CN121401372A

CN121401372A - Pharmaceutical Compositions and Their Application in the Preparation of Drugs for the Prevention and Treatment of Hyperuricemia and Gout

Info

Publication number: CN121401372A
Application number: CN202511712777.5A
Authority: CN
Inventors: 徐世军; 王平; 赵梅欢
Original assignee: Chengdu University of Traditional Chinese Medicine
Current assignee: Chengdu University of Traditional Chinese Medicine
Priority date: 2025-11-20
Filing date: 2025-11-20
Publication date: 2026-01-27

Abstract

This invention relates to the field of traditional Chinese medicine compound preparations, and discloses a pharmaceutical composition and its application in the preparation of drugs for the prevention and treatment of hyperuricemia and gout. The pharmaceutical composition uses Gentiana macrophylla as the principal ingredient, focusing on dispelling wind and dampness, clearing damp-heat, relieving pain, inhibiting uric acid production, and promoting renal excretion to lower blood uric acid levels. Gastrodia elata and Achyranthes bidentata are the assistant ingredients; Gastrodia elata dispels wind and unblocks the meridians, tonifies the liver and kidneys; Achyranthes bidentata promotes joint mobility, guides the medicine downwards, and promotes diuresis. Imperata cylindrica root is added as an adjuvant to clear heat, promote diuresis, and expel turbidity. Ligusticum chuanxiong is used as the guiding ingredient to invigorate blood circulation, promote qi circulation, dispel wind, and relieve pain. Through targeted adjustments to the proportions of each ingredient and scientific formulation, the composition achieves effects such as lowering serum uric acid levels, significantly increasing liver uricase activity, simultaneously improving renal function indicators, reducing liver and kidney pathological damage, and combating acute gout, demonstrating significant efficacy.

Description

Pharmaceutical composition and application thereof in preparation of medicines for preventing and treating hyperuricemia and gout

Technical Field

The invention relates to the technical field of traditional Chinese medicine compound preparations, in particular to a pharmaceutical composition and application thereof in preparing medicines for preventing and treating hyperuricemia and gout.

Background

Hyperuricemia (HUA) is a metabolic disease caused by in vivo purine metabolic disorder and/or uric acid excretion reduction, and not only directly causes the onset of gout, but also is an important cause of chronic diseases such as renal function impairment, hypertension, diabetes and the like. With the change of the dietary structure of people, the incidence rate of hyperuricemia and gout is continuously rising, and the hyperuricemia and gout become serious public health problems. At present, two types of medicines, namely medicines for inhibiting uric acid generation, such as allopurinol and febuxostat, and medicines for promoting uric acid excretion, such as phenylbromarone and probenecid, are mainly adopted in Western medicine for clinically treating hyperuricemia and gout. Although these drugs are able to lower serum uric acid levels faster, their long-term safety issues are becoming increasingly prominent. Allopurinol may cause severe skin allergic reactions, febuxostat has a potential cardiovascular risk, benzbromarone tablet has a potential hepatotoxicity hazard, and needs to be carefully used in patients with renal insufficiency. More importantly, the medicines can only temporarily control indexes, and serum uric acid level is easy to quickly rise after stopping medicines, so that the dilemma of effective medicine taking and medicine stopping recurrence is formed, and the treatment confidence and long-term prognosis of patients are seriously affected. In addition, the existing medicines are often focused on a single link (generation or excretion), and for complex inflammatory reactions in acute onset of gout, non-steroidal anti-inflammatory medicines or colchicine are often required to be additionally taken, so that the medicine burden of a patient is increased, and new side effect risks such as gastrointestinal damage and the like are brought.

Although there are some traditional Chinese medicine formulas for treating hyperuricemia, most of the traditional ideas of clearing heat and promoting diuresis, and relieving arthralgia and pain remain, and the formula ideas are relatively single. The medicinal material combination of the formulas often lacks pertinence, the specific link of uric acid metabolism is not fully considered, the generation of uric acid can not be effectively inhibited, the excretion of uric acid is difficult to promote, and the treatment effect is often not ideal. Meanwhile, the existing prescription mostly neglects the damage of hyperuricemia to important organs such as liver and kidney and lacks corresponding protection effect. Although these methods have a certain effect in promoting urination or alleviating symptoms, it is difficult to achieve a rapid, long-lasting and comprehensive therapeutic effect in terms of synergistically suppressing uric acid production, promoting renal excretion, and anti-inflammatory and analgesic effects.

Therefore, a scientifically compatible traditional Chinese medicine composition is researched, not only can uric acid synthesis be reduced, but also uric acid can be promoted to be discharged from kidneys efficiently, and effects of effectively regulating uric acid metabolism, inhibiting gout and protecting liver and kidney functions are achieved, so that the traditional Chinese medicine composition has important significance.

Disclosure of Invention

Aiming at the defects in the prior art, the invention aims to provide a pharmaceutical composition and application thereof in preparing medicines for preventing and treating hyperuricemia and gout. The pharmaceutical composition takes gentiana macrophylla as a monarch drug, gastrodia elata and cyathula officinalis as ministerial drugs, rhizoma imperatae as adjuvant drugs and ligusticum wallichii as conductant drugs, and through the proportion of the specific raw materials, scientific compatibility of the raw materials has the effects of inhibiting uric acid generation and promoting uric acid excretion, thereby realizing the effects of reducing serum uric acid level, obviously improving liver uricase activity, synchronously improving kidney function indexes, relieving liver and kidney pathological injury and resisting acute gout, and has obvious efficacy.

In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:

The invention provides a pharmaceutical composition, which is prepared from the following raw materials in parts by weight:

6-15 parts of gentiana macrophylla, 2-8 parts of gastrodia elata, 3-15 parts of cyathula root, 8-30 parts of raw cogongrass rhizome and 2-6 parts of ligusticum wallichii.

