A Chinese medicine composition for external use for treating dysmenorrhea and its preparation method and use

Abstract

The present invention provides a Chinese medicine composition for external use for treating dysmenorrhea, and a preparation method and use thereof, and belongs to the field of medicine. The present invention provides a Chinese medicine composition for external use for treating dysmenorrhea, which is a preparation prepared from the following raw materials in weight ratio: 40 parts of tetrahydropalmatine, angelica and chuanxiong volatile oil, based on the original medicinal materials of angelica and chuanxiong: 4000 parts, wherein the dosage ratio of angelica and chuanxiong is: 1:1. The present invention also provides a preparation method and use of the Chinese medicine composition for external use. The Chinese medicine composition for external use of the present invention is prepared from tetrahydropalmatine, angelica and chuanxiong volatile oil. The efficacy test proves that the efficacy of the Chinese medicine composition for external use is obvious in treating dysmenorrhea, and the raw materials can play a synergistic role, reducing the clinical dosage, reducing the cost of medication, and directly administering the drug for external use. The clinical use is more convenient and safe, and provides a new choice for the clinical treatment of dysmenorrhea.

Description

Technical Field

[0001] The invention relates to an external Chinese medicine composition for treating dysmenorrhea and a preparation method and application thereof, belonging to the field of medicines. Background Art

[0002] Dysmenorrhea refers to pain and distension in the lower abdomen before, during, or after menstruation, accompanied by backache or other discomfort. According to the unique physiological structure of women, dysmenorrhea is divided into two categories, namely primary dysmenorrhea and secondary dysmenorrhea. Among them, those who exclude organic lesions of the reproductive organs are called primary dysmenorrhea, and the incidence rate among women worldwide is about 50%-90%. In my country, the incidence rate of primary dysmenorrhea is as high as 74.03%, accounting for more than 90% of dysmenorrhea. With the current prevalence of irregular life, unhealthy diet, lack of sleep and other conditions, the incidence rate of dysmenorrhea is getting higher and higher. Long-term untreated dysmenorrhea will lead to aggravated pain, causing endometritis, endometriosis, etc., and even neuroimaging studies in recent years have found that patients with primary dysmenorrhea have abnormal brain function. Therefore, dysmenorrhea has always been one of the diseases that the obstetrics and gynecology community pays attention to and studies.

[0003] Western medicine believes that the pathophysiological process of primary dysmenorrhea is the production of prostaglandins (PGs) related to inflammation and pain by the cyclooxygenase (COX) pathway. Among them, PGF2a and PGE2 are the main culprits of primary dysmenorrhea. PGF2a not only causes uterine contraction and thus restricts blood flow, but also directly participates in arcuate vasoconstriction. Both mechanisms of action produce hypoxia, resulting in the accumulation of anaerobic metabolites that stimulate pain receptors. PGF2a also lowers the threshold of pain perception by sensitizing these neuroreceptors. Prostaglandin E2 has a dual mechanism of action that depends on receptor interaction, which can cause uterine muscle contraction or relaxation and uterine vasoconstriction or dilation. In addition, PGs may be involved in the formation of other chemokines and growth factors, and may also increase the migration of neutrophils and leukocytes to the endometrium. The peak time of dysmenorrhea is associated with the highest level of PGs, accompanied by gastrointestinal symptoms caused by PGs. Therefore, from the perspective of Western medicine, the current treatment of primary dysmenorrhea aims to exert analgesic effects by inhibiting the production of PGs. According to traditional Chinese medicine, dysmenorrhea is mostly caused by qi and blood, cold evil, and damp evil blocking the uterus, and pain occurs when it is blocked; or it is caused by insufficient qi and blood, liver and kidney deficiency, which leads to the emptiness of Chong and Ren meridians, and the uterus is not nourished, and pain occurs when it is not nourished. Dysmenorrhea is generally divided into five types: qi stagnation and blood stasis type, cold stagnation uterus type, damp-heat accumulation type, qi and blood deficiency type, and liver and kidney deficiency type. The clinical syndrome type is mainly cold stagnation and blood stasis type. Therefore, traditional Chinese medicine often uses prescriptions for nourishing blood and regulating menstruation, promoting qi and activating blood circulation, and dispersing blood stasis and relieving pain to treat dysmenorrhea, but Chinese medicine prescriptions are more cumbersome in use and processing. At present, there are also auxiliary therapies for the treatment of dysmenorrhea, such as Chinese medicine navel therapy, acupoint application, essential oil massage therapy, moxibustion, Chinese medicine foot bath, etc., but most of their effectiveness has not been scientifically explained. Therefore, the current first-line drugs for the clinical treatment of primary dysmenorrhea are hormonal contraception and non-steroidal anti-inflammatory drugs. But their use is limited by side effects, such as stomach irritation or ulcers in some women taking NSAIDs, and cardiovascular, liver, and kidney problems with long-term use. Similarly, oral contraceptives are associated with increased bleeding frequency, weight gain, or patients' underlying risk of venous thromboembolism.

