Traditional Chinese medicine composition, traditional Chinese medicine extract, preparation of traditional Chinese medicine extract and quality control method of traditional Chinese medicine extract

Abstract

The invention provides a traditional Chinese medicine composition, an extract, a preparation thereof and a quality control method. Specifically, the invention provides a traditional Chinese medicine composition, and the traditional Chinese medicine composition comprises the following components in parts by weight: 10-20 parts of lycopodium clavatum; 5-15 g of garden balsam stem; 5 to 15g of radix stemonae; 10-20 g of trichosanthes kirilowii maxim; 15-25g by weight of unprocessed radix aconiti; 15-25g of raw radix aconiti kusnezoffii 5 to 15g of caulis spatholobi; 5-15 parts by weight of dried ginger; 10-20 g by weight of Chinese ligusticum rhizome; and / or 5-15g of obscured homalomena rhizome.

Description

Technical Field

[0001] The present invention belongs to the field of traditional Chinese medicine, and particularly relates to a traditional Chinese medicine formula, an extract, its preparation and a quality control method. Background Art

[0002] Arthritis generally refers to inflammatory diseases that occur in human joints and their surrounding tissues, caused by inflammation, infection, degeneration, trauma or other factors, and can be divided into dozens of types. There are more than 100 million arthritis patients in China, and the number is increasing. The clinical manifestations are joint redness, swelling, heat, pain, dysfunction and joint deformity. In severe cases, it can lead to joint disability and affect the quality of life of patients. According to statistics, half of the people over 50 years old in China suffer from osteoarthritis, and 90% of women and 80% of men over 65 years old suffer from osteoarthritis. The prevalence rate in China is 0.34% - 0.36%, and the life span of severe patients is shortened by about 10 - 15 years. The etiology of arthritis is complex, mainly related to factors such as autoimmune reaction, infection, metabolic disorder, trauma, degenerative lesions, etc. According to the etiology, arthritis can be divided into osteoarthritis, rheumatic arthritis, rheumatoid arthritis, ankylosing spondylitis, reactive arthritis, gouty arthritis, etc. Most arthritis has a long course and is difficult to cure, and the treatment is quite difficult. Traditional Chinese medicine is characterized by the overall concept and syndrome differentiation and treatment. In the treatment of rheumatic and rheumatoid arthritis, multi-channel and multi-level combined medication shows its advantages. Commonly used effective traditional Chinese medicines such as: Strychnos nux-vomica, Asarum sieboldii, ants, etc., and commonly used Chinese herbal medicines include: anti-rheumatic drugs, blood-activating and stasis-removing drugs, heat-clearing and detoxifying drugs, liver and kidney-tonifying and bone-strengthening drugs, qi-tonifying and blood-nourishing and spleen-strengthening drugs, etc.

[0003] Rheumatoid arthritis (RA) accounts for 40% of bone and joint diseases in China. It is a disease with a high disability rate, which can cause serious harm in just a few years, and these harms are irreversible, accompanied by symptoms such as pannus, joint space narrowing, synovial tissue hyperplasia, etc. How to detect, diagnose and treat it early in a timely and effective manner, and then prevent the progression of arthritis and improve the prognosis of patients has always been an important issue faced by the medical community.

[0004] Therefore, there is an urgent need in this field to develop a preparation that can be used in a timely and effective manner to treat wind-cold-damp arthralgia, joint pain, rheumatism and rheumatoid arthritis. Summary of the Invention

[0005] The present invention aims to provide a traditional Chinese medicine composition, an extract, its preparation and a quality control method, and the traditional Chinese medicine composition or its extract has excellent therapeutic effects on wind-cold-damp arthralgia, joint pain, rheumatism and rheumatoid arthritis.

[0006] In the first aspect of the present invention, a traditional Chinese medicine composition is provided, and the traditional Chinese medicine composition includes:

[0007] 10 - 20 parts by weight of Lycopodium clavatum;

[0008] 5 - 15 parts by weight of Speranskia tuberculata;

[0009] 5 - 15 parts by weight of Stemona sessilifolia;

[0010] 10 - 20 parts by weight of Trichosanthes kirilowii;

[0011] 15 - 25 parts by weight of Aconitum carmichaelii var. wutaiense;

[0012] 15 - 25 parts by weight of Aconitum kusnezoffii;

[0013] 5 - 15 parts by weight of Spatholobus suberectus;

[0014] 5 - 15 parts by weight of Dry Ginger;

[0015] 10 - 20 parts by weight of Ligusticum sinense; and / or

[0016] 5 - 15 parts by weight of Homalomena occulta.

[0017] In another preferred embodiment, the traditional Chinese medicine composition comprises: 12 - 18 parts by weight of Lycopodium clavatum; 7 - 13 parts by weight of Speranskia tuberculata; 7 - 13 parts by weight of Stemona sessilifolia; 12 - 18 parts by weight of Trichosanthes kirilowii; 17 - 23 parts by weight of Aconitum carmichaelii var. wutaiense; 17 - 23 parts by weight of Aconitum kusnezoffii; 7 - 13 parts by weight of Spatholobus suberectus; 7 - 13 parts by weight of Dry Ginger; 12 - 18 parts by weight of Ligusticum sinense; and 7 - 13 parts by weight of Homalomena occulta.

[0018] In another preferred embodiment, the traditional Chinese medicine composition comprises: 15 parts by weight of Lycopodium clavatum; 10 g of Speranskia tuberculata; 10 g of Stemona sessilifolia; 15 g of Trichosanthes kirilowii; 20 g of Aconitum carmichaelii var. wutaiense; 20 g of Aconitum kusnezoffii; 10 g of Spatholobus suberectus; 10 parts by weight of Dry Ginger; 15 g of Ligusticum sinense; and 10 g of Homalomena occulta.

[0019] In the second aspect of the present invention, there is provided a traditional Chinese medicine extract, which is obtained by extracting the traditional Chinese medicine composition as described in the first aspect.

[0020] In another preferred embodiment, the traditional Chinese medicine extract contains components selected from the following group: 6 - gingerol, vanillic acid, linalool, aconitine diester alkaloids, chlorogenic acid, ferulic acid, ligustilide, or a combination thereof.

[0021] In another preferred embodiment, the traditional Chinese medicine extract is prepared by the following method:

[0022] (1) Extract Dry Ginger, Ligusticum sinense and Homalomena occulta with a first extraction solvent, filter the extract, collect the filtrate to obtain volatile oil, and collect the medicinal residues for later use;

[0023] (2) Add a second solvent to the medicinal residues obtained in step (1), and lycopodium clavatum, speranskia tuberculata, stemona japonica, trichosanthes kirilowii, radix aconiti carmichaeli preparata, radix aconiti kusnezoffii preparata, and caulis spatholobi, and decoct to obtain a decoction filtrate; combine the filtrates (including the filtrate in step (1)), concentrate to a concentrated filtrate with a relative density of 1.05 - 1.12, dilute with water, and let stand;

[0024] (3) Elute the supernatant obtained in step (2) on macroporous resin, first wash with water to remove impurities, and then elute with 40 v / v% - 60 v / v% ethanol and 80 v / v% - 95 v / v% ethanol respectively, collect the eluate, concentrate and dry to obtain an enriched product.

