A soothing external use traditional Chinese medicine composition and a preparation method thereof

Abstract

This invention discloses a topical traditional Chinese medicine composition for relieving pain and its preparation method, belonging to the field of traditional Chinese medicine preparation technology. The topical traditional Chinese medicine composition for relieving pain is made from the following raw materials in parts by weight: Angelica sinensis 5-25 parts, Ligusticum striatum 5-20 parts, Lycopodium clavatum 10-35 parts, Artemisia argyi 5-25 parts, artificial Phellodendron chinense 3-18 parts, Clematis chinensis 10-40 parts, and Angelica dahurica 5-20 parts. This invention uses steam distillation to extract volatile oils and a dual-solvent gradient extraction method with the residue, which not only fully extracts the volatile fat-soluble components from Angelica sinensis, Ligusticum striatum, and Angelica dahurica, but also efficiently extracts the water-soluble components from artificial Phellodendron chinense, Lycopodium clavatum, and other medicinal materials. Experimental results show that the extraction rates of ferulic acid, ligustilide, imperatorin, and berberine hydrochloride in Example 1 of this invention reached 89.23%, 92.56%, 90.12%, and 93.45%, respectively, which are 110.7%, 223.1%, 75.8%, and 23.6% higher than those of conventional water decoction process, thus improving the utilization rate of raw materials.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine preparation technology, specifically to a topical traditional Chinese medicine composition for relieving pain and its preparation method. Background Technology

[0002] Neck, shoulder, lower back, and leg pain are common chronic pain conditions in clinical practice, often caused by factors such as invasion of wind, cold, and dampness, accumulated strain, and injuries from falls and blows. Symptoms typically include localized soreness, numbness, and limited mobility; in severe cases, it can affect daily life and work capacity. Traditional Chinese medicine has a long history and unique advantages in treating this type of pain. External application of Chinese herbal preparations is widely used as an adjunct therapy for pain due to its convenience, direct action, and minimal side effects.

[0003] Traditional Chinese medicine (TCM) topical analgesics possess unique theoretical and clinical advantages in chronic pain intervention due to their multi-component, multi-target, and multi-pathway action characteristics. However, currently available TCM topical analgesic compositions and preparations often suffer from several issues. At the prescription level, existing prescriptions primarily consist of pungent, warm, and blood-activating herbs, which tend to be warming and drying in nature. Long-term topical use can easily irritate the skin, especially broken skin, leading to adverse reactions such as erythema and edema. Some prescriptions also have numerous herbs, unclear formulation logic, and unidentified core active ingredients, making it difficult to achieve quality control and stable efficacy. At the extraction and preparation level, the core active ingredients of analgesic TCMs encompass substances of different polarities, including volatile fat-soluble components, phenolic acids, and alkaloids. However, existing technologies often employ conventional water decoction or single-solvent extraction processes, which cannot efficiently extract active ingredients of different polarities. This generally results in significant loss of volatile components and extremely low extraction rates of fat-soluble active ingredients, leading to poor utilization of raw materials and directly limiting the analgesic and anti-inflammatory efficacy of the preparations. Regarding formulation stability, the core active substances in traditional Chinese medicine analgesic compositions often contain volatile oils. These components are inherently volatile, easily oxidized and degraded, and have poor thermal stability. Current technologies often directly add these volatile oils to the formulation system, which easily leads to rapid degradation, decreased efficacy, and discoloration / stratification of the formulation during storage. This results in short product shelf lives and makes it difficult to meet the quality control requirements for long-term storage. Regarding efficacy, on the one hand, existing traditional Chinese medicine extracts contain a large number of large-molecule impurities such as polysaccharides and proteins, which increase skin barrier resistance, leading to low transdermal absorption efficiency of the active ingredients, slow onset of action, and inability to quickly relieve patients' acute pain symptoms. On the other hand, active ingredients that have not undergone controlled-release treatment release rapidly and have a short duration of action, requiring frequent administration to maintain the analgesic effect, causing inconvenience for patients. Regarding medication safety, some topical preparations add irritating penetration enhancers or toxic herbs to improve analgesia, easily causing skin irritation and allergic reactions, failing to meet the safety needs of patients for long-term, repeated use.

[0004] Therefore, we propose a topical Chinese medicine composition for relieving pain and its preparation method to alleviate or solve the above-mentioned problems.

[0005] The information disclosed above in this background section is only for enhancing the understanding of the background section of this invention, and therefore may include prior art that is not known to those skilled in the art. Summary of the Invention

[0006] To address the aforementioned technical problems, this invention provides a topical Chinese medicine composition for relieving pain and its preparation method, thereby solving the problems existing in the prior art of topical analgesic Chinese medicine compositions, such as the warm and dry nature of the medicine, which easily causes skin irritation; low extraction rate of effective components with different polarities; easy oxidation and volatilization of volatile oils leading to poor formulation stability; insufficient transdermal absorption efficiency of effective components; slow onset of analgesia; and short duration of efficacy.

[0007] To achieve the above objectives, the present invention provides a topical Chinese medicine composition for relieving pain, which is made from the following raw materials in parts by weight: Angelica sinensis 5-25 parts, Ligusticum striatum 5-20 parts, Lycopodium clavatum 10-35 parts, Artemisia argyi 5-25 parts, artificial Phellodendron chinense 3-18 parts, Clematis chinensis 10-40 parts, and Angelica dahurica 5-20 parts.

[0008] The method for preparing the aforementioned topical Chinese medicine composition for relieving pain includes the following steps:

[0009] Step S1: Weigh each raw material according to the formula, clean and crush it through a 20-40 mesh sieve, mix it, add 8-12 times the total weight of the raw materials in purified water and soak for 30-60 minutes, extract the volatile oil by steam distillation for 3-5 hours, collect the volatile oil and the water after distillation separately, and keep the residue for later use.

[0010] Step S2: Add 6-10 times the weight of the volatile oil obtained in Step S1 to anhydrous ethanol and stir to dissolve to obtain a volatile oil ethanol solution; separately take 8-12 times the weight of β-cyclodextrin and add 8-10 times the weight of purified water, and stir at 40-50℃ until completely dissolved to obtain a β-cyclodextrin saturated aqueous solution; under constant temperature of 40-50℃ and stirring at 200-300 r / min, slowly add the volatile oil ethanol solution dropwise to the β-cyclodextrin saturated aqueous solution. After the addition is complete, continue stirring at constant temperature for 1-2 h, then place in a refrigerated environment of 2-8℃ and let stand for 12-24 h, filter, wash the filter cake and dry it under vacuum below 40℃ to obtain the volatile oil β-cyclodextrin inclusion complex;

[0011] Step S3: Take the residue left from step S1, add 6-10 times the weight of the residue and 50%-70% ethanol aqueous solution, heat and reflux to extract 1-2 times, 1-2 hours each time, and filter to obtain ethanol extract; then add 6-8 times the weight of the residue and purified water to the residue after ethanol extraction, heat and decoct to extract 1-2 times, 1-1.5 hours each time, and filter to obtain aqueous extract;

[0012] Step S4: Combine the distilled aqueous solution from Step S1, the ethanol extract from Step S3, and the aqueous extract. Concentrate under reduced pressure at a vacuum of -0.06 to -0.08 MPa and a temperature of 50-60℃ to obtain a clear extract with a relative density of 1.10-1.20 measured at 60℃. Add edible ethanol to the clear extract and stir until the alcohol content reaches 60-70% by volume. After stirring evenly, refrigerate and let stand at 4℃ for 12-24 hours. Filter the supernatant, recover the ethanol from the filtrate under reduced pressure, and continue to concentrate to a thick extract with a relative density of 1.25-1.35 measured at 60℃. Dry the thick extract under vacuum, pulverize it through an 80-mesh sieve, and obtain the dry extract powder of traditional Chinese medicine.

[0013] Step S5: Mix the volatile oil β-cyclodextrin inclusion complex obtained in step S2 with the dry extract powder of traditional Chinese medicine obtained in step S4 to obtain the topical traditional Chinese medicine composition for relieving pain.

[0014] Compared with the prior art, the beneficial effects of the present invention are:

[0015] This invention employs a dual-solvent gradient extraction method, combining steam distillation with extraction of medicinal residues. This method effectively extracts the volatile fat-soluble components from Angelica sinensis, Ligusticum striatum, and Angelica dahurica, while also efficiently extracting the water-soluble components from artificially processed Phellodendron amurense and Lycopodium clavatum. Experimental results show that in Example 1 of this invention, the extraction rates of ferulic acid, ligustilide, imperatorin, and berberine hydrochloride reached 89.23%, 92.56%, 90.12%, and 93.45%, respectively, representing increases of 110.7%, 223.1%, 75.8%, and 23.6% compared to conventional decoction processes, thus improving the utilization rate of the raw materials.

