CN116726117B

CN116726117B - A Chinese medicine composition for resisting gouty arthritis

Info

Publication number: CN116726117B
Application number: CN202310974318.9A
Authority: CN
Inventors: 张冰; 林志健; 王雨; 吴昊
Original assignee: Beijing University of Chinese Medicine
Current assignee: Beijing University of Chinese Medicine
Priority date: 2022-08-08
Filing date: 2023-08-03
Publication date: 2025-03-25
Anticipated expiration: 2043-08-03
Also published as: CN116726117A

Abstract

本发明提供了一种抗痛风性关节炎的中药组合物。该组合物的原料药组成为：菊苣、茯苓、土茯苓、槐米、秦艽，为祛除关节尿酸结晶进而抗痛风性关节炎提供一种新的药物选择。本发明所述的中药组合物可显著减少关节尿酸结晶，降低关节液尿酸水平，同时，明显改善痛风性关节炎鹌鹑关节肿胀度，降低血清中炎症因子白介素‑1β水平，并对肝、肾功能无影响，具抗痛风性关节炎的药理作用。The present invention provides a Chinese medicine composition for anti-gouty arthritis. The raw materials of the composition are composed of chicory, Poria cocos, Smilax glabra, Sophora japonica, and Gentiana macrophylla, which provides a new drug option for removing uric acid crystals in joints and anti-gouty arthritis. The Chinese medicine composition of the present invention can significantly reduce uric acid crystals in joints, reduce the level of uric acid in joint fluid, and at the same time, significantly improve the swelling degree of quail joints with gouty arthritis, reduce the level of inflammatory factor interleukin-1β in serum, and have no effect on liver and kidney function, and has a pharmacological effect of anti-gouty arthritis.

Description

Traditional Chinese medicine composition for resisting gouty arthritis

Technical Field

The invention relates to a traditional Chinese medicine composition, in particular to a novel traditional Chinese medicine composition for removing joint uric acid crystals and resisting gouty arthritis by compatibility of chicory, tuckahoe, glabrous greenbrier rhizome, pagodatree flower bud and gentiana macrophylla.

Background

Gouty arthritis is a progressive, disabling metabolic disease closely related to uric acid crystallization, and also has a tendency to develop and younger worldwide. Currently common drugs for gouty arthritis include non-steroidal anti-inflammatory drugs (NSAIDs), colchicine, and glucocorticoids. Among them, NSAIDs block Prostaglandin (PGE) production mainly by inhibiting Cyclooxygenase (COX) activity, colchicine reduces inflammatory substances production mainly by preventing neutrophil chemotaxis, and glucocorticoids exert anti-inflammatory and analgesic effects mainly by inhibiting arachidonic acid release. Although the medicine has definite action target points and stronger pharmacological activity, the medicine is mostly limited to the intervention of a single pathological stage, and is clinically used together to meet the comprehensive treatment requirement of gout. This not only increases the medication burden on the patient, but also potentially creates a safety risk for multi-drug combinations.

At present, the research on the acute gouty arthritis is temporarily free from radical medicines, mainly anti-inflammatory and analgesic medicines, but most medicines have side effects such as gastrointestinal reactions and the like, and the treatment effect is affected. The traditional operation for treating gouty arthritis is large in general operation traumas, the pathological change degree in the joint cannot be accurately judged, and meanwhile, the traditional operation is very easy to infect, has more adverse reactions and complications and has poor clinical effect when being used for operation treatment. The general consensus of the medical science of Chinese hyperuricemia related diseases indicates that 'gout is the occurrence of urate crystal deposition of hyperuricemia patients, which leads to gouty arthritis, uric acid nephropathy and kidney stones', corrects the limitation recognition that gout is only defined as inflammatory joint diseases, and defines the pathological characteristics of gout of urate deposition. This also indicates that the pathogenesis of gouty arthritis is that uric acid crystals appear at joints, and how to remove the uric acid crystals of joints to resist gouty arthritis is also a problem which needs to be solved in clinic at present.

The traditional Chinese medicine has advantages in preventing and treating advanced diseases related to uric acid metabolic disorders such as gouty arthritis and the like. The composition can remove joint uric acid crystals, further play the role of resisting gouty arthritis, invigorate spleen, remove dampness and turbid urine, promote joint circulation, regulate uric acid metabolism, remove wind-damp and arthralgia, intervene gouty inflammation and swelling and pain, clear heat and cool blood, diminish inflammation and relieve pain of the pagodatree flower bud. The whole formula has the effects of strengthening spleen, eliminating dampness, eliminating turbid pathogen, reducing acid, reducing swelling and relieving pain. Modern researches can be reported in the literature of partial medicines for preventing and treating gouty arthritis, however, the research report of the medicine combination for removing the uric acid crystals of joints to prevent gouty arthritis is not yet known at present.

In addition, the chicory, the poria cocos and the pagodatree flower bud in the traditional Chinese medicine compound combination have the characteristics of dual purposes of medicine and food, and are high in safety. Wherein, chicory, tuckahoe and pagodatree flower bud are all varieties collected in the article list which is published by the Ministry of health and is both food and medicine, and rhizoma smilacis glabrae is a variety collected in the article list which can be used for health food.

Disclosure of Invention

The invention aims to provide a traditional Chinese medicine composition for removing joint uric acid crystals and resisting gouty arthritis, and provides a new medicine choice for preventing and treating gouty arthritis.

The invention provides a traditional Chinese medicine composition for treating gouty arthritis, which is prepared from the following raw materials of chicory, poria cocos, smilax glabra, sophora flower bud and gentiana macrophylla.

Preferably, the traditional Chinese medicine composition is prepared from 6-18 parts by weight of chicory, 5-15 parts by weight of poria cocos, 10-60 parts by weight of rhizoma smilacis glabrae, 5-10 parts by weight of pagodatree flower bud and 3-10 parts by weight of gentiana macrophylla.

Further preferably, the traditional Chinese medicine composition is prepared from 6-10 parts by weight of chicory, 10-15 parts by weight of poria cocos, 20-40 parts by weight of rhizoma smilacis glabrae, 5-10 parts by weight of pagodatree flower bud and 3-10 parts by weight of gentiana macrophylla.

Most preferably, the traditional Chinese medicine composition is prepared from 9 parts by weight of chicory, 10 parts by weight of poria cocos, 30 parts by weight of rhizoma smilacis glabrae, 6 parts by weight of pagodatree flower bud and 9 parts by weight of gentiana macrophylla.

In the technical scheme, the traditional Chinese medicine composition can be in any form formed by or prepared from the raw materials, and comprises a composition prepared by respectively crushing the raw materials and then mixing the raw materials, or a composition prepared by crushing the raw materials after mixing the raw materials, or an extract obtained by extracting the raw materials according to a conventional extraction method after mixing the raw materials, wherein the extract is further subjected to a refining and purifying process to obtain an effective part, and the extract and the effective part are further prepared into a conventional oral preparation according to a conventional preparation process.

The conventional extraction method comprises soaking extraction, decoction extraction, reflux extraction, percolation extraction, ultrasonic extraction, microwave extraction and the like, wherein the extraction solvent comprises water or conventional organic solvents such as ethanol, methanol, ethyl acetate, petroleum ether, isopropanol and the like, and the refining and purifying process comprises extraction, column chromatography separation, high performance liquid chromatography separation and the like.