[0025] In the third aspect of the present invention, there is provided a preparation method of the traditional Chinese medicine extract as described in the second aspect, and the method includes the steps:

[0026] (1) Use a first extraction solvent to extract dried ginger, ligusticum sinense, and homalomena occulta, filter the extract, collect the filtrate to obtain volatile oil, and collect the medicinal residues for later use;

[0027] (2) Add a second solvent to the medicinal residues obtained in step (1), and lycopodium clavatum, speranskia tuberculata, stemona japonica, trichosanthes kirilowii, radix aconiti carmichaeli preparata, radix aconiti kusnezoffii preparata, and caulis spatholobi, and decoct to obtain a decoction filtrate; combine the filtrates (including the filtrate in step (1)), concentrate to a concentrated filtrate with a relative density of 1.05 - 1.12, dilute with water, and let stand;

[0028] (3) Elute the supernatant obtained in step (2) on macroporous resin, first wash with water to remove impurities, and then elute with 40 v / v% - 60 v / v% ethanol and 80 v / v% - 95 v / v% ethanol respectively, collect the eluate, concentrate and dry to obtain an enriched product;

[0029] (4) Mix the volatile oil obtained in step (1) and the enriched product obtained in step (3) to obtain the traditional Chinese medicine extract.

[0030] In another preferred example, the extraction in step (1) is carried out under slightly boiling conditions, and preferably the extraction temperature is 80 - 85 °C.

[0031] In another preferred example, the decocting temperature in step (2) is 80 - 90 °C.

[0032] In another preferred example, step (2) further includes concentrating the decoction filtrate to a concentrated filtrate with a relative density of 1.05 - 1.12.

[0033] In another preferred example, step (2) further includes diluting the concentrated filtrate with water and letting it stand.

[0034] In another preferred embodiment, step (3) further includes: concentrating the eluate to an eluate with a relative density of 1.05 to 1.20.

[0035] In another preferred embodiment, the method further includes: drying and pulverizing the obtained enriched solution to obtain a powdery enriched product.

[0036] In another preferred embodiment, the drying method is selected from the group consisting of: reduced pressure drying, vacuum drying, and freeze drying.

[0037] In another preferred embodiment, the reduced pressure drying is carried out under a vacuum degree of -0.07 to -0.1 MPa and a temperature ≤ 80 °C).

[0038] In another preferred embodiment, the drying temperature is ≤ 60 °C.

[0039] In a fourth aspect of the present invention, there is provided a use of the traditional Chinese medicine composition as described in the first aspect and / or the traditional Chinese medicine extract as described in the second aspect for preparing a pharmaceutical composition or preparation, wherein the pharmaceutical composition or preparation is used for: (i) preventing and / or treating arthritis; (ii) preventing and / or treating symptoms caused by arthritis.

[0040] In another preferred embodiment, the arthritis is selected from the group consisting of: wind-cold-damp arthralgia, joint pain, rheumatism, rheumatoid arthritis, or a combination thereof.

[0041] In another preferred embodiment, the arthritis is rheumatoid arthritis.

[0042] In a fifth aspect of the present invention, there is provided a pharmaceutical composition or preparation for preventing and / or treating arthritis, and / or preventing and / or treating symptoms caused by arthritis, comprising the traditional Chinese medicine composition as described in the first aspect or the pharmaceutical extract as described in the second aspect; and a pharmaceutically acceptable carrier.

[0043] In another preferred embodiment, the preparation is selected from the group consisting of: granules, capsules, dripping pills, or tablets.

[0044] In another preferred embodiment, the preparation method of the granules includes:

[0045] (a) Adding the enriched product to the conventional excipients of the granules and wet-mixing and granulating, drying and sizing to obtain the first granules;

[0046] (b) Adding the volatile oil to the first granules in step (1), mixing evenly, and packaging to obtain the granules.

[0047] In another preferred embodiment, in the pharmaceutical composition or preparation, the content of the traditional Chinese medicine composition or the traditional Chinese medicine extract is 0.1-99.9 wt%, preferably 1-99%, more preferably 5-95%, more preferably 10-90%, more preferably 20-80%, and most preferably 30-70%, based on the weight of the pharmaceutical composition.

[0048] In the sixth aspect of the present invention, there is provided a method for preventing and / or treating arthritis, and / or preventing and / or treating symptoms caused by arthritis, by administering to a subject in need a traditional Chinese medicine composition as described in the first aspect and / or a traditional Chinese medicine extract as described in the second aspect.

[0049] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features specifically described below (such as in the examples) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be elaborated one by one here. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 Shows the HPLC chromatograms (320 nm) of Shenjin Huoluo Formula and the control test sample; wherein, 1 chlorogenic acid, 2 ferulic acid, 3 kaempferol, 4 ligustilide, Figure 1 A in it is the HPLC chromatogram of Shenjin Huoluo Formula, Figure 1 B in it is the HPLC chromatogram of the control test sample.

[0051] Figure 2 Shows the HPLC chromatogram (210 nm) of Shenjin Huoluo Formula; wherein, 5 vanillic acid; 6 mesaconitine; 7 hypaconitine; 8 aconitine; 9 gingerol; 10 linalool, Figure 2 A in it is the HPLC chromatogram of Shenjin Huoluo Formula, Figure 2 B in it is the HPLC chromatogram of the control test sample. DETAILED DESCRIPTION OF THE INVENTION

[0052] Through extensive and in-depth research and a large number of experimental screenings, the present inventors unexpectedly discovered for the first time a traditional Chinese medicine composition or its extract and its uses for (i) preventing and / or treating arthritis; (ii) preventing and / or treating symptoms caused by arthritis; the traditional Chinese medicine composition includes one or more medicinal materials selected from the group consisting of Lycopodium clavatum, Speranskia tuberculata, Stemona japonica, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, Spatholobus suberectus, Zingiber officinale, Ligusticum sinense, and Homalomena occulta extract.

[0053] In the extraction process of the present invention, the method of first separately extracting Zingiber officinale, Ligusticum sinense, and Homalomena occulta to obtain volatile oils and then enriching and purifying the water extract can improve the active ingredients in the traditional Chinese medicine extract, especially 6-gingerol and ligustilide in the volatile oils, reduce the extract yield of the traditional Chinese medicine extract, thereby reducing the dosage and improving the compliance of patients. On this basis, the present invention was completed.

[0054] Term Explanation

[0055] Unless otherwise defined, all technical and scientific terms used herein shall have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0056] As used herein, the terms "comprising", "including", and "containing" are used interchangeably and include not only closed definitions but also semi - closed and open definitions. In other words, the said terms include "consisting of" and "consisting essentially of".

[0057] In the present invention, the term "prevention" refers to a method of preventing the onset of a disease and / or its attendant symptoms or protecting a subject from acquiring a disease. "Prevention" as used herein also includes delaying the onset of a disease and / or its attendant symptoms and reducing the risk of the subject getting the disease.

[0058] "Treatment" as described in the present invention includes delaying and terminating the progression of a disease, or eliminating the disease, and does not require 100% inhibition, eradication, and reversal. In some embodiments, compared to the levels observed in the absence of the compositions, kits, food kits, or health product kits, or combinations of active ingredients described in the present invention, the compositions or pharmaceutical compositions described in the present invention reduce, inhibit, and / or reverse arthritis and its complications by, for example, at least about 10%, at least about 30%, at least about 50%, or at least about 80%.

