CN110613816A

CN110613816A - Compound astragalus root traditional Chinese medicine composition and preparation method and application thereof

Info

Publication number: CN110613816A
Application number: CN201910871241.6A
Authority: CN
Inventors: 周长新; 甘礼社; 莫建霞; 张丽莎; 程凯航
Original assignee: Hangzhou Xuandun Technology Co Ltd
Current assignee: Hangzhou Xuandun Technology Co Ltd
Priority date: 2019-09-16
Filing date: 2019-09-16
Publication date: 2019-12-27

Abstract

The invention provides a compound astragalus Chinese medicinal composition and preparation and application thereof, wherein the composition is prepared from one or more of monarch drug astragalus, poria cocos, dried rehmannia root and rhodiola root as ministerial drug, one or more of dendrobium officinale, Chinese angelica and radix ophiopogonis as adjuvant drug, and galangal as conductant drug. Extracting with water/ethanol system to obtain soft extract, and mixing with adjuvant to obtain the final product. The invention starts from the aspects of supplementing qi and nourishing yin, enriching the blood and promoting blood circulation, uses astragalus root for supplementing qi and strengthening the body resistance, further matches tuckahoe, radix rehmanniae and rhodiola rosea, is matched with dendrobium, angelica and dwarf lilyturf tuber, combines the whole formula and is assisted with galangal for warming the spleen and the stomach, forms a new formula for nourishing yin and invigorating qi, moistening lung and clearing heart, enriching the blood and promoting blood circulation, strengthening the spleen and nourishing the stomach, has the effects of supplementing qi, blood and yin, can enhance the physique of a human body, improve the immunity of the human body, prevent the occurrence of diseases and effectively regulate the sub-health state, and is proved by in vitro cell experiments and in vivo animal experiments of pharmacology to have various effects of enhancing the immunity, benefiting intelligence, resisting oxidation.

Description

Compound astragalus root traditional Chinese medicine composition and preparation method and application thereof

Technical Field

The invention belongs to the technical field of traditional Chinese medicines, and relates to a compound astragalus Chinese medicinal composition, which is a Chinese medicinal compound composition with tonifying effect and taking astragalus as a main raw material, and a preparation method and application thereof.

Background

With the high-speed development of economy in China, social competition is increasingly violent, the work rhythm is accelerated, and the living pressure of people is sharply increased. This results in many people in sub-health states such as weakness, listlessness, fatigue, weakness, decreased appetite, sleep disturbance, or susceptibility to illness. From the perspective of traditional Chinese medicine, people in sub-health state mostly have the symptoms of deficiency of both qi and yin and insufficiency of essence and blood. The body resistance is insufficient, and the body fails to promote blood circulation, so that qi and blood can not be interconverted, the physiological function of the visceral organs is reduced, the disease resistance of the body is reduced, and the risk of diseases is increased. The basic number of sub-health people in China is large, and the people tend to rise year by year. And with the improvement of living standard, people pay more and more attention to their health. Therefore, the development of the health food capable of improving the immunity of the human body from the aspect of tonifying qi, blood and yin has certain research value.

Astragalus membranaceus is a qi-tonifying medicine in traditional Chinese medicines, is sweet in taste and slightly warm in nature, enters spleen and lung meridians, can strengthen body resistance and consolidate constitution, and has the effects of tonifying qi, raising yang and enhancing immune function [ Kim, C., Ha, H., Kim, J.S., Kim, Y.T., Kwon, S.C., Park, S.W. indication of growth hormone by the roots of Astragalus membranaceus inoculation cell culture. Arch.Pharm. Res.,2003,26:34-39 ]. The traditional Chinese medicine considers that qi is the commander of blood, blood is the mother of qi, and vital qi exists in interior, so that pathogenic factors cannot be dried. When the healthy qi is sufficient, the blood vessels are smooth, and all diseases are not born. Therefore, for sub-health people, qi is the first to invigorate. Modern pharmacological studies show that the pharmacological actions of the traditional Chinese medicine astragalus mainly comprise aspects of regulating immune function, improving cardiac function, controlling blood sugar, inhibiting viruses, resisting tumors, resisting aging and the like [ Yangqi, pharmacological action analysis and clinical application effect evaluation of the traditional Chinese medicine astragalus, Chinese sanatorium, 2019,28 (6): 660-.