The conventional oral dosage forms comprise tablets, capsules, granules, pills, powder and oral liquid. The preparation method comprises adding pharmaceutically acceptable adjuvants including filler, disintegrating agent, lubricant, suspending agent, binder, sweetener, correctant, antiseptic, matrix, etc. The filler comprises starch, pregelatinized starch, lactose, mannitol, chitin, microcrystalline cellulose, sucrose and the like, the disintegrating agent comprises starch, pregelatinized starch, microcrystalline cellulose, sodium carboxymethyl starch, cross-linked polyvinylpyrrolidone, low-substituted hydroxypropyl cellulose, cross-linked sodium carboxymethyl cellulose and the like, the lubricant comprises magnesium stearate, sodium dodecyl sulfate, talcum powder, silicon dioxide and the like, the suspending agent comprises polyvinylpyrrolidone, microcrystalline cellulose, sucrose, agar, hydroxypropyl methyl cellulose and the like, and the adhesive comprises starch slurry, polyvinylpyrrolidone, hydroxypropyl methyl cellulose and the like.

The traditional Chinese medicine composition can be added in the form of raw materials, and can also be added in the form of an extract or prepared into particles. Therefore, as another aspect of the application, the application further provides a traditional Chinese medicine composition for treating gouty arthritis, which is prepared from 6-18 parts by weight of chicory extract, 5-15 parts by weight of poria cocos extract, 10-60 parts by weight of smilax glabra extract, 5-10 parts by weight of pagodatree flower bud extract and 3-10 parts by weight of gentiana macrophylla extract, wherein the extracts are water extracts of the raw materials or granules prepared from the water extracts according to a conventional process.

As another aspect of the invention, the invention provides application of the traditional Chinese medicine composition in preparing a medicine for removing joint uric acid crystals and resisting gouty arthritis.

Pharmacological researches show that the traditional Chinese medicine composition with different concentrations can obviously reduce the crystallization of joint uric acid, reduce the uric acid level of joint fluid, obviously improve the swelling degree of quail joints in gouty arthritis, reduce the interleukin-1 beta level of inflammatory factors in serum, has no influence on liver and kidney functions, and has the pharmacological effect of resisting gouty arthritis.

Compared with the prior art, the invention provides a new medicine choice for preventing and treating gouty arthritis, the traditional Chinese medicine composition can obviously reduce the crystallization of joint uric acid, experimental researches show that each prescription does not have the effect after the prescription is disassembled, and the specific compatibility of the raw material medicines of each traditional Chinese medicine composition shows that the traditional Chinese medicine composition has a synergistic effect and has a specific technical effect in the aspect of removing the crystallization of joint uric acid and resisting gouty arthritis. The traditional Chinese medicine composition has the advantages of simple medicine taste, medicine and food containing two medicines, rich medicine sources and high safety, and has better clinical application prospect and economic utilization value.

Drawings

Fig. 1 is a flowchart of the method for extracting the Chinese medicinal composition.

Fig. 2 is a diagram showing uric acid crystallization of quail joints in example 2, wherein 2A is a normal joint rinse, 2B is a positive control suspension, 2C is a model joint rinse, 2D is a high-dose joint rinse, 2E is a medium-dose joint rinse, and 2F is a low-dose joint rinse.

FIG. 3 is a graph showing uric acid levels of joint fluid of quails in each group in example 2.

Fig. 4 is a graph showing the swelling degree of the left wrist joint at each time point of each group of quails in example 2.

Fig. 5 is a graph showing the swelling degree of the right wrist joint at each time point of each group of quails in example 2.

Fig. 6 is a graph showing the swelling degree of the left paw joint at each time point of each group of quails in example 2.

Fig. 7 is a graph showing the swelling degree of the joints of the right paw of each group of quails at each time point in example 2.

Fig. 8 is a graph showing the inhibition rate of left wrist joint swelling at each time point of each group of quails in example 2.

Fig. 9 is a graph showing the inhibition rate of swelling of the right wrist joint at each time point of each group of quails in example 2.

Fig. 10 is a graph showing the inhibition rate of swelling of the left paw joint at each time point of each group of quails in example 2.

Fig. 11 is a graph showing the inhibition rate of swelling of the joints of the right paw of the quails at each time point in example 2.

FIG. 12 is a model quail joint appearance diagram of example 2, wherein 12A is a side view and 12B and 12C are front views.

Fig. 13 is a morphology of quail paw appearance of each group in example 2, wherein 13A is a model group (left) versus a normal group (right), 13B is a high-dose group (left) versus a model group (right), 13C is a medium-dose group (left) versus a model group (right), and 13D is a low-dose group (left) versus a model group (right).

FIG. 14 is a graph showing IL-1β levels of quail serum for each group in example 2.

FIG. 15 is a chart showing the pathological changes of hematoxylin-eosin staining of joint tissues of quails in each group in example 2.

FIG. 16 is a graph of quail serum AST, ALT activity for each group of example 2.

FIG. 17 is a graph showing the serum CRE and BUN content of quail in example 2.

Detailed Description

The present invention will be further described with reference to the following specific embodiments, which are, however, not limited to the following embodiments.

Example 1 formulation for anti gouty arthritis preferred study

1. The experiment aims at exploring the pharmacological actions of different traditional Chinese medicine compositions for reducing uric acid and resisting gouty arthritis and optimizing the effective prescription.

2. Experimental materials

2.1 Experiment System

2.1.1 Laboratory animals, kunming mice, males, body weight 28+ -2 g, total 80.

2.1.2 Animal sources are supplied by si Bei Fu (Beijing) biotechnology limited company, [ laboratory Unit use license number: SYXK (Beijing) 2020-0033, license number: SCXK (Beijing) 2019-0010, laboratory animal ethical number: BUCM-4-2023030901-1033 ].

2.1.3 Raising conditions, namely, the experimental animal barrier system of Beijing Chinese medical university, the room temperature is 20-25 ℃, the relative humidity is 35-75% and the standard illumination is 12 hours.

2.1.4 Feed, supplied by si Bei Fu biotechnology Co.

2.2 Experimental drugs and reagents

Molding materials MSU, sigma Co., USA, lot number BCBQ2650V;

experimental decoction pieces:

the positive control drug is colchicine, kunka, inc., 210603-01.

Solvent deionized water

Reagent:

0.9% sodium chloride injection, shijia four-medicine Co., ltd., lot number 2208102011;

General tissue fixative, wohan Seville Biotechnology Co., ltd., lot number G1101-500ML, chloral hydrate, shanghai Michlin Biochemical Co., ltd., lot number C12204202

2.3 Detection kit

Mouse interleukin-1 beta (IL-I beta) ELISA kit, jiangsu Kort Biotech Co., ltd., lot number 20230240;

creatinine (CRE) determination kit, nanjing's institute of biological engineering, lot number 20230129;

urea Nitrogen (BUN) determination kit, set up bioengineering institute, south kyo, lot number 20230129;

glutamic Pyruvic Transaminase (GPT) kit, nanjing built institute of biological engineering, lot number 20230111;

glutamic-oxaloacetic transaminase (GOT) kit, nanjing institute of biological engineering, lot number 20230111.