[0059] As used herein, the term "effective amount" refers to an amount that produces a function or activity in a human and / or an animal and is acceptable to the human and / or the animal. Those of ordinary skill in the art should understand that the said "effective amount" may vary depending on the form of the pharmaceutical composition, the severity of the disease, and the co - administration of other drugs, etc.

[0060] As used herein, the terms "parts by weight" and "number of parts by weight" are used interchangeably, and the parts by weight can be any fixed weight expressed in milligrams, grams, or kilograms (such as 1 mg, 1 g, 2 g, or 1 kg, etc.). For example, a composition composed of 1 part by weight of component a and 9 parts by weight of component b can be a composition of 1 g of component a + 9 g of component b, or a composition of 10 g of component a + 90 g of component b, etc. In the said composition, the percentage content of a certain component = (the number of parts by weight of the component / the sum of the number of parts by weight of all components) × 100%. Therefore, in a composition composed of 1 part by weight of component a and 9 parts by weight of component b, the content of component a is 10% and the content of component b is 90%.

[0061] As used herein, the term "traditional Chinese medicine composition" refers to Chinese medicinal materials used as active medicinal materials in the present invention. The said medicinal materials can include various forms. For example, the said medicinal materials include (but are not limited to) dried medicinal materials, fresh medicinal materials, raw medicinal materials, processed medicinal materials, or combinations thereof. In addition, when the pharmaceutical composition or preparation of the present invention contains a certain traditional Chinese medicine extract, this term also includes the raw medicinal materials corresponding to the traditional Chinese medicine extract or the corresponding traditional Chinese medicine composition used for preparing the said extract.

[0062] As used herein, the terms "C1-C6 lower alcohols", "C1-C4 lower alcohols", or "C1-C3 lower alcohols" refer to alcohols having 1-6, 1-4, or 1-3 carbon atoms, such as methanol, ethanol, n-propanol, isopropanol, or n-butanol, etc.

[0063] arthritis

[0064] Arthritis generally refers to inflammatory diseases that occur in the human joints and their surrounding tissues and are caused by inflammation, infection, degeneration, trauma, or other factors, and can be divided into dozens of types. The causes of arthritis are complex and are mainly related to factors such as autoimmune reactions, infections, metabolic disorders, trauma, and degenerative lesions. According to the causes, arthritis can be divided into osteoarthritis, rheumatic arthritis, rheumatoid arthritis, ankylosing spondylitis, reactive arthritis, gouty arthritis, etc. Most types of arthritis have a long course and are difficult to cure, and the treatment is quite difficult. Traditional Chinese medicine is characterized by the overall concept and syndrome differentiation and treatment. In the treatment of rheumatic and rheumatoid arthritis, multi-channel and multi-level combined medication shows its advantages.

[0065] Traditional Chinese medicine composition and traditional Chinese medicine extract

[0066] The Shenjin Huoluo Formula (traditional Chinese medicine composition) of the present invention is a clinical experience formula of Dr. Liang Quanlong, which inherits modern medication experience and is gradually optimized and innovated in the formula. It is composed of ten medicinal materials such as Lycopodium clavatum, Speranskia tuberculata, and Ligusticum sinense, etc., and has the effects of dispelling wind and dampness, relaxing tendons and activating collaterals, and warming meridians to relieve pain. It is used for the treatment of wind-cold-damp arthralgia, joint pain, rheumatic and rheumatoid arthritis, etc., and is named Shenjin Huoluo Formula.

[0067] The main chemical components in each single medicinal material of the Shenjin Huoluo Formula are: Lycopodium clavatum (flavonoids, alkaloids, volatile oils), Speranskia tuberculata (calcium silicate, flavonoids, organic acids, etc.), Stemona japonica (alkaloids, organic acids), Trichosanthes kirilowii (flavonoids, triterpenoid saponins, organic acids, etc.), Homalomena occulta (volatile oils), Aconitum carmichaelii, Aconitum kusnezoffii (alkaloids), Spatholobus suberectus (organic acids, flavonoids), Zingiber officinale (volatile oils), Ligusticum sinense (organic acids, volatile oils), and they have the effects of antibacterial, analgesic, anti-inflammatory, etc.

[0068] The Shenjin Huoluo Formula prepared in the present invention has analgesic and anti-inflammatory effects and has a unique effect on the treatment of rheumatic and rheumatoid arthritis, etc.

[0069] The traditional Chinese medicine composition (Shenjin Huoluo Formula) in the present invention comprises Lycopodium clavatum, Speranskia tuberculata, Stemona japonica, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, Spatholobus suberectus, Drynaria fortunei, Ligusticum sinense, and / or Rhizoma seu Herba Homalomenae.

[0070] In a preferred embodiment of the present invention, the said traditional Chinese medicine composition comprises: 12 - 18 parts by weight of Lycopodium clavatum; 7 - 13 g of Speranskia tuberculata; 7 - 13 g of Stemona japonica; 12 - 18 g of Trichosanthes kirilowii; 17 - 23 g of Aconitum carmichaelii; 17 - 23 g of Aconitum kusnezoffii; 7 - 13 g of Spatholobus suberectus; 7 - 13 parts by weight of Drynaria fortunei; 12 - 18 g of Ligusticum sinense; and / or 7 - 13 g of Rhizoma seu Herba Homalomenae.

[0071] The traditional Chinese medicine extract in the present invention comprises extracts of Lycopodium clavatum, Speranskia tuberculata, Stemona japonica, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, Spatholobus suberectus, Drynaria fortunei, Ligusticum sinense, and / or Rhizoma seu Herba Homalomenae.

[0072] Preparation method of traditional Chinese medicine extract

[0073] Those skilled in the art can extract the said traditional Chinese medicine composition by conventional methods. The extraction includes but is not limited to: water extraction, alcohol extraction, water - alcohol solution extraction, CO2 supercritical extraction, aqueous solvent extraction, or a combination thereof.

[0074] The said traditional Chinese medicine raw materials can be mixed and then the effective components can be extracted by an appropriate method to obtain the extract; in addition, each raw material can also be separately extracted (such as by using the same or different extraction or processing methods respectively) to obtain the effective components (i.e., the extract of the raw material), and then combined to obtain the extract. It should be understood that the effective components obtained by extraction can exist in various forms, for example, the raw materials are extracted and purified to obtain single or mixed pure effective component powders, or the raw materials are extracted and concentrated by a solvent to obtain an extract (the effective components are present in the extract), or in other forms.

[0075] In a preferred embodiment of the present invention, the preparation method of the said traditional Chinese medicine extract comprises the steps:

[0076] (1) Use a first extraction solvent to extract Drynaria fortunei, Ligusticum sinense, and Rhizoma seu Herba Homalomenae, filter the extraction solution, collect the filtrate to obtain the volatile oil, and collect the medicinal residues for later use;

[0077] (2) Add a second solvent to the medicinal residues obtained in step (1), Lycopodium clavatum, Speranskia tuberculata, Stemona japonica, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, and Spatholobus suberectus for decoction to obtain a decoction filtrate; combine the filtrates (including the filtrate in step (1)), concentrate to a concentrated filtrate with a relative density of 1.05 - 1.12, dilute with water, and let stand;

[0078] (3) Elute the supernatant obtained in step (2) through macroporous resin. First, wash away impurities with water, and then elute with ethanol at 40 v / v% - 60 v / v% and 80 v / v% - 95 v / v% respectively. Collect the eluate, concentrate and dry it to obtain the enriched product.

