Compound for treating osteoarthritis and application thereof

Abstract

The invention belongs to the technical field of medicine, and discloses a compound for treating osteoarthritis and application thereof, the compound is named as 3, 8-dihydroxy-9-methyl-2-fenchene, and the molecular formula is C11H18O2. According to the application of the 3, 8-dihydroxy-9-methyl-2-fenchene disclosed by the invention, the 3, 8-dihydroxy-9-methyl-2-fenchene is used for preparing the medicine for treating osteoarthritis. The medicine for treating osteoarthritis is a tablet, an injection or a capsule.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and in particular relates to a compound for treating osteoarthritis and its use. Background Art

[0002] Osteoarthritis (OA) is the most common form of arthritis, with a prevalence of about 60% in men and about 70% in women after the age of 65. There are currently limited methods for treating osteoarthritis. They include symptomatic therapy with simple analgesics, nonsteroidal anti-inflammatory drugs (NSAIDs) or intra-articular (IA) injections of glucocorticoids and hyaluronic acid (HA) preparations. Non-drug means range from physical exercise and weight loss to joint lavage and ultimately surgical joint replacement.

[0003] There is a great unmet need in the treatment of osteoarthritis for osteoarthritis drugs that are not yet available to improve the disease, and for effective pain treatments with long-lasting effects. In addition, due to the chronic nature of the disease, it is often necessary to extend the treatment time, so there is an obvious unmet need for osteoarthritis treatment without any major side effects. Systemic drugs that can currently be used to relieve osteoarthritis pain, such as non-selective NSAIDs and selective cyclooxygenase 2 (COX-2) inhibitors, are effective in the early-mid stages of OA, but usually cannot provide enough pain relief as the joints deteriorate. In addition, NSAIDs cause gastrointestinal complications in many patients, and COX-2 inhibitors have recently caused concerns about cardiovascular side effects / risks, resulting in the withdrawal of COX-2 inhibitors Vioxx and Bextra from the U.S. market. A large number of patients still need drugs for the treatment or prevention of osteoarthritis. Summary of the invention

[0004] In view of the problems existing in the prior art, the present invention provides a compound for treating osteoarthritis and use thereof.

[0005] The present invention is achieved by providing a compound for treating osteoarthritis, the compound is named 3,8-dihydroxy-9-methyl-2-fennel, and the molecular formula is: 11 H 18 O 2。

[0006] Another object of the present invention is to provide a method for preparing the compound for treating osteoarthritis, which method specifically comprises:

[0007] S1: Take the dried whole herb of Viola chinensis, heat and reflux extract with 4-8 times the amount of 60-70% ethanol, and concentrate the extract under reduced pressure until there is no alcohol taste to obtain an extract;

[0008] S2: Take the extract, suspend it with 3-5 times the volume of water, and then extract it with half the volume of petroleum ether and ethyl acetate 2-3 times;

[0009] S3: Take the ethyl acetate extract, mix the sample and apply it to silica gel column chromatography, use chloroform-methanol (volume ratio 16:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the chloroform-methanol (volume ratio 4:1) elution part, use Sephadex LH-20 gel column chromatography for separation, use chloroform-methanol (volume ratio 4:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the chloroform-methanol (volume ratio 1:1) part;

[0010] S4: Separate by silica gel column chromatography, use petroleum ether-acetone (volume ratio 8:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the petroleum ether-acetone (volume ratio 4:1) elution portion, concentrate and recover the solvent, recrystallize with methanol to obtain a monomer compound, namely 3,8-dihydroxy-9-methyl-2-fenechten; the mass ratio of the sample loading amount to the filler amount of all the above column chromatography is 1:60-80.

[0011] Furthermore, the dried whole herb of Herba Violae chinensis is 10 kg; and the concentration of ethanol is 60%.

[0012] Furthermore, the chloroform-methanol was subjected to gradient elution, and the volume ratios were 16:1, 8:1, 4:1, 2:1, 1:2, and 1:4.

[0013] Further, the gradient elution specifically comprises:

[0014] S41: taking raw petroleum ether-acetone, passing it into an adsorbent column filled with modified activated carbon for adsorption treatment;

[0015] S42: passing the petroleum ether-acetone obtained in step a into an adsorbent column filled with ion exchange resin for adsorption treatment to obtain a gradient elution solvent petroleum ether-acetone with a purity of ≥99.99% (wt).