The first aspect of the invention provides a pharmaceutical composition which takes gentiana macrophylla as a monarch, and has the effects of dispelling wind-damp, clearing damp-heat, relieving arthralgia, inhibiting uric acid generation, promoting kidney excretion and reducing blood uric acid. Rhizoma Gastrodiae and radix Cyathulae are used as ministerial drugs, and has effects of dispelling pathogenic wind, dredging collaterals, nourishing liver and kidney, dredging joint, guiding downward and promoting urination, and promoting rhizoma Imperatae, clearing heat, promoting urination, and guiding turbid urine. Rhizoma Ligustici Chuanxiong is used for promoting blood circulation, activating qi-flowing, dispelling pathogenic wind and relieving pain. Through the specific adjustment of the proportion of the raw materials and scientific compatibility, the effects of reducing serum uric acid level, obviously improving liver uricase activity, synchronously improving kidney function indexes, relieving liver and kidney pathological injury, resisting acute gout and the like are realized, and the efficacy is obvious.

Preferably, the traditional Chinese medicine composition is prepared from the following raw materials, by weight, 8-10 parts of gentiana macrophylla, 2-4 parts of gastrodia elata, 4-6 parts of cyathula officinalis, 8-10 parts of raw cogongrass rhizome and 4-6 parts of ligusticum wallichii. More preferably, the traditional Chinese medicine composition is prepared from the following raw materials by weight part, 9 parts of gentiana macrophylla, 3 parts of gastrodia elata, 5 parts of medicinal cyathula root, 9 parts of raw cogongrass rhizome and 5 parts of szechuan lovage rhizome.

The second aspect of the present invention provides a method for preparing the above pharmaceutical composition, comprising the steps of:

Percolating radix Gentianae Marcrophyllae, rhizoma Gastrodiae and radix Cyathulae with ethanol water solution to obtain ethanol extractive solution;

steam distilling rhizoma Ligustici Chuanxiong, and separating to obtain volatile oil and distilled water solution;

extracting rhizoma Imperatae and residues generated after distilling rhizoma Ligustici Chuanxiong with water to obtain water extractive solution;

and combining the alcohol extract, the distilled water solution and the water extract, and concentrating to obtain paste.

Percolating with ethanol water solution to obtain radix Gentianae Marcrophyllae, rhizoma Gastrodiae and radix Cyathulae, extracting liposoluble active ingredients, distilling to obtain rhizoma Ligustici Chuanxiong volatile oil, retaining key medicinal materials for promoting blood circulation and activating qi-flowing, extracting residues and rhizoma Imperatae with water, and dissolving water-soluble diuretic components such as polysaccharide and potassium salt. The preparation method of the pharmaceutical composition provided by the invention has the advantages that the active substances of the raw materials are efficiently reserved, the test data show remarkable comprehensive curative effects, not only can serum uric acid level be effectively reduced, but also liver uricase activity can be remarkably improved, a definite liver and kidney protection function is shown, and joint swelling degree and pain sensitivity of gouty arthritis model animals can be inhibited.

Further, the coarse powder mixture of the large-leaf gentian root, the tall gastrodia tuber and the medicinal cyathula root is soaked for 18 to 36 hours by using the ethanol water solution with the volume concentration of 60 to 80 percent and then is percolated.

Further, the water extraction is carried out by adding water and decocting for 2-4 times, each time for 1-1.5h.

Further, the alcohol aqueous solution is 60% -80% of ethanol solution.

Further, the relative density of the paste is 1.3-1.5.

The third aspect of the invention provides a method for preparing the granule, which comprises the steps of mixing the paste prepared by the preparation method with pharmaceutically acceptable auxiliary materials, granulating, and adding the ligusticum wallichii volatile oil.

In a fourth aspect, the invention provides a method of preparing an instant oral powder comprising:

drying and crushing the paste prepared by the preparation method to obtain medicinal powder;

Mixing the medicine powder with the medicinal auxiliary materials subjected to superfine grinding treatment, wherein the average particle size of the mixture is less than or equal to 50 mu m;

adding the rhizoma Ligustici Chuanxiong volatile oil, and mixing.

The oral instant powder provided by the invention is prepared by drying and crushing paste obtained by extraction and concentration to obtain medicine powder with enriched active ingredients, mixing the medicine powder with crushed pharmaceutic adjuvant to obtain a mixture with the particle size of less than or equal to 50 mu m, adding the ligusticum wallichii volatile oil, and uniformly mixing to obtain the oral instant powder. The oral cavity instant dispersion can be completely dispersed and dissolved within 30 seconds when contacting saliva, thus effectively improving the dissolution rate and bioavailability of the medicine, and simultaneously, the ultrafine powder carrier effectively coats volatile active ingredients such as volatile oil of Ligusticum chuanxiong, thus obviously improving the storage stability of the preparation.

The fifth aspect of the invention provides an oral preparation, which contains the pharmaceutical composition and pharmaceutically acceptable auxiliary materials, and/or is granules, capsules, tablets, oral liquid, powder, oral instant powder or dripping pills.

In a fifth aspect, the present invention provides the use of a pharmaceutical composition as described above in the manufacture of a medicament for the treatment or prophylaxis of hyperuricemia, and/or gout, and/or gouty arthritis and/or uric acid nephropathy.

Further, the pharmaceutical composition can have any one of the following effects:

(a) Reducing blood uric acid level of hyperuricemia model animals;

(b) Inhibiting xanthine oxidase activity;

(c) Reducing joint swelling and pain sensitivity of gouty arthritis model animals;

(d) Inhibiting inflammatory factor expression associated with gout.