In order to better exert the qi tonifying effect of the astragalus, the formula references four basic famous formulas of the traditional famous formula, such as pulse-activating powder, decoction of four ingredients, decoction of four monarchs, decoction of liquid increasing and the like, references 101 traditional Chinese medicines for both medicine and food and 113 traditional Chinese medicines capable of being used for health food, and considers the effect and the safety, and references the research result of modern pharmacology to form an innovative formula which takes the astragalus as a monarch medicine, the poria cocos, the radix rehmanniae and the rhodiola rosea as ministerial medicines, the dendrobium officinale, the angelica sinensis and the radix ophiopogonis as adjuvant medicines and the galangal as a conductant medicine. In the formula, the tuckahoe is sweet, light and flat in taste, and has the functions of promoting diuresis, excreting dampness, strengthening spleen and calming heart, and the modern experimental study researches that the tuckahoe has the effects of inhibiting the growth of bacteria, enhancing the immunity of organisms, strengthening spleen and stomach, excreting dampness and promoting urination, calming the heart and tranquilizing the mind and reducing blood sugar; sheng Di Huang is sweet in flavor and cold in nature. The medicine has the effects of clearing heat, cooling blood, nourishing yin and promoting the production of body fluid, and is suitable for heart, liver and kidney channels; modern pharmacology considers that rhodiola rosea has the efficacy of tonifying qi and activating blood, wherein the pharmacological actions related to the efficacy of tonifying qi mainly comprise myocardial ischemia resistance, arrhythmia resistance, myocardial aging delay, fatigue resistance, immunity enhancement and the like, and the pharmacological actions related to the efficacy of invigorating blood mainly comprise reduction of pulmonary hypertension, improvement of hemorheology, angiogenesis promotion and the like [ Gong et Jun, Chen Su hong, Lugui source, research progress of pharmacological actions related to the efficacy of tonifying qi and activating blood of rhodiola rosea, Anhui medicine, 2008,12(4):292 and 294. ]; dendrobe is sweet in taste and slightly cold, enters kidney and stomach meridians, has the effects of benefiting stomach, promoting fluid production, nourishing yin, clearing heat and the like, and a large number of pharmacological studies prove that the dendrobium officinale has the effects of resisting oxidation, improving immunity, resisting cancer, resisting tumors and the like [ Ding Piao, Nidobei, Wang Meng Mao and the like, medicinal dendrobe research progress, Hubei agricultural science, 2015,54(11): 2561 one-time 2563 ]; dang Gui is recorded in Shen nong Ben Cao Jing (Shen nong's herbal), and its taste is sweet and heavy, so it is specially indicated for tonifying blood and promoting blood circulation; it is mild and pungent in qi, so it can move blood, tonify middle energizer and move middle energizer, and is good for qi in blood and holy herbs in blood. The traditional Chinese medicine is named as ' ten prescriptions and nine subspecies ', and the traditional efficacies of enriching blood and activating blood, regulating menstruation and relieving pain and relaxing bowel are known for people who have the traditional efficacies of ' Jianyali, Wanghui, Chinese angelica property and function test, Chinese medicine guidance and report, 2019,25(11):72-74 and 77. ]; mai Dong is sweet, slightly bitter and slightly cold in flavor, and enters heart, lung and stomach meridians. The ophiopogon polysaccharide has the effects of reducing blood sugar, enhancing immunity, resisting myocardial ischemia, resisting allergy, relieving asthma and increasing the hypoxia tolerance of organisms; galangal is pungent and warm in nature, mainly enters spleen channels and stomach channels, has the effects of warming spleen and stomach, preventing vomiting, stopping diarrhea, dispelling cold, relieving pain and the like, and is mainly used for treating diseases such as epigastric and abdominal psychroalgia, vomiting, diarrhea, dyspepsia and the like [ Modan, willow, Qinhui ice and the like, galangal and other 4 kinds of galangal belong to the herbal literature research of the drug property and the effect of traditional Chinese medicines, Chinese traditional medicine journal, 2018,43(13):2648 and 2653 ].

Disclosure of Invention

The invention aims to provide a compound astragalus root Chinese medicinal preparation which has the tonifying effect and consists of astragalus root, tuckahoe, root of rehmannia, rhodiola root, dendrobium officinale, angelica, dwarf lilyturf tuber and galangal. The preparation of each 1000 dosage units is composed of the following raw materials in proportion: 6-30 parts of monarch drug astragalus, preferably 9-18 parts; 4-20 parts of ministerial drugs of poria cocos, dried rehmannia root and rhodiola rosea respectively, and 6-16 parts of the ministerial drugs are preferred; 2-10 parts of adjuvant drug dendrobium officinale, angelica sinensis and ophiopogon root respectively, preferably 3-9 parts; 1-5 parts of galangal, preferably 1-2 parts.

The invention also aims to provide the application of the compound astragalus mongholicus traditional Chinese medicine composition in preparing medicines for resisting fatigue, resisting oxidation, benefiting intelligence and enhancing immunity. The preparation form of the medicine is oral dosage forms such as tablets, pills, granules or capsules.