[0004] Chuanxiong is a plant of the Umbelliferae family. Ligusticum chuanxiong Hort.'s dried root is one of the most important and commonly used blood-activating and stasis-removing drugs in TCM clinical practice. Angelica sinensis (Oliv.) Diels' dried root is a traditional TCM drug for treating gynecological diseases. It is mainly used in clinical practice to replenish blood, activate blood circulation, regulate menstruation and relieve pain.

[0005] Application number: CN200610078969.6, invention name: A Chinese medicine composition for external use for treating dysmenorrhea and its preparation method. A Chinese medicine composition for external use for treating dysmenorrhea and its preparation process are disclosed. The composition is mainly made of angelica volatile oil and total alkaloids of Corydalis yanhusuo, wherein the angelica volatile oil is determined by gas chromatography, and the content is greater than 20% in terms of ligustilide; the total alkaloids of Corydalis yanhusuo are determined by ultraviolet spectrophotometry, and the content is greater than 50% in terms of tetrahydropalmatine. The preparation process of the oral preparation is as follows: the raw material angelica volatile oil is included with β-cyclodextrin / HP-β-cyclodextrin, the inclusion compound is mixed with the total alkaloids of Corydalis yanhusuo, conventional excipients are added, and the oral preparation is prepared according to conventional methods; the preparation process of the injection is as follows: the angelica volatile oil is included with β-cyclodextrin, the inclusion compound and the total alkaloids of Corydalis yanhusuo are dissolved in water for injection, activated carbon is added for stirring, filtration is performed, pH value is adjusted, filtration is performed, filling is performed, and sterilization is performed. The present invention also provides the use of the external Chinese medicine composition in the preparation of a drug for treating dysmenorrhea. Application number 201210171840.5, invention name: an external Chinese medicine composition for preventing and treating menstrual diseases, its preparation method and application, relates to an external Chinese medicine composition for preventing and treating menstrual diseases, its preparation method and application, and its active ingredients are prepared from the following ingredients in parts by weight: 40-70 parts of angelica volatile oil and chuanxiong volatile oil in total, 5-40 parts of lithospermum percolate and 0.01-3 parts of berberine.

[0006] Most of the preparations reported so far are administered orally or by injection. Studies have found that long-term use of Chuanxiong volatile oil may have certain effects on the digestive and reproductive systems (Guan Yongmei, et al., Research Progress on Chemical Composition, Pharmacological Actions and Clinical Application of Chuanxiong Volatile Oil, Chinese Patent Medicine, 2024-03-20).

[0007] Tetrahydropalmatine is an alkaloid found in the tuber of the Chinese medicinal plant Corydalis yanhusuo, which has significant analgesic and sedative effects. It has a good therapeutic effect in treating patients who cannot fall asleep due to chronic persistent pain, pain caused by gastric ulcers and duodenal ulcers, menstrual pain, uterine contraction pain after childbirth, and tension insomnia. The chemical properties of Tetrahydropalmatine include being insoluble in water, soluble in chloroform, and able to form crystalline salts with various inorganic acids. CN02156854.5, Invention Name: Tetrahydropalmatine dispersible tablets for treating dysmenorrhea and preparation method thereof, discloses the use of Tetrahydropalmatine dispersible tablets to treat dysmenorrhea. Oral administration of Tetrahydropalmatine may cause side effects, such as occasional dizziness, fatigue, nausea, etc.; large doses may have an inhibitory effect on the respiratory center and may sometimes cause extrapyramidal symptoms; some patients may experience loss of appetite, abdominal distension, drowsiness, slow heart rate, etc. when taking larger doses (such as more than 10 grams each time). In rare cases, it may cause electrocardiogram changes (such as widening and elevation of T waves) (see the product instructions of "Tyridine Sulfate Tablets").

[0008] Microemulsion gel is a new dosage form that has emerged in recent years. It is a transparent, uniform and stable gel network structure formed by adding a microemulsion composed of an oil phase, an aqueous phase, a surfactant and a co-surfactant into a gel matrix. It can promote drug penetration, improve drug stability, has strong safety, and has a high local drug concentration. It effectively solves the problems of poor adhesion, short retention time and poor skin spreadability of microemulsions. There have been reports on the preparation of ibuprofen transdermal gel for the treatment of dysmenorrhea. The results show that ibuprofen gel has good skin permeability and sustained release effect through abdominal transdermal administration. The drug mainly enters the uterus, and a small amount of the drug enters the blood circulation, which has a certain targeting ability. In recent years, volatile oils of traditional Chinese medicine are often used as oil phases to prepare microemulsion gels. For example, volatile oils such as perilla, tangerine peel, cyperus, bergamot, fructus aurantii, fennel, prickly ash, and citron are made into microemulsion gels for different purposes. In recent years, microemulsion gels have been increasingly used in clinical diseases such as internal medicine, gynecology, and pediatrics, such as gynecological medical antibacterial hydrogel, Kangfuxin antibacterial gel, and children's antipyretic gel.

[0009] There is no report on the combination of Angelica sinensis volatile oil and Ligusticum chuanxiong volatile oil with corydaline for the treatment of dysmenorrhea, let alone preparing them into relevant external preparations. Summary of the invention

[0010] The technical solution of the present invention is to provide an external Chinese medicine composition for treating dysmenorrhea and a preparation method and use thereof.