In summary, due to the adoption of the technical scheme, the beneficial effects of the invention are as follows:

The invention provides a pharmaceutical composition and application thereof in preparing medicines for preventing and treating hyperuricemia and gout, wherein the pharmaceutical composition takes gentiana macrophylla as a monarch, and has the effects of dispelling wind dampness, clearing damp and heat, relieving arthralgia, inhibiting uric acid generation, promoting kidney excretion and reducing blood uric acid. Rhizoma Gastrodiae and radix Cyathulae are used as ministerial drugs, and has effects of dispelling pathogenic wind, dredging collaterals, nourishing liver and kidney, dredging joint, guiding downward and promoting urination, and promoting rhizoma Imperatae, clearing heat, promoting urination, and guiding turbid urine. Rhizoma Ligustici Chuanxiong is used for promoting blood circulation, activating qi-flowing, dispelling pathogenic wind and relieving pain. Through the specific adjustment of the proportion of the raw materials and scientific compatibility, the effects of reducing serum uric acid level, obviously improving liver uricase activity, synchronously improving kidney function indexes, relieving liver and kidney pathological injury, resisting acute gout and the like are realized, and the efficacy is obvious.

Drawings

FIG. 1 shows the change in serum uric acid levels of 10 patients suffering from hyperuricemia in example 1 before and after administration of the pharmaceutical composition of the invention.

FIG. 2 is a view of liver and kidney histopathological section (HE staining, 200X) of each group of mice in example 2, and the arrows in FIG. 2 indicate characteristic pathological change regions.

FIG. 3 is a graph showing the effect of JM administration on ankle morphology in mice with acute gouty arthritis model.

FIG. 4 shows the effect of JM administration on the ankle thickness variation of mice with acute gouty arthritis modelSEM), with #p <0.05 compared to Control group, and with Model group, # p <0.01.

FIG. 5 shows the effect of JM administration on the variation of foot thickness of mice with acute gouty arthritis modelSEM), # p <0.001 compared to Control group, and p <0.05 compared to Model group.

FIG. 6 shows the effect of JM administration on arthritis score in mice with acute gouty arthritis modelSEM), # p <0.001 compared to Control group, and # p <0.001 compared to Model group.

Fig. 7 shows the effect of JM administration on the change in left pain threshold (x±sem) in mice Model for acute gouty arthritis, # # p <0.001 compared to Control group and # p <0.001 compared to Model group.

FIG. 8 shows the effect of JM administration on the change in the right foot pain threshold (x+ -SEM) of mice Model for acute gouty arthritis, ^### p <0.001 compared to the Control group, and ^*** p <0.001 compared to the Model group.

FIG. 9 shows the effect of JM administration on serum TNF-. Alpha.content of mice model for acute gouty arthritisSD), # p <0.001 compared to the Control group, and # p <0.05 compared to the Model group.

Detailed Description

The present invention will be described in detail with reference to the accompanying drawings.

The present invention will be described in further detail with reference to the drawings and examples, in order to make the objects, technical solutions and advantages of the present invention more apparent. It should be understood that the specific embodiments described herein are for purposes of illustration only and are not intended to limit the scope of the invention.

The first aspect of the present embodiment provides a pharmaceutical composition, which is prepared from the following raw materials in parts by weight:

The first aspect of the present embodiment provides a pharmaceutical composition, which uses gentiana macrophylla as a monarch drug, and has the effects of dispelling wind-damp, clearing damp-heat, relieving arthralgia, inhibiting uric acid generation, and promoting kidney excretion so as to reduce blood uric acid. Rhizoma Gastrodiae and radix Cyathulae are used as ministerial drugs, and has effects of dispelling pathogenic wind, dredging collaterals, nourishing liver and kidney, dredging joint, guiding downward and promoting urination, and promoting rhizoma Imperatae, clearing heat, promoting urination, and guiding turbid urine. Rhizoma Ligustici Chuanxiong is used for promoting blood circulation, activating qi-flowing, dispelling pathogenic wind and relieving pain. Through the specific adjustment of the proportion of the raw materials and scientific compatibility, the effects of reducing serum uric acid level, obviously improving liver uricase activity, synchronously improving kidney function indexes, relieving liver and kidney pathological injury, resisting acute gout and the like are realized, and the efficacy is obvious.

A second aspect of the present embodiment provides a method for preparing the pharmaceutical composition described above, comprising the steps of:

Percolating with ethanol water solution to obtain radix Gentianae Marcrophyllae, rhizoma Gastrodiae and radix Cyathulae, extracting liposoluble active ingredients, distilling to obtain rhizoma Ligustici Chuanxiong volatile oil, retaining key medicinal materials for promoting blood circulation and activating qi-flowing, extracting residues and rhizoma Imperatae with water, and dissolving water-soluble diuretic components such as polysaccharide and potassium salt. The preparation method of the pharmaceutical composition provided by the embodiment efficiently retains the active substances of the raw materials, and the test data show remarkable comprehensive curative effects, so that the serum uric acid level can be effectively reduced, the liver uricase activity can be remarkably improved, and the definite liver and kidney protection function is shown.

In some embodiments, the aqueous alcohol solution is 60% -80% ethanol solution.

In some embodiments, the relative density of the paste is 1.3 to 1.5.

In a third aspect of this embodiment, a method for preparing a granule is provided, which comprises mixing the paste prepared by the above preparation method with pharmaceutically acceptable adjuvants, granulating, and adding rhizoma Ligustici Chuanxiong volatile oil.

For example, in some embodiments, the detailed preparation process of the granule formulation is:

Pulverizing radix Gentianae Marcrophyllae, rhizoma Gastrodiae and radix Cyathulae into coarse powder, soaking in 70% ethanol for 24 hr, percolating, and collecting percolate 2000 mL. Extracting volatile oil from rhizoma Ligustici Chuanxiong by steam distillation, collecting distilled water solution, decocting the residue and rhizoma Imperatae with water for three times each for 1 hr, filtering decoction, mixing filtrates, mixing with the above water solution, concentrating to about 500ml, mixing with the above percolate, recovering ethanol, and concentrating to obtain soft extract with relative density of 1.39+ -0.1 (60-65deg.C). Mixing soft extract 1g, sucrose powder 2.5 g and dextrin 1.5 g, granulating, drying, spraying volatile oil of rhizoma Ligustici Chuanxiong, and mixing.