The invention further aims to provide a preparation method of the medicine, which adopts a two-step extraction method and is realized by the following steps:

(1) soaking the medicinal materials in the composition in 6-30 times of 50-95% ethanol for 2-12h, reflux-extracting for 1-2 times (2-3 h each time), and filtering to obtain ethanol extract, wherein the solvent is 8-12 times of the amount, the ethanol concentration is 65-85% by volume, and the soaking time is 4-8 h;

(2) decocting the medicinal materials in the composition extracted with alcohol in step 1 with 6-30 times of water for 1-2 times, each time for 2-3h, filtering to remove residue to obtain water extractive solution, wherein the amount of water is preferably 10-15 times, and the decocting temperature is preferably 80-100 deg.C;

(3) concentrating the water extractive solution and the ethanol extractive solution at 50-70 deg.C under reduced pressure to about 1.2 times of the medicinal materials (weight/volume ratio) to obtain soft extract, mixing with pharmaceutically acceptable adjuvants, and making into medicine.

In the composition provided by the invention, the astragalus is a monarch drug, is sweet in taste and slightly warm, enters spleen and lung channels, can strengthen the body resistance and consolidate the constitution, and has the effects of tonifying qi, invigorating yang and enhancing the immune function; the ministerial drugs can be one or more of radix rehmanniae, Poria and radix Rhodiolae, wherein Poria is selected for invigorating spleen and tranquilizing mind, radix rehmanniae is selected for nourishing yin and promoting fluid production, radix Rhodiolae is selected for invigorating qi and promoting blood circulation, and the three drugs can be used together for invigorating qi and strengthening body resistance of radix astragali, nourishing stomach and invigorating spleen, and promoting the generation and production of qi and blood; the adjuvant drugs can be one or more of dendrobium officinale, Chinese angelica and dwarf lilyturf tuber, wherein the dendrobium officinale can tonify stomach and promote fluid production, nourish yin and clear heat, the Chinese angelica can enrich blood and activate blood, the dwarf lilyturf tuber can nourish yin and promote fluid production, moisten lung and clear heart, and the three drugs can nourish yin and moisten dryness, enhance the transportation and transformation of spleen and stomach and assist the circulation of qi and blood. When the seven medicines are used together, because the rehmannia root, the dendrobium officinale and the radix ophiopogonis are cold in nature and taste, yin is nourished, the spleen and stomach yang qi is prevented from being damaged, and the galangal is added as a guiding medicine to warm the spleen and stomach, so that the effects of tonifying qi and invigorating yang, nourishing yin and promoting the production of body fluid, enriching and activating blood and balancing yin and yang are achieved. On the basis of the recipe, the ministerial drugs and the adjuvant drugs are partially deleted, so that partial effects of the recipe can be obtained.

The invention starts from the aspects of supplementing qi and nourishing yin, enriching the blood and promoting blood circulation, takes the astragalus root which can tonify qi and strengthen the body resistance as a monarch drug, and further combines the tuckahoe, the radix rehmanniae and the rhodiola rosea, is supplemented with the dendrobium, the angelica and the radix ophiopogonis, and the galangal is used for warming the spleen and the stomach in the whole formula, so that an innovative prescription which can nourish yin and tonify qi, moisten the lung and clear the heart, enrich the blood and promote blood circulation, invigorate the spleen and nourish the stomach is formed.

Drawings

FIG. 1 shows the ConA-induced splenic lymphocyte proliferation assay (N3, 24hr) with the compound extract (DNAS).

FIG. 2 shows the ConA-induced splenic lymphocyte proliferation assay (N3, 48hr) with the compound extract (DNAS).

FIG. 3 shows the ConA-induced splenic lymphocyte proliferation assay (N3, 72hr) with the compound extract (DNAS).

FIG. 4 shows the optimal concentration and time of ConA-induced splenic lymphocyte proliferation in mice by the compound extract (DNAS).

FIG. 5 shows the effect of the combination of the extract of Fufang (DNAS) with the protein of mitogen Canavalia (ConA) and the lipopolysaccharide of mitogen (LPS) on lymphocyte proliferation.

Detailed Description

The invention will now be described in further detail with reference to the accompanying drawings and specific examples, which are set forth to illustrate, but are not to be construed to limit the invention.

EXAMPLE 1 preparation of tablets

The prescription of the composition is as follows: astragalus (300g), tuckahoe (200g), dried rehmannia root (200g), rhodiola (200g), dendrobium (100g), angelica (100g), ophiopogon root (100g) and galangal (50 g).

The preparation process comprises the following steps: soaking 1250g of the raw materials in 12 times of 75% ethanol for 6h, heating and reflux-extracting for 2 times, each time for 3h, and filtering to remove residue to obtain alcoholic extract; boiling the medicinal materials with 20 times of water for 2 times, each for 3 hr, and filtering to remove residue to obtain water extractive solution. Concentrating the water extractive solution and the ethanol extractive solution at 60 deg.C to about 1.2 times of the medicinal materials (weight/volume ratio) to obtain soft extract. Mixing the soft extract with 600g soluble starch, 1000g lactose, and 300g mannitol, adding appropriate amount of 50% ethanol as wetting agent, making into soft material, sieving, granulating, oven drying, adding 1% magnesium stearate, tabletting, and packaging.