2.4 Laboratory apparatus and devices

HH-2 digital display constant temperature water bath, nangsu Jiujinta Ind. Nanghua instruments manufacturing Co., ltd;

DH5-3 low-speed desk-top centrifuge, beijing era North centrifuge Co., ltd;

MTW120 Lemei electronic balance, shenzhen Mobil electronic Co., ltd;

TLE303E/02 electronic balance, metrele-Torisuo instruments (Shanghai) Co., ltd;

KQ5200E type ultrasonic cleaner, kunshan ultrasonic instruments Co., ltd;

SPECROstar Nano microplate reader, BMG, germany;

Reichert Histo STAT paraffin tissue microtomes, AO corporation, usa;

Olmpus BX53 microscope, DP72CCD camera, olympin bas, japan;

RE-501 rotary evaporator, beijing shentai Wei industry instruments and equipment Co., ltd;

D2F-6050ABF type electric heating vacuum drying oven, shanghai Kun Tian laboratory instruments Co., ltd

3. Experimental method

3.1 Experimental design basis

The experimental scheme and the administration dosage are determined by referring to the pharmaceutical research technical guidelines (trial) of each stage of the research of the new traditional Chinese medicine and the earlier-stage research results of the laboratory issued by the national drug administration.

3.2 Preparation of test drugs

3.2.1 Extract of Juling prescription

The chrysanthemum and poria cocos formula comprises 9g of chicory, 10g of poria cocos, 30g of smilax glabra, 6g of pagodatree flower bud and 9g of gentiana macrophylla;

The preparation method of the chrysanthemum-poria prescription extract comprises weighing a proper amount of medicinal materials according to the prescription proportion of the chrysanthemum-poria prescription, adding 12 times of deionized water for soaking for 30min, extracting for 1.5h by adopting a decoction method, collecting the extract, repeating for 3 times, and merging the extracts. Evaporating and concentrating in a rotary evaporator to obtain compound water decoction concentrate containing crude drug content of 1mg/ml, steaming in water bath at 80deg.C until no fluidity, drying in vacuum drying oven at 80deg.C for 7 days, taking out, standing at room temperature, and pulverizing to obtain compound extract powder.

3.2.2 Preparation of extract from formula (de-gentiana macrophylla), extract from formula (de-pagodatree flower bud), extract from formula (de-tuckahoe), extract from formula (de-smilax glabra)

The preparation method comprises the steps of removing gentiana macrophylla in a chrysanthemum and poria cocos formula based on the chrysanthemum and poria cocos formula, extracting other medicinal materials according to the chrysanthemum and poria cocos formula, removing sophora flower bud in the chrysanthemum and poria cocos formula, extracting other medicinal materials according to the chrysanthemum and poria cocos formula, removing poria cocos in the chrysanthemum and poria cocos formula, extracting other medicinal materials according to the chrysanthemum and poria cocos formula, removing rhizoma smilacis glabrae in the chrysanthemum and poria cocos formula, and extracting other medicinal materials according to the chrysanthemum and poria cocos formula.

3.2.3 Colchicine solution

Colchicine is selected as a positive control drug in the experiment. When in use, a proper amount of colchicine is taken and dissolved in deionized water to be prepared into corresponding dosage.

3.3 Grouping and dosing amount:

the experiment was performed in 8 groups. The mice are respectively a normal group, a model group, a colchicine group, a chrysanthemum and poria cocos formula group, a gentiana macrophylla group, a pagodatree flower bud group, a poria cocos group and a glabrous greenbrier rhizome group, and 10 Kunming mice are arranged in each group. Each of the administration groups was administered prophylactically and therapeutically by gastric lavage, and the amounts of the crude drugs in each group were converted in equal proportions to give the doses shown in table 1.

Table 1 design of dosing for mice of each group during the experiment

Group of	Medicament	Dosage for administration	Stomach filling volume (ml/100 g)
				Normal group	——	——	1
Model group	——	——	1
				Colchicine group	Colchicine	1mg/kg	1
Chrysanthemum and poria cocos square group	Chrysanthemum and poria cocos prescription	12.86g/kg	1
				Gentiana macrophylla removing group	Gentiana macrophylla removing prescription	11.05g/kg	1
Sophora flower bud removing group	Sophora flower bud removing prescription	11.65g/kg	1
				Poria removing group	Poria removing prescription	10.85g/kg	1
Rhizoma smilacis glabrae removing group	Rhizoma smilacis glabrae removing prescription	6.83g/kg	1

3.4 Method of administration:

Administration route is gastric lavage administration (consistent with the clinical planned administration route of human).

The administration volume is 1ml/100g.

The administration time is 1 time per day in the morning according to the weight of the stomach.

The administration period is 7 days after continuous administration.

3.5 Preparing the tested medicine, namely weighing a certain amount of the tested medicine, adding the weighed tested medicine into quantitative aqueous solution, and uniformly mixing by ultrasonic for later use.

3.6 Preparation and administration of the Molding agent A quantity of MSU crystals and proper physiological saline are taken for irradiation under ultraviolet light for 6 hours, and then MSU is dissolved in sterile physiological saline to prepare MSU suspension of 50 mg/ml.

3.7 Method of modeling 1h after administration of experiment 5d, 25. Mu.l of MSU suspension was injected into ankle cavity of mice in model group and each administration group, and 25. Mu.l of 0.9% sterile physiological saline was injected into ankle cavity of mice in normal group using 1ml of sterilized insulin syringe. Ankle injections were normalized to the success of the injection with the bulge on the opposite side of the articular cavity.

3.8 Sample collection and treatment the foot thickness, ankle circumference of mice were measured before (0 h) and 4h, 8h, 12h, 24h, 48h after injection of MSU and scored for inflammation. The material was taken at experiment 7 d. Collecting eyeball blood, separating serum for detection of biochemical indexes, preserving at-20deg.C, collecting joint tissue, fixing with 4% paraformaldehyde for 48 hr, decalcification with 10% formic acid solution, embedding paraffin, slicing (thickness of 4 μm), spreading, and taking out slices at 37deg.C overnight.

3.9 Detection index

3.9.1 General state indexes, namely ankle joint swelling degree, foot thickness swelling degree and inflammation score.

1) Ankle joint/foot thickness swelling was measured with vernier calipers before (0 h) and after (4 h, 8h, 12h, 24h, 48h mice were tested for ankle joint circumference and foot thickness and repeated three times. The swelling degree at each time point was calculated according to formula (1).

2) Inflammation scoring

The grading standard of the inflammation index is as follows, 0 score, normal, 2 score, skin erythema of the joint under test, slight swelling, visible osseous signs, 4 score, obvious redness and swelling of the joint under test, disappearance of osseous signs, but limitation of swelling to joint parts, and 6 score, limb swelling beyond the joint under test.

3.9.2 Inflammation related index, interleukin-1 beta (IL-1 beta) level, and the kit detects serum IL-1 beta level.

3.9.3 Renal function related indexes, namely serum creatinine (SCre) and urea nitrogen (BUN) levels, and detecting serum Cre and BUN levels by using the kit.

3.9.4 Liver function related indexes, namely serum glutamic pyruvic transaminase (ALT) and glutamic oxaloacetic transaminase (AST), and detecting serum ALT and AST levels by using the kit.

4 Statistics and analysis of results

Statistical analysis using SPSS27.0 software, data were taken as mean.+ -. Standard deviationAs shown, the data comparison between the groups is based on whether each group is normal or not, a one-way anova or a Kruskal-Wallis H nonparametric test is selected. The comparison between the normal distribution data sets is based on whether the variance is uniform or not, LSD test or Dunnett's T test is selected, the non-normal distribution data is selected from the Kruskal-Wallis H test, the average rank is multiple, and the difference is P < 0.05.

5. Results

5.1 General state index

5.1.1 Ankle swelling degree

The ankle swelling degree was calculated in this experiment by the formula (1).

Model mice had significantly increased ankle swelling levels (P < 0.01) at each observation time point compared to the normal group.

