripheral blood of a mouse caused by cyclophosphamide and can enhance the phagocytic function of the reticuloendothelial system of the mouse with low immunity. In addition, 2 dosage groups of the Laiyang straight ladybell stem and leaf extract can obviously increase the thickness of the foot sole of a mouse, and the function is enhanced along with the dosage increase, which shows that the Laiyang straight ladybell stem and leaf extract has the function of promoting the delayed allergy of the mouse. The extract of stem and leaf of radix Glehniae (Dahongpao variety) also has humoral immunity regulating effect.

The immunoregulation function of astragalus: the astragalus glycoprotein reduces Th17 transcription factor ROR gamma while inhibiting JAK-STAT3 signals, down-regulates Th17, activates a JAK-STAT5 passage, and improves the expression of a Treg transcription factor Foxp 3; tregs are upregulated to regulate the Th17/Treg balance. The occurrence of allergic rhinitis is mainly involved by immune cells and factors thereof, the astragaloside can regulate PD-L1 to resist tumors, and researches prove that the balance of Th17/Treg can be also relieved by the influence of PD-L1 and receptor actions thereof, so that the astragaloside also has great possibility of treating the allergic rhinitis by the PD-L1 route. Influencing macrophage is another way of regulating immunity, and radix astragali extract can inhibit autoimmune encephalomyelitis model mouse CD4+, CD11b + macrophage, CD11c + cell percentage, and inhibit peripheral immune cell activation to inhibit further inflammation. The astragalus polysaccharide regulates the cellular immunity, improves the thymus index and the spleen index of a model mouse by increasing the phagocytosis of splenic lymphocytes and peritoneal macrophages of the model mouse, recovers damaged thymus and spleen tissues, and prevents damaged organs from being damaged by anticancer drugs due to the action on immune organs. The regulation effect of astragalus to macrophages is bidirectional, and the response of normal cultured macrophages RAW264.7 and LPS stimulated cultured macrophages to the effect of astragalus total flavonoids is opposite and dose-dependent. The powerful immunoregulation action of astragalus root suggests that it may improve functions by such as activating immune effect-related cell activation and reducing harmful adhesion molecule.

Disclosure of Invention

The invention provides a traditional Chinese medicine composition with an immunoregulation function and a preparation method thereof, aiming at effectively preventing cancers through the immunoregulation function.

The medicine of the invention is researched on the action mechanism aspects of improving immunity and preventing cancer, and the action mechanism of the medicine is clarified, and the prescription of the invention consists of 8 traditional Chinese medicines of rehmannia root, tree peony bark, salvia miltiorrhiza, astragalus root, finger citron, thunberg fritillary bulb, wild buckwheat rhizome and coastal glehnia root; in the formula, rehmanniae radix is the monarch drug, which is bitter and cold in nature, is the essential drug for clearing heat and cooling blood, and sweet and cold in nature and moist in nature, and can clear heat and nourish yin, promote the production of body fluid and quench thirst. Cortex moutan is bitter, pungent and slightly cold, enters heart, liver and blood, and can clear heat and cool blood and clear yin and latent heat; zhejiang fritillaria is bitter and cold, enters lung and heart channels, has strong actions of bitter clearing, clearing heat and removing toxicity, and can resolve phlegm, dissipate stagnation and cure carbuncle; the two are used together to clear heat and cool blood, and the assistant drugs clear heat, nourish yin and cool blood are used as ministerial drugs. Radix Glehniae has effects in tonifying stomach yin, clearing away stomach heat, nourishing yin, invigorating stomach, promoting salivation, and quenching thirst; the wild buckwheat rhizome is pungent and cool, can clear away heat and toxic materials, and has the effects of tonifying spleen and promoting digestion; the salvia miltiorrhiza enters heart channels, can clear heat and cool blood, can relieve restlessness and calm nerves, and can promote blood circulation and nourish blood to calm the nerves and calm mind; astragalus root, radix astragali, being sweet and warm, is good at entering spleen and stomach, is the essential drug for tonifying middle-jiao and Qi, invigorating qi and strengthening spleen, and qi moving is the blood moving, and qi is used as adjuvant drug for assisting blood circulation. Fingered citron, fructus Citri Sarcodactylis, with the action of fragrant and spleen-enlivening, bitter and warm properties, dampness-drying, spleen-invigorating and phlegm-resolving, pungent flavor-moving and bitter-purging, liver-soothing and qi-regulating, is a guiding drug. The whole formula has the effects of nourishing yin, clearing heat, cooling blood and activating blood. The prescription consists of 8 traditional Chinese medicines such as radix rehmanniae, moutan bark, thunberg fritillary bulb, radix glehniae, wild buckwheat rhizome, salvia miltiorrhiza, fingered citron, astragalus and the like, and is mainly used for treating the syndrome of yin deficiency caused by excessive fire-heat toxin and consumption of yin fluid. The cancer is caused by stasis in vivo, the blood and the channels are blocked, the spleen and the stomach are the acquired root and the source of the generation of qi and blood, the medicines achieve the effect of preventing the cancer by regulating the spleen and the stomach, activating blood and dissolving stasis, nourishing yin and clearing heat, cooling blood and activating blood, and the results of modern pharmacological research show that the traditional Chinese medicine has the function of immunoregulation and can be used for preventing the cancer.