EXAMPLE 2 preparation of pellets

The prescription of the composition is as follows: astragalus root (200g), poria cocos (160g), dried rehmannia root (120g), rhodiola root (100g), dendrobium (40g), Chinese angelica root (80g) and ophiopogon root (60 g).

The preparation process comprises the following steps: soaking the raw materials (760g) in 15 times of 50% ethanol for 8hr, heating and reflux-extracting for 2 times (each for 3 hr), and filtering to remove residue to obtain ethanol extract; boiling the medicinal materials with 10 times of water for 2 times, each for 3 hr, and filtering to remove residue to obtain water extractive solution. Concentrating the water extractive solution and the ethanol extractive solution at 50 deg.C to about 1.2 times of the medicinal materials (weight/volume ratio) to obtain soft extract, drying, pulverizing into fine powder, adding refined honey 120g, 50% ethanol and hydroxypropyl methylcellulose 200g, mixing, making into honeyed pill with YUJ-16A full-automatic numerical control Chinese medicinal pill making machine, drying, and polishing.

EXAMPLE 3 preparation of granules

The prescription of the composition is as follows: astragalus root (120g), poria cocos (60g), dried rehmannia root (40g), dendrobium stem (40g), Chinese angelica root (20g) and ophiopogon root (60 g).

The preparation process comprises the following steps: soaking 340g of the raw materials in 15 times of 50% ethanol for 8hr, heating and reflux-extracting for 2 times (each for 3 hr), and filtering to remove residue to obtain alcoholic extract; boiling the medicinal materials with 10 times of water for 2 times, each for 3 hr, and filtering to remove residue to obtain water extractive solution. Concentrating the water extractive solution and the ethanol extractive solution at 50 deg.C to about 1.2 times of the medicinal materials (weight/volume ratio) to obtain soft extract. Mixing the soft extract with 100g microcrystalline cellulose, 50g soluble starch and 80g mannitol, adding appropriate amount of 50% ethanol as wetting agent, making into soft material, sieving, granulating, oven drying, and packaging to obtain granule.

EXAMPLE 4 preparation of capsules

The prescription of the composition is as follows: astragalus root (240g), poria cocos (200g), dried rehmannia root (160g), dendrobium stem (80g) and ophiopogon root (80 g).

The preparation process comprises the following steps: soaking the raw materials (760g) in 15 times of 50% ethanol for 8hr, heating and reflux-extracting for 2 times (each for 3 hr), and filtering to remove residue to obtain ethanol extract; boiling the medicinal materials with 10 times of water for 2 times, each for 3 hr, and filtering to remove residue to obtain water extractive solution. Concentrating the water extractive solution and the ethanol extractive solution at 50 deg.C to about 1.2 times of the medicinal materials (weight/volume ratio) to obtain soft extract. Mixing the soft extract with 300g soluble starch, 400g lactose, and 150g mannitol, adding appropriate amount of 50% ethanol as wetting agent, making into soft material, sieving, granulating, oven drying, and encapsulating in No. 1 capsule.

Example 5 fatigue resistance test

1. Material

Experimental animals: kunming mice (Swiss line), 6 weeks old, male, purchased at Nanjing institute of biomedical sciences, tested for animal quality certification: SCXK (threo) 2015-0001. The drugs were prepared by the laboratory in the same manner as in examples 1-4, and administered to the high dose group at 200mg/kg, the medium dose group at 100mg/kg, and the low dose group at 50 mg/kg.

2. Experimental methods

2.1 animal screening and formal test 60 mice were weighed, loaded at 10% of the weight, and placed in a constant temperature bucket at 26 ℃ with a water depth of 22cm and 3 mice per bucket. The swimming time of the mice (5 s during swimming to the head is taken as a mark) is recorded, and the mice are rejected when the time is shorter than 4min and longer than 15 min. The selected 40 mice were divided into 4 groups by swimming time and body weight at screening, 10 mice per group, and administered at 9 am daily for 28 days. And weighing at 28d, administering for 1h, performing a weight swimming test, screening under the same test conditions, and recording the swimming time of the mice.

2.2 animals were divided into 4 groups of 40 mice randomly, each group was a placebo group and a drug high, medium and low dose group.

2.3 statistical methods all data are expressed in x. + -.s, and t-tests are compared between the two groups.

3. As a result: the swimming time of the mice can be obviously improved in the medium and high drug dose groups compared with the blank group, and the results are shown in table 1.

TABLE 1 Effect of the combination on swimming time of mice: (n＝10)

Example 6 enhanced immunization experiments

1. Materials and instruments:

RPMI1640 cell culture solution, calf serum, 2-mercaptoethanol (2-ME), penicillin, streptomycin, concanavalin A (ConA), hydrochloric acid, isopropanol, MTT, Hank's solution, and PBS buffer solution (pH 7.2-7.4).