[0016] Furthermore, the raw material petroleum ether-acetone flows through an adsorbent column filled with modified activated carbon at a rate of 0.5 to 30 column volumes per hour.

[0017] Furthermore, the modified activated carbon is activated carbon with an oxidizing acid attached to the surface.

[0018] Further, petroleum ether-acetone flows through an adsorbent column filled with ion exchange resin at a rate of 0.5 to 15 column volumes per hour.

[0019] Furthermore, it is used for the preparation of drugs for treating osteoarthritis.

[0020] Furthermore, the drug for treating osteoarthritis is in the form of tablets, injections or capsules.

[0021] Another object of the present invention is to provide a compound for treating osteoarthritis, wherein the compound is 3,8-dihydroxy-9-methyl-2-fennel, which is used to inhibit the activity of NLRP3 inflammasome and alleviate the symptoms of osteoarthritis by regulating the inflammatory pathway. The compound can also inhibit the generation of ROS associated with osteoarthritis, thereby reducing joint damage caused by oxidative stress.

[0022] Another object of the present invention is to provide a method for preparing 3,8-dihydroxy-9-methyl-2-fennel, comprising the following steps: S1, extracting raw materials from purple grass; S2, separating and purifying the extract using an organic solvent series; S3, further refining and purifying by chromatography technology to obtain a target compound. The method also includes chemically modifying 3,8-dihydroxy-9-methyl-2-fennel to improve its therapeutic effect and targeting for osteoarthritis symptoms.

[0023] Another object of the present invention is to provide a pharmaceutical composition for treating osteoarthritis, characterized in that it comprises an effective amount of 3,8-dihydroxy-9-methyl-2-fennel and at least one excipient. The pharmaceutical composition is further characterized in that 3,8-dihydroxy-9-methyl-2-fennel can affect the expression or activity of SOX transcription factors in the in vivo metabolic pathway, and is used to regulate gene expression and cell differentiation related to osteoarthritis.

[0024] In combination with the above technical solutions and the technical problems solved, the advantages and positive effects of the technical solutions to be protected by the present invention are as follows:

[0025] First, the purification method of the gradient elution solvent petroleum ether-acetone provided by the present invention can obtain petroleum ether-acetone with a purity of ≥99.99% (wt), which meets the high purity requirements of the gradient elution solvent, so that the purified petroleum ether-acetone can be applied to the field of gradient elution, expanding the scope of application. The purification method provided by the present invention fills the production gap of high-purity petroleum ether-acetone in China, reduces the cost required for import, and reduces the degree of being constrained by foreign technology. The purification method provided by the present invention has a product yield of ≥98%, a high yield, and also has the advantages of low energy consumption, simple production process, stable operation, and suitability for industrial production.

[0026] The application of 3,8-dihydroxy-9-methyl-2-fenchene in the present invention is used for preparing a drug for treating osteoarthritis. The drug for treating osteoarthritis is in the form of tablets, injections or capsules.

[0027] Second, the present invention utilizes 3,8-dihydroxy-9-methyl-2-fennel as a compound for treating osteoarthritis, and its preparation method and application provide the following significant technical advances:

[0028] 1) Innovation in targeted therapeutic mechanism: By inhibiting the activity of NLRP3 inflammasome, this compound provides a new mechanism to treat osteoarthritis. Traditional treatments often focus on relieving symptoms, while this new approach directly acts on the key pathways of the inflammatory process, providing more effective therapeutic effects.

[0029] 2) Ability to reduce oxidative stress: By reducing the generation of ROS, 3,8-dihydroxy-9-methyl-2-fennel can effectively reduce joint damage caused by oxidative stress. This not only helps relieve symptoms, but also slows the progression of the disease.

[0030] 3) Optimization of pharmacogenetics and pharmacodynamics: In-depth studies of the metabolic pathways and mechanisms of action of compounds provide opportunities to improve drug design and optimize their efficacy, including improving the bioavailability of drugs and reducing side effects.

[0031] 4) Multidimensional drug effects: This compound plays a role in regulating osteoarthritis-related gene expression and cell differentiation by affecting the expression or activity of SOX transcription factors. This multidimensional mode of action provides a new perspective for the treatment of osteoarthritis.