The technical scheme adopted by the invention is that the traditional Chinese medicine is prepared from the following raw materials in parts by weight:

1-6 parts of rehmannia, 2-5 parts of tree peony bark, 2-5 parts of salvia miltiorrhiza, 1-6 parts of astragalus membranaceus, 2-5 parts of fingered citron, 2-5 parts of thunberg fritillary bulb, 2-5 parts of wild buckwheat rhizome and 1-6 parts of radix glehniae.

In one embodiment of the invention, the food is prepared from the following raw materials in parts by weight:

5 parts of rehmannia root, 4 parts of tree peony bark, 2 parts of salvia miltiorrhiza, 1 part of astragalus root, 4 parts of fingered citron, 3 parts of thunberg fritillary bulb, 2 parts of wild buckwheat rhizome and 1 part of radix glehniae.

4 parts of rehmannia root, 2 parts of tree peony bark, 4 parts of salvia miltiorrhiza, 4 parts of astragalus root, 2 parts of fingered citron, 5 parts of thunberg fritillary bulb, 3 parts of wild buckwheat rhizome and 6 parts of radix glehniae.

4 parts of rehmannia root, 3 parts of tree peony bark, 3 parts of salvia miltiorrhiza, 4 parts of astragalus root, 3 parts of fingered citron, 2 parts of thunberg fritillary bulb, 3 parts of wild buckwheat rhizome and 4 parts of coastal glehnia root.

5 parts of rehmannia root, 3 parts of tree peony bark, 5 parts of salvia miltiorrhiza, 3 parts of astragalus root, 5 parts of fingered citron, 3 parts of thunberg fritillary bulb, 4 parts of wild buckwheat rhizome and 4 parts of coastal glehnia root.

4 parts of rehmannia root, 5 parts of tree peony bark, 3 parts of salvia miltiorrhiza, 5 parts of astragalus root, 3 parts of fingered citron, 4 parts of thunberg fritillary bulb, 3 parts of wild buckwheat rhizome and 5 parts of coastal glehnia root.

The invention also provides a preparation method of the traditional Chinese medicine composition, which comprises the following steps:

decocting and extracting with water: decocting rehmannia root, tree peony bark, red sage root, astragalus root, fingered citron, thunberg fritillary bulb, wild buckwheat rhizome and coastal glehnia root in water for 1-4 times, filtering, combining filtrates, and concentrating to obtain an extract;

drying the extract to obtain the Chinese medicinal composition.

Preferably, in the step of water decoction and extraction, the adding amount of the aqueous solution is 4-10 times of the total weight of the medicinal materials, the extraction times are 1-4 times, and the time for each extraction is 0.5-3 hours.

The invention also provides a traditional Chinese medicine preparation which comprises the traditional Chinese medicine composition and pharmaceutically acceptable auxiliary materials.

Preferably, the dosage form of the Chinese medicinal preparation is capsules, tablets, granules, dripping pills or oral liquid.