[0079] In another preferred embodiment, the extraction in step (1) is carried out under slightly boiling conditions, and preferably the extraction temperature is 80 - 85 °C.

[0080] In another preferred embodiment, the decocting temperature in step (2) is 80 - 90 °C.

[0081] In another preferred embodiment, step (2) further includes concentrating the decocted filtrate to a concentrated filtrate with a relative density of 1.05 - 1.12.

[0082] In another preferred embodiment, step (2) further includes diluting the concentrated filtrate with water and allowing it to stand.

[0083] In another preferred embodiment, step (3) further includes concentrating the eluate to an eluate with a relative density of 1.05 - 1.20.

[0084] In another preferred embodiment, the method further includes: drying and pulverizing the obtained enriched product solution to obtain the enriched product.

[0085] In another preferred embodiment, the drying method is selected from the group consisting of: vacuum drying, freeze drying.

[0086] In another preferred embodiment, the vacuum drying has a vacuum degree of -0.07 to -0.1 MPa and a temperature ≤ 80 °C).

[0087] In another preferred embodiment, the drying temperature is ≤ 60 °C.

[0088] In the present invention, it should be understood that the extract not only includes the concentrated solution (such as extract) or active ingredient (such as powder) obtained by extraction after mixing the raw medicinal materials, but also includes the concentrated solution (such as extract) or the mixture of active ingredients (including the mixture between concentrated solutions, between active ingredients, or between concentrated solution and active ingredient) obtained by extracting individual raw medicinal materials. The active ingredients of the raw medicinal materials include single components or mixed components.

[0089] Preparation method of the preparation

[0090] 1. Powder

[0091] Take the enriched product prepared in "2.2.5" of Example 2, add the extracted volatile oil thereto, mix well, and package to obtain the powder.

[0092] 2. Granules

[0093] The prepared enriched product is respectively added with conventional excipients for granules and wet-mixed and granulated, dried, sized, then the extracted volatile oil is added, mixed evenly, and packaged to obtain the granules.

[0094] 3. Other dosage forms

[0095] The prepared volatile oil and enriched product are taken for pharmaceutical preparation research, and capsules, soft capsules, dripping pills, etc. can be prepared.

[0096] Use

[0097] The present invention provides a use of a traditional Chinese medicine composition and / or a traditional Chinese medicine extract for preparing a pharmaceutical composition or preparation, and the pharmaceutical composition or preparation is used for: (i) preventing and / or treating arthritis; (ii) preventing and / or treating symptoms caused by arthritis.

[0098] In another preferred example, the arthritis is selected from the group consisting of wind-cold-damp arthralgia, joint pain, rheumatism, rheumatoid arthritis, or a combination thereof.

[0099] In another preferred example, the arthritis is rheumatoid arthritis.

[0100] Pharmaceutical composition

[0101] In a preferred example of the present invention, the raw medicinal materials of Lycopodium clavatum, Speranskia tuberculata, Stemona sessilifolia, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, Spatholobus suberectus, Dry Ginger, Ligusticum sinense, and Homalomena occulta can be directly added to the composition or added to the pharmaceutical composition after being pulverized. In addition, those skilled in the art can also directly use the traditional Chinese medicine extract of the present invention for processing to prepare a pharmaceutical composition. Moreover, those skilled in the art can extract the active ingredients from each raw medicinal material and mix them into the pharmaceutical composition. In a preferred embodiment of the present invention, the pharmaceutical composition of the present invention is prepared by mixing the extract of raw medicinal materials with a carrier, and the extract of medicinal materials can be formed by mixing the extracts of each raw medicinal material.

[0102] There is no particular limitation on the dosage form of the pharmaceutical composition of the present invention, and it can be any dosage form suitable for mammals. Preferably, the dosage form may include tablets, capsules, granules, pills, powders, oral liquids, buccal tablets, or aerosols; most preferably, they are tablets, capsules or granules. From the perspective of easy preparation, administration or taking, the preferred composition is a solid composition. Oral administration is preferred.

[0103] Various conventional carriers and / or excipients required for preparing different dosage forms can be added to the composition of the present invention, such as fillers (such as starch, cellulose, dextrin), disintegrants (sodium carboxymethyl starch), lubricants (magnesium stearate), solvents (purified water), solubilizers (ethanol), coating materials. It can be prepared into any common dosage form by conventional traditional Chinese medicine preparation methods, such as tablets, capsules, granules, capsules, pills, powders.

[0104] As used herein, the components of the term "pharmaceutically acceptable carrier" refer to substances that are suitable for humans and / or animals without excessive adverse side effects (such as toxicity, irritation, and allergic reactions), that is, substances with a reasonable benefit / risk ratio. Some examples of pharmaceutically acceptable carriers are cellulose and its derivatives (such as methylcellulose, ethylcellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as Tween), wetting agents (such as sodium lauryl sulfate), buffers, chelating agents, thickening agents, pH regulators, transdermal promoters, coloring agents, flavoring agents, stabilizers, antioxidants, preservatives, bacteriostatic agents, pyrogen-free water, etc.

[0105] Administration routes and treatment methods

[0106] The traditional Chinese medicine composition and its extract of the present invention can be used to prepare a pharmaceutical composition or preparation for preventing and / or treating arthritis and / or preventing and / or treating the symptoms caused by arthritis. The pharmaceutical composition or preparation of the present invention may also contain other optional medicinal materials or their extracts or contain other drugs effective for treating and / or preventing the symptoms caused by arthritis.

[0107] The pharmaceutical composition or preparation is matched with the administration route. When using the pharmaceutical composition or preparation, a safe and effective amount of the drug is administered to the desired subject (such as a human or non-human mammal). Usually, the pharmaceutical composition or preparation of the present invention is administered at a dose of 0.001 - 20 g / kg of the animal body weight per day, preferably in 1 - 4 separate doses per day, or in a sustained-release form. For most mammals, the total daily dose is 0.05 - 600 g. Of course, the specific dose also varies with the administration mode, dosage form, and severity of the disease to be treated, as well as the administration route, which are all within the skills of a skilled physician. Administration can be carried out through conventional routes, including (but not limited to): oral, intramuscular, cutaneous, or topical administration. Oral administration is preferred.

[0108] The present invention also provides a method for preventing and / or treating pneumonia and / or preventing and / or treating the symptoms caused by pneumonia, by administering the traditional Chinese medicine composition and / or the traditional Chinese medicine extract of the present invention to the desired subject.

[0109] In another preferred example, the subject is a human or non-human mammal (such as a dog, pig, cat, monkey, sheep, horse, cow, etc.).

[0110] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0111] (1) The traditional Chinese medicine composition and its extract of the present invention can effectively prevent / treat arthritis.

[0112] (2) In the extraction process of the present invention, the method of first separately extracting volatile oil and then enriching and purifying the water extract can improve the active ingredients in the traditional Chinese medicine extract, especially 6-gingerol and ligustilide in the volatile oil, reduce the paste yield of the traditional Chinese medicine extract, thereby reducing the dosage and improving the compliance of patients.

[0113] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. The experimental methods without specific conditions noted in the following embodiments are usually carried out under conventional conditions or according to the conditions recommended by the manufacturer. Unless otherwise stated, percentages and parts are by weight.

[0114] Example 1 Prescription and Dosage

[0115] In this embodiment, a prescription and dosage of a traditional Chinese medicine composition are provided.