A fourth aspect of the present embodiment provides a method of preparing an instant oral powder comprising:

adding the rhizoma Ligustici Chuanxiong volatile oil, and mixing.

The oral instant powder provided by the embodiment is prepared by drying and crushing paste obtained by extraction and concentration to obtain medicine powder with enriched active ingredients, mixing the medicine powder with crushed medicinal auxiliary materials to obtain a mixture with the particle size of less than or equal to 50 mu m, adding the ligusticum wallichii volatile oil, and uniformly mixing to obtain the oral instant powder. The oral cavity instant dispersion can be completely dispersed and dissolved within 30 seconds when contacting saliva, thus effectively improving the dissolution rate and bioavailability of the medicine, and simultaneously, the ultrafine powder carrier effectively coats volatile active ingredients such as volatile oil of Ligusticum chuanxiong, thus obviously improving the storage stability of the preparation.

For example, in some embodiments, the detailed preparation process of the oral instant powder is as follows:

Pulverizing radix Gentianae Marcrophyllae, rhizoma Gastrodiae and radix Cyathulae into coarse powder, soaking in 70% ethanol for 24 hr, percolating, and collecting percolate 2000 mL. Extracting volatile oil from rhizoma Ligustici Chuanxiong by steam distillation, collecting distilled water solution, decocting the residue and rhizoma Imperatae with water for three times each for 1 hr, filtering decoction, mixing filtrates, mixing with the above water solution, concentrating to about 500ml, mixing with the above percolate, recovering ethanol, and concentrating to obtain soft extract with relative density of 1.39+ -0.1 (60-65deg.C). Taking 1g of thick paste, drying under reduced pressure, pulverizing into fine powder, spraying volatile oil of rhizoma Ligustici Chuanxiong, mixing and grinding lactose powder 1g and glucose powder 1g into powder with particle size of 0.3-5 μm, mixing and grinding xylitol 1.5g to 0.2-3 μm, mixing the medicine and adjuvant powder in vibration mill, and obtaining the instant oral powder.

The fifth aspect of the present embodiment provides an oral preparation, which contains the pharmaceutical composition and pharmaceutically acceptable excipients, and/or the oral preparation is a granule, a capsule, a tablet, an oral liquid, a powder, an oral instant powder or a dripping pill.

In some embodiments, the auxiliary materials comprise, but are not limited to, sucrose, lactose, microcrystalline cellulose or mannitol as a filling agent, starch slurry, polyvinylpyrrolidone or hydroxypropyl methylcellulose as a binding agent, croscarmellose sodium or sodium carboxymethyl starch as a disintegrating agent, magnesium stearate or talcum powder as a lubricant, aspartame or fruit essence as a flavoring agent and hydroxypropyl methylcellulose or acrylic resin as a coating material, wherein the auxiliary materials can be used singly or in combination according to the production process requirements of different dosage forms, for example, granules can comprise sucrose and a flavoring agent, tablets can comprise microcrystalline cellulose, croscarmellose sodium and magnesium stearate, and the powder can use superfine crushed lactose and glucose to prepare oral instant powder so as to ensure good stability, dissolution characteristics and administration compliance of the preparation.

A fifth aspect of this embodiment provides the use of a pharmaceutical composition as described above in the manufacture of a medicament for the treatment or prophylaxis of hyperuricemia, and/or gout, and/or gouty arthritis and/or uric acid nephropathy.

In some embodiments, the pharmaceutical composition can have any one of the following effects:

(a) Reducing blood uric acid level of hyperuricemia model animals;

(b) Inhibiting xanthine oxidase activity;

(d) Inhibiting inflammatory factor expression associated with gout.

In order to more clearly explain the foregoing example embodiments, the following specific embodiments are provided for further explanation:

Example 1

Raw materials of the medicine comprise 9 parts of gentiana macrophylla, 3 parts of gastrodia elata, 5 parts of medicinal cyathula root, 9 parts of raw cogongrass rhizome and 5 parts of szechuan lovage rhizome.

30 Patients taking the standard decoction of the drug ratio of example 1 were statistically analyzed for serum uric acid levels before and after treatment using a retrospective study method. The proportion of men and women, the age and the like of 30 outpatients have statistical significance.

Treatment prescription:

the standard decoction prepared in example 1 was taken 30 d a day after one dose.

As shown in fig. 1 and table 1, patients with hyperuricemia are treated for 1 month by taking the standard decoction prepared in example 1 (other uric acid lowering drugs are not used together), and serum uric acid is obviously reduced after treatment, which suggests that the pharmaceutical composition of the invention has good therapeutic effect of reducing serum uric acid.

TABLE 1 influence of serum uric acid treatment on 30 patients±SD)

As shown in Table 1 and FIG. 1, the pharmaceutical composition provided by the invention has good effect of reducing serum uric acid level clinically. Furthermore, during the administration period, no significant adverse reactions and toxic side effects associated with the drug were observed in the clinical patients.

Example 2

Purpose of experiment

The uric acid-lowering efficacy of the standard decoction of the drug ratio of example 2 was evaluated by using an adenine and potassium oxazinate-induced KM mouse hyperuricemia model.

Materials and methods

Mouse strain

54 KM mice (SPF, all male, 4-6 weeks old, 28-32 g) were purchased from Sichuan Tonglihua laboratory animal technologies Co., ltd, production license number SCXK (Sichuan) 2023-0040, animal pass No.511215600024062. The experimental animals are raised in the environment with room temperature (22+/-2) DEG C, humidity of 50% -60% and light circulation (12 hours of light/12 hours of darkness). The animal experiment development place is SPF animal raising room of university of Chinese medicine and college of medicine, single cage raising, using license SYXK (Chuan) 2025-0294. The experiments are approved by the animal welfare ethics committee of the university of Chengdu traditional Chinese medicine, and the ethics number is 2025122.