Compared with a model group, the colchicine group mice ankle joint swelling degree is obviously reduced (P < 0.05) after molding, no obvious difference exists at other time points, the chrysanthemums side group mice ankle joint swelling degree is obviously reduced (P < 0.05) after molding, no obvious difference exists at other time points, the gentiana macrophylla group mice ankle joint swelling degree is obviously reduced (P < 0.05) after molding, no obvious difference exists at other time points, the pagodatree flower bud group mice ankle joint swelling degree is obviously reduced (P < 0.05) after molding, no obvious difference exists at other time points, and the poria cocos group and the smilax glabra group mice ankle joint swelling degree are respectively different at all observation points.

Compared with the Juling prescription group, the ankle joint swelling degree of the mice in the Qinjiao group has no significant difference at all observation points, the ankle joint swelling degree of the mice in the Qinjiao group has no significant difference at all time points after 4 hours and 48 hours after molding (P < 0.05), the ankle joint swelling degree of the mice in the Qinjiao group has no significant difference at all time points after 48 hours after molding (P < 0.05), the ankle joint swelling degree of the mice in the Qinjiao group has no significant difference at all time points after molding (P < 0.05). See table 2 below.

Table 2 mice of each group were given time ankle joint swelling degree (%,n=10)

Note that P <0.05, P <0.01 compared to the normal group, #p <0.05 compared to the model group, ^& P <0.05 compared to the chrysanthemums square group.

5.1.2 Foot thickness swelling degree

The experiment calculates the foot thickness swelling degree with formula (2).

The model group mice had significantly elevated foot thickness swelling levels (P <0.05 or P < 0.01) at each observation time point compared to the normal group.

Compared with a model group, the colchicine group mice foot thickness swelling degree has no significant difference at all observation time points, the chrysanthemum and poria cocos formula group mice foot thickness swelling degree is significantly reduced (P < 0.05) after molding for 4 hours, the rest time points have no significant difference, the pagodatree flower bud group mice foot thickness swelling degree is significantly increased (P < 0.01) after molding for 12 hours, the rest time points have no significant difference, and the gentiana macrophylla group, the poria cocos group and the smilax glabra group mice foot thickness swelling degree have no significant difference at all observation time points.

Compared with the chrysanthemum and poria cocos formula group, the radix gentianae macrophyllae removing group has no significant difference at all observation time points, the flos sophorae removing group has no significant difference at all other time points after molding for 48 hours (P < 0.05), the poria cocos removing group has significant difference at all other time points after molding for 8 hours and 48 hours (P <0.05 or P < 0.01), and the rhizoma smilacis glabrae removing group has no significant difference at all observation time points. See table 3.

Table 3 mice of each group had a thick enough swelling (%,n=10)

Note that P <0.05, # P <0.01 compared to the normal group, #p <0.05, # P <0.01 compared to the model group, ^&P＜0.05,^&& P <0.01 compared to the chrysanthemic group.

5.1.3 Mice inflammation score

The model group mice had significantly elevated inflammation scores (P < 0.01) at each observation time point compared to the normal group.

Compared with the model group, the colchicine group mouse inflammation scores are obviously reduced after 12 hours of molding (P < 0.05), all other time points have no obvious difference, the chrysanthemum and poria cocos group mouse inflammation scores are obviously reduced after 4 hours and 12 hours of molding (P <0.05 or P < 0.01), all other time points have no obvious difference, and the gentiana macrophylla group, the pagodatree flower bud group, the poria cocos group and the smilax glabra group mouse inflammation scores have no obvious difference at all observation time points.

Compared with the Juling prescription group, the inflammation scores of the mice in the gentiana macrophylla group have no significant difference at all observation time points, the inflammation scores of the mice in the sophora flower bud group have no significant difference at all other time points after 12 hours of molding (P < 0.05), the inflammation scores of the mice in the poria cocos group have significant difference at all other time points after 12 hours of molding (P < 0.05), the inflammation scores of the mice in the smilax glabra group have significant difference at all other time points after 12 hours of molding (P < 0.05), and the inflammation scores of the mice in the sophora flower bud group have no significant difference at all other time points. See table 4.

TABLE 4 inflammation score for each time point for each group of micen=10)

5.2 Mouse serum interleukin-1 beta (IL-1 beta) levels

The serum IL-1β levels were significantly elevated in the mice of the model group compared to the normal group (P < 0.01).

The serum IL-1 beta levels were significantly reduced in mice from each of the dosing groups compared to the model group (P <0.01 or P < 0.05).

Compared with the chrysanthemum and poria prescription group, the serum IL-1 beta level of the mice with gentiana macrophylla, poria cocos and smilax glabra is obviously increased (P <0.01 or P < 0.05), and the serum IL-1 beta level of the mice with sophora flower bud has a reducing trend but no obvious difference. See table 5.

Table 5 serum IL-1 beta content (ng/L,)

Note that P <0.05, # P <0.01 compared to the normal group, #p <0.05, # P <0.01 compared to the model group, ^& P <0.05 compared to the chrysanthemums square group.

5.3 Pathological changes in ankle joints in mice

The pathological section results show that the ankle joint structure of the normal group mice is clear, the joint surface is smooth, the synovial tissue does not have obvious pathological changes, the ankle joint synovial tissue of the model group mice has obvious hyperplasia with a large amount of inflammatory cell infiltration, the ankle joint structure of the colchicine group mice is clear, the synovial tissue does not have obvious pathological changes, the ankle joint structure of the chrysanthemum and poria cocos group mice is clear, the joint surface is smooth, the synovial tissue does not have obvious pathological changes, the ankle joint structure of the gentiana macrophylla group mice is clear, the joint surface is smooth, the synovial tissue has small hyperplasia, the ankle joint structure of the pagodatree flower bud group mice is clear, the synovial tissue has small hyperplasia, the ankle joint synovial tissue of the poria cocos group mice has small hyperplasia with small inflammatory cell infiltration, and the ankle joint synovial tissue of the smilax glabra group mice has small hyperplasia with small inflammatory cell infiltration.

5.4 Serum creatinine (Cre), urea Nitrogen (BUN) levels

Compared with the normal group, the serum Cre and BUN levels of the model group have no significant difference, and the serum Cre and BUN levels of each administration group have no significant difference.

5.5 Glutamic-pyruvic transaminase (ALT) and glutamic-oxaloacetic transaminase (AST) expression levels

Compared with the normal group, the model group has no significant difference in serum ALT and AST activities of mice after 48 hours of molding, and the colchicine group, the chrysanthemum and poria formula group, the gentiana macrophylla group, the pagodatree flower bud group, the poria cocos group and the smilax glabra group have no significant difference in serum ALT and AST activities of mice.

Discussion 6

The chrysanthemum and poria cocos compound prescription is a crystal of a drug experience prescription for treating arthralgia syndrome, acute and chronic gouty arthritis by Zhang Bing professors, and is combined with the study of the subject group for more than 20 years, and the whole prescription has the effects of dredging collaterals and relieving pain, clearing heat and resolving phlegm, strengthening spleen and eliminating dampness. The aim of this study was to observe and compare the effect of the Juling prescription and its prescription on the mouse model of acute gouty arthritis.

In the overall animal characterization, the experiment uses the ankle joint swelling degree and the foot thickness swelling degree of the mice as main evaluation indexes and uses the inflammation score as an auxiliary evaluation index.

At each observation time point of the experiment, ankle swelling, foot thickness swelling, inflammation score, serum IL-1β levels were all significantly elevated (P <0.05 or P < 0.01) in the model group mice compared to the normal group.