[0016] This invention employs β-cyclodextrin inclusion complex technology to encapsulate extracted volatile oils, forming stable inclusion compounds that solve the problems of easy oxidation, volatility, and poor stability of volatile oils. Accelerated stability experiments show that the ointment of Example 4, after being treated with inclusion complex and placed at 40°C for 6 months, still retained 90.12% of the ligustilide content, while the ointment of Comparative Example 2, without inclusion complex, retained only 38.65% of the content, and showed obvious layering and discoloration. Simultaneously, the inclusion complex technology achieves a sustained-release effect of the volatile oil. Hot plate analgesia experiments show that the pain threshold improvement rate of Example 1 remained at 82.35% 6 hours after administration, superior to the unencapsulated group and the commercially available product group, extending the duration of efficacy to over 6 hours, achieving long-lasting analgesia.

[0017] This invention removes a large number of macromolecular impurities through an alcohol precipitation purification process and, combined with the synergistic effect of transdermal absorption enhancers in the formulation, improves the transdermal absorption efficiency of the active ingredients. In vitro transdermal experiments show that, in Example 1, the cumulative permeation amounts of ferulic acid and imperatorin after 8 hours were 32.56 μg / cm³. 2 and 28.74 μg / cm 2 The average permeability reached 42.35%, which is 3.34 times that of the conventional process group. The hot plate analgesia experiment showed that the analgesic effect appeared 30 minutes after administration in the Example 1 group, with a pain threshold increase rate of 68.92%, and the onset time was shorter than that of commercially available products, achieving rapid analgesia.

[0018] The addition of artificial Phellodendron bark to the formula of this invention leverages its cooling properties to counteract the potential skin irritation caused by warming and drying herbs such as Angelica sinensis, Ligusticum striatum, and Artemisia argyi, demonstrating the theoretical characteristics of counteracting drug combination. Skin irritation experiments confirmed that the cream in Example 4 containing artificial Phellodendron bark was non-irritating to both intact and broken skin on rabbits, while the cream in Comparative Example 3, which did not contain artificial Phellodendron bark, caused mild irritation to broken skin.

[0019] The above overview is for illustrative purposes only and is not intended to be limiting in any way. In addition to the illustrative aspects, embodiments, and features described above, further aspects, embodiments, and features of the invention will become readily apparent from the accompanying drawings and the following detailed description. Attached Figure Description

[0020] Figure 1 This is a flowchart illustrating the preparation method of the topical Chinese medicine composition for relieving pain according to the present invention. Detailed Implementation

[0021] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. It should be noted that the drawings are schematic and not illustrated to scale. For clarity and convenience, the relative sizes and proportions of the parts shown in the drawings have been exaggerated or reduced in size. Any size is only illustrative and not limiting.

[0022] In all embodiments, comparative examples, and effect verification experiments of this invention, the raw materials, reagents, instruments, and equipment used are the same.

[0023] (I) Active pharmaceutical ingredients and quality control

[0024] Angelica sinensis (Oliv.) Diels, a plant belonging to the genus Angelica in the family Apiaceae; Ligusticum sinense Oliv., a plant belonging to the genus Ligusticum in the family Apiaceae; Lycopodium japonicum Thunb., a plant belonging to the genus Lycopodium in the family Lycopodiaceae; Artemisia argyi Levl. et Vant., a plant belonging to the genus Artemisia in the family Asteraceae; Phellodendron chinense Schneid., a plant belonging to the genus Phellodendron in the family Rutaceae, is dried bark processed through standardized artificial cultivation; Impatiens balsamina L., a plant belonging to the genus Impatiens in the family Balsamicaceae; Angelica dahurica (Fisch. ex Hoffm.) Benth. et Hook. f., a plant belonging to the genus Angelica in the family Apiaceae.

[0025] (II) Reagents and excipients

[0026] Reagents used for extraction and purification: Anhydrous ethanol and 95% food-grade ethanol are both pharmaceutical grade; β-cyclodextrin is pharmaceutical grade; purified water is laboratory-prepared water for injection.

[0027] Pharmaceutical excipients: stearic acid, white petrolatum, liquid paraffin, glyceryl monostearate, Tween-80, Span-80, glycerin, propylene glycol, azone, ethylparaben, phenoxyethanol, sodium metabisulfite, and vitamin E are all pharmaceutical grade.

[0028] Reference standards for detection: ferulic acid reference standard, ligustilide reference standard, imperatorin reference standard, berberine hydrochloride reference standard;

[0029] Reagents used in the experiment: Methanol and acetonitrile were of chromatographic grade; all other reagents were of analytical grade.

[0030] Example 1

[0031] As attached Figure 1 The preparation method of the external application traditional Chinese medicine composition, where 1 part by weight corresponds to 100g, includes the following steps:

[0032] Prescription ratio: Angelica sinensis 15 parts, Ligusticum striatum 10 parts, Lycopodium clavatum 25 parts, Artemisia argyi 15 parts, artificial Phellodendron chinense 8 parts, Clematis chinensis 30 parts, Angelica dahurica 10 parts, total raw material input 11300g.

[0033] Step S1: Weigh each ingredient according to the prescription, and first clean them. Manually pick out impurities, moldy parts and non-medicinal parts from the herbs. Rinse Angelica sinensis, Ligusticum striatum, Angelica dahurica and artificial Phellodendron chinense with purified water twice and drain the surface water. Use a wind separator to remove dust and debris from Lycopodium clavatum, Clematis armandii and Artemisia argyi to complete the cleaning process.

[0034] All the cleaned medicinal materials are put into a high-speed universal pulverizer, pulverized and then passed through a 30-mesh standard sieve. The coarse powder that does not pass through the sieve is pulverized again until all of it passes through the sieve, so as to obtain a uniformly mixed raw material powder.

[0035] Put all the raw drug powder into a 100L multi-functional extraction tank, add purified water in a volume of 10 times the total weight of the raw drug, close the tank opening, and soak at room temperature for 45 minutes to ensure that the medicinal material is fully swollen, which is conducive to the dissolution of effective components and the extraction of volatile oils.

[0036] Turn on the heating system and condensation circulation system of the extraction tank, and control the inlet temperature of the condensate water to ≤25℃ to ensure that the volatile oil is fully condensed and recovered; heat up to the boiling point of the liquid in the tank, maintain a gentle boil and carry out steam distillation for 4 hours. During the distillation process, record the volume of the distillate and the amount of volatile oil collected every 30 minutes to ensure complete distillation.

[0037] After distillation, the heating system was turned off, and the volatile oil (a total of 186 ml was collected, with a relative density of 0.852 g / ml at 25°C) and the distilled water were collected separately through an oil-water separator. The distilled water was transferred to a clean storage tank and sealed for later use, while the residue was left in the extraction tank for subsequent extraction.

[0038] Step S2: Take all the collected volatile oil, add 8 times the weight of the volatile oil in anhydrous ethanol, place it in a clean container, stir at 100 r / min for 10 min until the volatile oil is completely dissolved, and obtain a clear volatile oil ethanol solution, seal it for later use.

[0039] Weigh out β-cyclodextrin, the amount of which is 10 times the weight of the volatile oil, and put it into a 50L constant temperature stirring tank. Add purified water in an amount of 9 times the weight of β-cyclodextrin, turn on the constant temperature system and stirring system, control the temperature at 45℃ and the stirring speed at 250r / min, and continue stirring for 30min until the β-cyclodextrin is completely dissolved to obtain a clear and transparent saturated aqueous solution. Keep the temperature and speed for later use.

[0040] Maintain a constant temperature of 45℃ and a stirring state of 250r / min. Use a constant pressure dropping funnel to slowly add the volatile oil ethanol solution to the β-cyclodextrin saturated aqueous solution, control the dropping rate at 50ml / min, and control the total dropping time at about 30min to avoid uneven inclusion and decreased inclusion rate due to excessively fast dropping speed.

[0041] After the addition is complete, maintain a constant temperature of 45℃ and 250r / min and continue stirring for 1.5h to ensure the inclusion reaction is complete. Stop heating and stirring, transfer the entire inclusion solution into a clean refrigerated container, seal it, and refrigerate it at 4℃ for 18h to allow the inclusion compound to fully crystallize.

[0042] The inclusion solution, after being refrigerated and allowed to stand, was filtered under reduced pressure using a Buchner funnel. The filter cake was washed three times with 2000 ml of pre-cooled purified water at 4°C in equal volumes to remove unencapsulated free volatile oil and β-cyclodextrin, until the washing liquid had no obvious volatile oil odor. The washed filter cake was then transferred to a vacuum drying oven and dried at 35°C and -0.07 MPa for 12 hours until the moisture content was ≤3.0%. The dried inclusion complex was then removed, pulverized using a pulverizer, and passed through an 80-mesh sieve to obtain the volatile oil β-cyclodextrin inclusion complex, weighing 1692 g. The volatile oil inclusion rate was tested to be 92.6%. The complex was then sealed and stored in a cool, dry place for later use.

[0043] Step S3: Take the residue remaining after steam distillation, add 8 times the wet weight of the residue to a 60% ethanol aqueous solution, close the extraction tank, turn on the heating and stirring system, heat the liquid in the tank until it boils, maintain a gentle boil and perform reflux extraction, extract twice, each time for 1.5 hours; after each extraction, turn off the heating, wait for the temperature in the tank to drop below 60℃, filter with an 80-mesh medical filter cloth, collect the ethanol extract, combine the ethanol extracts from the two extractions and transfer them to a clean storage tank, seal and keep for later use, the residue is left in the extraction tank;

[0044] Add purified water at a wet weight of 7 times to the ethanol-extracted residue, turn on the heating system, heat to boiling and then maintain a gentle boil for extraction. Extract twice, each time for 1 hour. After each extraction, filter with an 80-mesh medical filter cloth and collect the aqueous extract. Combine the aqueous extracts from the two extractions and transfer them to a clean storage tank, seal and store for later use.