Design of experiment

After the KM mice were adaptively fed with 7 d, they were randomly divided into 7 groups according to body weight stratification, namely, a blank Control group (Control, n=10), a hyperuricic acid Model Control group (Model, n=10), a positive Control group (Febuxostat, feb, n=8), an instant dispersion low dose group of example 2 (JM-L, n=8) and an instant dispersion high dose group of example 2 (JM-H, n=8). Mice were prepared with 100 mg/kg adenine lavage compound 300 mg/kg potassium oxazinate intraperitoneal injection daily for hyperuricemia models, and the blank control group was given equal volumes of saline lavage and intraperitoneal injection. After molding 1h a day, the mice of each group were subjected to gastric lavage with an equal volume of 0.5% CMC-Na corresponding drug solution except for 10 mL/kg of the blank control group and the hyperuricic acid model control group, and 15 d a 1/d continuous administration was performed.

Sample preparation and index detection

(1) Determination of uric acid, namely, taking blood from the eye sockets at 0d (basic value) before molding, 5d, 10d and 15d after molding, preparing serum, collecting 24 mouse urine, and detecting the uric acid content in serum and urine samples by using a uric acid kit, wherein the serum uric acid change value = serum uric acid actual measurement value-basic value before administration.

(2) Serum creatinine and uric acid nitrogen content detection, namely, blood is taken from the eye socket for 0d before molding, 5d after molding, 10d and 15d respectively, serum is prepared, and creatinine and uric acid nitrogen content is detected by adopting a creatinine and uric acid nitrogen detection kit.

(3) And (3) measuring urease, namely taking liver tissues after the administration is finished, and detecting the activity of uric acid oxidase in the liver tissues by using a urease kit.

(4) Histopathological staining, namely fixing liver and kidney tissues in paraformaldehyde after the administration, dehydrating, embedding and slicing, staining hematoxylin-eosin, and observing pathological morphological changes of the liver and kidney tissues.

Statistical processing of data

Data were processed by GRAPHPAD PRISM 9.3.0 software and data were expressed as mean±sd. The average comparison among the groups adopts analysis of variance, and the comparison among the groups adopts t-test. p <0.05 indicates that the difference is statistically significant.

Experimental results

Effect of JM on serum uric acid content of hyperuricemia model mice

As shown in Table 2, the serum uric acid of mice in the Model group was significantly increased in comparison with the Control group, and had a good time dependence (p < 0.01), suggesting that the mice in the hyperuricemia Model were successfully prepared, and the serum uric acid increase values of mice in the Model group were significantly decreased (p <0.05 or p < 0.01) at the corresponding time points in the Feb and JM groups compared with the Model group, but there was no statistical difference (p > 0.05) in Feb at the time point of 15 d.

TABLE 2 JM influence on serum uric acid levels in mice with hyperuricemia models±SD)

Note that ^##p <0.01,^### p <0.001 compared to the Control group and ^*p <0.05,^** p <0.01 compared to the Model group.

Effect of JM on uric acid content in urine of hyperuricemia model mice

As shown in Table 3, the uric acid levels in the urine of mice in the Model group were extremely significantly reduced (p < 0.001) in comparison with the Control group, suggesting that the uric acid levels in mice in the Model group were significantly impaired by urine excretion, and the uric acid levels in urine at the corresponding time points were significantly increased (p <0.05 or p < 0.01) in the Feb-administered group compared with the Model group, whereas the uric acid levels in urine at the corresponding time points were significantly increased in each JM-administered group, and showed a certain dose correlation, but no significant statistical significance (p > 0.05), suggesting that JM has a certain effect of reducing serum uric acid by promoting uric acid urine excretion.

TABLE 3 JM influence on urine uric acid content of mice with hyperuricemia model±SD)

Note that ^### p <0.001 compared to the Control group and ^*p <0.05,^** p <0.01 compared to the Model group.

Effect of JM on liver uricase Activity of hyperuricase model mice

As shown in Table 4, the liver uricase activity of mice in the Model group was extremely significantly reduced (p < 0.001) compared with the Control group, suggesting that the uric acid oxidative metabolism of mice in the Model group was impaired, and the liver uricase activity of each of Feb and JM administration groups was significantly increased (p <0.05, p <0.01 or p < 0.001) compared with the Model group, suggesting that JM could promote uric acid oxidation by enhancing the uric acid oxidase activity in the liver, thereby achieving the effect of lowering serum uric acid.

TABLE 4 JM influence on liver uricase Activity of mice with hyperuricemia model±SD)

Note that ^## p <0.01 compared to the Control group and ^*p <0.05,^**p <0.01,^*** p <0.001 compared to the Model group.

Effect of JM on serum creatinine content in mice with hyperuricemia model

As shown in Table 5, the serum creatinine content of mice in the Model group was significantly increased (p <0.01 or p < 0.001) during the molding process compared with the Control group, suggesting that the hyperuricemia Model had significant kidney function impairment, and the JM-administered group was significantly decreased in serum creatinine content of 10d and 15 d, especially in serum creatinine content of 15 d compared with the Model group, and had significant statistical significance (p <0.05 or p < 0.001), suggesting that JM had good protective effect on kidney function impairment due to hyperuricemia.

TABLE 5 JM influence on serum creatinine content of mice with hyperuricemia model±SD)

Note that ^##p <0.01,^### p <0.001 compared to the Control group and ^*p <0.05,^**p <0.01,^*** p <0.001 compared to the Model group.

Effect of JM on serum urea nitrogen content of hyperuricemia model mice

As shown in Table 6, the serum urea nitrogen content in the Model mice during the molding process was also significantly increased (p <0.05 or p < 0.001) compared with the Control group, suggesting that the hyperuricemia Model kidney filtration and excretion function was impaired, and the JM-administered group showed significantly decreased serum urea nitrogen content at 15 d th dose compared with the Model group (p <0.05 or p < 0.001), suggesting that JM has a good protective effect against kidney function injury caused by potassium oxazinate.