The invention has the beneficial effects that:

the pharmaceutical composition is used for immunoregulation and cancer prevention, has concise and concise pharmaceutical composition and clear main components, regulates the spleen and stomach through various medicines, promotes blood circulation to remove blood stasis, treats the diseases by nourishing yin, clearing heat, cooling blood and promoting blood circulation to achieve the immunoregulation and the prevention and treatment effects on the cancer, has small toxic and side effects in the prescription, can comprehensively regulate the internal environment and improve the immunity, and can play a role in attenuation and synergism in radiotherapy, chemotherapy and surgical treatment of the cancer. The extraction technology of the water decoction technology is adopted, so that the traditional Chinese medicine medication characteristics are maintained, the effective components of the medicines in the prescription can be extracted and retained to the maximum extent, the comprehensive effect of the prescription is exerted, the preparation is safe and effective, the drug effect is clear, the preparation has positive functional significance for preventing and treating cancers, has the effects of inhibiting tumors, resisting metastasis and resisting recurrence, the life quality of tumor patients is greatly improved, and the preparation has a very wide prospect in the future.

Detailed Description

The invention discloses a traditional Chinese medicine composition for immunoregulation and cancer prevention, a preparation method thereof and a traditional Chinese medicine preparation. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the methods and applications of this invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications in the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of this invention without departing from the spirit and scope of the invention.

EXAMPLE 1 preparation of capsules of the invention

The feed is prepared from the following raw materials in parts by weight:

5 parts of rehmannia root, 4 parts of tree peony bark, 2 parts of salvia miltiorrhiza, 1 part of astragalus, 4 parts of fingered citron, 3 parts of thunberg fritillary bulb, 2 parts of wild buckwheat rhizome and 1 part of radix glehniae;

extracting the above 8 materials with 8 times of water for 2 times, each for 2 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder;

adding 1.5 times of dextrin, mixing, making into granule, and making into capsule.

EXAMPLE 2 preparation of tablets of the invention

The feed is prepared from the following raw materials in parts by weight:

4 parts of rehmannia root, 2 parts of tree peony bark, 4 parts of salvia miltiorrhiza, 4 parts of astragalus root, 2 parts of fingered citron, 5 parts of thunberg fritillary bulb, 3 parts of wild buckwheat rhizome and 6 parts of radix glehniae;

extracting the above 8 materials with 6 times of water for 3 times, each for 1.5 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder;

mixing the above extracts with 1 time of starch, granulating, and tabletting.

Example 3 preparation of granules according to the invention

The feed is prepared from the following raw materials in parts by weight:

4 parts of rehmannia root, 3 parts of tree peony bark, 3 parts of salvia miltiorrhiza, 4 parts of astragalus root, 3 parts of fingered citron, 2 parts of thunberg fritillary bulb, 3 parts of wild buckwheat rhizome and 4 parts of radix glehniae;

extracting the above 8 materials with 10 times of water for 3 times, each for 1 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder;

adding 0.8 times of dextrin, mixing, and making into granule.

EXAMPLE 4 preparation of dropping pills of the present invention

The feed is prepared from the following raw materials in parts by weight:

5 parts of rehmannia root, 3 parts of tree peony bark, 5 parts of salvia miltiorrhiza, 3 parts of astragalus, 5 parts of fingered citron, 3 parts of thunberg fritillary bulb, 4 parts of wild buckwheat rhizome and 4 parts of radix glehniae;

extracting the above 8 materials with 6 times of water for 3 times, each for 2 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder;

adding 1 time of polyethylene glycol into the above extract, heating and melting, mixing, and making into dripping pill.

EXAMPLE 5 preparation of oral liquid of the present invention

The feed is prepared from the following raw materials in parts by weight:

4 parts of rehmannia root, 5 parts of tree peony bark, 3 parts of salvia miltiorrhiza, 5 parts of astragalus, 3 parts of fingered citron, 4 parts of thunberg fritillary bulb, 3 parts of wild buckwheat rhizome and 5 parts of radix glehniae;

extracting the above 8 medicinal materials with 8 times of water for 1 time, each time for 1.5 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder;

adding 0.02 times of sweetener into the above extract, and dissolving in water to obtain oral liquid.

Example 6

The feed is prepared from the following raw materials in parts by weight:

1 part of rehmannia root, 2 parts of tree peony bark, 2 parts of salvia miltiorrhiza, 1 part of astragalus, 2 parts of fingered citron, 2 parts of thunberg fritillary bulb, 2 parts of wild buckwheat rhizome and 1 part of radix glehniae;

extracting the above 8 materials with 1 times of water for 1 time, each time for 0.5 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder.