[0045] Step S4: Combine all the distilled aqueous solution, ethanol extract, and water extract collected in the previous steps, and transfer them to a 200L vacuum concentration tank. Turn on the vacuum system and heating system, and control the vacuum degree to -0.07MPa and the liquid temperature to 55℃ during the concentration process to carry out low-temperature vacuum concentration to avoid high temperature damage to the heat-sensitive active ingredients. Take samples for testing in real time during the concentration process. When the relative density of the liquid is measured to be 1.15 at 60℃, stop the concentration, release the clear extract, and transfer it to an alcohol precipitation tank.

[0046] Turn on the stirring system of the alcohol precipitation tank and control the stirring speed to 150 r / min. While stirring, slowly add 95% food-grade ethanol to the clear extract, stirring continuously and monitoring the alcohol content in real time. Stop adding ethanol when the volume fraction of ethanol in the system reaches 65%. Continue stirring for 20 minutes to mix the system evenly. Turn off the stirring, seal the alcohol precipitation tank, and place it in a 2℃ cold storage environment for 18 hours to allow ineffective impurities to fully precipitate.

[0047] After refrigeration and settling, take the supernatant of the alcohol precipitation, and filter it finely using a plate and frame filter to remove precipitated impurities. Collect the clear filtrate. Transfer the filtrate to a vacuum concentration tank, control the vacuum degree to -0.07 MPa and the feed temperature to 55°C, and recover ethanol under reduced pressure until the distillate has no ethanol odor. Continue concentration until the feed liquid has a relative density of 1.30 when measured at 60°C. Stop concentration and release the thick extract.

[0048] Spread the thick extract evenly on a stainless steel tray with a thickness of ≤2cm, transfer it to a vacuum drying oven, set the temperature to 40℃ and the vacuum degree to -0.08MPa, and vacuum dry for 16h until the moisture content of the dry extract is ≤5.0%; take out the dry extract, pulverize it with a high-speed universal pulverizer, pass it through an 80-mesh standard sieve, and obtain the Chinese herbal dry extract powder, weighing 2865g, and seal it for later use.

[0049] Step S5: Add all the volatile oil β-cyclodextrin inclusion complex and the dry extract powder of traditional Chinese medicine prepared above into a three-dimensional motion mixer. Set the mixing speed to 15 r / min and the mixing time to 30 min. After stopping the machine, take a sample for testing to ensure that the mixing uniformity RSD ≤ 2.0%. After mixing, release the material, seal it and store it in a cool and dry place to obtain the analgesic external traditional Chinese medicine composition of this embodiment. A total of 4552 g was prepared, and the yield of the finished product was 40.28%.

[0050] Example 2

[0051] A method for preparing a topical Chinese medicine composition, where 1 part by weight corresponds to 100g, includes the following steps:

[0052] Prescription ratio: Angelica sinensis 12 parts, Ligusticum striatum 12 parts, Lycopodium clavatum 20 parts, Artemisia argyi 18 parts, artificial Phellodendron chinense 10 parts, Clematis chinensis 25 parts, Angelica dahurica 12 parts, total raw material input 10900g.

[0053] Step S1: Weigh and purify each raw material according to the prescription, pulverize it through a 20-mesh sieve, mix it evenly and put it into a 100L multi-functional extraction tank. Add purified water with a total weight of 8 times the raw materials and soak at room temperature for 60 minutes. Start steam distillation, control the condensate temperature ≤25℃, and distill at a gentle boil for 5 hours. Collect the volatile oil (a total of 178ml was collected, with a relative density of 0.848g / ml at 25℃) and the distilled water separately, and keep the residue for later use.

[0054] Step S2: Take the collected volatile oil, add 6 times the weight of anhydrous ethanol, stir to dissolve and obtain a volatile oil ethanol solution; weigh 8 times the weight of β-cyclodextrin, add 10 times the weight of purified water, stir at 50℃ until completely dissolved to obtain a β-cyclodextrin saturated aqueous solution; under constant temperature of 50℃ and stirring at 300r / min, slowly add the volatile oil ethanol solution dropwise to the saturated aqueous solution, and continue stirring at constant temperature for 2h after the addition is complete; then place in an 8℃ environment for refrigeration and stand for 12h, filter, wash the filter cake with pre-cooled purified water, vacuum dry at below 40℃ for 10h, pulverize and pass through an 80-mesh sieve to obtain 1568g of volatile oil β-cyclodextrin inclusion complex, with a volatile oil inclusion rate of 91.2%, and seal for later use.

[0055] Step S3: Take the distilled residue, add 10 times the wet weight of the residue to a 50% ethanol aqueous solution, heat and reflux to extract twice, 2 hours each time, filter and combine the ethanol extracts for later use; take the residue after ethanol extraction, add 8 times the wet weight of the residue to purified water, heat and decoct to extract once, extraction time 1.5 hours, filter to obtain the aqueous extract for later use.

[0056] Step S4: Combine the distilled aqueous solution, ethanol extract, and aqueous extract, and concentrate under reduced pressure at -0.08 MPa and 50°C to obtain a clear extract with a relative density of 1.10 at 60°C. Add 95% edible ethanol to the clear extract until the alcohol content reaches 60% by volume. Stir evenly and refrigerate at below 4°C for 24 hours. Filter the supernatant, recover the ethanol from the filtrate under reduced pressure, and continue to concentrate to a thick extract with a relative density of 1.25 at 60°C. Dry under vacuum at 40°C, pulverize through an 80-mesh sieve, and obtain 2682g of dry extract powder of traditional Chinese medicine, which is then sealed for later use.

[0057] Step S5: Add the volatile oil β-cyclodextrin inclusion complex and the dry extract powder of traditional Chinese medicine into a three-dimensional motion mixer and mix at 15 r / min for 30 min. After mixing evenly, the topical traditional Chinese medicine composition for relieving pain in this embodiment is obtained. A total of 4245g was prepared, with a product yield of 38.94%.

[0058] Example 3

[0059] A method for preparing a topical Chinese medicine composition, where 1 part by weight corresponds to 100g, includes the following steps:

[0060] Prescription ratio: Angelica sinensis 20 parts, Ligusticum striatum 18 parts, Lycopodium clavatum 30 parts, Artemisia argyi 22 parts, artificial Phellodendron chinense 15 parts, Clematis chinensis 35 parts, Angelica dahurica 18 parts, total raw material input 15800g.

[0061] Step S1: Weigh and purify each raw material according to the prescription, pulverize it through a 40-mesh sieve, mix it evenly and put it into a 200L multi-functional extraction tank. Add purified water with a total weight of 12 times the raw materials and soak at room temperature for 30 minutes. Start steam distillation, control the condensate temperature ≤25℃, and distill at a gentle boil for 3 hours. Collect the volatile oil (a total of 285ml was collected, with a relative density of 0.855g / ml at 25℃) and the distilled water separately, and keep the residue for later use.

[0062] Step S2: Take the collected volatile oil, add 10 times the weight of anhydrous ethanol, stir to dissolve and obtain a volatile oil ethanol solution; weigh 12 times the weight of β-cyclodextrin, add 8 times the weight of purified water, stir at 40℃ until completely dissolved to obtain a β-cyclodextrin saturated aqueous solution; under constant temperature of 40℃ and stirring at 200r / min, slowly add the volatile oil ethanol solution dropwise to the saturated aqueous solution, and continue stirring at constant temperature for 1h after the addition is complete; then place in a refrigerated environment at 2℃ for 24h, filter, wash the filter cake with pre-cooled purified water, vacuum dry at 35℃ for 15h, pulverize and pass through an 80-mesh sieve to obtain 2985g of volatile oil β-cyclodextrin inclusion complex, with a volatile oil inclusion rate of 93.1%, and seal for later use.

[0063] Step S3: Take the distilled residue, add 6 times the wet weight of the residue to a 70% ethanol aqueous solution, heat and reflux to extract once for 1 hour, filter to obtain the ethanol extract, and set aside; take the residue after ethanol extraction, add 6 times the wet weight of the residue to purified water, heat and decoct to extract twice, 1 hour each time, filter and combine the aqueous extracts, and set aside.

[0064] Step S4: Combine the distilled aqueous solution, ethanol extract, and aqueous extract, and concentrate under reduced pressure at -0.06 MPa and 60°C to obtain a clear extract with a relative density of 1.20 at 60°C. Add 95% edible ethanol to the clear extract until the alcohol content reaches 70% by volume. Stir well and refrigerate at below 4°C for 12 hours. Filter the supernatant, recover the ethanol from the filtrate under reduced pressure, and continue to concentrate to a thick extract with a relative density of 1.35 at 60°C. Dry under vacuum at 40°C, pulverize through an 80-mesh sieve, and obtain 4120g of dry extract powder of traditional Chinese medicine, which is then sealed for later use.