TABLE 6 JM influence on serum urea nitrogen content of mice with hyperuricemia model±SD)

Note that ^##p <0.05,^### p <0.001 compared to the Control group and ^*p <0.05,^** p <0.01 compared to the Model group.

Effect of JM on liver and kidney pathological form damage in hyperuricemia model mice

As shown in the HE staining results of the liver, compared with the Control group, the liver tissue of the mice in the Model group has obvious balloon-like change, which is shown by obvious increase of the number of liver cell vacuoles, concentration of cytoplasm, fragmentation of nucleus, obvious increase of damaged cells forming eosinophils, obvious glomerular atrophy, tubular expansion, inflammatory cell infiltration, unclear cell boundaries of proximal tubules, thinning of tubular wall epithelial cells, degeneration of cell vacuoles, particle degeneration and even necrosis shedding, and obvious reduction of vacuoles of liver cells and the number of cells forming eosinophils of mice in the Feb and JM administration groups, obvious reduction of pathological changes of the kidney, obvious reduction of inflammatory cell infiltration, and prompt that JM has good protection effect on liver and kidney cell injury of mice in the hyperuricic Model.

Hyperuricemia is a chronic metabolic disease caused by purine metabolic disorder, caused by reduced uric acid excretion or increased uric acid production, and clinically manifested as elevated serum uric acid, is a key cause of gout, and is often accompanied by obesity and abnormal glycolipid metabolism. The treatment is mainly to inhibit uric acid generation and/or promote uric acid excretion, and long-term hyperuricemia can lead to liver and kidney damage, further worsen uric acid metabolism and aggravate hyperuricemia. According to the clinical manifestations of the traditional Chinese medicine, the disease is caused by damp-heat blockage at the heart of the disease, and the theory of wind-damp-heat blockage is called "Zhou Zhu Zhong, strong and supporting pain, acute muscle contraction in the neck, and qi of liver and gallbladder at the location of the foot chilly wind Xue Zhi. The treatment is to clear heat and remove dampness and promote joint circulation.

According to the application, the adenine and potassium oxazinate combined induction KM mice hyperuricemia model is used for evaluating the therapeutic effect of the pharmaceutical composition provided by the application on the model. Compared with a normal group, the pharmaceutical composition can obviously reduce the serum uric acid content of a model mouse, promote the discharge of urea in the urine, improve the activity of liver uricase, reduce the content of serum creatinine and urea nitrogen, and improve the cell damage of liver and kidney of the model mouse. In conclusion, the pharmaceutical composition provided by the application has good effects of reducing serum uric acid, reducing serum uric acid level mainly by inhibiting uric acid generation and promoting uric acid excretion, protecting liver and kidney functions, and playing a comprehensive treatment role on hyperuricemia model mice constructed by adenine and potassium oxazinate.

Example 3

All examples and comparative examples used a preparation process exactly identical to that of inventive example 1.

Example 3-1 gentiana macrophylla 6 parts, gastrodia elata 2 parts, cyathula officinalis 3 parts, rhizoma imperatae 8 parts, and ligusticum wallichii 2 parts.

Example 3-2 gentiana macrophylla 15 parts, gastrodia tuber 8 parts, cyathula root 15 parts, cogongrass rhizome 30 parts, ligusticum wallichii 6 parts.

Example 3-3 gentiana macrophylla 8 parts, gastrodia elata 4 parts, cyathula root 4 parts, cogongrass rhizome 10 parts, ligusticum wallichii 4 parts.

Comparative example 1 gentiana macrophylla was subtracted from the formulation of examples 3-3.

Comparative example 2 Cyathula root was subtracted from the formulation of examples 3-3.

Comparative example 3 raw lalang grass rhizome was subtracted from the formulation of example 3-3.

Comparative example 4 tall gastrodia tuber was subtracted from the formulation of example 3-3.

Comparative example 5 Ligusticum wallichii was subtracted from the formulation of examples 3-3.

Comparative example 6 based on the formulation of examples 3-3, gentiana macrophylla was replaced with an equivalent amount of radix clematidis.

Comparative example 7 based on the formulation of examples 3-3, fresh cogongrass rhizome was replaced with an equivalent amount of plantain herb.

Comparative example 8 the amount of gentiana macrophylla was reduced to 4 parts based on the formulation of examples 3-3.

Comparative example 9 gentiana macrophylla 3 parts, gastrodia elata 1 part, cyathula root 3 parts, rhizoma imperatae 4 parts, and Ligusticum wallichii 1 part.

Comparative example 10, 20 parts of gentiana macrophylla, 10 parts of gastrodia elata, 20 parts of medicinal cyathula root, 40 parts of raw cogongrass rhizome and 8 parts of szechuan lovage rhizome.

3. Evaluation of pharmacodynamics

Animal model and method

Model adenine (100 mg/kg) +Potassium oxazinate (300 mg/kg) induced mouse hyperuricemia model.

The groups were a blank control group, a model control group, a positive drug (febuxostat) group, an example 3-1 group, an example 3-2 group, an example 3-3 group, and a comparative example 1-10 group.

The core indicators are shown in Table 7 as Serum Uric Acid (SUA), liver Uricase (UOX) activity, 24-hour Urine Uric Acid (UUA) excretion, and serum creatinine (SCr) following 15 days of administration in the high dose group of example 2.

TABLE 7 comparison of pharmacodynamic indicators for groups (x-SD ± SD)

Note that ^*p <0.05,^**p <0.01,^*** p <0.001 compared to the model control group.

Example 4

In example 4, the following formulation of the gentiana macrophylla gout formula is consistent with the formulations of examples 1 and 2. 9 parts of gentiana macrophylla, 3 parts of gastrodia elata, 5 parts of medicinal cyathula root, 9 parts of raw cogongrass rhizome and 5 parts of szechuan lovage rhizome.

1. Purpose of investigation

Based on clinical value as guidance and on the clinical curative effect of the gentian and hemp gout prescription, an MSU crystal injection induced acute gouty arthritis model is adopted to examine the effectiveness and potential action mechanism of the gentian and hemp gout prescription, and a modern pharmacological basis is provided for clinical and industrial transformation.