Example 7

The feed is prepared from the following raw materials in parts by weight:

6 parts of rehmannia root, 5 parts of tree peony bark, 5 parts of salvia miltiorrhiza, 6 parts of astragalus root, 5 parts of fingered citron, 5 parts of thunberg fritillary bulb, 5 parts of wild buckwheat rhizome and 6 parts of radix glehniae;

extracting the above 8 materials with 10 times of water for 4 times, each for 3 hr, filtering, mixing extractive solutions, concentrating, drying, and pulverizing into fine powder.

The following experiments on the immunoregulatory function are further provided to illustrate the effects of the present invention

Examples of the experiments

1.1 Experimental animals

Public mice of clean grade ICR provided by animal experiment center of bayan medical school of jilin university, production license number: SCXK- (gib) 20200004. The feed is provided by Yisi experimental animal technology Limited liability company in Changchun city, and the production license number is as follows: SCXK- (Ger) 2020-. The experimental animal environment facility certificate is 10-1005 in a lucky movement, and the experimental animal uses a license number: SYXK- (Ji) 2020-. Choosing healthThe healthy male mice 48, weight 18 g-22 g, divided into 4 groups, each group of 12, as an immune group, organ/body weight ratio determination, half of the hemolysis value (HC)₅₀) And antibody-producing cell detection; selecting 48 healthy male mice with the weight of 18-22 g, dividing the mice into 4 groups, and taking 12 mice in each group as two groups of immunity to perform a carbon clearance experiment; selecting 48 healthy male mice with the weight of 18-22 g, dividing the mice into 4 groups with 12 mice in each group as three immune groups, and performing ConA-induced mouse lymphocyte transformation experiments and NK cell activity determination; selecting 48 healthy male mice with the weight of 18-22 g, dividing the mice into 4 groups, and taking 12 mice in each group as four immune groups to perform a delayed allergy experiment; selecting 48 healthy male mice with the weight of 18 g-22 g, dividing the mice into 4 groups, and taking 12 mice in each group as five immune groups to carry out an experiment of phagocytosis of chicken red blood cells by abdominal macrophages of the mice.

1.2 dosage selection and formulation

Example 1, 2 times/day, 10 grams/time, i.e., 10.0g/60kg BW, equivalent to 0.165g/kg BW, the gavage dosages, i.e., 0.165, 1.65, 4.95g/kg BW, were set at 1, 10, and 30 times the daily recommended intake by the manufacturer, with each dosage group diluted with purified water. Pure water is used as a negative control group, the gavage amount of each group of mice is 0.2mL/10g BW, and each immune index is measured after continuous gavage for 30 days.

1.3 Instrument and reagents:

an electronic balance (0.1g), an analytical balance, a clean bench, a carbon dioxide incubator, a centrifuge, a 722 spectrophotometer, a constant temperature water bath, an elisa instrument, a microscope and the like; sterile surgical instruments, vernier calipers (precision 0.02mm), microsyringes (25 μ L), cytometers, 24-well and 96-well flat-bottom cell culture plates, 96-well U-shaped cell culture plates, glass plates, gauze, test tubes, slide racks, 200-mesh screens, timers, hemoglobin pipettes, slides, and the like; sheep erythrocytes (SRBC), physiological saline, Hank's solution (pH 7.2-7.4), RPMI1640 culture solution, calf serum, streptomycin, concanavalin A (ConA), L% glacial acetic acid, 1mol/L HCl solution, acidic isopropanol (96mL isopropanol plus 4mL hydrochloric acid), MTT, PBS buffer (pH 7.2-7.4), complement (guinea pig serum), SA buffer, agarose, Dushi reagent (sodium bicarbonate 1.0g, potassium ferricyanide 0.2g, potassium cyanide 0.05g, distilled water to 1000mL), YAC-1 cells, sodium lactate, nitrotetrazolium chloride, phenazine dimethyl sulfate, oxidized coenzyme I, 0.2mol/L Tris-HCl buffer (pH8.2), L% NP40, India ink, Na2CO3, chicken erythrocytes, methanol, Giemsa dye solution, and the like.

1.4 Experimental methods

1.4.1 organ/body weight ratio measurement

Weighing mice, dislocating and killing the mice, taking spleen and thymus, removing fascia, sucking blood stains on the surfaces of visceral organs by using filter paper, weighing, and calculating the ratio of spleen/body weight and the ratio of thymus/body weight.