[0032] 5) Environmentally friendly extraction and preparation methods: The active ingredients are extracted from the natural plant Viburnum odoratum and purified by environmentally friendly methods, providing a sustainable way for the production of the compound.

[0033] The application of 3,8-dihydroxy-9-methyl-2-fennel in the treatment of osteoarthritis provides an innovative mechanism with the potential advantages of improved efficacy and reduced side effects, while showing environmental friendliness and sustainability in drug extraction and preparation.

[0034] Third, the preparation method of the purple grass extract provided by the present invention adopts refined chemical extraction and separation technology, and achieves high-purity extraction of the target compound-3,8-dihydroxy-9-methyl-2-fennel by controlling specific parameters such as solvent concentration, extraction times, elution volume ratio, mass ratio of sample loading amount and filler amount of column chromatography, etc. These parameters have a key impact on achieving technical effects, which are specifically reflected in the following aspects:

[0035] 1. Improve extraction efficiency: Using 4-8 times the amount of 60-70% ethanol as the extraction solvent not only ensures sufficient extraction efficiency, but also avoids the destruction of target compounds caused by excessively high concentrations of solvent. Precise control of ethanol concentration helps to effectively extract target components while reducing unnecessary impurity dissolution.

[0036] 2. Extraction selectivity: By sequentially extracting the suspension with petroleum ether and ethyl acetate, the target compound can be effectively separated and enriched. The selection and use order of the extractants increase the selective extraction of the target compound and improve the purity of the extraction.

[0037] 3. Accurate gradient elution: Gradient elution of silica gel column chromatography and Sephadex LH-20 gel column chromatography allows effective separation of compounds of different polarities in complex mixtures by precisely controlling the volume ratio of the eluent (from chloroform-methanol 16:1 to 1:4). This gradient elution technology significantly improves the separation purity and yield of the target compound.

[0038] 4. Optimized column chromatography conditions: By controlling the mass ratio of sample loading to filler usage to 1:60-80, the sample is fully separated during column chromatography and the decrease in separation efficiency caused by overloading is avoided. This precise ratio control ensures efficient and uniform separation of target compounds.

[0039] 5. Purification and recrystallization: Finally, the target compound was further purified by petroleum ether-acetone gradient elution and methanol recrystallization to obtain high-purity 3,8-dihydroxy-9-methyl-2-fenechten. This step not only improves the purity of the compound, but also contributes to its structural stability.

[0040] Through these finely controlled parameters, this method can not only efficiently and selectively extract the target compound from purple grass, but also ensure the high purity and stability of the obtained compound, providing high-quality raw materials for further chemical analysis and drug development. In addition, this technical process can provide a reference for the purification of similar plant extracts and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 is a flow chart of a method for preparing a compound for treating osteoarthritis provided in an embodiment of the present invention;

[0042] Figure 2 It is a flow chart of the gradient elution method provided in an embodiment of the present invention. DETAILED DESCRIPTION

[0043] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0044] The present invention provides a compound for treating osteoarthritis. The compound is named 3,8-dihydroxy-9-methyl-2-fennel, and has the molecular formula: C 11 H 18 O 2。

[0045] like Figure 1 As shown, the embodiment of the present invention provides a method for preparing the compound for treating osteoarthritis, and the method is specifically:

[0046] S1: Take the dried whole herb of Viola chinensis, heat and reflux extract with 4-8 times the amount of 60-70% ethanol, and concentrate the extract under reduced pressure until there is no alcohol taste to obtain an extract;

[0047] S2: Take the extract, suspend it with 3-5 times the volume of water, and then extract it with half the volume of petroleum ether and ethyl acetate 2-3 times;

[0048] S3: Take the ethyl acetate extract, mix the sample and apply it to silica gel column chromatography, use chloroform-methanol (volume ratio 16:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the chloroform-methanol (volume ratio 4:1) elution part, use Sephadex LH-20 gel column chromatography for separation, use chloroform-methanol (volume ratio 4:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the chloroform-methanol (volume ratio 1:1) part;

[0049] S4: Separate by silica gel column chromatography, use petroleum ether-acetone (volume ratio 8:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the petroleum ether-acetone (volume ratio 4:1) elution portion, concentrate and recover the solvent, recrystallize with methanol to obtain a monomer compound, namely 3,8-dihydroxy-9-methyl-2-fenechten; the mass ratio of the sample loading amount to the filler amount of all the above column chromatography is 1:60-80.