Experimental materials

2.1 Experimental animal

Male C57BL/6J mice (7-8 weeks old, 18-22 g, 60) purchased from Style Bei Fu (Beijing) biotechnology Co., ltd., production license number SCXK (Beijing) 2024-0001, animal eligibility No.110324251105458174.

2.2 Experimental drugs and reagents

Table 8 experimental drug and reagent information table

2.3 Experimental instrument

TABLE 9 Main laboratory instrument information Table

Experimental method

3.1 Preparation of medicine and standard water decoction

The Marcrophyllae and hemp gout prescription (JM) consists of gentiana macrophylla 9 g, gastrodia elata 3g, cyathula officinalis 5 g, rhizoma imperatae 9 g and ligusticum wallichii 5 g, and the JM standard decoction is prepared by adopting a decoction method and is dissolved in 0.5% CMC-Na solution, and the final concentration is 0.93 g/mL respectively for standby.

3.2 Grouping and administration of animals

After 7 days of adaptive feeding, C57BL/6J mice were randomly divided into 6 groups according to body weight stratification, namely, a blank Control group (Control, n=10), an acute gouty arthritis Model group (Model, n=10), a colchicine positive Control group (Col, n=10), and a gentian gout side low, medium and high dose administration group (JM-L, JM-Z, JM-H, n=10). The Control and Model groups were given respective volumes of 0.5% CMC-Na solution by gavage, the remaining groups were given respective medications by gavage, the gavage volumes were 10 mL/kg,1 time/d, and 9 d consecutive doses were given. After administration of 7 d and intragastric administration of 1, 1h, model under anesthesia and posterior ankle of the administration group mice were respectively injected with 0.15 mL of a 20 mg/mL monosodium urate (MSU) suspension, and Control group was injected with an equal dose of physiological saline.

3.3 Index evaluation and detection

(1) Joint photograph recording right ankle photographs of each group of mice were photographed before and after molding 2, 6, 10, 24 and 48 h.

(2) Foot and ankle swelling measurements the thickness of the left foot and right ankle was marked using digital vernier caliper measurements before and after molding 2, 6, 10, 24 and 48 h. Each measurement was repeated 3 times and averaged.

(3) Arthritis scoring bilateral hind paw joint clinical scores were performed in parallel on each group of mice with reference to the arthritis index criteria of the table below at 3, 8, 24 and 48 h before and after molding.

TABLE 10 arthritis index criteria

Note that the final score was a double-sided hindfoot score running up, with a maximum of 8 scores available.

Von Frey test mechanical pain mice were measured for pain threshold using Von Frey before and after molding, 3, 8, 24 and 48 h, respectively. Each measurement was repeated 3 times and averaged.

Serum inflammatory factor detection (TNF-alpha) the mouse serum was collected on the day of sampling, and the content of the serum TNF-alpha was detected by referring to the kit instructions.

4. Experimental results

4.1 Effect of JM administration on the extent of ankle and foot swelling in mice model for acute gouty arthritis

As shown in fig. 3, after MSU or physiological saline was injected into the ankle joints of mice, various degrees of swelling were observed, and compared with the Control group, the ankle joints 2h to 48h of the Model group mice continued to be swollen, and slight erythema and moderate swelling extended to the toe joints, and the toe appeared stiff, suggesting that the acute gouty arthritis Model mice were successfully established. The ankle swelling was significantly reduced by the JM-L and JM-Z groups at 10H, by 24H, by the Col and JM-H groups, and by 48H, the low, medium, and high dose of Col and JM administration significantly inhibited joint swelling. JM administration was suggested to improve MSU-induced gout joint swelling.

As shown in table 11 and fig. 4, the Model group mice showed a significant increase in ankle swelling thickness change (p < 0.05) compared to the Control group, suggesting that the acute gouty arthritis Model mice were successfully established. In contrast to the Model group, the ankle swelling thickness variation value of the Col group showed a decreasing trend after 6 hours, but no significant difference (p > 0.05), the ankle swelling thickness variation value of the JM group showed a decreasing trend after 6 hours, the JM-L and JM-Z groups showed no significant difference (p > 0.05), and the JM-H group showed a significantly decreased change value (p < 0.01). JM administration was suggested to reduce MSU-induced ankle swelling thickness in mice.

TABLE 11 influence of JM administration on ankle thickness variation in mice model for acute gouty arthritis± SEM)

Note that, #p <0.05 compared to Control group and, #p <0.01 compared to Model group.

As shown in table 12 and fig. 5, the Model group mice showed a significant increase in foot thickness variation (p < 0.001) compared to the Control group, suggesting successful establishment of acute gouty arthritis Model mice. While the Col foot thickness variation was reduced at 48H with no significant difference overall (p > 0.05) compared to the Model group, the JM-Z foot thickness variation was significantly reduced (p < 0.05), and the JM-H group showed a trend of reduction with no significant difference (p > 0.05). JM administration was suggested to reduce MSU-induced swelling of the feet of mice to some extent.

TABLE 12 JM influence of drug administration on the variation of foot thickness in mice model for acute gouty arthritis± SEM)

Note that # # p <0.001 compared to Control group and p <0.05 compared to Model group.

4.2 Effect of JM administration on clinical score of arthritis in mice model for acute gouty arthritis

As shown in table 13 and fig. 6, model mice developed significant arthritic symptoms after 3h molding, erythema and swelling on both ankle, foot and toe, and significantly increased arthritic index (p < 0.001), suggesting that mice developed moderate arthritic symptoms after molding, as compared to Control mice. In contrast, the Col group showed a slight arthritic symptom, the arthritic index was significantly decreased (p < 0.001), and the arthritic index of the JM group was also significantly decreased (p < 0.001). JM administration was suggested to alleviate the clinical manifestations of MSU-induced arthritis in mice.