Compared with a model group, serum IL-1 beta level of mice in each administration group is obviously reduced (P <0.05 or P < 0.01), ankle joint swelling degree of mice in colchicine group is obviously reduced (P < 0.05) at 24h and 48h after molding, inflammation scores are obviously reduced (P < 0.05) at 12h after molding, indexes at other time points are not obviously different, ankle joint swelling degree of mice in the chrysanthemum tuckahoe group is obviously reduced (P < 0.05) at 24h and 48h after molding, foot thickness swelling degree is obviously reduced (P < 0.05) at 4h after molding, inflammation scores are obviously reduced (P <0.05 or P < 0.01) at other time points, ankle joint swelling of mice in the large leaf-removing group is obviously reduced (P < 0.05) at other time points, indexes at other time points are not obviously different, ankle joint swelling degree of mice in the chrysanthemum tuckahoe group is obviously reduced (P < 0.05) at 24h after molding, and the rest time points are not significantly different at other time points are observed, and the rest indexes are not significantly different after the four-point of the lung-removing mice are subjected to experiment.

Compared with the chrysanthemum and poria prescription group, the regulation and control effect of the gentiana macrophylla removing group, the poria cocos removing group and the smilax glabra removing group on the serum IL-1 beta level of the mice is reduced (P <0.05 or P < 0.01), the pagodatree flower bud removing group has no obvious difference, and the other indexes of the gentiana macrophylla removing group mice have no obvious difference at each observation time point of the experiment. The ankle joint swelling of the mice with the pagodatree flower bud group is obviously higher than Yu Ju poria cocos square group (P < 0.05) after 4 hours and 48 hours after molding, the foot thickness swelling degree is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding, the inflammation score is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding, the ankle joint swelling of the mice with the pagodatree flower bud group is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding, the foot thickness swelling degree is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding and 48 hours, the inflammation score is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding, the ankle joint swelling of the mice with the smile group is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding, the inflammation score is obviously higher than Yu Ju poria cocos square group (P < 0.05) after molding, and the rest time points have no obvious difference;

compared with the normal group, the model group mice show remarkable hyperplasia of ankle synovial tissue accompanied by massive inflammatory cell infiltration. Compared with the model group, the colchicine group and the chrysanthemum and poria cocos formula group have clear ankle joint structure, the synovium tissue has no obvious pathological change, and the mice with the gentiana macrophylla group, the pagodatree flower bud group, the poria cocos group and the smilax glabra group have clear ankle joint structure and the synovium tissue has little hyperplasia.

During the experiment period, the serum CRE, BUN, ALT, AST levels of all groups of experimental animals have no significant difference, which indicates that the traditional Chinese medicine test object has no obvious influence on the liver and kidney functions of mice.

The results show that colchicine is used as a clinically common anti-gout drug, and the colchicine is used as a positive drug to exert an anti-inflammatory effect in the experiment, thereby proving that the establishment of the acute gouty arthritis mouse model is successful. The chrysanthemum and poria cocos formula plays a good role in inhibiting the ankle joint swelling degree and the foot thickness swelling degree of the mice, and each formula can also inhibit the ankle joint swelling degree and the foot thickness swelling degree of the mice. The IL-1 beta results in serum prove that the chrysanthemum indicum prescription and all the prescription have better effect of resisting acute gouty arthritis, the anti-gout effect of the chrysanthemum and poria cocos formula is obviously better than that of each formula.

In conclusion, the chrysanthemum and poria cocos formula and all the disassembly formulas thereof can improve the acute gouty arthritis induced by MSU of mice, and the chrysanthemum and poria cocos formula has the best curative effect, so that the result that the traditional Chinese medicine compound is the synergistic effect of all the components is reflected.

EXAMPLE 2 study of the efficacy of Chinese medicinal composition against gouty arthritis

1. The experimental aim is to explore the pharmacological action of the traditional Chinese medicine composition on resisting gouty arthritis on the basis of the embodiment 1, and provide a new medicine choice for preventing and treating gouty arthritis.

2. Experimental method

2.1 Grouping of animals

72 Quails of 40-day-old Di-Fa-ke quails are selected and fed adaptively for 3d, and are randomly divided into 6 groups according to the mass, namely a normal group, a model group, a benzbromarone group (20.00 mg/kg), a high-dose group of the extract of the inula japonica (corresponding to 10g/kg of crude drug), a medium-dose group of the extract of the inula japonica (corresponding to 7.5g/kg of crude drug) and a low-dose group of the extract of the inula japonica (corresponding to 3.75g/kg of crude drug), wherein each group is 12.

2.2 Modeling

Experiments 1-50 d, model groups and each administration group are given high-purine feed (basal feed: yeast extract powder is 8:2), experiments 51-83 d, model groups and each administration group are given high-protein high-calcium feed (basal feed: yeast extract powder: bone powder is 5:2:3) and 10% fruit sugar water (limit of 15 mL/d). During the experiment, the normal group was given sufficient basal feed, clear water. Controlling the room temperature of the animal house to be 20-25 ℃, humidity is 40-60%.

2.3 Preparation method of composition (extract of Chrysanthemum Indicum)

The formula comprises 9g of chicory, 10g of tuckahoe, 30g of glabrous greenbrier rhizome, 6g of pagodatree flower bud and 9g of gentiana macrophylla;

The preparation method comprises soaking in deionized water with a material-to-liquid ratio of 12 times for 30min, extracting for 1.5 hr by decocting method, collecting extractive solution, repeating for 3 times, and mixing extractive solutions. Concentrated to a concentration of 10g/kg, 7.5g/kg, 3.75g/kg by evaporation in a rotary evaporator and kept at 4℃for further use. The specific operation flow is shown in figure 1.

2.4 Experimental dosing

The high, medium and low dosage groups of the composition have the administration concentrations of 10g/kg, 7.5g/kg and 3.75g/kg respectively. The experiment selects benzbromarone as a uric acid-reducing positive control drug. When in use, a proper amount of benzbromarone is dissolved in deionized water to be prepared into corresponding dosage. The administration concentration of the benzbromarone is 20mg/kg. The administration mode is intragastric administration, and the normal group and the model group are filled with the ultrapure water with the same volume of the intragastric administration every day.

2.5 Materials selection

Taking 1.0-1.5 mL of blood from the jugular vein of each group of quails, and taking the blood without taking water for 12 hours after fasting. The serum sample is centrifugated for 10min at normal temperature of 3500r/min, and the supernatant is used for detecting serum biochemical indexes. And respectively flushing the left wrist joint cavities of the quails by adopting 0.2mL of physiological saline, coating the obtained wrist joint flushing liquid on a glass slide, and observing uric acid crystals in the wrist joint flushing liquid under a polarized light microscope. And collecting joint tissues of quails of each group, performing HE staining, and observing inflammatory infiltration.

2.6 Joint swelling index Collection

Experiments 50d, 60d, 70d and 80d, the circumferences of the left and right wrist joints and the left and right claw joints of each group of quails were measured by using a flexible ruler. Taking the measurement result of the experiment 50d as the circumference of the wrist and jaw joint at the beginning of the experiment, calculating the swelling degree of the wrist and jaw joint of each group of quail of the experiment 60d, 70d and 80d according to the formula (1), and calculating the swelling inhibition rate of the wrist and jaw joint of each group of quail of the experiment 60d, 70d and 80d according to the formula (2).

2.7 Joint flushing fluid Collection and treatment

In experiment 83d, the left wrist joint cavities of all groups of quails are respectively washed by adopting 0.2mL of physiological saline, the obtained wrist joint washing liquid is coated on a glass slide, and uric acid crystals in the wrist joint washing liquid are observed under a polarized light microscope.