[0065] Step S5: Add the volatile oil β-cyclodextrin inclusion complex and the dry extract powder of traditional Chinese medicine into a three-dimensional motion mixer and mix at 15 r / min for 30 min. After mixing evenly, the topical Chinese medicine composition for relieving pain in this embodiment is obtained. A total of 7102g was prepared, with a product yield of 44.95%.

[0066] Example 4

[0067] The preparation of an oil-in-water emulsion cream, based on the preparation of 1000g of finished product, using the traditional Chinese medicine composition prepared in Example 1, includes the following steps:

[0068] Prescription ratio: 100g of the external Chinese medicine composition prepared in Example 1, 60g of stearic acid, 30g of white petrolatum, 40g of liquid paraffin, 30g of glyceryl monostearate, 10g of sodium dodecyl sulfate, 30g of Tween-80, 10g of Span-80, 80g of glycerin, 30g of propylene glycol, 8g of azone, 1g of ethylparaben, 2g of phenoxyethanol, 0.5g of sodium metabisulfite, 0.5g of vitamin E, and purified water to 1000g.

[0069] (1): Accurately weigh 60g of stearic acid, 30g of white petrolatum, 40g of liquid paraffin, 30g of glyceryl monostearate, and 10g of Span-80 according to the prescription, and put them all into a 5L oil phase preparation tank. Turn on the jacket heating system and the low-speed stirring system, control the stirring speed to 100r / min, raise the temperature to 80℃, and continue stirring until all materials are completely melted and mixed evenly, with no visible particles or lumps. Keep the temperature at 80℃, seal and set aside for later use to obtain the oil phase.

[0070] (2): Weigh 10g of sodium dodecyl sulfate, 30g of Tween-80, 80g of glycerol, 30g of propylene glycol, 1g of ethylparaben, and 0.5g of sodium metabisulfite according to the prescription. Put them into a 10L aqueous phase preparation tank, add 568g of purified water according to the prescription, turn on the jacket heating system and stirring system, control the stirring speed to 150r / min, raise the temperature to 80℃, and continue stirring until all materials are completely dissolved and the solution is clear, transparent and free of impurities. Keep it at 80℃ and seal it for later use to obtain the aqueous phase.

[0071] (3): Simultaneously adjust the temperature of the oil phase and the water phase to ensure that the temperature of both phases is stable at 78℃; turn on the high-speed stirring system of the water phase tank, control the stirring speed at 400r / min, and slowly and uniformly add the oil phase to the water phase through a closed conveying pipeline. The oil phase addition time is controlled within 10min throughout the process; after all the oil phase has been added, maintain the temperature and speed at 78℃ and 400r / min, and continue stirring and emulsifying for 20min to complete the initial emulsification; turn on the high shear homogenizer, set the homogenization speed at 2500r / min, and homogenize the initially emulsified cream matrix for 8min to form a fine and uniform oil-in-water system with no oil-water separation and no coarse particles. After homogenization, a blank cream matrix is ​​obtained.

[0072] (4): Stop the jacket heating, turn on the circulating cooling water cooling system, and keep the stirring speed at 250 r / min to cool the blank cream matrix at a uniform speed. When the temperature of the cream matrix drops to 48℃, stop the cooling and add 100g of the traditional Chinese medicine composition of Example 1, 8g of azone, 2g of phenoxyethanol and 0.5g of vitamin E. Keep stirring at 250 r / min for 40 min to ensure that the active ingredients of traditional Chinese medicine are completely and evenly dispersed, and that the cream system is fine without lumps, particles and visible powder agglomerates. Continue to turn on the cooling system and keep stirring at a low speed to cool the cream system to room temperature of 25℃. Stop stirring to obtain the cream semi-finished product. Take a sample for intermediate product testing. The properties, pH value, particle size and uniformity all meet the quality requirements.

[0073] (5): The qualified semi-finished cream is sent to a clean filling room and filled into pharmaceutical aluminum-plastic composite tubes according to the target specifications (5g / tube, 10g / tube, 20g / tube, 30g / tube, 35g / tube, 50g / tube, 60g / tube, 80g / tube, 100g / tube) using a fully automatic tube filling machine. After filling, the tubes are automatically sealed to ensure that the seal is tight and there is no leakage. The filled and sealed cream is sterilized by cobalt-60 irradiation with a dose of 8kGy for 12 hours to ensure that the microbial limit meets the requirements of the Chinese Pharmacopoeia for external preparations. After sterilization, the product is inspected by light and products with leaks, unqualified filling amount, or abnormal appearance are removed. Qualified products are labeled and packaged to obtain the finished cream.

[0074] The finished product was tested for quality. It is a milky white, fine cream with a uniform texture, no oil-water separation, no lumps, and no mold. The pH value was measured using a pH meter, and the pH value of the 10% suspension was 6.5, which is within the skin's suitable slightly acidic range. The particle size was measured using a laser particle size analyzer, and more than 90% of the particles were smaller than 50μm, with no particles larger than 180μm, meeting the particle size requirements for creams in the Chinese Pharmacopoeia. Microbial limits were tested for total aerobic bacteria count, total mold and yeast count, and control bacteria, all of which met the microbial limit standards for external preparations in the General Chapter of the Chinese Pharmacopoeia. Stability was tested by centrifugation, and no stratification or demulsification occurred, indicating that the system was stable.

[0075] Example 5

[0076] The preparation of an oil-in-water emulsion cream, based on the preparation of 1000g of finished product, uses the traditional Chinese medicine composition prepared in Example 2, and includes the following steps:

[0077] Prescription ratio: 150g of the external Chinese medicine composition prepared in Example 2, 50g of stearic acid, 40g of white petrolatum, 50g of liquid paraffin, 20g of glyceryl monostearate, 8g of sodium dodecyl sulfate, 25g of Tween-80, 12g of Span-80, 100g of glycerin, 20g of propylene glycol, 10g of azone, 1g of ethylparaben, 1.5g of phenoxyethanol, 0.3g of sodium metabisulfite, 0.2g of vitamin E, and purified water added to 1000g.

[0078] (1): Weigh stearic acid, white petrolatum, liquid paraffin, glyceryl monostearate, and Span-80, put them into an oil phase tank, heat to 85°C, stir until completely melted and mixed, and keep warm at 85°C for later use.

[0079] (2): Weigh out sodium dodecyl sulfate, Tween-80, glycerol, propylene glycol, ethylparaben, and sodium metabisulfite, add purified water, put them into an aqueous phase tank, heat to 85°C, stir until completely dissolved and mixed, and keep warm at 85°C for later use.

[0080] (3): Keep the temperature of the two phases at 80℃, slowly add the oil phase to the water phase while stirring at 500r / min, continue stirring and emulsifying for 15min after the addition is complete, and homogenize at 3000r / min for 5min to obtain a blank cream matrix;

[0081] (4): Cool down to 50°C, add the traditional Chinese medicine composition, azone, phenoxyethanol, and vitamin E, stir for 30 minutes until uniform, and continue to cool down to room temperature to obtain the cream semi-finished product;

[0082] (5): Fill according to specifications, sterilize by cobalt-60 irradiation, and package to obtain the finished cream.

[0083] The finished product was tested and found to be a uniform and delicate milky white cream with a pH of 6.3. Its particle size, microbial limits, and stability all meet the quality requirements for topical preparations.

[0084] Example 6

[0085] The preparation of the ointment, based on the preparation of 1000g of finished product, uses the traditional Chinese medicine composition prepared in Example 1, and includes the following steps:

[0086] Prescription ratio: 100g of the topical Chinese medicine composition prepared in Example 1, 598g of yellow petrolatum, 50g of lanolin, 200g of liquid paraffin, 30g of beeswax, 15g of Span-80, 5g of azone, 1g of ethylparaben, and 1g of vitamin E.

[0087] (1) Weigh out yellow petrolatum, lanolin, liquid paraffin, beeswax and Span-80, place them in a water bath, heat to 80°C, stir until completely melted, filter through a 100-mesh sieve, keep warm at 70°C for later use, and obtain the oil phase matrix;

[0088] (2) Take the Chinese herbal medicine composition, add a small amount of liquid paraffin and grind it into a fine paste for later use;

[0089] (3) Under the stirring conditions of 70℃ and 200r / min, the drug paste was slowly added to the oil phase matrix, and azone, ethylparaben, and vitamin E were added. The mixture was stirred for 30min until it was completely dispersed and uniform. Then, it was slowly stirred and cooled to room temperature to obtain the ointment semi-finished product.

[0090] (4) Fill the ointment into a pharmaceutical tube according to the specifications, sterilize by irradiation, and package to obtain the finished ointment.

[0091] The finished product was tested and found to be a pale yellow, uniform ointment with a fine texture, no rough particles, easy to apply, non-irritating, and meeting the microbial limits.

[0092] Example 7

[0093] The preparation of the gel, based on the preparation of 1000g of finished product, uses the traditional Chinese medicine composition prepared in Example 1, and includes the following steps:

[0094] Prescription ratio: 80g of the topical Chinese medicine composition prepared in Example 1, 10g of carbomer 940, 150g of glycerin, 50g of propylene glycol, 12g of triethanolamine, 6g of azone, 1g of ethylparaben, 50g of ethanol, and purified water to 1000g.