[0011] The present invention provides an external Chinese medicine composition for treating dysmenorrhea, which is a preparation prepared from the following raw materials in weight ratio:

[0012] 40 parts of tetrahydropalmatine, volatile oils of Chinese angelica and Chuanxiong, based on the original Chinese angelica and Chuanxiong materials: 4000 parts, wherein the dosage ratio of Chinese angelica to Chuanxiong is 1:1;

[0013] Among them, in the ethanol extracts of Angelica sinensis and Chuanxiong, the content of ligustilide in the volatile oil of Angelica sinensis is not less than 30% w / w; in the volatile oil of Chuanxiong, the content of ligustilide A is not less than 10% w / w, and the content of ligustilide is not less than 15% w / w.

[0014] The external Chinese medicine composition of the present invention is prepared from tetrahydropalmatine, angelica sinensis and volatile oil of chuanxiong as active ingredients, and is added with pharmaceutically acceptable auxiliary materials or auxiliary ingredients to form a commonly used external preparation in medicine.

[0015] Wherein, the external preparation is a patch, an ointment, a paint, a gel, or a liniment.

[0016] The gel is a microemulsion gel, which is prepared from an oil phase, an aqueous phase, an emulsifier and an emulsifier, and then added to a gel matrix to prepare a microemulsion gel; wherein the oil phase is prepared from a tetrahydropalmatine oleic acid solution, angelica sinensis and chuanxiong volatile oil; the emulsifier is Tween-80; the emulsifier is glycerol, 1,2-propylene glycol, and PEG-400;

[0017] The mass ratio Km value of emulsifier to co-emulsifier is (1-3): (1-3); the mass ratio of oil phase to emulsifier and co-emulsifier is 1: (1-9).

[0018] The gel matrix is ​​Carbomer 940.

[0019] Wherein, the auxiliary emulsifier is PEG-400; the mass ratio Km value of the emulsifier to the auxiliary emulsifier is 1:1; the mass ratio of the oil phase to the emulsifier and the auxiliary emulsifier is 1:9.

[0020] The present invention provides a method for preparing an external Chinese medicine composition for treating dysmenorrhea, wherein the method for preparing the microemulsion gel comprises the following steps:

[0021] a. Dissolve tetrahydropalmatine in oleic acid by ultrasonication, and use it together with volatile oil as the oil phase;

[0022] b. Evenly mix the emulsifier, the co-emulsifier and the oil phase, stir them at a speed of 100 r / min using a magnetic stirrer, and add deionized water while stirring to prepare a W / O type microemulsion;

[0023] c. Continue to drop deionized water until the system forms a gel phase. Continue stirring and add deionized water to allow the system to quickly pass through the gel phase. The system changes from turbid to clear, forming an O / W microemulsion.

[0024] d. Dissolve Carbomer 940 in water to prepare a 1.5% solution, and mix the microemulsion and the Carbomer solution in a ratio of 2:1 to form a microemulsion gel.

[0025] The present invention also provides use of the external Chinese medicine composition in preparing a medicine for treating dysmenorrhea.

[0026] Wherein, the medicine is a medicine with analgesic effect.

[0027] The administration method of the drug of the present invention is: take the prepared microemulsion gel and apply it around the lower abdomen below the navel or on the local pain point of the lower abdomen, and gently apply it with the fingertips until it is absorbed.

[0028] The beneficial effects of the present invention are:

[0029] The external Chinese medicine composition of the present invention is prepared from tetrahydropalmatine, angelica and chuanxiong volatile oil. The efficacy test proves that the composition has obvious efficacy in treating dysmenorrhea, the raw materials can play a synergistic role, the clinical administration dosage is reduced, the medication cost is reduced, the composition is directly externally administered, the clinical use is more convenient and safe, and a new choice is provided for the clinical treatment of dysmenorrhea. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is a diagram of the Tyndall phenomenon of drug-containing microemulsion under laser pen irradiation (from left to right: drug-containing microemulsion, drug-containing microemulsion, deionized water);

[0031] Figure 2 This is the appearance picture of drug-containing microemulsion gel;

[0032] Figure 3 Pseudo-ternary phase diagram of different co-emulsifiers (Tween 80 / PEG-400);

[0033] Figure 4 Pseudo-ternary phase diagram for different Km values;

[0034] Figure 5 is the transmission electron microscopy image of microemulsion (scale bar = 200 nm);

[0035] Figure 6 is the particle size distribution diagram of microemulsion;

[0036] Figure 7 are chromatograms of each sample (among which, A is the chromatogram of blank microemulsion gel; B is the chromatogram of paeonol and blank microemulsion gel; C is the chromatogram of drug-containing microemulsion gel; D is the chromatogram of paeonol and drug-containing microemulsion gel; peak 1, peak 2, and peak 3 in the figure are the characteristic peaks of paeonol, ligustilide A, and ligustilide, respectively). DETAILED DESCRIPTION

[0037] The raw material of the invention, chuanxiong, is the dried root of Ligusticum chuanxiong Hort. of the Umbelliferae family, purchased from Pengzhou, Sichuan; the angelica is the dried root of Angelica sinensis (Oliv.) Diels of the Umbelliferae family, purchased from Minxian County, Gansu Province, and both are identified by researcher Shu Guangming of Sichuan Academy of Traditional Chinese Medicine.