1.4.2 delayed type hypersensitivity (thickening of the foot sole DTH)

Sheep blood was taken, washed with physiological saline, and mice were immunized intraperitoneally with 2% (V/V) SRBC, 0.2 mL/mouse. The thickness of the left hind foot plantar region was measured 4 days after immunization, 20% (V/V) SRBC was subcutaneously injected at the measurement site at 20 μ L/body, and the thickness of the left hind foot plantar region was measured three times 24h after injection, and the average value was calculated.

1.4.3 ConA-induced mouse lymphocyte transformation experiment (MTT method)

Spleens were removed aseptically, ground to single cell suspensions in Hank's solution, washed 2 times with Hank's solution, and centrifuged 10min (1000r/min) each time. Suspending the cells in 1mL of complete culture medium, staining with Tulip blue to count the number of viable cells (more than 95%), and adjusting the cell concentration to 3X 10⁶one/mL. Each cell suspension was added to a 24-well plate in two wells, 1mL per well, 75. mu.L of LCoA solution (equivalent to 7.5. mu.g/mL) in one well, and 5% CO in the other well as a control₂ 37 ℃CO₂Culturing for 72h in an incubator. 4 hours before the end of the culture, 0.7mL of the supernatant was aspirated from each well, and 0.7mL of calf serum-free RPMI1640 culture medium was added thereto together with 50. mu.L of MTT (5mg/mL) per well, and the culture was continued for 4 hours. After the culture is finished, lmL acidic isopropanol is added into each hole, the mixture is beaten and uniformly mixed by blowing, purple crystals are completely dissolved, and OD is determined_570nm。

1.4.4 detection of antibody-producing cells (Jerne modified slide method)

Placing sheep blood into sterilized conical flask containing glass beads, shaking in one direction to remove fiber, and heating at 4 deg.CThe refrigerator is kept for standby (can be kept for 2 weeks). 2% (V/V) cell suspension is prepared by the packed SRBC with normal saline, and 0.2 mL/mouse is injected into the abdominal cavity. Killing mice which are immunized by SRBC for 4-5 days by dislocation, taking spleens, putting the spleens into Hank's solution, grinding the spleens into cell suspension, filtering the cell suspension by gauze, washing the Hank's solution for 2 times, centrifuging the solution (1000r/min) for 10min each time, adding the spleen cell suspension into RPMI1640 culture solution, counting cells, and adjusting the cell concentration to 5 multiplied by 10⁶one/mL. Heating and dissolving agarose, preserving heat in a water bath at 45-50 ℃, mixing with Hank's solution with the same amount of PH 7.2-7.42 times, subpackaging into small test tubes with 0.5mL per tube, adding 10% (V/V, prepared by SA solution) into the tubes, pressing to accumulate 50 muL of SRBC and 20 muL of spleen cell suspension, pouring the mixture on a glass slide with an agarose thin layer, making parallel sheets, horizontally buckling the glass slide on a slide rack after the agar is solidified, and placing the glass slide on a CO rack₂After incubation in the incubator for 1.5h, complement diluted with SA buffer (1:8) was added to the slide rack wells and after incubation for a further 1.5h, the number of lyso-plaques was counted.

1.4.5 half maximal hemolysis value (HC)₅₀) Measurement of (2)

Sheep blood was washed 3 times with physiological saline, centrifuged (2000r/min) for 10min each time, and injected intraperitoneally with 2% (V/V, prepared with physiological saline) of packed SRBC 0.2 mL/mouse for immunization. After 4 days, taking blood from the centrifugal tube, placing for about 1h, stripping coagulated blood from the tube wall to fully separate out serum, centrifuging at 2000r/min for 10min, and collecting serum. The serum was diluted (400-fold) with SA buffer, and 1mL of the diluted serum was placed in a test tube, and 0.5mL of 10% (V/V) SRBC and 1mL of complement were added in this order (diluted 1:8 with SA buffer). A control tube without serum was provided (SA buffer was used instead). And (4) preserving the heat for 15-30 min in a constant-temperature water bath at 37 ℃, and stopping the reaction in an ice bath. Centrifuging at 2000r/min for 10min, collecting supernatant 1mL, and adding Duchen reagent 3 mL. Simultaneously taking 0.25mL of 10% (V/V) SRBC, adding Dushi reagent to 4mL, mixing well, standing for 10min, using reference as blank, and respectively determining OD of each tube_540nmThe amount of hemolysin is expressed as half the hemolysin value (HC)₅₀) Representing, calculating:

HC₅₀sample optical density value/optical density value at half SRBC hemolysis × dilution factor