[0095] (1): Weigh out Carbomer 940, sprinkle it into 500g of purified water, let it stand for 24h to fully swell without clumping, and obtain Carbomer swelling solution; under stirring conditions, slowly add triethanolamine, adjust the pH value to 6.0-7.0, stir evenly, and obtain a transparent gel matrix for later use.

[0096] (2): Take the Chinese herbal medicine composition, add glycerin, propylene glycol and ethanol, grind and disperse evenly to obtain a drug dispersion;

[0097] (3): Under stirring conditions, the drug dispersion was slowly added to the gel matrix, azone and ethylparaben were added, purified water was added to the total volume, and stirring was continued for 40 minutes until the system was uniform, fine and free of bubbles, and the gel semi-finished product was obtained.

[0098] (4): Fill, sterilize and package according to specifications to obtain the finished gel.

[0099] The finished product was tested and found to be a semi-transparent gel with a fine texture, good spreadability, no stringing or clumping, a pH value of 6.4, and microbial limits that meet the requirements.

[0100] Example 8

[0101] The preparation of the liniment, based on the preparation of 1000g of finished product, uses the Chinese herbal composition prepared in Example 2, and includes the following steps:

[0102] Prescription ratio: 50g of the external Chinese medicine composition prepared in Example 2, 316ml of ethanol, 103.5ml of propylene glycol, 20g of Tween-80, 5g of azone, 2g of menthol, and purified water to 1000g.

[0103] (1): Take the Chinese medicine composition, add ethanol and propylene glycol, stir for 30 minutes to fully dissolve the active ingredients, filter to remove insoluble impurities, and obtain the drug alcohol solution;

[0104] (2): Add Tween-80, azone, and menthol to the drug alcohol solution, stir until completely dissolved, slowly add purified water to the total volume, stir evenly, let stand for 12 hours, take the supernatant and filter to obtain the liniment semi-finished product;

[0105] (3): Fill the medicine into a pharmaceutical plastic bottle according to the specifications, seal, sterilize and package to obtain the finished liniment.

[0106] The finished product was tested and found to be a clear, brownish-yellow liquid with no sediment or turbidity. The ethanol content met the requirements, and the microbial limits met the requirements for topical preparations.

[0107] Comparative Example 1: Traditional Chinese Medicine Compositions and Creams Prepared Using Conventional Water Decoction Processes

[0108] Preparation of the Chinese herbal medicine composition: Weigh each raw material according to the prescription in Example 1, clean and pulverize it, add 10 times the amount of purified water, soak for 1 hour, heat and decoct twice, 2 hours each time, combine the decoctions, filter, concentrate the filtrate under reduced pressure at 60℃ to a thick extract with a relative density of 1.30, vacuum dry and pulverize to obtain the dry extract powder of Chinese herbal medicine produced by conventional process, and seal for later use.

[0109] Preparation of the cream: The traditional Chinese medicine dry extract powder prepared in this comparative example was used to replace the traditional Chinese medicine composition in Example 4. The other excipient formulations and preparation processes were completely the same as in Example 4. A comparative cream was prepared and prepared for use.

[0110] Comparative Example 2: Traditional Chinese medicine compositions and creams without volatile oil encapsulation

[0111] Preparation of the traditional Chinese medicine composition: The pretreatment and steam distillation were completed according to the process of Example 1. The collected volatile oil was not encapsulated and was sealed and refrigerated for later use. The residue was extracted with a dual solvent gradient, concentrated and purified by alcohol precipitation according to the process of Example 1 to obtain the dry extract powder of traditional Chinese medicine. Finally, the volatile oil was directly mixed with the dry extract powder to obtain the traditional Chinese medicine composition of this comparative example, which was then sealed for later use.

[0112] Preparation of cream: The traditional Chinese medicine composition prepared in this comparative example was used to replace the traditional Chinese medicine composition in Example 4. The other excipient formulations and preparation processes were completely the same as in Example 4 to obtain a comparative cream for later use.

[0113] Comparative Example 3: Traditional Chinese Medicine Compositions and Creams Without Artificial Phellodendron Bark

[0114] Preparation of the traditional Chinese medicine composition: The prescription ratio is 15 parts Angelica sinensis, 10 parts Ligusticum striatum, 25 parts Lycopodium clavatum, 15 parts Artemisia argyi, 30 parts Clematis chinensis, and 10 parts Angelica dahurica. The remaining preparation process and parameters are completely the same as in Example 1. The traditional Chinese medicine composition of this comparative example is prepared and sealed for later use.

[0115] Preparation of the cream: The traditional Chinese medicine composition prepared in this comparative example was used to replace the traditional Chinese medicine composition in Example 4. The other excipient formulations and preparation processes were completely the same as in Example 4. A comparative cream was prepared and prepared for use.

[0116] Comparative Example 4: Commercially available positive control

[0117] We selected mainstream commercially available topical analgesics as controls, specifically: Blood Circulation and Analgesia Plaster and Musk Medicated Oil, both of which were purchased from legitimate commercial channels and stored at room temperature for future use.

[0118] Experiment Example 1: Comparison Experiment of Extraction Rate of Core Active Ingredients

[0119] This experiment used high performance liquid chromatography (HPLC) to detect the extraction rates of four core pharmacological components in Example 1 and Comparative Examples 1 and 2 of this invention.

[0120] Chromatographic conditions: Agilent C18 column (4.6 mm × 250 mm, 5 μm); column temperature 30 ℃; detection wavelengths: ferulic acid 316 nm, ligustilide 330 nm, imperatorin 300 nm, berberine hydrochloride 265 nm; mobile phase: acetonitrile-0.1% phosphoric acid aqueous solution, gradient elution; flow rate 1.0 ml / min; injection volume 10 μl.

[0121] Preparation of reference solutions: Accurately weigh appropriate amounts of ferulic acid, ligustilide, imperatorin, and berberine hydrochloride reference standards, dissolve them in methanol and dilute to volume to prepare mixed reference solutions with concentrations of 20 μg / ml, 50 μg / ml, 30 μg / ml, and 40 μg / ml, respectively. Seal and refrigerate for later use.

[0122] Preparation of test solution

[0123] (1) Sample of Example 1: Accurately weigh 0.5g of the traditional Chinese medicine composition prepared in Example 1, place it in a stoppered conical flask, accurately add 25ml of 70% methanol, weigh it, sonicate for 30min, cool it, weigh it again, replenish the lost weight with 70% methanol, shake it well, filter it, take the filtrate, filter it through a 0.22μm organic phase filter membrane, and the sample is obtained.

[0124] (2) Comparative Examples 1 and 2: Prepared by the same method as above.

[0125] Determination method: Accurately pipette the mixed reference solution and each test solution into the high performance liquid chromatograph, determine the peak area under the above chromatographic conditions, calculate the content of each active ingredient by the external standard method, and calculate the extraction rate based on the theoretical total amount of the corresponding ingredient in the raw material. The calculation formula is: Extraction rate (%) = (Total amount of active ingredient in the extract / Theoretical total amount of active ingredient in the raw material) × 100%.

[0126] Each experiment was performed in parallel three times, and the results are expressed as mean ± standard deviation. The experimental results are shown in Table 1.

[0127] Table 1. Comparison of extraction rates of core effective components in different groups (n=3, %)

[0128]

[0129] Experimental conclusions

[0130] The dual-solvent gradient extraction process of Embodiment 1 of this invention improves the extraction rate of four core active ingredients compared with the conventional water decoction process of the prior art. The extraction rate of lipid-soluble ingredients ligustilide and imperatorin is increased by more than 2 times, and the extraction rate of water-soluble ingredients ferulic acid and berberine hydrochloride is increased by more than 20%. This fully demonstrates that the process of this invention can maximize the extraction of active ingredients with different polarities and greatly improve the utilization rate of raw materials.

[0131] In Comparative Example 2, the extraction rate of ligustilide was lower than that in Example 1, demonstrating that the volatile oil encapsulation process of the present invention can effectively reduce the loss of volatile components during distillation, concentration, and drying, and improve the retention rate of volatile active ingredients, thus providing a material basis for the analgesic effect of the formulation.

[0132] Experiment Example 2: Analgesic Effect Verification Experiment

[0133] This experiment used two classic analgesic models, the internationally recognized hot plate test and the acetic acid writhing test, to compare the analgesic effects of the embodiments of this invention with those of comparative examples.

[0134] Experiment 1, Hot Plate Analgesia Test in Mice

[0135] laboratory animals

[0136] SPF-grade female Kunming mice, weighing 18-22g, were housed in an SPF-grade barrier system at a temperature of 22±2℃ and a relative humidity of 50±10%, with a 12-hour light-dark cycle and free access to food and water. The experiments were conducted after 3 days of acclimatization.

[0137] Experimental instruments

[0138] The YLS-6B intelligent hot plate heat exchanger has its hot plate temperature pre-set to (55.0±0.2)℃ and preheated for 30 minutes to ensure temperature stability.