[0038] Example 1 Preparation of the microemulsion gel of the present invention

[0039] 40 mg of tetrahydropalmatine was dissolved in 0.6 g of oleic acid by ultrasonication, and mixed with 178 mg of volatile oil, wherein the volatile oil was composed of 40 mg of angelica volatile oil + 138 mg of Chuanxiong volatile oil (converted into the amount of medicinal materials, 4 g of angelica and 2 g of Chuanxiong medicinal materials, respectively) as the oil phase, and 3.681 g of Tween-80 and PEG-400 were added, mixed, and stirred at 100 r / min with a magnetic stirrer, and deionized water was added while stirring. With the addition of deionized water, the system changed from a translucent state to a clear and transparent state, and a W / O type microemulsion was formed.

[0040] Continue to add deionized water, the system forms a gel phase with poor fluidity, continue to stir and add deionized water, let the system quickly pass through the gel phase, the system changes from turbid to clear, and O / W type microemulsion is formed. Prepare a 1.5% solution of Carbomer 940 with deionized water, and mix them in a ratio of 2:1 (v / v) of microemulsion: Carbomer solution to obtain a microemulsion gel (see Figure 1 , Figure 2 ).

[0041] Example 2 Screening test of the matrix of microemulsion in the microemulsion gel of the present invention

[0042] 1. Drawing of pseudo-ternary phase diagram

[0043] The oil phase (volatile oil and tetrahydropalmatine oleic acid solution are the oil phase) and the emulsifier (including the co-emulsifier) ​​are mixed in different proportions, and the microemulsion is prepared according to the above-mentioned microemulsion preparation method. The mass percentages of the oil phase, the water phase, and the emulsifier (including the co-emulsifier) ​​corresponding to the critical point are calculated, and a pseudo-ternary phase diagram is drawn with each phase as the three vertices of the pseudo-ternary phase diagram. The critical points are connected to obtain the microemulsion area under the corresponding conditions. The larger the microemulsion area, the easier it is to form a microemulsion.

[0044] 2. Screening of emulsifier

[0045] Microemulsions were prepared with volatile oil and tetrahydropalmatine oleic acid solution as oil phase, Tween-80 as emulsifier, deionized water as aqueous phase, glycerol, 1, 2-propylene glycol and PEG-400 as co-emulsifiers, respectively. The Km was fixed at 1:1 and the mass ratios of oil phase to emulsifier (including co-emulsifier) ​​were 1:9, 1:8, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2 and 1:1. The transparency, color and other aspects of the microemulsions were observed and compared. The effects of the three co-emulsifiers were compared, the pseudo-ternary phase diagrams were drawn, and the sizes of the microemulsion areas formed in the pseudo-ternary phase diagrams drawn with different co-emulsifiers were compared.

[0046] From Table 1, Figure 3 It can be seen that PEG-400 can form microemulsion when used as an emulsifier, and the area of ​​the microemulsion region is S PEG-400=1.8223. Glycerol and 1-2-propylene glycol as co-emulsifiers cannot form microemulsions, so PEG-400 is selected as the co-emulsifier.

[0047] Table 1 Screening results of emulsifiers

[0048]

[0049] 3. Determination of Km value

[0050] With volatile oil and tetrahydropalmatine oleic acid solution as oil phase, Tween-80 as emulsifier, PEG-400 as co-emulsifier, deionized water as water phase, the proportion screening was carried out with Km values ​​of 3: 1, 2: 1, 1: 1, 1: 2, and 1: 3, and the mass ratios of oil phase to emulsifier (including co-emulsifier) ​​were 1: 9, 1: 8, 1: 7, 1: 6, 1: 5, 1: 4, 1: 3, 1: 2, and 1: 1 to prepare microemulsions, and the appearance transparency, color, etc. of the microemulsions of various proportions were observed and compared, and the pseudo-ternary phase diagram was drawn ( Figure 2 ), compare the size of the microemulsion area formed in the pseudo-ternary phase diagram drawn with different Km values.

[0051] From Table 2 and Figure 4 It can be seen that 3:1, 2:1, 1:1, and 1:2 can all form microemulsions, and the area of ​​the microemulsion region S 3:1 =1.0815, S 2:1 =1.7693, S 1:1 =1.8223, S 1;2 =0.9748. When the Km value is 1:1, the area of ​​the microemulsion zone is the largest and the microemulsion properties are optimal. At the same time, in order to reduce the use of excipients, the Km value is determined to be 1:1.

[0052] Table 2 Km value screening results

[0053]

[0054] 4. Ratio of oil phase to emulsifier (including co-emulsifier)

[0055] With volatile oil and tetrahydropalmatine oleic acid solution as oil phase, Tween-80 as emulsifier, PEG-400 as co-emulsifier, deionized water as water phase, Km value of 1:1, the mass ratio of oil phase to emulsifier (including co-emulsifier) ​​was 1:9, 1:8, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2, 1:1 respectively, and microemulsions were prepared by mixing evenly. The appearance transparency, color, etc. of the microemulsions in different ratios were observed and compared.