[0116] 1.1 Prescription and dosage of the traditional Chinese medicine composition:

[0117] Herba Lycopodii 15g, Herba Speranskiae tuberculatae 10g, Radix Stemonae 10g, Fructus Trichosanthis 15g, Rhizoma Homalomenae 10g

[0118] Radix Aconiti 20g, Radix Aconiti Kusnezoffii 20g, Caulis Spatholobi 10g, Rhizoma Zingiberis 10g, Rhizoma Ligustici 15g

[0119] The above prescription is the Shishen Jinhuo Recipe.

[0120] 1.2 Usage of the traditional Chinese medicine composition

[0121] Soak feet, one prescription per day, and one course of treatment for seven days.

[0122] Example 2 Method for Preparing the Drug

[0123] 2.1 Medicinal powder

[0124] All the prescription drugs in Example 1 were powdered and passed through an 80-mesh sieve.

[0125] 2.2 Preparation method of the extract of the traditional Chinese medicine composition

[0126] In this embodiment, the traditional Chinese medicine composition is extracted using the processes of water extraction, ethanol extraction, water extraction followed by ethanol precipitation, and ethanol extraction followed by water extraction, and the process steps are optimized.

[0127] 2.2.1 Extract powder: water extraction

[0128] Provide the prescription drug in Example 1, add 10 times the amount of water and decoct twice, each time for 1.5 hours, filter, concentrate the filtrate to an extract with a relative density of 1.05 - 1.20 (50 - 60 °C), dry under reduced pressure (-0.07 - -0.1 MPa, ≤60 °C), pulverize, and pass through a 60-mesh sieve to obtain the extract powder.

[0129] 2.2.2 Extract powder: water extraction followed by ethanol precipitation

[0130] Provide the prescription drug in Example 1, add 10 times the amount of water and decoct twice, each time for 1.5 hours, filter, concentrate the filtrate to a concentrated filtrate with a relative density of 1.08 - 1.12 (50 - 60 °C). Add ethanol to the concentrated filtrate to make the ethanol content reach 50%, refrigerate overnight, filter, concentrate the filtrate to an extract with a relative density of 1.05 - 1.20 (50 - 60 °C), dry under reduced pressure (-0.07 - -0.1 MPa, ≤60 °C), pulverize, and pass through a 60-mesh sieve to obtain the extract powder.

[0131] 2.2.3 Extract powder: ethanol extraction

[0132] Provide the prescription drug in Example 1, add 10 times the amount of 50% ethanol and decoct twice, each time for 1.5 hours, filter, concentrate the filtrate to an extract with a relative density of 1.05 - 1.20 (50 - 60 °C), dry under reduced pressure (-0.07 - -0.1 MPa, ≤60 °C), pulverize, and pass through a 60-mesh sieve to obtain the extract powder.

[0133] 2.2.4 Extract powder: ethanol extraction followed by water precipitation

[0134] Provide the prescription drug in Example 1, add 10 times the amount of 50% ethanol and decoct twice, each time for 1.5 hours, filter, concentrate the filtrate to a concentrated filtrate with a relative density of 1.05 - 1.10 (50 - 60 °C), refrigerate overnight, concentrate the supernatant to an extract with a relative density of 1.05 - 1.20 (50 - 60 °C), dry under reduced pressure (-0.07 - -0.1 MPa, ≤60 °C), pulverize, and pass through a 60-mesh sieve to obtain the extract powder.

[0135] 2.2.5 Volatile oil + enriched substance

[0136] (1) Provide the prescription drug in Example 1. First, add 6 times the amount of water to dried ginger, ligusticum sinense, and homalomena occulta and extract with gentle boiling for 4 hours to obtain the volatile oil; filter the extract and collect the filtrate;

[0137] (2) The medicinal residues after filtration in step (1) and the remaining herbs in the prescription are decocted with 10 times the amount of water twice, each time for 2 hours. After filtration, the filtrates are combined (including the filtrate in step (1)). The filtrate is concentrated to a concentrated filtrate with a relative density of 1.05 - 1.12, diluted with water, and allowed to stand.

[0138] (3) Take the supernatant after standing in step (2) and pass it through a macroporous resin column. Wash it with water and discard the eluate. Then, elute it successively with 50% ethanol solution and 90% ethanol solution. Collect the eluate, recover ethanol under reduced pressure, concentrate the eluate to a relative density of 1.05 - 1.20 (50 - 60 °C), concentrate, dry, pulverize to obtain the enriched product.

[0139] 2.3 Confirmation of drug preparation

[0140] The medicinal powder (extract powder) is directly used in medicine, with less loss of active ingredients, but the content of active ingredients is relatively low and the dosage is relatively large. It can be made into powder for external use. The disadvantage is that the dissolution of active ingredients is relatively slow, which is not conducive to the utilization of active ingredients. Since the herbs such as dried ginger, ligusticum sinense, and homalomena occulta in this prescription contain volatile oils, there is loss of volatile oils during the drying process, and the extract yield is relatively large and the dosage is also relatively large. Therefore, the preparation process of separately extracting volatile oils and enriching and purifying the water extract is selected.

[0141] Preparation method of the preparation in Example 3

[0142] 3.1 Powder

[0143] Take the enriched product prepared in "2.2.5" in Example 2, add the extracted volatile oil to it, mix well, and package to obtain the powder.

[0144] 3.2 Granules

[0145] Take the enriched product prepared in "2.2.5" in Example 2, add the conventional excipients for granules respectively, mix and granulate by wet method, dry, screen, then add the extracted volatile oil, mix well, and package to obtain the granules.

[0146] 3.3 Other dosage forms

[0147] Take the volatile oil and enriched product prepared in "2.2.5" in Example 2, conduct preparation research, and it can be made into capsules, soft capsules, dropping pills, etc.

[0148] Quality control method in Example 4

[0149] In this example, the content determination methods of diester diterpenoid alkaloids (mesaconitine, hypaconitine, aconitine), chlorogenic acid, ferulic acid, quercetin, linalool, vanillic acid, ligustilide, and gingerol in Shenjin Huoluo Prescription were explored. Chlorogenic acid and luteolin were not detected due to their low contents; alkaloids belong to end absorption, with difficult separation of response values and need further research, so they are not controlled temporarily.

[0150] In the experiment, ferulic acid, a representative water-soluble component with a legal reference substance, and 6-gingerol, a volatile oil component, were selected as the investigation indicators. The HPLC-PDA method was used to establish a method for determining the content of effective index components in Shenjin Huoluo Formula, which was used to establish the quality control system of Shenjin Huoluo Formula.

[0151] 4.1 Instruments and reagents

[0152] 4.1.1 Instruments

[0153] The instruments used in this experiment are shown in Table 1

[0154] Table 1: Instruments

[0155]

[0156]

[0157] 4.1.2 Reagents

[0158] The reagents used in this experiment are shown in Table 2

[0159] Table 2: Reagents

[0160]

[0161] 4.2 Methods

[0162] 4.2.1 Preparation of test solution

[0163] Take an appropriate amount of this product (0.2 g for granules) (equivalent to 2.5 g of crude drug), weigh accurately, place it in a stoppered conical flask, add 50 ml of 50% methanol, weigh, ultrasonically treat for 45 minutes, let it stand at room temperature, make up the lost weight with 50% methanol, shake well, centrifuge, and take the supernatant to pass through a 0.45 μm filter membrane to obtain the test solution.