[0050] The dried whole herb of Herba Violae chinensis is 10 kg; and the concentration of ethanol is 60%.

[0051] The chloroform-methanol was subjected to gradient elution, and the volume ratios were 16:1, 8:1, 4:1, 2:1, 1:2, and 1:4.

[0052] like Figure 2 As shown, the gradient elution specifically includes:

[0053] S41: taking raw petroleum ether-acetone, passing it into an adsorbent column filled with modified activated carbon for adsorption treatment;

[0054] S42: passing the petroleum ether-acetone obtained in step a into an adsorbent column filled with ion exchange resin for adsorption treatment to obtain a gradient elution solvent petroleum ether-acetone with a purity of ≥99.99% (wt).

[0055] The raw material petroleum ether-acetone flows through an adsorbent column filled with modified activated carbon at a speed of 0.5 to 30 column volumes per hour.

[0056] Modified activated carbon is activated carbon with oxidizing acid attached to its surface.

[0057] Petroleum ether-acetone flows through an adsorbent column filled with ion exchange resin at a rate of 0.5 to 15 column volumes per hour.

[0058] The invention discloses an application of a compound for treating osteoarthritis, and is used for preparing a drug for treating osteoarthritis.

[0059] The medicine for treating osteoarthritis is in the form of tablets, injections or capsules.

[0060] The following are two specific examples of compounds for treating osteoarthritis with 3,8-dihydroxy-9-methyl-2-fennel, including detailed implementation schemes:

[0061] Example 1: Extraction and purification of 3,8-dihydroxy-9-methyl-2-fennel

[0062] 1) Raw material preparation: collect the dried whole herb of Herba Violae chinensis.

[0063] 2) Extraction process: Mix the dried whole herb of Viola chinensis with 4-8 times the amount of 60-70% ethanol, and heat and reflux for extraction. The extraction time is adjusted according to the amount of raw materials, usually 2-3 hours.

[0064] 3) Concentration and preliminary purification: The extract is concentrated under reduced pressure until there is no alcohol taste to obtain an extract.

[0065] 4) Further extraction: suspend the extract with 3-5 times the volume of water, and then extract with half the volume of petroleum ether and ethyl acetate 2-3 times in sequence.

[0066] 5) Column chromatography separation: Take the ethyl acetate extract, mix the sample and apply it to silica gel column chromatography, use chloroform-methanol gradient elution, and collect the eluted fractions of a specific ratio.

[0067] 6) Purification: Further purification was performed using Sephadex LH-20 gel column chromatography, and the chloroform-methanol fraction with a specific ratio was collected.

[0068] 7) Crystallization: The obtained product is concentrated to recover the solvent, and then recrystallized with methanol to obtain the target monomer compound.

[0069] Example 2: Evaluation and optimization of the efficacy of 3,8-dihydroxy-9-methyl-2-fennel

[0070] 1) Pharmacological activity test: Use cell culture and molecular biology techniques to evaluate the effect of the compound on the activity of NLRP3 inflammasome. The anti-inflammatory effect is evaluated by detecting inflammatory cytokines.

[0071] 2) ROS inhibition experiment: Evaluate the effect of compounds on ROS generation in an in vitro model and monitor changes in ROS levels through specific chemical detection methods.

[0072] 3) Drug metabolism research: Use high performance liquid chromatography-mass spectrometry (HPLC-MS) and other methods to study the metabolic pathways of compounds in the body.

[0073] 4) Regulation of SOX transcription factors: Study the effects of compounds on SOX family transcription factors through gene expression analysis and transcription factor activity determination.

[0074] 5) Drug improvement: Based on the above research results, explore chemical modifications or ligand design to optimize the therapeutic effect and targeting of the compounds.

[0075] 6) Animal model testing: Evaluate the therapeutic efficacy and safety of the compound in an osteoarthritis animal model to provide data support for preclinical research.

[0076] These two examples cover the entire process from raw material extraction, purification to efficacy evaluation and optimization, and fully illustrate the research and development process of the compound as a drug for treating osteoarthritis. The above is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any modification, equivalent substitution and improvement made by any technician familiar with the technical field within the technical scope disclosed by the present invention and within the spirit and principle of the present invention should be covered within the protection scope of the present invention.