[0056] As shown in Table 3, when the Km value is 1:1, the microemulsions prepared with the mass ratio of oil phase to emulsifier (including co-emulsifier) ​​of 1:9, 1:8, 1:7, 1:6, and 1:5 have different appearances. The microemulsion with the mass ratio of oil phase to emulsifier (including co-emulsifier) ​​of 1:9 has better properties, is clear and transparent, and does not separate after centrifugation. Therefore, the ratio of oil phase to emulsifier (including co-emulsifier) ​​is selected to be 1:9.

[0057] Table 3 Screening results of the ratio of oil phase to emulsifier

[0058]

[0059] Therefore, the optimal formula for preparing microemulsion is: emulsifier is Tween-80, co-emulsifier is PEG-400, the ratio of oil phase to emulsifier (including co-emulsifier) ​​is 1:9, and the Km value is 1:1.

[0060] 5. Evaluation of microemulsion morphology and particle size

[0061] The microemulsion was diluted with deionized water to an appropriate concentration, a small amount of microemulsion was added to the surface of the copper mesh, spread, and stained with 2% phosphotungstic acid. After the water evaporated, the size and morphology of the microemulsion droplets were observed by transmission electron microscopy.

[0062] The results of transmission electron microscopy observation ( Figure 5 ) It can be seen that after dilution, the droplets of the microemulsion appear as round or quasi-round particles with a relatively uniform particle size of about 140nm.

[0063] Take a small amount of microemulsion and add deionized water to dilute to an appropriate concentration, and use a nanoparticle tracking analyzer to measure the particle size of the microemulsion. The average particle size of the microemulsion measured by the nanoparticle tracking analyzer is 146.1 nm, which is consistent with the particle size observed by transmission electron microscopy and meets the requirements for the particle size of the microemulsion (see Figure 6 ).

[0064] Example 3 Test of properties of microemulsion gel of the present invention

[0065] 1. HPLC determination of encapsulation efficiency

[0066] Chromatographic conditions: Waters 2695-2487 UV detection system, chromatographic column: Sepax; C18-H (250×4.6 mm, 5 μm); mobile phase: methanol: acetonitrile: water = 15:35:50; detection wavelength: 280 nm; flow rate: 1.0 mL / min; column temperature: 35 °C; injection volume: 20 μL.

[0067] Detection method: Volatile oil is unstable, so the functional substances ligustilide and ligustilide A in volatile oil are mainly detected, and the encapsulation efficiency of microemulsion for drugs is evaluated with ligustilide and ligustilide A as evaluation indicators. The one-test-multiple-evaluation method is adopted, with paeonol as the internal standard, and relative retention time and relative correction factor are used to qualitatively and quantitatively determine ligustilide and ligustilide A.

[0068] Table 4 Correction factors for the components to be measured

[0069] Component to be measured (peak) Relative retention time Correction Factor (F) Paeonol 1.00 1.0000 Cnidium sutchuenolide A 1.88 4.5147 Ligustilide 3.04 2.2506

[0070] F=(peak area of ​​paeonol × concentration of the component to be tested) / (peak area of ​​the component to be tested × concentration of paeonol)

[0071] 2. Methodological investigation

[0072] Previous studies have determined that the instrument specificity, precision, and recovery rate all meet the requirements for the detection of ligustilide and ligustilide A (see Figure 7 ).

[0073] 3. Determination of encapsulation efficiency

[0074] Accurately weigh 0.45 g of microemulsion gel from different batches in 3 portions, dissolve in 3 mL of anhydrous ethanol respectively, vortex and shake for 2 min to break the microemulsion and fully dissolve the volatile oil in anhydrous ethanol. Take 100 μL of the mixed ethanol solution, add 900 μL of mobile phase, mix well, filter with a 0.22 um microporous membrane, and detect the drug content (W1). Accurately weigh 1.5 g of microemulsion gel, dilute with water to 10 mL, take 9 mL of the diluted solution and centrifuge in an ultrafiltration tube (100 kDa) (4000 r / min, 30 min), take the lower filtrate to detect the content of ligustilide A and ligustilide (W2). Calculate the encapsulation efficiency (EE%) according to the following formula.

[0075] Encapsulation efficiency (EE%) = (W1-W2) / W1 × 100%

[0076] 4. Stability study

[0077] The stability of microemulsion gel was evaluated by studying the effect of storage time on the drug content. Three batches of microemulsion gel were stored at room temperature. The day of preparation of microemulsion gel was recorded as day 0, and stability tests were conducted on days 0, 15, and 30. First, the appearance, stratification, color change, etc. of the microemulsion gel were observed, and then the content of ligustilide A and ligustilide was determined. The drug content on day 0 was taken as the initial content, and the percentage of the measured value to the initial content, i.e., the relative content, was calculated based on 100%.

[0078] As can be seen from Table 5, after 30 days, the encapsulation rates of ligustilide A and ligustilide in the microemulsion gel were 86.88% and 96.45%, respectively, both greater than the 80% specified in the pharmacopoeia.