[0164] 4.2.2 Preparation of mixed reference solution

[0165] Take appropriate amounts of ferulic acid and 6-gingerol reference substances, dissolve them with 50% methanol and dilute to the mark to obtain a solution containing 15 μg of ferulic acid and 50 μg of 6-gingerol per 1 ml, that is, the mixed reference solution.

[0166] 4.2.3 Chromatographic conditions and system suitability test

[0167] Chromatographic column: Thermo Hypersil GOLD (4.6 mm × 250 mm, 5 μm) column; Mobile phase: Acetonitrile - 0.1% phosphoric acid gradient elution, see Table 3; Flow rate: 0.9 ml / min; Column temperature: 30 °C; Detection wavelength: 320 nm, 210 nm; The number of theoretical plates calculated by ferulic acid peak should be not less than 10,000; The chromatographic conditions are shown in Table 3.

[0168] Table 3: Elution conditions

[0169]

[0170] 4.2.4 Determination method

[0171] Respectively pipette 10 μl of the test solution and the mixed reference solution that have passed through a 0.45 μm microporous filter membrane, inject them into the liquid chromatograph for determination, and that's it. The determination results are as shown in A in Figure 1 and Figure 1 B in Figure 2 A of Figure 2 and B in

[0172] Comparative study on the index components of the water extraction process and the refining process in Example 5

[0173] 5.1 Preparation of powder samples

[0174] In this example, the medicinal materials in the prescription are subjected to a comminution process study, the best medicinal powder is selected, mixed evenly, and packaged to obtain the powder.

[0175] 5.2 Preparation of water extraction process samples

[0176] Take the ten medicinal materials of Shishen Jinhuo Formula, add 10 times the amount of water and decoct for 2 times, each time for 1.5 hours, filter, concentrate the filtrate to an extract with a relative density of 1.10 - 1.30 (50 - 60 °C), dry under reduced pressure (-0.07 - -0.1 MPa, ≤ 60 °C), pulverize, and pass through a 60-mesh sieve to obtain the dry extract powder.

[0177] 5.3 Preparation of refined process samples

[0178] Prepare the extract of the traditional Chinese medicine composition according to the prescription of Example 1 and the preparation method of Example 2.

[0179] 5.4 Detection method

[0180] Take the samples obtained in 5.1 and 5.2 and determine them according to the method of Example 4.

[0181] 5.5 Detection results

[0182] 5.5.1 Research on powder

[0183] 5.5.1.1 Selection of comminution degree

[0184] Simulate the clinical foot bath application (20 times the amount of water, 45 °C), and investigate the influence of the degree of comminution (chinese herbal pieces, coarse powder, medium powder, fine powder, ultrafine powder) on the dissolution of the index components during the use process. According to the experimental results, considering comprehensively, fine powder (passing through No. 5 sieve) is selected, as shown in Table 4.

[0185] Table 4: Results of the influence of the degree of comminution on the peak area of the index components

[0186]

[0187] 5.5.1.2 Selection of foot bath temperature

[0188] Simulate the clinical foot bath application, and select fine powder to optimize the foot bath temperature (commonly 40 - 45 °C). According to the experimental results: Considering comprehensively, 42 - 45 °C is selected, as shown in Table 5.

[0189] Table 5: Influence of foot bath temperature on the peak area of the index components

[0190]

[0191] 5.5.1.3 Study on the stability of the foot bath process

[0192] Simulate the clinical foot bath application, and conduct a stability study on the index components in the foot bath process (adding boiling water and stirring, cooling to 45 °C). According to the experimental results: The stability is good, as shown in Table 6 and Table 7.

[0193] Table 6: Influence of foot bath temperature (room temperature) on the stability of the peak area of the index components

[0194]

[0195] Table 7: Influence of foot bath temperature (heat preservation) on the stability of the peak area of the index components

[0196]

[0197] 5.5.1.4 Determination of the content of index components

[0198] Crush the prescription medicinal materials into fine powder, mix evenly, package, and detect the content of the index components.

[0199] According to the experimental results: Select 6-gingerol with a relatively high content as the representative volatile component, and select ferulic acid with a relatively high content as the representative water-soluble component, as shown in Table 8.

[0200] Table 8: Determination of the content of index components in the samples

[0201]

[0202] 5.5.2 Study on the extraction and purification process

[0203] Investigate the effects of water extraction, alcohol extraction, water extraction followed by alcohol precipitation, and alcohol extraction followed by water extraction processes on the index components. Since the extract powder and preparations involve concentration and drying, the volatile components are greatly damaged. Moreover, volatile oils have antibacterial, analgesic, anti-inflammatory and other effects. For example, ginger extract can effectively improve the mechanical pain of patients with joint inflammation, and the effect is comparable to that of ibuprofen.

[0204] According to the experimental results: the transfer rates of the volatile components 6-gingerol, linalool and ligustilide in the extract powder are all lower than 30%. Therefore, this formula is not suitable for high-temperature extraction and drying. See Tables 9 and 10.

[0205] Table 9: Contents of Index Components

[0206]

[0207] Table 10: Transfer Rates of Index Components

[0208]

[0209] 5.5.3 Study on Volatile Oil + Enrichment

[0210] Since volatile oils are easily lost and damaged, the volatile oils in this study were collected separately. At the same time, due to the large prescription and high extract yield of the traditional process, an enrichment process was adopted to reduce the extract yield and increase the content of index components.

[0211] According to the experimental results: the contents of 6-gingerol and ligustilide in the volatile oil are relatively high, and the contents of other index components in the enrichment are also greatly increased. See Tables 11 and 12.

[0212] Table 11: Contents of Index Components

[0213]

[0214] Table 12: Transfer Rates of Index Components

[0215]

[0216] Note: "*" is the oil yield of the three medicinal materials of dried ginger, ligusticum rhizome and homalomena occulta.

[0217] 5.6 Result Analysis

[0218] According to the above results, since the formula contains volatile components, the volatile components are severely lost during the extraction, concentration and purification processes. Moreover, the alkaloids contained in aconite are highly toxic and there are safety hazards for oral administration. Therefore, a safe, effective and innovative external dosage form is temporarily selected for this process.

[0219] Meanwhile, the centralized collection of volatile oil and resin enrichment and purification were investigated. On the basis of ensuring the easy preservation of index components, the dissolution of active components during use was enhanced, so as to interpret the classics with modern technology.

[0220] In summary, the traditional innovative process of Shenjin Huoluo Prescription was determined: on the basis of traditional foot bath, on the premise of ensuring volatile oil, the enrichment process was adopted to increase the content of active components, reduce the dosage of medicine, and improve the compliance of patients.

[0221] Example 6 Pharmacodynamic effects of Shenjin Huoluo Prescription on rats with rheumatoid arthritis

[0222] In this example, the used Shenjin Huoluo Prescription is the clinical experience prescription of Dr. Liang Quanlong, which is used to treat wind-cold-dampness arthralgia, joint pain, rheumatism, rheumatoid arthritis, etc. There are many kinds of animal models currently used for the study of the pathogenesis of inflammation. The adjuvant arthritis model of rats induced by complete Freund's adjuvant (CFA) is often used in the study of RA. In the experiment, the effects of Shujin Huoluo Prescription on female SD rats were explored, the appropriate dose of CFA for establishing the RA model was determined, and its therapeutic effect on RA was verified.