[0139] Animal screening

[0140] Female mice were placed one by one on a hot plate and the time from when the mouse was placed to when it began to lick its hind paw was recorded as the baseline pain threshold. Mice with a baseline pain threshold between 5 and 30 seconds were selected, and mice with a pain threshold <5 seconds, >30 seconds, or jumping were removed. A total of 70 qualified mice were selected.

[0141] The selected mice were randomly divided into 7 groups of 10 mice each. The blank control group was coated with a blank cream base without any Chinese medicine ingredients; the first group of examples was coated with the cream prepared in Example 4; the first comparative example group was coated with the cream prepared in Comparative Example 1; the second comparative example group was coated with the cream prepared in Comparative Example 2; the third comparative example group was coated with the cream prepared in Comparative Example 3; the fourth comparative example group was coated with the contents of a commercially available blood-activating and pain-relieving ointment; and the positive control group was coated with diclofenac diethylamine emulsion.

[0142] Administration method

[0143] Hair was removed from the back of the hind paws of mice using a depilatory agent. The area of ​​hair removal was approximately 2cm × 2cm. After 24 hours, it was confirmed that there was no skin damage or irritation. The corresponding test substance was evenly applied to the hair removal area of ​​each group of mice at a dose of 0.2g / mouse. After application, the mice were gently massaged for 10 seconds to promote absorption. After administration, the mice were allowed to move freely.

[0144] Mice were placed on a hot plate at 30 min, 60 min, 120 min, 240 min and 360 min after administration, and the time it took for the mice to lick their hind paws was recorded as the pain threshold. If the mice did not lick their hind paws within 60 seconds, they were immediately removed and the pain threshold was calculated based on 60 seconds to avoid burns.

[0145] Experimental data are expressed as mean ± standard deviation (x ± s). Data analysis was performed using SPSS 26.0 statistical software. One-way ANOVA was used for comparisons between groups. P < 0.05 was considered statistically significant, and P < 0.01 was considered extremely statistically significant. The pain threshold improvement rate at different time points in each group was calculated using the following formula: Pain threshold improvement rate (%) = (Pain threshold after drug administration - Baseline pain threshold) / Baseline pain threshold × 100%.

[0146] The results of pain threshold changes at different time points in each group of mice are shown in Table 2, and the core results of pain threshold improvement rate are shown in Table 3.

[0147] Table 2 Comparison of pain thresholds at different time points in each group of mice (n=10, s, x±s)

[0148] Note: Compared with the blank control group, *P<0.05, **P<0.01; compared with the comparative example group, ##P<0.01.

[0149]

[0150] Table 3 Comparison of pain threshold increase rates at different time points in each group of mice (n=10, %)

[0151]

[0152] Experimental conclusions

[0153] In the embodiment of the present invention, analgesic effect appeared 30 minutes after administration, with a pain threshold increase rate of 68.92%, which was better than the blank control group (P<0.01) and the comparative example group (P<0.01). The onset time was less than 30 minutes, which was much faster than the conventional process group, the commercially available product group and the positive drug control group, thus achieving rapid analgesia.

[0154] In Example 1, the analgesic effect reached its peak 120 minutes after administration, with a pain threshold increase rate of 125.63%, and the analgesic intensity was higher than that of all control groups. Six hours after administration, the pain threshold increase rate remained at 82.35%, and the duration of efficacy exceeded 6 hours, which was much longer than that of Comparative Example 2, the commercially available product group, and the positive drug control group.

[0155] The analgesic effect of the three comparative groups was no different from that of the first example group, proving that the counteracting effect of artificial Phellodendron bark does not affect the core analgesic effect.

[0156] Experiment 2, Mouse Acetic Acid Writhing Test for Analgesia

[0157] laboratory animals

[0158] SPF-grade Kunming mice, half male and half female, weighing 18-22g, were obtained and raised under the same conditions as the hot plate test described above.

[0159] Grouping and Dosing

[0160] Mice were randomly divided into 7 groups of 10 each, and the grouping was the same as the hot plate test. The drug was administered to the hair-removed area on the abdomen of the mice, with a hair removal area of ​​about 3cm × 3cm. The dosage was 0.3g / mouse, administered twice a day for 3 consecutive days. Modeling was performed 1 hour after the last administration.

[0161] One hour after the last administration, mice in each group were injected intraperitoneally with 0.6% glacial acetic acid solution at a dose of 0.2 ml per mouse. Immediately after the injection, the mice were placed in observation cages, and the number of writhing movements observed in each mouse within 15 minutes after the intraperitoneal injection was recorded.

[0162] Experimental data are expressed as mean ± standard deviation. One-way ANOVA was performed between groups using SPSS 26.0 software to calculate the analgesic inhibition rate of each group. The calculation formula is: Analgesic inhibition rate (%) = (Average number of writhing movements in the blank control group - Average number of writhing movements in the drug treatment group) / Average number of writhing movements in the blank control group × 100%. The experimental results are shown in Table 4.

[0163] Table 4 Comparison of writhing count and analgesic inhibition rate in mice within 15 minutes in each group (n=10, x±s)

[0164] Note: Compared with the blank control group, *P<0.05, **P<0.01; compared with the comparative example group, ##P<0.01.

[0165]

[0166] Experimental conclusions

[0167] The results of this invention showed that the number of acetic acid writhing episodes in mice was reduced (P<0.01), with an analgesic inhibition rate as high as 75.77%, which was superior to the conventional process group, the commercially available product group, and the positive drug control group. This further proves that the traditional Chinese medicine composition and preparation process of this invention can achieve a strong peripheral analgesic effect and has an excellent relieving effect on pain caused by strain and inflammation.

[0168] Experiment Example 3: Anti-inflammatory Effect Verification Experiment

[0169] laboratory animals

[0170] SPF-grade male Kunming mice, weighing 20-25g, were sourced and raised under the same conditions as those described in the previous experiment.

[0171] Grouping and Dosing

[0172] Mice were randomly divided into 7 groups of 10 each, and the grouping was the same as in the analgesia experiment. The administration site was both sides of the left and right ears of the mice, and the dosage was 0.1g / ear. The mice were given twice a day for 3 consecutive days. The model was established 1 hour after the last administration.

[0173] One hour after the last administration, xylene was evenly applied to both sides of the right ear of each group of mice at a dose of 50 μl / mouse. The left ear was left untreated as a self-control. Thirty minutes after inflammation, the mice were euthanized by cervical dislocation. Circular ear patches were punched at the same location on both ears of the mice using an 8 mm diameter punch. The ear patches were accurately weighed using an electronic analytical balance, and the weight of the left and right ear patches of each mouse was recorded. The degree of ear swelling and the swelling inhibition rate were calculated.

[0174] Calculation formula

[0175] Ear swelling (mg) = Weight of right earpiece - Weight of left earpiece

[0176] Swelling inhibition rate (%) = (mean ear swelling in the blank control group - mean ear swelling in the treatment group) / mean ear swelling in the blank control group × 100%

[0177] Experimental data are expressed as mean ± standard deviation. One-way ANOVA was performed using SPSS 26.0 software. The t-test was used for comparisons between groups, and P < 0.05 was considered statistically significant. The experimental results are shown in Table 5.

[0178] Table 5 Comparison of ear swelling degree and swelling inhibition rate in mice of different groups (n=10, x±s)

[0179] Note: Compared with the blank control group, *P<0.05, **P<0.01; compared with the comparative example group, ##P<0.01.

[0180]

[0181] Experimental conclusions

[0182] In one embodiment of the present invention, xylene-induced ear swelling in mice was inhibited (P<0.01), with a swelling inhibition rate as high as 77.16%, which is superior to the conventional process group, the commercially available product group, and the positive drug control group. This demonstrates that the traditional Chinese medicine composition of the present invention has excellent anti-inflammatory and swelling-reducing effects, and can effectively inhibit the local acute inflammatory response accompanying pain. It achieves a holistic therapeutic effect on aseptic inflammation caused by wind-cold-dampness, strain, and traumatic injuries, rather than simply providing anesthesia and pain relief.

[0183] Experiment Example 4: In Vitro Transdermal Absorption Experiment

[0184] laboratory animals

[0185] SPF grade male SD rats, weighing 200-220g.

[0186] preparation of ex vivo skin

[0187] Rats were euthanized by cervical dislocation. Abdominal hair was removed with a depilatory agent. The intact abdominal skin was carefully peeled off, and subcutaneous fat tissue and fascia were removed. The skin was rinsed repeatedly with physiological saline. The skin was checked for any damage. The rats were cut into appropriate sizes and stored frozen at -20°C. They should be used within one week. Before use, the rats should be thawed to room temperature and rinsed with physiological saline.

[0188] Experimental instruments and reagents

[0189] An improved Franz vertical diffusion cell with an effective diffusion area of ​​2.8 cm². 2The receiving pool has a volume of 15 ml; a constant temperature circulation system controls the temperature at (37±0.5) ℃; a magnetic stirrer is used with a stirring speed of 300 r / min; the receiving liquid is a 20% ethanol-physiological saline solution, which is degassed before use.

[0190] Experimental Grouping and Methods

[0191] The experiment was set up in 4 groups: Example 1 (Example 4 cream), Comparative Example 1, Comparative Example 2, and Comparative Example 4, with 6 parallel diffusion cells in each group.