2.8 Joint histopathological observations

Paraffin sections were prepared from 10% formalin-fixed joint tissue and stained with hematoxylin eosin. An inverted microscope was used to collect images for histopathological observation.

3. Data analysis

Statistical analysis was performed using GraphPad prism7.0 software, and data were taken as mean ± standard deviation The data comparison between the groups shows that whether the normal and variance of each group are uniform or not is selected for single-factor analysis of variance or Kruskal-Wallis anecdotal test, the comparison between the groups is selected for the variance of each group according to the variance of each group, dunnett-t test or Dunnett's T test is adopted, and P <0.05 is taken as a difference to have statistical significance.

4. Main instruments and equipment (see Table 6 below)

TABLE 6 Main instruments and apparatus

Name of the name	Model number	Manufacturer' s
			Electromagnetic oven	HY-221	Guangdong hemisphere Utility company
Rotary evaporator	RE-501	Beijing shentai wei industry instrument Equipment Co Ltd
			Circulating water type multipurpose vacuum pump	SHB-III	Beijing shentai wei industry instrument Equipment Co Ltd
Electronic balance	TLE303E	Metrele-Tolyduo instruments (Shanghai) Inc
			Desk type centrifugal machine	DT5-3 type	Beijing era North centrifuge Co., ltd
Freezing embedding machine	KH-BL	Hubei Xiaozhi broad sea medical science and technology Co., ltd
			Paraffin tissue slicer	Reichert Histo STAT	AO Co Ltd
Microscope	Olmpus BX53	Orinbas Corp Japan
			Camera with camera body	DP72CCD	Orinbas of Japan
Electrothermal blowing drying box	101-1AB type	Test instruments Inc. of Tianjin City
			Enzyme label instrument	sunrise	TeCAN company Switzerland
Water bath kettle	HH-1 high-end type	Jintan city and west sense, laboratory instrumentation factory

5. Main reagents and drugs (see Table 7 below)

TABLE 7 Main reagents and drugs

6. Results

6.1 Effect of Chinese medicinal composition on Joint uric acid crystallization

A 0.375% sodium urate suspension was prepared and applied to a glass slide as a positive control, and the positive control and the quail wrist joint washes of each group were observed under a 200-fold polarized light microscope. The results are shown in FIG. 2.

Under the black visual field, needle-shaped urate crystals are not seen in the normal group. In contrast to the normal group, the positive control suspensions were seen as bright, blue or yellow urate crystals, mostly clustered into "spherical" clusters, with large needles scattered around.

Compared with the normal group, the model group is mostly scattered, bright, blue or yellow urate crystals, the shape of the crystal is small in head and tail or small in sharp needle shape at two ends, the crystal clusters are occasionally aggregated into feather-shaped crystal clusters, and the length of each single needle crystal is about 1-20 mu m.

In contrast to the model group, the high (10 g/kg), medium (7.5 g/kg), low (3.75 g/kg) dose groups of the chrysanthemum indicum showed little scattering distribution of needle-like urate crystals, and little bundles of needle-like crystals arranged approximately in parallel, or bundles of needle-like crystals crossed and overlapped to form an X-shaped crystal bundle, and no aggregation of the crystal clusters was found.

6.2 Uric acid levels in joint fluid

The joint cavities of quails were rinsed with 0.2ml of physiological saline, ankle fluids of each group were collected, and uric acid levels of joint fluids of each group were measured. The results are shown in FIG. 3. The joint fluid uric acid levels of model group quails were significantly elevated compared to normal group (P < 0.01).

Compared with the model group, the high (10 g/kg) and medium (7.5 g/kg) dosage group of the poria extract has significantly reduced uric acid level (P < 0.05) of quail joint fluid.

6.3 Joint swelling degree

6.3.1 Left wrist joint swelling degree

The left wrist joint swelling degree was calculated in this experiment by formula (1). The results are shown in Table 8 and FIG. 4. Experiments 60d, 70d, 80d showed a significant increase in swelling of the left wrist joint of the model group quail (P < 0.01) compared to the normal group.

Compared with the model group, the swelling degree of the left wrist joint of the quail is obviously reduced (P <0.05 or P < 0.01) in the group with high (10 g/kg), medium (7.5 g/kg) and low (3.75 g/kg) dosage of the poria extract in the experiment 70 d.

Table 8 the left wrist joint swelling degree (%, n=12,)

Note that P <0.01 compared to the normal group, # P <0.05, # P <0.01 compared to the model group.

6.3.2 Degree of swelling of the right wrist

The present experiment calculates the degree of swelling of the right wrist joint with equation (1). The results are shown in Table 9 and FIG. 5. Experiments 60d, 70d, 80d, model group quail right wrist joint swelling increased significantly compared to normal group (P < 0.01).

Compared with the model group, the swelling degree of the right wrist joint of the quail in the group with the dosage of 7.5g/kg in the poria cocos extract is obviously reduced (P < 0.05), the swelling degree of the right wrist joint of the quail in the group with the dosage of high (10 g/kg) and low (3.75 g/kg) in the poria cocos extract is reduced (P=0.059 and P=0.052), and the swelling degree of the right wrist joint of the quail in the group with the dosage of 10g/kg in the poria cocos extract is reduced (P=0.092) in the experiment of 80 d.

Table 9 the swelling degree (%, n=12,)

Note that P <0.01 compared to the normal group and #p <0.05 compared to the model group.

6.3.3 Left paw joint swelling degree

The present experiment calculates the left paw joint swelling degree with formula (1). See table 10, fig. 6. Compared with the normal group, the swelling degree of the left paw joint of the model group quail has an increasing trend (P=0.069), and the swelling degree of the left paw joint of the model group quail has a significant increase (P <0.05 or P < 0.01) in the experiments 70d and 80 d.

Compared with the model group, the swelling degree of the left paw joint of the quail in the tribenuron-methyl group and the poria cocos extract low (3.75 g/kg) dosage group has a tendency of reducing (P=0.076, P=0.064), the swelling degree of the left paw joint of the quail in the tribenuron-methyl group and the poria cocos (7.5 g/kg) dosage group is obviously reduced (P <0.05 or P < 0.01), and the swelling degree of the left paw joint in the tribenuron-methyl group and the poria cocos extract low (7.5 g/kg) dosage group is obviously reduced (P <0.05 or P < 0.01).

Table 10 Each time point for each group of quails left paw joint swelling degree (%, n=12 and,)

Note that P <0.05, # P <0.01 compared to the normal group, and # P <0.05, # P <0.01 compared to the model group.

6.3.4 Degree of swelling of the right paw joint

The present experiment calculates the degree of swelling of the right paw joint with formula (1). The results are shown in Table 11 and FIG. 7. Experiment 80d, model group quail right paw joint swelling was significantly increased (P < 0.01) compared to normal group.

Compared with the model group, the swelling degree of the joints of the right paw of the quail in the experiment 80d, the tribenuron-methyl group, the chrysanthemum indicum high (10 g/kg), the middle (7.5 g/kg) and the low (3.75 g/kg) dose group is obviously reduced (P < 0.01).

Table 11 quail groups time points swelling degree of right paw joint (%, n=12 and,)

Note that P <0.01 compared to the normal group and ^## P <0.01 compared to the model group.

6.3.5 Inhibition rate of left wrist joint swelling

The experiment calculates the left wrist joint swelling inhibition rate by the formula (2). The results are shown in Table 12 and FIG. 8. In experiment 60d, the highest inhibition rate of the swelling degree of the left wrist joint of the quail in the dosage group (7.5 g/kg) in the poria cocos extract is 65.77%, and the dosage groups of low (3.75 g/kg) and high (10 g/kg) are sequentially reduced, but are higher than those of the tribenuron-methyl group.