[0079] Table 5 Microemulsion stability test results

[0080] time Cnidium monnieri lactone A (mg) Relative content Ligustilide (mg) Relative content Day 0 5.32 100% 16.29 100% Day 15 5.42 102% 16.32 100% Day 30 5.11 96% 15.94 98%

[0081] The beneficial effects of the present invention are demonstrated by the following efficacy test.

[0082] Test Example 1 Screening and verification test of the raw material drug compatibility of the present invention

[0083] 1. Optimizing the best combination of the raw materials of the drug of the present invention by orthogonal design

[0084] Tetrahydropalmatine was used as factor A. On the basis of previous studies, Angelica sinensis volatile oil and Chuanxiong volatile oil (original drug 1:1) were used as factor B. A two-factor three-level orthogonal experimental design (Table 6) was used to prepare microemulsion gel according to the corresponding ratio combination dose in Table 7 and the screened optimal microemulsion formula (the total amount of oil phase was based on the maximum dosage A3B3, and the drug combination with insufficient oil phase was supplemented with oleic acid to an oil-emulsion ratio of 1:9). The analgesic effect of the drug combination was evaluated using the mouse acetic acid writhing model. The optimal ratio of volatile oil and tetrahydropalmatine was screened.

[0085] Table 6 Orthogonal design factor level table

[0086]

[0087] Table 7 Orthogonal design table

[0088]

[0089] After adaptive feeding, 18-22 g male KM mice were randomly divided into a model group (blank microemulsion gel without drug), aspirin 100 mg / kg, and microemulsion gel A1B1, A1B2, A1B3, A2B1, A2B2, A2B3, A3B1, A3B2, and A3B3, a total of 11 groups, with 8 animals in each group. Except for the aspirin group, the other groups had a 3*3 cm abdominal area punctured before administration. 2 The area was depilated for easy application and medication. The volume of aspirin gavage was 10 mL / kg. The corresponding drug was applied to the skin of the model group and microemulsion gel group at 60 uL / mouse, twice a day for 3 consecutive times. 40 minutes after the last administration, 0.8% glacial acetic acid solution (0.1 mL / 10g) was injected intraperitoneally to establish the model. The number of twisting times of mice in each group within 5 to 20 minutes after modeling was observed and recorded.

[0090] It can be seen from Table 8 that the range R value in the dose screening is R A >RB Therefore, the factors affecting the number of twists are mainly tetrahydropalmatine > volatile oil. Combining the optimal levels of each factor, it can be seen that K3 is the smallest in both columns A and B (the fewer the number of twists, the better). Therefore, the optimal ratio is A3B3, that is, the dose of A (tetrahydropalmatine) is 40 mg / kg, and the dose of B (volatile oil) is 4g / kg. From the variance analysis in Table 9, it can be seen that A and B have an interaction.

[0091] Table 8 Range analysis results

[0092] Experiment No. A B Number of twists (times) 1 1 1 17.0 2 1 2 15.0 3 1 3 6.5 4 2 1 11.3 5 2 2 9.1 6 2 3 10.9 7 3 1 8.1 8 3 2 8.0 9 3 3 6.3 <![CDATA[K1]]> 38.5 36.4 — <![CDATA[K2]]> 31.3 32.1 — <![CDATA[K3]]> 22.4 23.6 — <![CDATA[k1]]> 12.8 12.1 — <![CDATA[k2]]> 10.4 10.7 — <![CDATA[k3]]> 7.5 7.9 — R 5.4 2.8 — Optimal level 3 3

[0093] Table 9 ANOVA Interaction

[0094] Sources of variation sum of squares Degrees of Freedom Mean Square F-number P-value Calibration model <![CDATA[896.111 a ]]> 8 112.014 3.732 0.001 intercept 7544.014 1 7544.014 251.351 0.000 A 347.861 2 173.931 5.795 0.005 B 242.528 2 121.264 4.040 0.022 A*B 305.722 4 76.431 2.547 0.048 error 1890.875 63 30.014 total 10331.000 72 Corrected Total 2786.986 71

[0095] a. R-square = 0.414 (adjusted R-square = 0.340)

[0096] In the above test, ANOVA (Analysis of Variance) is a statistical method used to analyze whether the differences in the means of three or more samples are significant. In ANOVA, interaction refers to the joint influence of two or more factors (independent variables) on the dependent variable. This influence is not a simple additive relationship, but there is a certain degree of interaction. The analysis results show that when the dosage of corydalis butylene is 40 mg and the dosage of volatile oil is 4g (based on the original dosage), it can play a synergistic role.

[0097] 2. Synergistic Verification Test of the Compatibility of the APIs of the Present Invention

[0098] 1. Comparison between the drug of the present invention and the single-use group

[0099] After adaptive feeding, 18-22 g male KM mice were randomly divided into a model group (blank microemulsion gel without drug), aspirin 100 mg / kg, 4 g native drug / kg volatile oil microemulsion gel group, 40 mg / kg tetrahydropalmatine microemulsion gel group, 4 g native drug / kg volatile oil + 40 mg / kg tetrahydropalmatine microemulsion gel group, a total of 5 groups, 8 animals in each group. Except for the aspirin group, the other groups had a 3*3 cm abdominal area punctured before administration. 2 The area was depilated for easy application and medication. The volume of aspirin gavage was 10 mL / kg. The corresponding drug was applied to the skin of the model group and microemulsion gel group at 60 uL / mouse, twice a day for 3 consecutive times. 40 minutes after the last administration, 0.8% glacial acetic acid solution (0.1 mL / 10g) was injected intraperitoneally to establish the model. The number of twisting times of mice in each group within 5 to 20 minutes after modeling was observed and recorded.