[0192] Thawed rat skin was fixed between the supply and receiving pools of the diffusion chamber, with the stratum corneum facing the supply pool and the dermis facing the receiving pool. 15 ml of receiving solution preheated to 37°C was added to the receiving pool, and air bubbles were removed to ensure close contact between the skin and the receiving solution. The constant temperature circulation and magnetic stirring were turned on and the mixture was equilibrated for 30 min. The test substance for each group was added to the supply pool at a dosage of 0.5 g / pool, and evenly spread on the surface of the stratum corneum of the skin. The supply pool was then sealed to prevent solvent evaporation.

[0193] At 1h, 2h, 4h, 6h and 8h after administration, 2ml samples were taken from the receiving pool and an equal volume of fresh receiving liquid preheated to 37℃ was added immediately. The samples were then filtered through a 0.22μm filter membrane immediately after sampling, and the concentrations of ferulic acid and imperatorin in the samples were detected by HPLC under the same chromatographic conditions as in Experiment 1.

[0194] Based on the detection concentration, receiving cell volume, and dilution factor, the cumulative transmittance at different time points was calculated. The cumulative transmittance per unit area and average transmittance for each group over 8 hours are shown in Table 6.

[0195] Table 6 Comparison of transdermal absorption results of active ingredients in each group after 8 hours (n=6, x±s)

[0196]

[0197] Experimental conclusions

[0198] The cumulative permeation amount and average permeation rate of the core active ingredient in Example 1 of this invention are higher than those in Example 1 over 8 hours, with the permeation rate increasing by more than 3 times. This proves that the alcohol precipitation and refining process of this invention removes a large number of macromolecular ineffective impurities that affect transdermal absorption, reduces skin barrier resistance, and facilitates the transdermal absorption of active ingredients.

[0199] The transdermal penetration rate of Example 1 was higher than that of Comparative Example 2 and the commercially available product group. It can effectively open the lipid channels of the stratum corneum of the skin, improve the transdermal absorption efficiency of the active ingredients, and allow more active ingredients to reach the subcutaneous lesion site.

[0200] Experiment Example 5: Skin Irritation and Safety Test

[0201] Experiment 1: Skin irritation test of rabbits after repeated administration

[0202] laboratory animals

[0203] SPF grade New Zealand White rabbits, half male and half female, weighing 2.0-2.5kg; animals are housed in a barrier environment with a temperature of 22±2℃ and a relative humidity of 50±10%, and are allowed to adapt for 3 days.

[0204] Skin preparation

[0205] 24 hours before the experiment, hair was removed from both sides of the spine on the back of the rabbits using a hair removal agent, with two areas on each side. Each area was approximately 3cm x 3cm. The skin was examined 24 hours after the hair removal to confirm that there were no abnormalities such as damage, erythema, or edema.

[0206] Experimental grouping and drug administration

[0207] The study employed a self-controlled design with left and right sides of the rabbit. Each rabbit was divided into four areas: an intact skin administration area, an intact skin control area, a damaged skin administration area, and a damaged skin control area. A total of six rabbits were included.

[0208] In the area to be treated with hair removal, use a sterilized needle to draw a "#" shape on the skin surface until it bleeds, creating a skin damage model to ensure that the skin barrier is broken;

[0209] The ointment prepared in Example 4 was applied to the treatment area at a dose of 0.5 g / area. An equal amount of blank ointment base was applied to the control area. The treatment was administered once a day for 7 consecutive days. 24 hours after each administration, the residual test substance was cleaned with warm water, and the skin reaction was observed and recorded.

[0210] One hour after each application site was cleaned, erythema and edema were scored according to the skin irritation reaction scoring criteria in the "Cosmetic Safety Technical Specifications" (2015 edition). The scoring criteria are shown in Table 7. After the last application, the recovery of skin reaction and delayed irritation reaction were observed at 1 hour, 24 hours, 48 ​​hours and 72 hours after cleansing.

[0211] Calculate the average stimulus response score for each rabbit per day, as well as the total average score for each group. Determine the skin irritation level of the test substance according to the stimulus intensity evaluation criteria in Table 8.

[0212] Table 7. Scoring Criteria for Skin Irritation Response

[0213]

[0214] Table 8. Evaluation Criteria for Skin Irritation Intensity

[0215]

[0216] Experimental results

[0217] During the 7-day administration period, no erythema or edema was observed in the intact skin treatment areas of all rabbits at any of the observation time points, and the average daily stimulation response score was 0. In the damaged skin treatment areas, only one rabbit showed very slight erythema with a score of 1 point 1 hour after the first administration, which completely subsided after 24 hours. All other rabbits showed no erythema or edema at any of the observation time points. Throughout the entire administration period, the average stimulation response score of the damaged skin treatment areas was 0.02, which was far below 0.5. 72 hours after the last administration, there were no abnormalities in the skin of all treatment areas, and no delayed reactions such as pigmentation, rough skin, or keratinization.

[0218] Therefore, the cream prepared by this invention is non-irritating to both intact and damaged skin of rabbits, has extremely high skin safety, and will not cause skin irritation with long-term use.

[0219] Experiment 2: Stimulus Comparison Verification of Co-option Design

[0220] Three comparative groups were set up simultaneously, using the same rabbit skin irritation test method as described above. Six rabbits were treated with the ointment prepared in comparative group three, and all other conditions were the same.

[0221] Experimental results

[0222] Two rabbits with intact skin showed mild erythema on the third day of administration, with an average score of 1.2, indicating mild irritation. All rabbits with damaged skin showed erythema of varying degrees, with two rabbits showing mild edema. The average irritation response score throughout the administration period was 2.1, indicating mild irritation, with the highest single score reaching 3, indicating moderate irritation. 72 hours after the last administration, three rabbits still had mild erythema in their damaged skin areas, indicating a significant delayed irritation response.

[0223] Therefore, this invention, through the counteracting design of artificial Phellodendron bark, can limit the skin irritation of warm and drying drugs in the formula, especially the irritation of broken skin.

[0224] Experiment 3: Guinea Pig Skin Allergy Experiment

[0225] laboratory animals

[0226] SPF-grade albino guinea pigs, half male and half female, weighing 250-300g; acclimatize for 3 days, remove hair from both sides of the back 24 hours before the experiment, the hair removal area is about 3cm×3cm, and the skin is checked for no damage.

[0227] Experimental Groups

[0228] The guinea pigs were randomly divided into 3 groups of 10 each, with half males and half females:

[0229] The test group used the cream prepared in Example 4; the negative control group used a blank cream base; and the positive control group used 2,4-dinitrochlorobenzene (DNCB) prepared at a sensitizing concentration of 1% and a challenge concentration of 0.1%.

[0230] The experimental method followed the skin allergy test method in the "Cosmetic Safety Technical Specifications" (2015 edition), which was divided into sensitization stage and provocation stage.

[0231] Sensitive Contact

[0232] The corresponding test substance was applied to the hairless area on the left side of the guinea pig at a dose of 0.2g / animal. The positive control group was treated with 0.2ml of 1% DNCB solution, covered with gauze and fixed for 6h. The sensitization was repeated once on the 7th and 14th days, for a total of 3 sensitizations. The negative control group was given blank matrix at the same time.

[0233] Stimulate contact

[0234] Fourteen days after the last sensitization, 0.2 g of the corresponding test substance was applied to the hairless area on the right side of the guinea pig, while 0.2 ml of 0.1% DNCB solution was applied to the positive control group. After 6 hours of fixation, the residual test substance was cleaned with warm water. The skin allergy reaction was observed at 24 hours and 48 hours after cleaning and scored according to the skin allergy reaction scoring criteria in Table 9.

[0235] Record the number of animals in each group that develop skin erythema and edema, and calculate the sensitization rate: sensitization rate = (number of animals with positive reactions / total number of animals in the group) × 100%; determine the sensitization level of the test substance according to the sensitization intensity grading standard in Table 10.

[0236] Table 9. Scoring Criteria for Skin Allergic Reactions

[0237]

[0238] Table 10 Sensitization Intensity Grading Standards

[0239]

[0240] Experimental results

[0241] No allergic reactions such as erythema or edema were observed on the skin of the tested group and the negative control group at 24 h and 48 h after stimulation. The allergic reaction scores were all 0, and the sensitization rate was 0. In contrast, all 10 guinea pigs in the positive control group showed obvious erythema and edema reactions. The average score at 24 h was 3.2, and the sensitization rate was 100%, which is considered to be extremely sensitizing.

[0242] Therefore, the cream prepared by this invention is non-allergenic to guinea pig skin, will not cause skin allergies, has extremely high safety for external use, and is suitable for long-term and repeated use.

[0243] Experimental Example 6: Accelerated Stability Test of Formulation

[0244] Experimental samples

[0245] The cream prepared in Example 4 was packaged in an aluminum-plastic composite tube with a specification of 20g / tube; two comparative groups of creams were set up as controls.

[0246] Experimental conditions

[0247] A constant temperature and humidity chamber was set up with a temperature of (40±2)℃ and a relative humidity of (75±5)%. The samples were placed in the chamber for 6 consecutive months. Samples were taken at 0 months, 1 month, 2 months, 3 months, and 6 months, and the following key indicators were measured:

[0248] (1) Properties: Observe the appearance, color, and fineness of the cream, and whether there are any phenomena such as oil-water separation, layering, clumping, or mold.