Gauze, 200-mesh screen, 24-hole culture plate, 96-hole culture plate (flat bottom), surgical instruments, large-size injector inner core, carbon dioxide incubator, microplate reader, spectrophotometer, super clean bench, autoclave and sterile filter.

2. The experimental method comprises the following steps: ConA-induced splenic lymphocyte transformation experiment (MTT method) for mice

2.1 preparation of reagents

Complete culture solution: RPMI1640 culture solution is sterilized by filtration, and 10% calf serum, 1% glutamine (200mmol/L), penicillin (100U/mL), streptomycin (100ug/L) and 5 × 10 are added before use^-5The pH of the 2-mercaptoethanol solution is adjusted to 7.0-7.2 by sterile 1mol/L HCl or 1mol/L NaOH, and the culture solution is completely cultured.

ConA liquid: preparing 100ug/mL solution with double distilled water, filtering for sterilization, and storing in a low temperature refrigerator (-20 deg.C).

Sterile Hank's solution: before use, the pH is adjusted to 7.2-7.4 with 3.5% sterile NaHCO 3.

MTT solution: 5mg of MTT was dissolved in 1mL of PBS (pH7.2) and used as it is.

Acidic isopropanol solution: to 96mL of isopropanol was added 4mL of 1mol/L HCl and the mixture was prepared just before use.

2.2 spleen cell suspension preparation

The spleen was aseptically removed and placed in a dish containing an appropriate amount (3-5ml) of sterile Hank's solution, and a piece of gauze was placed over the spleen, which was gently ground with a large syringe plunger to make a single cell suspension. Filtered through a 200-mesh screen and washed 2 times with Hank's solution, centrifuged 10min (1000r/min) each time. Then, the cells were suspended in 2mL of complete culture medium, and spleen cells were counted by a full-automatic cell counter, or viable cells were counted by staining with Tulipan (which should be 95% or more), and the cell concentration was adjusted to 3X 10⁶one/mL.

2.2.3 investigation of the optimal concentration and time of the compound extract (DNAS) on the proliferation of splenic lymphocytes of mice.

The 96-well plate is taken and added with the liquid medicine DNAS with gradient concentration (1.28mg/ml, 640ug/ml, 320ug/ml, 160ug/ml, 80ug/ml, 40ug/ml, 20ug/ml, 10ug/ml and 5 ug/ml). Adding 1ml of mouse spleen lymphocytes, and adjusting the cell density to 5 × 10⁶ML/ML. Setting 3 parallel experiments at the same time in each concentration of 2 multiple wells, culturing at 37 ℃ for 24, 48 and 72 hours respectively under the condition of 5% CO2, adding 20 mu L (5g/L) of MTT into each well, continuously incubating for 4 hours, discarding supernatant, adding 150 mu L of MTT into each well to dissolve MTT formazan precipitate, shaking and mixing uniformly by a micro-oscillator, and measuring absorbance A570nm at the wavelength of 570nm by an enzyme-linked immunosorbent assay after color change to serve as an index of spleen lymphocyte proliferation. The results are shown in FIGS. 1-4.

The results show that the optimal concentration of the compound extract (DNAS) on mouse spleen lymphocyte proliferation is 40, 80 and 160ug/ml, and the optimal time is 48 h.

2.4 Compound extract (DNAS) on lymphocyte proliferation reaction

The experimental groups include resting-stage cell control group (DNAS), mitogen canavalin (ConA) -stimulated T cell + DNAS drug group, and mitogen Lipopolysaccharide (LPS) -stimulated B cell + DNAS drug group.

The cell suspension was added to a 24-well plate at 1mL per well, and 75. mu.l of ConA solution (equivalent to 7.5. mu.g/mL) was added to one well, and the other well was used as a control, and incubated in a 5% CO2 incubator at 37 ℃ and CO2 for 48 hours. 4 hours before the end of the culture, 0.7mL of the supernatant was gently aspirated into each well, and 0.7mL of RPMI1640 medium containing no calf serum was added thereto together with 50. mu.l/well of MTT (5mg/mL), and the culture was continued for 4 hours. After the culture is finished, 1mL of acidic isopropanol is added into each hole, and the mixture is uniformly stirred by ultrasonic oscillation (2 seconds) or manually blown to dissolve the purple crystals completely. Then, the cells were individually plated in 96-well plates, each well was plated in 3 wells as a parallel sample, and the optical density was measured at a wavelength of 570nm using an enzyme-linked immunoassay instrument. The solution can also be transferred directly into a 2mL cuvette and the OD measured at 570nm on a spectrophotometer. The results of the experiment are shown in FIG. 5.