[0100] As shown in Table 10, when used alone, the two drugs have analgesic effects, and the volatile oil has better efficacy than tetrahydropalmatine. However, the orthogonal analysis results in Table 8 show that tetrahydropalmatine is the leading factor in analgesia. The reason may be that when the two drugs are used together, the volatile oil promotes the absorption of tetrahydropalmatine, making it play the maximum efficacy. When the two drugs are used together, their efficacy is better than when used alone.

[0101] Factorial analysis is a statistical method that can be used to determine the synergistic effect of drugs. In drug research, when two or more drugs are used at the same time, factorial design can study the effects of each of these drugs and the possible interactions between them. For example, assuming there are drugs A and B, factorial analysis can determine whether A and B have a synergistic effect by comparing the effects of using A alone, B alone, and using A and B at the same time. Its basic principle is to analyze the main effects of each factor (different drugs) and the interaction effects between factors. Through experimental design and corresponding statistical analysis, the relationship between drugs when used in combination can be determined more accurately. From the results of the factorial analysis in Table 11, it can be seen that A and B have an interaction, indicating that the two have a synergistic effect.

[0102] Table 10 Effects of microemulsion gel on the number of writhing times of mice ( ,n=8)

[0103] Grouping dose Number of twists (times) Model Group — 22.4±6.6 aspirin 100 mg / kg <![CDATA[9.6±6.4 ＊＊ ]]> Essential oil microemulsion gel 4g / kg <![CDATA[9.9±4.3 ＊＊ ]]> Tetrahydropalmatine microemulsion gel 40mg / kg <![CDATA[13.3±5.0 ＊＊ ]]> Volatile oil + tetrahydropalmatine microemulsion gel 4g / kg+40mg / kg <![CDATA[8.4±4.4 ＊＊ ]]>

[0104] Note: Compared with the model group, **P<0.01, the same below.

[0105] Table 11 Results of factorial analysis

[0106] Sources of variation Type III Sum of Squares Degrees of Freedom Mean Square F-number P-value Calibration model <![CDATA[945.844 a ]]> 3 315.281 11.863 0.000 intercept 5805.031 1 5805.031 218.432 0.000 A 225.781 1 225.781 8.496 0.007 B 603.781 1 603.781 22.719 0.000 A*B 116.281 1 116.281 4 0.046 error 744.125 28 26.576 total 7495.000 32 Corrected Total 1689.969 31

[0107] a. R-square = 0.510 (adjusted R-square = 0.513)

[0108] 2. Comparison of the drug of the present invention with the single-use group at the same dose

[0109] Due to the solubility of tetrahydropalmatine in oleic acid, it was impossible to prepare tetrahydropalmatine microemulsion gel with a dosage of 218 mg / kg. In addition, this dosage of monomer drug was too high for clinical use, which may have toxic side effects. Therefore, when comparing the efficacy of the same drug dose, only the efficacy of volatile oil microemulsion gel alone and volatile oil + tetrahydropalmatine microemulsion gel was set.

[0110] According to the optimal dose ratio A3B3 (volatile oil was converted according to the extraction rate, i.e. A (tetrahydropalmatine) 40 mg, B (volatile oil) 178 mg, a total of 218 mg / kg), single and combined use tests were conducted at equal doses. After adaptive feeding, 18-22 g male KM mice were randomly divided into a model group (blank microemulsion gel without drug), aspirin 100 mg / kg, 218 mg / kg volatile oil microemulsion gel group, 178 mg volatile oil + 40 mg / kg tetrahydropalmatine microemulsion gel group, a total of 5 groups, 10 animals in each group. Except for the aspirin group, the other groups had a 3*3 cm large area of ​​​​abdomen removed before administration. 2 The area was depilated for easy application and medication. The volume of aspirin gavage was 10 mL / kg. The corresponding drug was applied to the skin of the model group and microemulsion gel group at 60 uL / mouse, twice a day for 3 consecutive times. 40 minutes after the last administration, 0.8% glacial acetic acid solution (0.1 mL / 10g) was injected intraperitoneally to establish the model. The number of twisting times of mice in each group within 5 to 20 minutes after model establishment was observed and recorded.

[0111] As shown in Table 12, compared with the model group, the volatile oil microemulsion gel alone and the volatile oil + tetrahydropalmatine microemulsion gel can significantly reduce the number of body twisting in mice; compared with the volatile oil microemulsion gel alone, the volatile oil and tetrahydropalmatine microemulsion gel can further significantly reduce the number of body twisting in mice. Therefore, the combination of volatile oil and tetrahydropalmatine has a synergistic effect.