TABLE 13 influence of JM dosing on the arthritis score in mice with acute gouty arthritis model± SEM)

Note that # # p <0.001 compared to the Control group and # p <0.001 compared to the Model group.

4.3 Effect of JM administration on mechanical pain threshold in mice model for acute gouty arthritis

As shown in tables 14 and 16 and fig. 7 and 8, the pain threshold change amount of the left and right feet of the Model mice was significantly reduced (p < 0.001) compared with the Control group, suggesting that the acute gouty arthritis Model mice were reduced in tolerance to mechanical stimulus. In contrast, the changes in the left and right pain threshold of Col mice were significantly increased (p < 0.001) compared to the Model mice, and the changes in the left and right pain threshold of JM mice were significantly increased (p < 0.001) for each of the administration groups. JM dosing was suggested to restore the model mice' ability to tolerate mechanical stimuli.

TABLE 14 JM influence of dosing on the change in left foot pain threshold in mice model for acute gouty arthritis± SEM)

TABLE 15 JM influence of dosing on the change in threshold right foot pain in mice model for acute gouty arthritis± SEM)

4.5 Effect of JM administration on serum inflammatory factor TNF- α content in mice model of acute gouty arthritis

As shown in Table 16 and FIG. 9, the TNF-. Alpha.content was significantly elevated in the serum of mice from the Model group (p < 0.001) as compared to the Control group. In contrast, the serum levels of TNF- α were significantly reduced in the Col mice (p < 0.05) compared to the Model mice, and were significantly increased in both JM-Z and JM-H mice (p <0.001 or p < 0.05), with the maximum change in TNF- α levels in the JM dosed sera. JM administration was suggested to reduce the levels of TNF- α in the serum of model mice.

TABLE 16 JM influence of dosing on serum TNF-alpha levels in mice model for acute gouty arthritis± SD)

Note that # # p <0.001 compared to the Control group and # p <0.05 compared to the Model group.

5. Knot (S)

The present study systematically evaluated the therapeutic efficacy of JM against acute episodes of gout by constructing a model of monosodium urate (MSU) crystallization-induced acute gouty arthritis in mice. The results show that JM intervention can remarkably inhibit the swelling reaction of ankle joints and toes of model mice, effectively up-regulate the mechanical pain threshold of the model mice, and remarkably reduce the level of proinflammatory cytokine TNF-alpha in serum. The research result provides a reliable experimental basis for the transformation application of JM in the clinical treatment of acute gouty arthritis.

The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, and alternatives falling within the spirit and principles of the invention.

Claims

1. A pharmaceutical composition, characterized in that it is made from the following raw materials in parts by weight:

Gentian root 6-15 parts, Gastrodia elata 2-8 parts, Achyranthes bidentata 3-15 parts, Imperata cylindrica 8-30 parts, Ligusticum chuanxiong 2-6 parts.

2. The pharmaceutical composition according to claim 1, characterized in that it is made from the following raw materials in parts by weight:

Gentian root 8-10 parts, Gastrodia elata 2-4 parts, Achyranthes bidentata 4-6 parts, Imperata cylindrica 8-10 parts, Ligusticum chuanxiong 4-6 parts.

3. The pharmaceutical composition according to claim 2, characterized in that it is made from the following raw materials in parts by weight:

Gentian root 9 parts, Gastrodia elata 3 parts, Achyranthes bidentata 5 parts, Imperata cylindrica root 9 parts, Ligusticum chuanxiong 5 parts.

4. A method for preparing the pharmaceutical composition according to any one of claims 1-3, characterized in that it comprises the following steps:

Gentiana macrophylla, Gastrodia elata, and Achyranthes bidentata were percolated with an alcohol-water solution to obtain an alcohol extract;

The volatile oil of Ligusticum chuanxiong and the distilled aqueous solution were obtained by steam distillation.

Aqueous extracts were obtained by water extraction of raw Imperata cylindrica root and the residue produced after distillation of Ligusticum chuanxiong.

The alcohol extract, the distilled aqueous solution, and the water extract were combined and concentrated to obtain a paste.

5. The preparation method according to claim 4, characterized in that the crude powder mixture of Gentiana macrophylla, Gastrodia elata, and Achyranthes bidentata is soaked in an ethanol aqueous solution with a volume concentration of 60%-80% for 18-36 hours and then percolated; and/or, the water extraction is carried out by boiling with water 2-4 times, each time for 1-1.5 hours; and/or, the alcohol aqueous solution is a 60%-80% ethanol solution; and/or, the relative density of the paste is 1.3-1.5.

6. A method for preparing granules, characterized in that it comprises mixing the ointment obtained by the preparation method according to claim 4 or 5 with pharmaceutically acceptable excipients, granulating, and adding volatile oil of Ligusticum chuanxiong.

7. A method for preparing an oral instant powder, characterized in that it comprises:

The ointment prepared by the method described in claim 4 or 5 is dried and pulverized to obtain drug powder;

The drug powder is mixed with pharmaceutical excipients that have undergone ultrafine pulverization, wherein the average particle size of the mixture is ≤50μm;

Add the volatile oil from Ligusticum chuanxiong and mix well.

8. An oral formulation, characterized in that it contains a pharmaceutical composition as described in any one of claims 1-3 and a pharmaceutically acceptable excipient; and/or, the oral formulation is a granule, capsule, tablet, oral liquid, powder, oral instant powder, or drop pill.

9. The use of the pharmaceutical composition according to any one of claims 1-3 in the preparation of a medicament for the treatment or prevention of hyperuricemia, and/or gout, and/or gouty arthritis and/or uric acid nephropathy.

10. The application according to claim 9, characterized in that the pharmaceutical composition can have any one of the following effects:

(a) Reduce serum uric acid levels in hyperuricemia model animals;

(b) Inhibits xanthine oxidase activity;

(c) Reduce joint swelling and pain sensitivity in animal models of gouty arthritis;

(d) Inhibit the expression of inflammatory factors associated with gout.