The above results indicate that, compared with the resting-stage cell control group (DNAS), at a concentration of 20ug/ml, mitogen Lipopolysaccharide (LPS) stimulates the B cell + DNAS drug group to significantly promote the proliferation of lymphocytes (p < 0.05), and at a concentration of 40ug/ml, mitogen canavalin (ConA) stimulates the T cell + DNAS drug group and mitogen Lipopolysaccharide (LPS) stimulates the B cell + DNAS drug group to significantly promote the proliferation of lymphocytes (p < 0.05). It can be seen that DNAS has a significant synergistic effect on LPS and ConA induced B lymphocyte proliferation.

Example 7 Effect on phagocytosis of Chicken erythrocytes by macrophages in mouse peritoneal Cavity (in vivo method)

1. Experimental methods

Mice were weighed and randomized into 5 groups, (1) normal group: equal volume of CMC-Na solution; (2) lentinan group: 0.01 g/kg^-1(ii) a (3) Low dose group: 0.3 g.kg^-1(corresponding to 5 times of the recommended dosage for human body) (4) middle dose group: 0.6 g.kg^-1(ii) a (5) high dose group: 0.12 g/kg^-1. Each group of animals was administered 10d by continuous gavage.

Each mouse was injected with 1ml of 20% chicken red blood cell suspension. At an interval of 30min, the animal is sacrificed by dislocation of cervical vertebrae, fixed on a mouse plate in an upward position, the abdominal wall skin is cut off at the center, 2ml of physiological saline is injected into the abdominal cavity, and the mouse plate is rotated for 1 min. Then sucking out 1ml of abdominal cavity washing solution, evenly dropping on 2 glass slides, placing in an enamel box padded with wet gauze, and moving out of an incubator at 37 ℃ for incubation for 30 min. After incubation, the cells were rinsed in physiological saline to remove non-adherent cells. Drying in air, and mixing the raw materials in a ratio of 1: fixing with 1 acetone methanol solution, staining with 4% (v/v) Giemsa-phosphate buffer solution for 3min, rinsing with distilled water, and air drying. Macrophages were counted under oil lens, 100 per tablet, and percent phagocytosis and phagocytosis index were calculated as follows.

Percent (%) phagocytosis ═ number of macrophages engulping chicken erythrocytes × number of macrophages per count

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

2. Results of the experiment

The phagocytic function of macrophages can be judged by the ability of abdominal macrophages to phagocytose chicken erythrocytes. The results show that the phagocytic indexes of abdominal cavity macrophages of the mice in the medium dose group on chicken erythrocytes are higher than those of the mice in the normal group (P < 0.05), and the medicaments in the medium dose group have the effect of promoting the phagocytic function of the macrophages of the mice (see table 2). Meanwhile, when the degree of digestion of the chicken red blood cells is observed, the cytoplasm of the normal group of chicken red blood cells is light yellow green, the nucleus is condensed to be purple blue, and the chicken red blood cells belong to II-grade mild digestion; the cytoplasm of the chicken red blood cells of each administration group is lightly dyed, and the nucleus is light grey, and belongs to III-level severe digestion; only vacuoles with the shape similar to the size of chicken erythrocyte are seen in a small amount of macrophages, the edges are neat, the chicken erythrocyte can be seen in the hidden nucleus, and the digestion is complete in IV level.

TABLE 2 influence of DNAS on phagocytosis of chicken erythrocytes by macrophages of mouse peritoneal cavity

Group of	Dosage (g/kg)	Animal number (only)	Percentage of phagocytosis (%)	Phagocytic index
					Normal group	—	20	31.9±4.5	0.84±0.141
Lentinan	0.01	20	48.3±5.8	1.039±0.101^*
					Low dose group	0.30	20	37.6±9.6	0.925±0.203
Middle dose group	0.60	20	45.5±6.6	1.095±0.131^*
					High dose group	1.20	20	41.6±4.7	1.006±0.151

P < 0.05 compared to normal group

Example 8: carbon clearance test of mice

1. Instruments and reagents:

spectrophotometer, full-automatic enzyme marker, timer, hemoglobin straw, India ink and Na₂CO₃

2. Experimental procedure

2.1 preparation of the solution

The ink for injection is prepared by diluting india ink stock solution by 3-4 times with normal saline.

Na₂CO₃Taking 0.1g of Na₂CO₃Distilled water was added to 100 mL.

2.2 Experimental methods

Randomly dividing mice (18-22 g, each half of male and female) into 5 groups, namely a control group, a positive group (lentinan), a DNAS group (0.31g/kg, 0.62g/kg, 1.24g/kg), performing intragastric administration for three days, and injecting India ink 0.05-0.1 ml/10g of body weight into each rat tail vein 30 minutes after the last administration for 1min (t₁) And 5min (t)₂) Then, 20. mu.l of blood was collected from orbital vein of each mouse, and 2ml of 0.1% NaCO was added₃Shaking the solution evenly, measuring the colorimetric density (OD) under 680nm of a spectrophotometer, and calculating the clearance index (K).