In experiment 70d, the inhibition rate of the swelling degree of the left wrist joint of the quail in the dosage group of 7.5g/kg in the extract of the chrysanthemum indicum is 77.95 percent at the highest, the dosages of the extract of the chrysanthemum indicum are equivalent to each other in high (10 g/kg) and low (3.75 g/kg) dosages, but are higher than that of the tribenuron-methyl group,

In experiment 80d, the inhibition rate of the swelling degree of the left wrist joint of the quail in the dosage group (7.5 g/kg) of the chrysanthemum indicum extract is up to 24.61%, and the dosage group (7.5 g/kg) and the dosage group (3.75 g/kg) of the tribenuron-methyl are weaker.

Table 12 inhibition of left wrist joint swelling (%, n=12) for each time point of quail administration group

6.3.6 Inhibition rate of swelling of Right wrist joint

The present experiment calculates the right wrist joint swelling inhibition rate by formula (2). The results are shown in Table 8 and FIG. 9. Experiment 60d, the inhibition rate of the swelling degree of the right wrist joint of the quail in the high (10 g/kg), medium (7.5 g/kg) and low (3.75 g/kg) dosage groups is higher than that of the phenylbromarone group, and the highest inhibition rate of the swelling degree of the left wrist joint of the quail in the high (10 g/kg) dosage group is 28.86%.

In experiment 70d, the inhibition rate of the swelling degree of the right wrist joint of the quail in high (10 g/kg), medium (7.5 g/kg) and low (3.75 g/kg) dosage groups is higher than that of the phenylbromarone group, and the inhibition rate of the swelling degree of the left wrist joint of the quail in medium (7.5 g/kg) dosage groups is up to 57.87%.

In experiment 80d, the inhibition rate of the swelling degree of the right wrist joint of the quail in high (10 g/kg), medium (7.5 g/kg) and low (3.75 g/kg) dosage groups is higher than that of the phenylbromarone group, and the inhibition rate of the swelling degree of the left wrist joint of the quail in high (10 g/kg) dosage groups is 28.40% at most.

Table 8 inhibition of swelling of the right wrist at each time point (%, n=12) for each group of quails

6.3.7 Left paw joint swelling inhibition rate

The experiment calculates the inhibition rate of left paw joint swelling by the formula (2). The results are shown in Table 9 and FIG. 10. Experiment 60d, the highest inhibition rate of the swelling degree of the left paw joint of the quail in the low-dose (3.75 g/kg) group of the chrysanthemum indicum extract is 36.73 percent higher than that of the tribenuron-methyl group.

Experiment 70d, the highest inhibition rate of the swelling degree of the left paw joint of the quail in the dosage group (7.5 g/kg) of the chrysanthemum indicum extract is 34.23% higher than that of the benzbromarone group.

Experiment 80d, the inhibition rate of the swelling degree of the left paw joint of the quail in the dosage group (7.5 g/kg) in the poria cocos extract is equal to that of the benzbromarone group, and the inhibition rate is 38.66% and 39.78% respectively.

Table 9 inhibition of left paw joint swelling (%, n=12) for each time point of quail group

6.3.8 Inhibition rate of swelling of the joints of the right foot paw

The experiment calculates the inhibition rate of the swelling of the joints of the right paw by the formula (2). The results are shown in Table 10 and FIG. 11. Experiment 60d, the highest inhibition rate of the swelling degree of the right foot paw joint of quail in a dosage group (7.5 g/kg) in the poria cocos extract is 32.59%, the high (10 g/kg) dosage group of the poria cocos extract is equivalent to the tribenuron-methyl, the swelling degree of the low (3.75 g/kg) dosage group is the lowest, and the dosage groups are 28.81% and 28.97% respectively.

Experiment 70d, the highest inhibition rate of the swelling degree of the right foot paw joint of the quail in the dosage group (7.5 g/kg) of the chrysanthemum indicum extract is 32.82 percent, which is higher than that of the benzbromarone group, and the dosage group with high (10 g/kg) of the chrysanthemum indicum extract is equivalent to the dosage group with low (3.75 g/kg) of the chrysanthemum indicum extract, which are 25.52 percent and 27.63 percent respectively.

Experiment 80d, the low (3.75 g/kg) dosage group of the poria extract is equivalent to that of the benzbromarone group, which are 50.19% and 52.03% respectively, and the medium and low dosage groups are lower.

Table 10 inhibition of right paw joint swelling (%, n=12) for each time point of quail group

6.4 Appearance and shape changes of wrist joints and feet paws

During the experiment, the wrist joint and the paw of the quail are normal, and the movement is flexible. Compared with the normal group, the model group quails have different degrees of swelling and local erythema at the wrist joint, obvious swelling at the paw and toe joint, or a plurality of swelling nodules (tophus) with different sizes, and the swelling of the individual quails has the phenomena of purple stasis, suppuration and crumbling, as shown in figure 12.

Compared with the model group, the high, medium and low dosage group quail of the chrysanthemum indicum extract has slight swelling of the wrist joint and the paw joint, as shown in figure 13.

6.5 Inflammatory factor expression levels

The results are shown in FIG. 14. Compared with the normal group, the IL-1 beta level of the quail serum of the model group is obviously increased, and compared with the model group, the IL-1 beta level of the quail serum of the model group with high, medium and low doses of the chrysanthemum indicum of the experiment 83d is obviously reduced (P < 0.01).

6.6 Pathological observation of joint tissue

The results are shown in FIG. 15. The HE staining of the quail wrist joint tissue can be seen, the normal quail wrist joint tissue structure is clear, the joint surface is smooth and flat, the synovial cells are loose in arrangement and normal in morphology, and the model group wrist joint tissue can be seen to be fibroplasia of the synovial membrane, pannus formation and massive inflammatory cell infiltration. Compared with the model group, the high-dose group of the chrysanthemum and the middle-dose group of the chrysanthemum and the poria have obviously reduced inflammatory cell infiltration and reduced synovial fibrous tissue proliferation.

6.7 Liver and kidney function index

6.7.1 Serum AST, ALT activity

The results are shown in Table 11 and FIG. 16. Compared with the normal group, the experimental group 83d has no obvious difference in quail serum AST and ALT activities.

Compared with the model group, in the experiment 83d, the quail serum AST activity of the high-dose (10 g/kg) of the poria extract has a reduced trend but no significant difference (P=0.052), and the quail serum AST and ALT activity of the other administration groups have no significant difference.

Table 11 quail serum AST, ALT viability (U/L, n=12,)

6.7.2 Serum CRE, BUN content

The results are shown in Table 12 and FIG. 17. Compared with the normal group, the experiment 83d shows that the serum CRE content of the model group quail has no significant difference, the serum BUN content is significantly increased (P < 0.05), and the serum CRE content and the BUN content of the administration group quail have no significant difference.

Compared with a model group, in the experiment 83d, the CRE content of the quail serum of a high (10 g/kg) dosage group of the chrysanthemum is obviously reduced (P < 0.05), the CRE content of the quail serum of the rest administration group is not obviously different, the BUN content of the quail serum of a high (10 g/kg), middle (7.5 g/kg) and low (3.75 g/kg) dosage group of the chrysanthemum is obviously reduced (P < 0.01), and the dosage dependency is presented, and the BUN content of the quail serum of the phenylbromarone group is not obviously different.