1.4.6 mouse carbon clearance test

Intravenous injection of Indian ink at 1:4 times dilution into mouse tail, timing immediately, collecting 20 μ L blood from angular venous plexus at 2 min and 10min later, and adding 2mL of 0.1% Na₂CO₃In solution with Na₂CO₃As a control, OD was measured_600nm. Mice were sacrificed and livers and spleens were removed and weighed. Calculation of phagocytic index a:

k＝(lgOD1-lgOD2)/(t2-t1)

1.4.7 experiment of phagocytosis of chicken red blood cells by macrophages in abdominal cavity of mouse

Injecting 20% (V/V prepared by normal saline) into abdominal cavity of mouse, pressing and accumulating chicken red blood cell (2000r/min, 10min) suspension 1mL, spacing 30min, dislocating and killing, taking 1mL of abdominal cavity macrophage washing liquid, dripping on glass slide, incubating in incubator at 37 ℃ for 20min, rinsing in normal saline to remove non-patch cells. Air-drying, fixing with methanol, staining with 4% (V/V) Giemsa-phosphate buffer, rinsing with distilled water, and air-drying. Counting under oil lens, counting 100 macrophages per tablet, calculating phagocytic rate and phagocytic index:

percent phagocytosis%

Phagocytosis index-total number of phagocytosed chicken red blood cells/number of macrophages counted

1.4.8 NK cell Activity assay (lactate dehydrogenase LDH assay)

Subculturing target cell YAC-l 24h before experiment, washing with Hank's solution for 3 times before application, adjusting cell concentration to 4 × 10 in RPMI1640 complete culture solution containing 10% calf serum⁵one/mL. The test mouse is dislocated and killed, the spleen is taken out to prepare spleen cell suspension, Hank's solution is washed for 3 times, the centrifugation is carried out for 10min at 1500r/min each time, 2mL RPMI1640 complete culture solution containing 10% calf serum is used for resuspension, the number of viable cells is counted by staining Taifenglan (the number of the viable cells is more than 95%), the cell concentration is adjusted to be 2 multiplied by 10⁷seed/mL, such that the effective target ratio is 50: 1. taking 100 mu L of target cells and effector cells respectively, and adding the target cells and the effector cells into a U-shaped 96-hole culture plate; target cell natural release pore target cell adding and culturing100 μ L of each solution, 100 μ L of each target cell maximum release pore plus target cell and L% NP 40; all the above-mentioned materials are equipped with three parallel holes, 37 deg.C, 5% CO₂Culturing for 4h in incubator, centrifuging 96-well plate at 1500r/min for 5min, sucking supernatant 100 μ L per well, placing in enzyme-linked plate, adding LDH matrix solution 100 μ L, reacting for 10min, adding lmol HCl solution 30 μ L per well to terminate reaction, and determining OD_490nmAnd calculating the NK activity:

NK cell activity% (reaction well OD-Natural Release well OD)/(maximum Release well OD-Natural Release well OD) × 100%

Preparation of LDH matrix solution:

sodium lactate 5X 10^-2mol/L, nitro tetrazole chloride 6.6 multiplied by 10^-4mol/L phenazine dimethyl sulfate 2.8 xl 0^-4mol/L, oxidized coenzyme I1.3X 10^-3mol/L；

Dissolving the reagent in 0.2mol/L Tris-HCl buffer solution (pH8.2);

1.5 data statistics

Performing mean comparison by adopting single-factor variance analysis in SPSS11.5 statistical software, and performing LSD (least squares decomposition) method for pairwise comparison among groups when the variances are uniform; when the variance is uneven, the Tamhane method is adopted for pairwise comparison among the groups.

2. Results

2.1 Effect of test substances on mouse body weight, see Table 1:

TABLE 1 weight effects of test substances on immunized groups of mice

P value: each experimental group was compared with a negative control group

As can be seen from Table 1, after statistical treatment, the body weight and weight gain of mice in each dose group are not significant (P >0.05) when the test object is orally administered for 30 days, compared with the negative control group, i.e. the test object has no obvious influence on the body weight and weight gain of the mice.

TABLE 2 weight effects of test substances on immunized mice

P value: each experimental group was compared with a negative control group

As can be seen from Table 2, statistically treated, the body weight and weight gain of mice in each dose group are not significantly different (P >0.05) compared with those of the negative control group after oral administration of the test substance for 30 days, i.e., the test substance has no obvious influence on the body weight and weight gain of the mice.