3. Results of the experiment

The carbon clearance index of the model group mouse is obviously reduced; the clearance index of carbon granules in the positive medicine group is obviously increased; the carbon clearance index of mice in each dose group of the drug is increased, and the carbon clearance index of mice in the medium dose group is the highest.

TABLE 3 influence of DNAS on mouse carbon clearance index

Group of	Dosage (g/kg)	Animal number (only)	Index of clearance
				Normal group	—	20	4.045±1.16
Lentinan	0.01	20	5.27±1.10^*
				Low dose group	0.31	20	4.76±1.35
Middle dose group	0.62	20	5.13±1.51
				High dose group	1.24	20	5.08±1.21^*

P < 0.05 compared to normal group

Example 9: experiment for developing intelligence

1. Material

Mouse open field devices (shanghai chunlong science and technology development ltd); SLY-WMS model Morris Water maze (Beijing major Aster science, Inc.); FSH-2A adjustable high-speed electric homogenizer (Jinnan Instrument plant, gold Tan, Inc.); LXJ-IIB Low speed high capacity Multi-tube centrifuge (Najing Xiaoxiao Instrument Equipment Co., Ltd.); powerwave X340 full-wavelength microplate reader (Bio-Tek, USA); d-galactose (Shanghai Baoman Biometrics, Inc.); clean-grade Kunming mouse, certification number: SCXK (Su) 2009-.

2. Method and results

2.1 Experimental methods

KM male mice, weight 20 ~ 24 g. Groups were randomized into 4 groups of 10 individuals each. Namely a blank control group and a positive control group (oxiracetam 0.6g kg)^-1) Compound preparation large dose group (0.3 g.kg)^-1) Small dose group (0.15 g.kg)^-1). The model group and the administration group are injected with D-galactose 500 mg/kg intraperitoneally every morning^-1·d^-1The blank group was injected with equal amount of normal saline, the administration group in the afternoon was separately gavaged with drugs of corresponding concentration, the blank group and the model group were separately gavaged with equal amount of normal saline, and the dosage was 0.1ml·10g^-1And continues for 21 d.

Morris Water maze experiment

Positioning navigation experiment: training was performed daily after injection and gavage 1 time each in the morning and afternoon for 5 days, starting from the 14 th day of modeling and administration. And (3) facing the wall of the pool, fixing a water inlet point from the opposite angle quadrant of the platform, putting the water inlet point into water, recording the time for finding the platform within 120s, if the platform cannot be found, leading the platform to the upper side for staying for 10s, putting the platform back into the cage, and recording the latency as 120 s. Space exploration experiment: and 24h after the positioning navigation experiment is finished, moving out the platform, fixing the mouse into the water from the opposite angle quadrant of the platform, putting the mouse into the water facing the pool wall, and recording the times of passing through the original platform position within 120s so as to measure the acquisition capacity of the mouse on the water maze memory.

The results show that: the latency and crossing times of each experimental group are different, and the latency of the mice in the model group is obviously increased (P is less than 0.01) and the crossing times are reduced compared with those in the control group. The incubation period was significantly reduced and the number of crossings (P < 0.05, P < 0.01) was significantly increased in each of the other administration groups compared to the model group (see Table 4).

TABLE 4 Effect of FUFANGS on learning and memory of Alzheimer's mice (Morris water maze) (n ═ 10)

Group of	Dosage (mg/kg)	Mean incubation period(s)	Number of passes through original platform
				Control group	--	73.2±24.8^▲▲	5.12±1.76^▲▲
Model set	--	97.2±44.0	3.80±1.65
				Compound small dose group	150	85.5±39.4^▲	4.05±1.87^▲
Compound large dose group	300	78.2±25.1^▲▲	4.76±1.98^▲▲
				Oxiracetam group	600	78.6±30.1^▲▲	4.83±2.77^▲▲

Note: in comparison to the set of models,^▲P＜0.05，^▲▲P＜0.01；

example 10: oxidation resistance test

1. Experimental Material

1.1 pharmaceutical products and their preparation

The drug was prepared in the laboratory by the method of examples 1-4, with DMSO dissolved, and the positive control vitamin C (Vc) as a product of national drug group chemical reagents, Inc.

During primary screening, Vc is prepared into 1mg/ml by double distilled water, a medicine is prepared into 1mg/ml by dimethyl sulfoxide (DMSO), and the medicine and a positive control are added into a reaction system according to the proportion of 1%. According to the result of primary screening, selecting a test object with an inhibition rate of more than 50% and a Vc solution, and diluting the test object and the Vc solution into a series of solutions with equal specific concentrations by respective solvent multiple ratio before use. Determination of IC₅₀In values, drug and Vc concentrations were 1.0, 0.5, 0.25, 0.13, and 0.0625mg/ml, respectively.