Table 12 quail serum CRE and BUN content for each group (n=12,)

Note that P <0.05 compared to the normal group and ^#P<0.05,^## P <0.01 compared to the model group.

7. Analysis of results

The study was conducted on the use of the pharmaceutical composition to remove the uric acid crystals of joints against gouty arthritis. The results show that the high, medium and low dosage groups of the traditional Chinese medicine composition can obviously reduce the uric acid crystallization of quail joints of a model group, reduce the uric acid level of joint fluid, obviously improve the swelling degree of the quail joints of gouty arthritis, reduce the interleukin-1 beta level of inflammatory factors in serum, and define the efficacy of the traditional Chinese medicine composition in removing the uric acid crystallization of the joints and resisting gouty arthritis.

In the case of joint uric acid crystals, the experiment shows that needle-shaped, bright and blue-yellow joint uric acid crystals are visible in the wrist joint flushing liquid of the model group quails through microscopic observation of the wrist joint flushing liquid of each group of quails by polarized light. The uric acid crystal in the joint synovial fluid is used as a gold index for clinically diagnosing gouty arthritis, and can effectively prove the modeling success of a gouty arthritis model. The experimental result shows that uric acid crystals deposited in the model group quail wrist joint flushing liquid are in the shape of scattered needles or gathered into feathers, and have the unique needle-shaped, blue/yellow and strong negative birefringence morphology and optical characteristics of uric acid crystals under polarized light.

In the condition of inflammatory expression, the experiment uses joint swelling degree as a main evaluation index and joint appearance form as an auxiliary evaluation index, and discovers the typical inflammatory manifestation of swelling and redness when the local wrist and paw joints of the model group quail have acute attacks of gouty arthritis. The joint swelling results show that the swelling of the left and right wrist joints of the quails in the model group is significantly increased at different time points compared with the model group. The joint appearance morphology observation shows that compared with the joints with normal morphology in a normal group, the toe joints of the model group quail can see a plurality of swelling nodules with different sizes, the swelling places of the model group quail can see the swelling places to be broken and run out of pus, the wrist joints can also see obvious swelling, and the skin at the swelling places is red purple.

The experiment evaluates the anti-gout efficacy of the poria cocos extract in terms of the swelling degree of the wrist joint, the swelling degree of the paw joint and the crystal shape of uric acid deposited in joint flushing liquid. Firstly, in the anti-joint urate deposition condition, compared with the large-scale urate crystal clusters scattered or gathered into a feather shape in the model group quail wrist joint flushing liquid, the urate crystals existing in the high, medium and low dose group quail wrist joint flushing liquid of the chrysanthemums are scattered, single needle crystals or crystal bundles which are arranged in two-three parallel or overlapped in an X shape in a crossing way. Secondly, under the condition of resisting inflammation caused by joint urate deposition, the swelling degree of the wrist and paw joints of the chrysanthemum indicum extract can be obviously reduced at the dosage of 3.75-10 g/kg, which indicates that the chrysanthemum indicum extract has better effect of resisting gouty arthritis.

The research results show that the traditional Chinese medicine composition has obvious effect of resisting gouty arthritis, the efficacy of the traditional Chinese medicine composition is possibly related to removing joint uric acid crystals and improving joint injury of model animals, and a new medicine choice is provided for preventing and treating gouty arthritis.

Example 3 Chinese herbal Compound composition and method of preparing the same

The traditional Chinese medicine composition is prepared by extracting 9g of chicory, 10g of poria cocos, 30g of rhizoma smilacis glabrae, 6g of pagodatree flower bud and 9g of gentiana macrophylla with water.

S1, adding deionized water with the mass ratio of 12 times, extracting for 1h by adopting a decoction method, collecting an extracting solution, repeating for 3 times, and combining the extracting solutions;

s2, concentrating, namely concentrating the obtained mixed liquid medicine at minus 0.09Mpa and 80 ℃ under reduced pressure to obtain thick paste;

S3, preparing medicinal powder, namely drying the obtained thick paste at the temperature of-0.09 Mpa and 80 ℃ under reduced pressure to obtain dry extract, taking the dry extract, crushing the dry extract, and sieving the crushed dry extract with a 60-mesh sieve to obtain the traditional Chinese medicine composition.

Auxiliary materials including lactose, starch, dextrin, sorbitol and mannitol

The dry extract powder of the traditional Chinese medicine composition is added with a proper amount of auxiliary materials and prepared into the dosage forms of granules, capsules or oral liquid by a conventional method.

Example 4 Chinese herbal Compound composition and method for preparing the same

The traditional Chinese medicine composition is prepared by extracting 7g of chicory, 13g of poria cocos, 20g of rhizoma smilacis glabrae, 9g of pagodatree flower bud and 5g of gentiana macrophylla with water.

The extraction method is the same as in example 3

Auxiliary materials including lactose, starch, dextrin, sorbitol and mannitol

Example 5 Chinese herbal Compound composition and method for preparing the same

The traditional Chinese medicine composition is prepared by extracting 15g of chicory, 6g of poria cocos, 50g of rhizoma smilacis glabrae, 5g of pagodatree flower bud and 8g of gentiana macrophylla with water.

The extraction method is the same as in example 3

Auxiliary materials including lactose, starch, dextrin, sorbitol and mannitol

Claims

1. Use of a traditional Chinese medicine composition in the preparation of an anti-gouty arthritis drug, characterized in that the traditional Chinese medicine composition is made of the following raw materials: 6-18 parts by weight of chicory, 5-15 parts by weight of Poria cocos, 10-60 parts by weight of Smilax glabra, 5-10 parts by weight of Sophora japonica seeds, and 3-10 parts by weight of Gentiana macrophylla.

2. The use according to claim 1, characterized in that the traditional Chinese medicine composition is made of the following raw materials: 6-10 parts by weight of chicory, 10-15 parts by weight of Poria cocos, 20-40 parts by weight of Smilax glabra, 5-10 parts by weight of Sophora japonica seeds, and 3-10 parts by weight of Gentiana macrophylla.

3. The use according to claim 2, characterized in that the traditional Chinese medicine composition is made of the following raw materials: 9 parts by weight of chicory, 10 parts by weight of Poria cocos, 30 parts by weight of Smilax glabra, 6 parts by weight of Sophora japonica seeds, and 9 parts by weight of Gentiana macrophylla.

4. The use according to any one of claims 1-3, characterized in that the Chinese medicine composition is: a composition formed by mixing the raw materials after they are crushed separately; or, a composition obtained by mixing the raw materials and then crushing them; or, the raw materials are mixed and then extracted according to a conventional extraction method to obtain an extract, the extract is further subjected to a refining and purification process to obtain an effective part, and the above extract and effective part are further prepared according to a conventional preparation process to obtain a conventional oral dosage form.

5. The use according to claim 4, characterized in that the conventional oral dosage form is tablets, capsules, granules, pills, powders, and oral liquids.

6. The use according to any one of claims 1 to 3, characterized in that the Chinese medicine composition is prepared by the following method:

S1, take each raw material drug in proportion and add water to extract;

S2, concentrating the extract into a thick paste;

S3, drying the thick paste under reduced pressure to obtain a dry extract; crushing and sieving to obtain a Chinese medicine composition.

7. The use according to any one of claims 1 to 3, characterized in that the Chinese medicine composition is used in the preparation of a drug for removing uric acid crystals in joints.

8. The use according to any one of claims 1 to 3, characterized in that the Chinese medicine composition is used in the preparation of a medicament for inhibiting inflammatory response in gouty arthritis.