TABLE 3 weight effects of test substances on immunized three groups of mice

P value: each experimental group was compared with a negative control group

As can be seen from Table 3, after statistical treatment, the body weight and weight gain of mice in each dose group are not significant (P >0.05) when the test object is orally administered for 30 days, compared with the negative control group, i.e. the test object has no obvious influence on the body weight and weight gain of the mice.

TABLE 4 weight effects of test substances on immunized four groups of mice

P value: each experimental group was compared with a negative control group

As can be seen from Table 4, after statistical treatment, the body weight and weight gain of mice in each dose group are not significant (P >0.05) when the test object is orally administered for 30 days, compared with the negative control group, i.e. the test object has no obvious influence on the body weight and weight gain of the mice.

TABLE 5 weight effects of test substances on five groups of immunized mice

P value: each experimental group was compared with a negative control group

As can be seen from Table 5, after statistical treatment, the body weight and weight gain of mice in each dose group are not significant (P >0.05) when the test object is orally administered for 30 days, compared with the negative control group, i.e. the test object has no obvious influence on the body weight and weight gain of the mice.

2.2 Effect of test substances on the organ/body weight ratio of mice

TABLE 6 Effect of test substances on the organ/body weight ratio of mice

Spleen/body weight ratio P₁The value: comparing each experimental group with a negative control group;

thymus/body weight ratio P₂The value: comparing each experimental group with a negative control group;

as can be seen from Table 6, when the test substance is orally administered to the mice at different doses for 30 days, the spleen/body weight ratio and the thymus/body weight ratio of each dose group have no significant difference (P >0.05) compared with the negative control group, i.e., the test substance has no significant effect on the organ/body weight ratio of the mice.

2.3 Effect of test substances on the immune function of mouse cells

2.3.1 Effect of test Agents on mouse body weight and delayed type allergy (DTH)

TABLE 7 Effect of test substances on mouse body weight and delayed type allergy (DTH)

P value: p < 0.05 for each experimental group compared with negative control group

As can be seen from table 7, the mice in each dose group had no significant difference in body weight and weight gain compared with the negative control group (P >0.05) after being administered orally to the test subject for 30 days by statistical treatment; the swelling degree of the foot plantar swelling of the low-dose group has no significance (P is more than 0.05) compared with that of the negative control group, and the difference between the medium-high dose group and the negative control group has significance (P is less than 0.05), namely, the medium-high dose of the test substance can enhance the delayed type allergic reaction of the mice.

2.3.2 Effect of test Agents on ConA-induced murine lymphocyte transformation experiments

TABLE 8 Effect of test substances on ConA-induced mouse lymphocyte transformation experiments

P value: each experimental group was compared with a negative control group

As can be seen from Table 8, when the test substances are orally administered to the mice at different doses for 30 days, the proliferation capacity of the lymphocytes in each dose group is not significantly different from that in the negative control group (P >0.05) by statistical treatment, i.e., the test substances have no obvious influence on the ConA-induced lymphocyte transformation capacity of the mice.

2.4 Effect of test substances on humoral immunity

2.4.1 Effect of test substance on the number of antibody-producing cells

TABLE 9 Effect of test substances on the number of mouse antibody-producing cells

P value: each experimental group was compared with a negative control group

As can be seen from Table 9, the number of antibody-producing cells of each dose group was not significantly different from that of the negative control group (P >0.05) when the test substance was orally administered to the mice at different doses for 30 days, i.e., the test substance had no significant effect on the number of antibody-producing cells of the mice.

2.4.2 test substance to half-maximal hemolysis value (HC) of mice₅₀) Influence of (2)

TABLE 10 test substance to half-maximal hemolytic value HC of mice₅₀Influence of (2)

P value: each experimental group was compared with a negative control group

As can be seen from Table 10, when the test substance is orally administered to the mice for 30 days at different doses, the difference between the half hemolysis value of each dose group and the comparison between the negative control groups is not significant (P >0.05) after statistical treatment, i.e., the test substance has no significant effect on the half hemolysis value of the mice.