2.2 Primary reagents

DPPH was purchased from SIGMA-ALDRICH, and was sold as STBB0510A 0.

2.3 instruments

Desk type enzyme-labeling instrument (Elx 800 type, Bio-Tek company)

3. Experimental methods

3.1 principles and methods of the experiment

DPPH is a very stable nitrogen-centered radical and is widely used to measure the antioxidant capacity of biological samples, phenolics and foods. The method is based on the characteristic that DPPH free radical has single electron, strong absorption is provided at 517nm, and alcoholic solution is purple. In the presence of a free radical scavenger, its absorption gradually disappears by pairing with its single electron, and its degree of discoloration is quantitatively related to the number of electrons received, so that a rapid quantitative analysis can be performed with a spectrophotometer.

2ul of the sample solution was added to 200ul of 25. mu.g/ml DPPH methanol solution, mixed well, and allowed to stand at room temperature for 30 min. The absorbance was measured at 517nm using methanol as a blank. The same experiment was repeated 3 times.

3.2 data processing and statistical methods

The radical inhibition ratio was calculated as follows:

inhibition (%) - (A517 solvent-A517 sample)/A517 solvent X100%

Radical scavenging activity was expressed as the half inhibition (IC 50). Calculating IC by Loit method according to the percentage of free radical inhibition at the corresponding concentration of each compound₅₀The value is obtained. Mean ± sd for dataAnd (4) showing.

4. Results of the experiment

The clearance rates of DPPH of reference vitamin C (Vc)10 mu g/ml and compound vitamin C10 mu g/ml are 81.3% and 72.7%, respectively. See table 5.

TABLE 5 DPPH scavenging action of the Compound

Test article	Assay concentration (μ g/ml)	Maximum inhibition ratio (%)
			Vc	10	81.3
Compound recipe	10	72.7

Compound experiment and calculating IC₅₀The value is obtained.The experimental results show that the removing effect of the reference Vc and the compound on DPPH is enhanced along with the increase of the concentration, and IC₅₀The values were 2.64. mu.g/ml and 2.35. mu.g/ml, respectively. See table 6.

Table 6 IC50 values for DPPH scavenging (n ═ 3)

Test article	IC₅₀(μg/ml)
		Vc	2.64±0.03
NXF	2.35±0.03

5. Conclusion of the experiment

The compound has strong antioxidant effect, and can remove DPPH IC₅₀The value was 2.35. mu.g/ml, and the radical scavenging titer was comparable to Vc (2.64. mu.g/ml).

The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the present invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention.

Claims

1. The compound astragalus Chinese medicinal composition is characterized in that each 1000 dosage units of the preparation consists of the following raw materials in proportion: 6-30 parts of astragalus membranaceus, 4-20 parts of poria cocos, radix rehmanniae and rhodiola rosea respectively, 2-10 parts of dendrobium officinale, angelica sinensis and radix ophiopogonis respectively and 1-5 parts of galangal.

2. The compound astragalus mongholicus traditional Chinese medicine composition according to claim 1, wherein a preparation of each 1000 dosage units of the composition is composed of the following raw materials in proportion: 9-18 parts of astragalus membranaceus, 6-16 parts of poria cocos, radix rehmanniae and rhodiola rosea respectively, and 3-9 parts of dendrobium officinale, angelica sinensis and radix ophiopogonis respectively; 1-2 parts of galangal.

3. The use of the compound astragalus mongholicus traditional Chinese medicine composition according to claim 1 or 2 in the preparation of medicines for resisting fatigue, resisting oxidation, benefiting intelligence and enhancing immunity.

4. The use according to claim 3, wherein the medicament is formulated in the form of tablets, pills, granules or capsules.

5. The use according to claim 3, wherein the medicament is prepared by the following method of preparation:

(1) soaking the medicinal materials in the composition in 6-30 times of 50-95% ethanol for 2-12h, reflux-extracting for 1-2 times (2-3 hr each time), and filtering to obtain ethanol extract;

(2) decocting the medicinal materials in the composition extracted by the alcohol in the step 1 with 6-30 times of water for 1-2 times, each time for 2-3h, and filtering to remove residues to obtain water extract;

6. The use according to claim 5, wherein the solvent in the amount of 8-12 times in the step (1), the ethanol concentration is 65-85% by volume, and the soaking time is 4-8 hours.

7. The use of claim 5, wherein the amount of water used in step (2) is 10-15 times and the decocting temperature is 80-100 ℃.

8. The compound astragalus mongholicus traditional Chinese medicine composition according to claim 1, which can be prepared into various oral dosage forms including tablets, pills, granules, capsules and the like by adopting a corresponding preparation technology after extraction.

9. The compound astragalus mongholicus traditional Chinese medicine composition of claim 1 is proved to have remarkable effects of resisting fatigue, resisting oxidation, benefiting intelligence and enhancing immunity through pharmacological experiments.