Claims

1. A compound for treating osteoarthritis, characterized in that: The compound is named 3,8-dihydroxy-9-methyl-2-fennel, and its molecular formula is: 11 H 18 O 2。 2. A method for preparing a compound for treating osteoarthritis according to claim 1, characterized in that: The method is as follows: S1: Take the dried whole herb of Viola chinensis, heat and reflux extract with 4-8 times the amount of 60-70% ethanol, and concentrate the extract under reduced pressure until there is no alcohol taste to obtain an extract; S2: Take the extract, suspend it with 3-5 times the volume of water, and then extract it with half the volume of petroleum ether and ethyl acetate 2-3 times; S3: Take the ethyl acetate extract, mix the sample and apply it to silica gel column chromatography, use chloroform-methanol (volume ratio 16:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the chloroform-methanol (volume ratio 4:1) elution part, use Sephadex LH-20 gel column chromatography for separation, use chloroform-methanol (volume ratio 4:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the chloroform-methanol (volume ratio 1:1) part; S4: Separate by silica gel column chromatography, use petroleum ether-acetone (volume ratio 8:1→1:4) for gradient elution, each gradient elution is 2-3 times the column volume, collect the petroleum ether-acetone (volume ratio 4:1) elution portion, concentrate and recover the solvent, recrystallize with methanol to obtain a monomer compound, namely 3,8-dihydroxy-9-methyl-2-fenechten; the mass ratio of the sample loading amount to the filler amount of all the above column chromatography is 1:60-80.

3. The method for preparing a compound for treating osteoarthritis according to claim 2, characterized in that: The compound is 3,8-dihydroxy-9-methyl-2-fennel, which is used to inhibit the activity of NLRP3 inflammasome and alleviate the symptoms of osteoarthritis by regulating the inflammatory pathway. The compound can also inhibit the generation of ROS associated with osteoarthritis, thereby reducing joint damage caused by oxidative stress.

4. The method for preparing a compound for treating osteoarthritis according to claim 2, characterized in that: The following steps are involved: S1. Extracting raw materials from purple grass; S2. Separating and purifying the extract using a series of organic solvents; S3. Further refining and purifying by chromatography technology to obtain the target compound. The method also includes chemically modifying 3,8-dihydroxy-9-methyl-2-fennel to improve its therapeutic effect and targeting for osteoarthritis symptoms.

5. The method for preparing a compound for treating osteoarthritis according to claim 2, characterized in that: The pharmaceutical composition comprises an effective amount of 3,8-dihydroxy-9-methyl-2-fennel and at least one auxiliary material. The pharmaceutical composition is further characterized in that 3,8-dihydroxy-9-methyl-2-fennel can affect the expression or activity of SOX transcription factors in the in vivo metabolic pathway, and is used to regulate gene expression and cell differentiation related to osteoarthritis.

6. The method for preparing the compound for treating osteoarthritis according to claim 1, characterized in that: The dried whole herb of Viola flos-sinica is 10 kg; the concentration of ethanol is 60%; the chloroform-methanol is used for gradient elution, and the volume ratios are 16:1, 8:1, 4:1, 2:1, 1:2, and 1:4, respectively.

7. The method for preparing a compound for treating osteoarthritis according to claim 1, characterized in that: The gradient elution specifically comprises: S41: taking raw petroleum ether-acetone, passing it into an adsorbent column filled with modified activated carbon for adsorption treatment; S42: passing the petroleum ether-acetone obtained in step a into an adsorbent column filled with ion exchange resin for adsorption treatment to obtain a gradient elution solvent petroleum ether-acetone with a purity of ≥99.99% (wt).

8. The method for preparing a compound for treating osteoarthritis according to claim 5, characterized in that: The raw material petroleum ether-acetone flows through an adsorbent column filled with modified activated carbon at a rate of 0.5 to 30 column volumes per hour; the modified activated carbon is activated carbon with oxidizing acid attached to the surface; and the petroleum ether-acetone flows through an adsorbent column filled with ion exchange resin at a rate of 0.5 to 15 column volumes per hour.

9. The use of a compound for treating osteoarthritis according to claim 1, characterized in that: Used for the preparation of drugs for treating osteoarthritis.

10. The use according to claim 9, characterized in that The medicine for treating osteoarthritis is in the form of tablets, injections or capsules.

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