2.5 Effect of test Agents on phagocytic function of mouse mononuclear-macrophages

2.5.1 Effect of test Agents on mouse monocyte-macrophage carbon clearance function

TABLE 11 Effect of test substances on mouse monocyte-macrophage carbon clearance function

P value: p < 0.05 for each experimental group compared to the negative control group; p < 0.01

As can be seen from Table 11, after the test substances are orally administered to the mice at different dosages for 30 days, the differences of the carbon clearance function of the low-dose group and the negative control group are not significant (P is more than 0.05) and the differences of the medium-high dose group and the negative control group are significant (P is less than 0.05) after the statistical treatment, namely, the test substances in the medium-high dose group can improve the carbon clearance capacity of the mouse mononuclear-macrophage.

2.5.2 Effect of test Agents on mouse body weight and ability of macrophages to phagocytose chicken erythrocytes

TABLE 12 Effect of test substances on mouse body weight and ability of macrophages to phagocytose chicken erythrocytes

Phagocytosis ratio (%) P₁The value: each experimental group was compared with a negative control group

Phagocytic index P₂The value: each experimental group was compared with a negative control group

As can be seen from Table 12, the mice in each dose group had no significant difference in body weight and weight gain (P >0.05) compared with the negative control group after being administered orally to the test object for 30 days by statistical treatment; the phagocytosis rate of each dose group has no significance (P is more than 0.05) compared with that of a negative control group, and the phagocytosis index of each dose group has no significance (P is more than 0.05) compared with that of the negative control group, namely the test object has no obvious influence on the ability of macrophage of a mouse to phagocytize chicken erythrocyte.

2.6 Effect of test Agents on NK cell Activity in mice

TABLE 13 Effect of test substances on NK cell Activity in mice

P value: each experimental group was compared with a negative control group

As can be seen from Table 13, when the test substances are orally administered to the mice at different doses for 30 days, the comparison between the NK cell activity of each dose group and the negative control group has no significance (P is more than 0.05) after statistical treatment, i.e., the test substances have no obvious influence on the NK cell activity of the mice.

3. Small knot

The delayed allergy of the mouse can be enhanced by orally administering different doses of the test substance to the mouse for 30 days, namely the cellular immune function can be enhanced; can improve the carbon clearance capability of mouse mononuclear-macrophage, namely can enhance the function of the mononuclear-macrophage; the composition has no influence on the half hemolysis value of a mouse, the number of antibody-producing cells of the mouse, the weight gain of the mouse, the organ/weight ratio, the chicken erythrocyte phagocytosis capacity of macrophages in the abdominal cavity of the mouse, the spleen lymphocyte transformation capacity of the mouse induced by ConA and the NK cell activity. Thus, example 1 was judged to have an effect of enhancing the immunity function.

Claims

1. A traditional Chinese medicine composition with an immunoregulation function is characterized by being prepared from the following raw materials in parts by weight:

2. The traditional Chinese medicine composition with the immunoregulation function according to claim 1 is characterized by being prepared from the following raw materials in parts by weight:

3. The traditional Chinese medicine composition with the immunoregulation function according to claim 1 is characterized by being prepared from the following raw materials in parts by weight:

4. The traditional Chinese medicine composition with the immunoregulation function according to claim 1 is characterized by being prepared from the following raw materials in parts by weight:

5. The traditional Chinese medicine composition with the immunoregulation function according to claim 1 is characterized by being prepared from the following raw materials in parts by weight:

6. The traditional Chinese medicine composition with the immunoregulation function according to claim 1 is characterized by being prepared from the following raw materials in parts by weight:

7. The preparation method of the traditional Chinese medicine composition with the immunoregulation function, disclosed by any one of claims 1-6, is characterized by comprising the following steps:

drying the extract to obtain the Chinese medicinal composition.

8. The preparation method of the traditional Chinese medicine composition with immunoregulation function according to claim 7, characterized in that: in the step of water decoction and extraction, the adding amount of the water solution is 4-10 times of the total weight of the medicinal materials, the extraction times are 1-4 times, and the extraction time is 0.5-3 hours each time.

9. The traditional Chinese medicine preparation for preparing the traditional Chinese medicine composition with the immunoregulation function, which is disclosed by any one of claims 1 to 6, is characterized in that: comprises a traditional Chinese medicine composition and pharmaceutically acceptable auxiliary materials.

10. The traditional Chinese medicine preparation of the traditional Chinese medicine composition with the immunoregulation function, according to claim 9, is characterized in that: the dosage form of the traditional Chinese medicine preparation is capsules, tablets, granules, dripping pills or oral liquid.