[0223] 6.1 Materials

[0224] 6.1.1 Experimental animals

[0225] 30 female SD rats were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. (Production License: SCXK(Jing)2016-0006, Use License No.: SYXK(Su)2018-0034); after purchase, they were conventionally raised in the animal house with sufficient light and good ventilation in this laboratory, drank tap water, and were fed with standard rat feed. The application for the management and use of animals in this study has been approved by the Animal Management and Use Committee of this institution.

[0226] 6.1.2 Drugs and main reagents

[0227] Shenjin Huoluo Prescription (self-made, and the Chinese herbal pieces were all purchased from Shandong Shangyao Chinese Herbal Pieces Co., Ltd.); Shangshi Zhitong Plaster (Batch No. 220303, Henan Lingrui Pharmaceutical Co., Ltd.)

[0228] Freund's complete adjuvant (product number: F5881, Sigma-Aldrich, USA); chloral hydrate (batch number 20210307, Sinopharm Chemical Reagent Co., Ltd.); hematoxylin staining solution (batch number CR2102133, Wuhan Sevier Biotechnology Co., Ltd.); eosin staining solution (batch number CR2101094, Wuhan Sevier Biotechnology Co., Ltd.); 4% paraformaldehyde fixative (product number BL539A, Lan Jie Ke Technology Co., Ltd.); tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), vascular endothelial growth factor A (VEFG-A) kits, Wuhan Elitre Biotechnology Co., Ltd.

[0229] 6.1.3 Main instruments

[0230] Electric thermostatic incubator (ICV-450 type, ASONE, Japan); digital vernier caliper (PD-151 type, Shanghai Baogong Tools Co., Ltd.); multifunctional microplate reader (FlexStation 3 type, Molecular Devices); electric blast drying oven (101-1A type, Cangzhou Jinghong Engineering Instruments Co., Ltd.); vacuum drying oven (DZ-1BC type, Tianjin Test Instrument Co., Ltd.); refrigerator (BCD-455WLDPC type, Qingdao Haier Co., Ltd.); TD5B automatic de-capping centrifuge (TD5B type, Yancheng Kate Experimental Instrument Co., Ltd.), etc.

[0231] 6.2 Methods

[0232] 6.2.1 Preparation of Shenjin Huoluo formula

[0233] In this example, the Shenjin Huoluo formula was prepared using the optimal process optimized in the above example.

[0234] Shenjin Huoluo formula: According to the prescription and dosage ratio in Example 1, and following the processes in 2.2.5 of Example 2 and 3.2 of Example 3, the granules were prepared and packaged to obtain the Shenjin Huoluo formula.

[0235] 6.2.2 Establishment of rat RA model

[0236] Each rat was injected with 0.1 mL of Freund's complete adjuvant intradermally into the right hind paw toe for inflammation induction. Rats in the control group were injected with the same volume of 0.9% sodium chloride solution at the same site, and were boosted with an injection on the 7th day.

[0237] 6.2.3 Animal grouping and intervention

[0238] The modeled rats were randomly divided into a model group, a blank preparation group, a Shangshi Zhitong Plaster (positive drug) group, and a Shenjin Huoluo Formula group. Another group of normal rats was used as the control group, with 6 rats in each group. Administration started after the second immunization. Before administration, hair removal was performed on the right hind leg joints of the rats. The rats in the Shenjin Huoluo Formula group were soaked in the Shenjin Huoluo Formula (containing 0.55% 6-gingerol and 0.16% ferulic acid, and the anti-RA effect of the Shenjin Huoluo Formula was obvious under this drug loading amount as shown by the pre-pharmacodynamic experiment) for half an hour under the right hind thigh joint. The positive drug group had the Shangshi Zhitong Plaster attached to the right hind thigh joint of the rats. The blank preparation group was soaked in plain water without drugs. Administration was given once a day for 21 consecutive days. The model group and the control group were raised normally without any treatment.

[0239] 6.2.4 General condition observation and body weight measurement

[0240] Macroscopically observe the changes in local inflammatory reactions, systemic symptoms, behavioral states, and diet of the rats in each group before and on the 5th, 10th, 15th, and 20th days after modeling, and record the changes in body weight.

[0241] 6.2.5 Determination of toe swelling degree of rats in each group

[0242] On the 5th, 10th, 15th, and 20th days after modeling, measure the thickness of the right hind toes of the rats in each group, calculate the toe swelling degree of the rats, and keep the measurement site consistent each time. Toe swelling degree (mm) = thickness of the right toe after inflammation induction (mm) - thickness of the right toe before inflammation induction (mm)

[0243] 6.2.6 Specimen collection and index detection

[0244] On the 21st day after modeling, the rats were sacrificed by taking blood from the abdominal aorta. The right knee joints of the rats in each group were separated, fixed with 4% paraformaldehyde solution, decalcified with ethylenediaminetetraacetic acid (EDTA), dehydrated with ethanol, embedded in paraffin, sectioned, stained with hematoxylin-eosin (HE), photographed and analyzed under an optical microscope. The blood samples were allowed to stand for 1 h, centrifuged at 3000 r·min-1 for 15 min to obtain the supernatant, aliquoted and stored at -80 °C until detection. The expression levels of IL-6, TNF-α, VEGF-A, and IL-17 in the serum of the rats were detected by enzyme-linked immunosorbent assay (ELISA) according to the instructions of the kit.

[0245] 6.2.7 Statistical methods

[0246] SPSS 21.0 statistical software was used for data analysis. Measurement data were expressed as and the t-test was used for comparison between two groups; count data were expressed as rates, and the χ 2 test was used for comparison between groups.

[0247] 6.3 Results

[0248] 6.3.1 General Observation and Body Mass Detection Results

[0249] Before modeling, the rats in each group were in good health, with smooth and shiny hair. Compared with the control group, the rats in the model group showed reduced activity and diet, slightly yellowish hair, swollen toes, and acute inflammatory manifestations in the joints, with the surface skin being red, shiny, and congested, and limited activity, but no deaths. The conditions of the rats in the Shenjin Huoluo Formula group and the Shangshi Zhitong Plaster group improved.

[0250] The changes in the body mass of the rats in each group showed that there was no significant difference in the body mass changes among the groups. However, the rats in the control group had normal food and water intake and the fastest body mass growth. After modeling, the body mass growth of the rats in the other groups was slow. After treatment, the body mass of the rats in the Shenjin Huoluo Formula group and the Shangshi Zhitong Plaster group increased faster compared with the model group. The body mass growth of the rats in the model group and the blank preparation group was the slowest, as shown in Table 13.

[0251] Table 13: Changes in the Body Mass of Rats in Each Group( n = 6)

[0252]

[0253] 6.3.2 Results of Determination of Toe Swelling Degree in Rats

[0254] The results of the determination of toe swelling degree in the rats of each group showed that the toe thickness of the rats in the control group hardly changed during the whole experiment. The toes of the rats in the other groups swelled after the first inflammation induction. After the second booster immunization on the 7th day, the rats in each group could not bear weight on the modeled side and even ulcerated. On the 5th, 10th, 15th, and 20th days after modeling, compared with the control group, the toe swelling degree of the rats in the model group increased significantly, and the difference was statistically significant (P < 0.05). Compared with the model group, the toe swelling degree of the rats in the Shenjin Huoluo Formula group and the Shangshi Zhitong Plaster group decreased significantly, and the difference was statistically significant (P < 0.05), as shown in Table 14.