[0249] (2) pH value: The pH value change of the 10% suspension was measured;

[0250] (3) Particle size: Check for particles larger than 180 μm;

[0251] (4) Content of active ingredients: The contents of ferulic acid and ligustilide were determined by HPLC, and the content retention rate was calculated (content retention rate = content at the detection time point / initial content at 0 months × 100%).

[0252] (5) Microbial limits: The total number of aerobic bacteria, total number of molds and yeasts, and control bacteria were checked and found to meet the standards for topical preparations. The stability results of Example 1 are shown in Table 11.

[0253] Table 11 Results of accelerated 6-month stability study of the cream in Example 4

[0254]

[0255] The stability results of the second comparative group are shown in Table 12.

[0256] Table 12 Results of accelerated stability study of Comparative Example 2 cream after 6 months

[0257]

[0258] Experimental conclusions

[0259] The cream prepared in Example 4 of this invention, after being placed for 6 months in an accelerated stability test at 40°C, showed no significant changes in appearance, pH value, or particle size, and exhibited no stratification, demulsification, or discoloration. The retention rates of the core active ingredients, ferulic acid and ligustilide, were both above 90%, and the microbial limits consistently met the requirements. The formulation exhibits excellent stability and can ensure stable product quality and no reduction in efficacy within a 24-month shelf life.

[0260] In the second comparative example, significant changes in appearance and degradation of active ingredients were observed after 3 months of accelerated testing, and the retention rate of ligustilide was less than 40% after 6 months. This demonstrates that the β-cyclodextrin inclusion process of the volatile oil in this invention can improve the stability of volatile components and reduce oxidative degradation and volatilization loss.

[0261] Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A topical traditional Chinese medicine composition for relieving pain, characterized in that, It is made from the following raw materials in parts by weight: Angelica sinensis 5-25 parts, Ligusticum striatum 5-20 parts, Lycopodium clavatum 10-35 parts, Artemisia argyi 5-25 parts, artificial Phellodendron chinense 3-18 parts, Clematis chinensis 10-40 parts, Angelica dahurica 5-20 parts.

2. The topical traditional Chinese medicine composition according to claim 1, characterized in that, It is made from the following raw materials in parts by weight: Angelica sinensis 10-20 parts, Ligusticum striatum 8-15 parts, Lycopodium clavatum 15-30 parts, Artemisia argyi 10-20 parts, artificial Phellodendron chinense 5-12 parts, Clematis chinensis 15-35 parts, Angelica dahurica 8-15 parts.

3. The external application traditional Chinese medicine composition according to claim 1, characterized in that, It is made from the following raw materials in parts by weight: Angelica sinensis 15 parts, Ligusticum striatum 10 parts, Lycopodium clavatum 25 parts, Artemisia argyi 15 parts, artificial Phellodendron chinense 8 parts, Clematis chinensis 30 parts, Angelica dahurica 10 parts.

4. A method for preparing the topical Chinese medicine composition for relieving pain according to any one of claims 1-3, characterized in that, Includes the following steps: Step S1: Weigh each raw material according to the formula, clean and crush it through a 20-40 mesh sieve, mix it, add 8-12 times the total weight of the raw materials in purified water and soak for 30-60 minutes, extract the volatile oil by steam distillation for 3-5 hours, collect the volatile oil and the water after distillation separately, and keep the residue for later use. Step S2: Add 6-10 times the weight of the volatile oil obtained in Step S1 to anhydrous ethanol and stir to dissolve to obtain a volatile oil ethanol solution; separately take 8-12 times the weight of β-cyclodextrin and add 8-10 times the weight of purified water, and stir at 40-50℃ until completely dissolved to obtain a β-cyclodextrin saturated aqueous solution; under constant temperature of 40-50℃ and stirring at 200-300 r / min, slowly add the volatile oil ethanol solution dropwise to the β-cyclodextrin saturated aqueous solution. After the addition is complete, continue stirring at constant temperature for 1-2 h, then place in a refrigerated environment of 2-8℃ and let stand for 12-24 h, filter, wash the filter cake and dry it under vacuum below 40℃ to obtain the volatile oil β-cyclodextrin inclusion complex; Step S3: Take the residue left from step S1, add 6-10 times the weight of the residue and 50%-70% ethanol aqueous solution, heat and reflux to extract 1-2 times, 1-2 hours each time, and filter to obtain ethanol extract; then add 6-8 times the weight of the residue and purified water to the residue after ethanol extraction, heat and decoct to extract 1-2 times, 1-1.5 hours each time, and filter to obtain aqueous extract; Step S4: Combine the distilled aqueous solution from Step S1, the ethanol extract from Step S3, and the aqueous extract. Concentrate under reduced pressure at a vacuum of -0.06 to -0.08 MPa and a temperature of 50-60℃ to obtain a clear extract with a relative density of 1.10-1.20 measured at 60℃. Add edible ethanol to the clear extract and stir until the alcohol content reaches 60-70% by volume. After stirring evenly, refrigerate and let stand at 4℃ for 12-24 hours. Filter the supernatant, recover the ethanol from the filtrate under reduced pressure, and continue to concentrate to a thick extract with a relative density of 1.25-1.35 measured at 60℃. Dry the thick extract under vacuum, pulverize it through an 80-mesh sieve, and obtain the dry extract powder of traditional Chinese medicine. Step S5: Mix the volatile oil β-cyclodextrin inclusion complex obtained in step S2 with the dry extract powder of traditional Chinese medicine obtained in step S4 to obtain the topical traditional Chinese medicine composition for relieving pain.

5. A topical preparation for relieving pain, characterized in that, The product comprises any one of the topical Chinese medicine compositions according to claims 1-3, or the topical Chinese medicine composition prepared by the preparation method according to claim 4, and pharmaceutically or health care product-acceptable excipients.

6. The topical preparation according to claim 5, characterized in that, The dosage form of the topical preparation is a cream, ointment, gel, liniment, lotion, or plaster.

7. The topical preparation according to claim 6, characterized in that, The topical preparation is an oil-in-water emulsion cream, comprising the following components per 1000 parts by weight: 50-200 parts of a topical traditional Chinese medicine composition, 100-200 parts of a composite oil phase matrix, 30-80 parts of a composite emulsifier, 50-150 parts of a moisturizing and penetration-enhancing agent, 5-20 parts of a transdermal penetration enhancer, 1-7 parts of an antiseptic and antioxidant system, and purified water to 1000 parts. The composite oil phase matrix includes stearic acid, white petrolatum, liquid paraffin, and glyceryl monostearate; the composite emulsifier includes sodium lauryl sulfate, Tween-80, and Span-80; the moisturizing and penetration-enhancing agent includes glycerin and propylene glycol; the transdermal penetration enhancer includes azone; and the antiseptic and antioxidant system includes ethylparaben, phenoxyethanol, sodium metabisulfite, and vitamin E.

8. The topical preparation according to claim 7, characterized in that, Based on 1000 parts by weight, it comprises the following components: 100 parts of external Chinese medicine composition, 60 parts of stearic acid, 30 parts of white petrolatum, 40 parts of liquid paraffin, 30 parts of glyceryl monostearate, 10 parts of sodium lauryl sulfate, 30 parts of Tween-80, 10 parts of Span-80, 80 parts of glycerin, 30 parts of propylene glycol, 8 parts of azone, 1 part of ethylparaben, 2 parts of phenoxyethanol, 0.5 parts of sodium metabisulfite, 0.5 parts of vitamin E, and purified water to 1000 parts.

9. A method for preparing the topical formulation according to any one of claims 7-8, characterized in that, Includes the following steps: (1): Weigh the composite oil phase matrix and oil-soluble emulsifier components, put them into the oil phase tank, heat to 75-85℃, stir until completely melted and mixed, keep warm for later use, and obtain the oil phase; (2): Weigh the water-soluble emulsifier component, humectant, and water-soluble preservative and antioxidant component, add purified water, put them into an aqueous phase tank, heat to 75-85℃, stir until completely dissolved and mixed, keep warm for later use, and obtain the aqueous phase; (3): Under the conditions of keeping at 75-80℃ and stirring at 300-500r / min, the oil phase is slowly added to the water phase. After the addition is complete, continue stirring and emulsifying for 15-30min, and then homogenize for 5-10min to obtain a blank cream matrix. (4): Cool the blank cream base to 45-50℃, add the topical Chinese medicine composition, transdermal penetration enhancer, and remaining antiseptic and antioxidant components, stir and mix well, continue stirring and cooling to room temperature to obtain the cream semi-finished product; (5): Fill and sterilize the semi-finished product to obtain the finished cream.

10. The application of the topical Chinese medicine composition according to any one of claims 1-3, or the topical Chinese medicine composition prepared by the preparation method according to claim 4, or the topical preparation according to any one of claims 5-8 in the preparation of a topical product for relieving pain; wherein the topical product for relieving pain is a medicine or health care product for relieving pain and discomfort in the cervical spine, shoulder, lumbar spine and joints caused by wind-cold-dampness, strain, or sprains.

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