[0255] Table 14: Effect of Shenjin Huoluo Formula on Toe Swelling Degree in RA Rats n = 6)

[0256]

[0257]

[0258] Compared with the control group: *P < 0.05; compared with the model group: #P < 0.05

[0259] 6.3.3 Results of Pathological Sections of Knee Joints in Rats of Each Group

[0260] The pathological sections of the knee joints showed that the joint structures of the rats in the control group were normal, and no pannus formation or inflammatory cell infiltration was observed. In the model group and the blank preparation group, synoviocyte hyperplasia, inflammatory cell infiltration, and pannus formation were observed in the knee joints of the rats, and the cartilage was eroded. Compared with the model group, the conditions of cartilage erosion, inflammatory cell infiltration, and pannus formation in the ankle joints of the rats in the Shenjin Huoluo Prescription group and the Shangshi Zhitong Plaster group were improved to varying degrees.

[0261] 6.3.4 Determination results of IL-6, TNF-α, VEGF-A, and IL-17 factors in the peripheral blood of rats in each group

[0262] Compared with the control group, the levels of IL-6, TNF-α, VEGF-A, and IL-17 in the serum of the rats in the model group were significantly increased, and the difference was statistically significant (P < 0.05). Compared with the model group, the levels of IL-6, TNF-α, VEGF-A, and IL-17 in the serum of the rats in the Shangshi Zhitong Plaster group and the Shenjin Huoluo Prescription group were significantly decreased, and the difference was statistically significant (P < 0.05), as shown in Table 15.

[0263] Table 15: Levels of IL-6, TNF-α, VEGF-A, and IL-17 in the serum of rats in each group ( n = 6)

[0264]

[0265] Compared with the control group: *P < 0.05; compared with the model group: #P < 0.05

[0266] 6.3.5 Analysis and evaluation

[0267] A pre-experiment on the pharmacodynamic effects was conducted to explore the best anti-RA effect of the Shenjin Huoluo Prescription.

[0268] In this experiment, after successful modeling, different intervention measures were given to different groups. During the administration period, the daily status, body weight changes, and toe swelling changes of the rats in each group were observed. It was found that as the number of administration days increased, the rats in the Shenjin Huoluo Prescription group (granules) were more active in their daily activities, their body weight increased, and the degree of toe swelling decreased compared with the rats in the model group. The therapeutic effect was comparable to that of the positive drug group.

[0269] Through joint pathological observation, after administration of the Shenjin Huoluo Prescription, the degree of knee joint lesions in RA rats was improved. It was also found that after treatment with the Shenjin Huoluo Prescription, the expression of inflammatory factors in the serum could be correspondingly reduced.

[0270] The above results suggest that the Shenjin Huoluo Prescription has good anti-inflammatory and arthritis-relieving effects, and has an ideal therapeutic effect on RA.

[0271] All documents mentioned in this invention are cited herein by reference as if each individual document was cited by reference. In addition, it should be understood that after reading the above teachings of this invention, those skilled in the art can make various changes or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims of this application.

Claims

1. A traditional Chinese medicine composition, characterized in that, The traditional Chinese medicine composition comprises: 10 - 20 parts by weight of Lycopodium clavatum; 5 - 15 parts by weight of Speranskia tuberculata; 5 - 15 parts by weight of Stemona japonica; 10 - 20 parts by weight of Trichosanthes kirilowii; 15 - 25 parts by weight of Aconitum carmichaelii; 15 - 25 parts by weight of Aconitum kusnezoffii; 5 - 15 parts by weight of Spatholobus suberectus; 5 - 15 parts by weight of Dry Ginger; 10 - 20 parts by weight of Ligusticum sinense; and / or 5 - 15 parts by weight of Homalomena occulta.

2. A traditional Chinese medicine extract, characterized in that, The traditional Chinese medicine extract is obtained by extracting the traditional Chinese medicine composition as described in claim 1.

3. The traditional Chinese medicine extract according to claim 2, characterized in that, The traditional Chinese medicine extract contains components selected from the following group: 6 - gingerol, vanillic acid, linalool, aconitine diester alkaloids, chlorogenic acid, ferulic acid, ligustilide, or a combination thereof.

4. The traditional Chinese medicine extract according to claim 2, characterized in that, The traditional Chinese medicine extract is prepared by the following method: (1) Extract Dry Ginger, Ligusticum sinense and Homalomena occulta with a first extraction solvent, filter the extract, collect the filtrate and obtain volatile oil, and collect the medicinal residues for later use; (2) Add a second solvent to the medicinal residues obtained in step (1), Lycopodium clavatum, Speranskia tuberculata, Stemona japonica, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, and Spatholobus suberectus for decoction to obtain a decoction filtrate; combine the filtrates (including the filtrate in step (1)), concentrate to a concentrated filtrate with a relative density of 1.05 - 1.12, dilute with water, and let stand; (3) Elute the supernatant obtained in step (2) in macroporous resin, first wash with water to remove impurities, and then elute with 40v / v% - 60v / v% ethanol and 80v / v% - 95v / v% ethanol respectively, collect the eluate, concentrate and dry to obtain an enriched product.

5. A preparation method for preparing the traditional Chinese medicine extract according to claim 2, characterized in that, The method comprises the steps of: (1) Extract Dry Ginger, Ligusticum sinense and Homalomena occulta with a first extraction solvent, filter the extract, collect the filtrate, obtain volatile oil, and collect the medicinal residues for later use; (2) Add a second solvent to the medicinal residues obtained in step (1), Lycopodium clavatum, Speranskia tuberculata, Stemona japonica, Trichosanthes kirilowii, Aconitum carmichaelii, Aconitum kusnezoffii, and Spatholobus suberectus for decoction to obtain a decoction filtrate; combine the filtrates (including the filtrate in step (1)), concentrate to a concentrated filtrate with a relative density of 1.05 - 1.12, dilute with water, and let stand; (3) Elute the supernatant obtained in step (2) in macroporous resin, first wash with water to remove impurities, and then elute with 40v / v% - 60v / v% ethanol and 80v / v% - 95v / v% ethanol respectively, collect the eluate, concentrate and dry to obtain an enriched product; (4) Mix the volatile oil obtained in step (1) and the enriched product obtained in step (3) to obtain the traditional Chinese medicine extract.

6. The method according to claim 5, characterized in that, Step (3) further comprises: concentrating the eluate to an eluate with a relative density of 1.05 - 1.

20.

7. The method according to claim 5, characterized in that, The method further comprises: drying and pulverizing the obtained enriched product solution to obtain a powdery enriched product.

8. The use of the traditional Chinese medicine composition according to claim 1 and / or the traditional Chinese medicine extract according to claim 2, characterized in that, For preparing a pharmaceutical composition or preparation, the pharmaceutical composition or preparation is used for: (i) preventing and / or treating arthritis; (ii) preventing and / or treating symptoms caused by arthritis.

9. The use according to claim 8, characterized in that, The arthritis is selected from the following group: wind - cold - damp arthralgia, joint pain, rheumatism, rheumatoid arthritis, or a combination thereof.

10. A pharmaceutical composition or preparation for preventing and / or treating arthritis, and / or preventing and / or treating symptoms caused by arthritis, characterized in that, Comprises the traditional Chinese medicine composition as described in claim 1 or the pharmaceutical extract as described in claim 2; and a pharmaceutically acceptable carrier.

Images