[0112] Table 12 Effects of equal doses of microemulsion gel on the number of writhing times of mice ( ,n=10)

[0113] Grouping dose Number of twists (times) Model Group — 21.1±5.5 aspirin 100 mg / kg <![CDATA[8.1±3.6 ＊＊ ]]> Essential oil microemulsion gel 218mg / kg <![CDATA[11.4±3.5 ＊＊ ]]> Volatile oil + tetrahydropalmatine microemulsion gel 218mg / kg <![CDATA[7.4±4.5 ＊＊# ]]>

[0114] Note: Compared with volatile oil + tetrahydropalmatine microemulsion gel, # P<0.05.

[0115] Test Example 2 Pharmacological efficacy test of the microemulsion gel of the present invention (external efficacy verification (acetic acid writhing model + dysmenorrhea model))

[0116] Microemulsion gels were prepared according to the optimal drug ratio obtained by orthogonal design, and the gels with this drug content were used as large-dose (A, 40 mg / kg; B, 4g / kg). Medium-dose (A, 20 mg / kg; B, 2g / kg) and small-dose (A, 10 mg / kg; B, 1g / kg) microemulsion gels were also prepared for external efficacy verification.

[0117] Acetic acid writhing model: 18-22 g male KM mice were randomly divided into a model group (microemulsion gel without drug), a western medicine positive drug aspirin group (100 mg / kg) and three microemulsion gel groups, with 10 mice in each group. Except for the aspirin group, the other groups had a 3*3 cm 2 The area was depilated for easy application and medication. Aspirin, a positive Western medicine, was administered intragastrically at 10 mL / kg. The corresponding drug was applied to the skin of the model group and microemulsion gel group at 60 uL / mouse, twice a day for 3 consecutive times. 0.8% glacial acetic acid solution (0.1 mL / 10 g) was injected intraperitoneally 40 minutes after the last administration to establish the model. The number of twists of mice in each group within 5 to 20 minutes after modeling was observed and recorded.

[0118] As shown in Table 13, both the medium-dose and high-dose groups of microemulsion gel can significantly reduce the number of twisting times of mice. Compared with the model group, the difference is statistically significant (P<0.05), indicating that the topical analgesic effect of microemulsion gel is good.

[0119] Table 13 Effect of microemulsion gel on the number of writhing times induced by acetic acid in mice ( ,n=10)

[0120] Grouping dose Number of twists (times) Model Group — 21.7±7.5 aspirin 100 mg / kg <![CDATA[9.0±7.9 ＊＊ ]]> Microemulsion gel A, 10 mg / kg; B, 1 g / kg 15.6±12.2 Microemulsion gel A, 20 mg / kg; B, 2 g / kg <![CDATA[12.6±8.4 ＊ ]]> Microemulsion gel A, 40 mg / kg; B, 4 g / kg <![CDATA[10.7±7.8 ＊＊ ]]>

[0121] Note: Compared with the model group, *P<0.05; the same below.

[0122] Dysmenorrhea model: 8-week-old female KM mice were randomly divided into a model group (microemulsion gel without drug), a western medicine positive drug aspirin group (100 mg / kg) and three microemulsion gel groups, with 10 mice in each group. Except for the aspirin group, the other groups were given a 3*3 cm 2 The area was depilated for easy application and medication. Western medicine positive drug aspirin was gavaged at 10mL / kg, and the corresponding drug was applied to the skin of the model group and microemulsion gel group at 60uL / mouse, twice a day, once in the morning and once in the afternoon, for 4 consecutive days. Estradiol valerate (Progynova) combined with oxytocin was used for modeling: the model was built for 4 days, and Progynova 4mg / kg was gavaged on the morning of the 1st and 4th days, and Progynova 3mg / kg was gavaged on the 2nd and 3rd days. Oxytocin injection was intraperitoneally injected at 5U / mouse in the afternoon of the 4th day, and the number of twists within 30 minutes after the injection of oxytocin was observed and recorded.

[0123] As shown in Table 14, the medium and large doses of microemulsion gel can significantly reduce the number of body twisting of mice with dysmenorrhea, indicating that the microemulsion gel has a good therapeutic effect on dysmenorrhea.

[0124] Table 14 Effects of microemulsion gel on the number of twisting times in dysmenorrhea model mice ( ,n=10)

[0125] Grouping dose Number of twists (times) Model Group — 13.3±3.9 aspirin 100 mg / kg <![CDATA[6.9±4.0 ＊＊ ]]> Microemulsion gel A, 10 mg / kg; B, 1 g / kg 10.4±4.1 Microemulsion gel A, 20 mg / kg; B, 2 g / kg <![CDATA[9.0±4.1 ＊ ]]> Microemulsion gel A, 40 mg / kg; B, 4 g / kg <![CDATA[8.1±3.9 ＊＊ ]]>

Claims

1. A Chinese medicine composition for external use for treating dysmenorrhea, characterized in that: It is a preparation made from the following raw materials in the following weight ratio: 40 parts of tetrahydropalmatine, volatile oils of Chinese angelica and Chuanxiong, based on the original Chinese angelica and Chuanxiong materials: 4000 parts, wherein the dosage ratio of Chinese angelica to Chuanxiong is 1:1; Among them, the volatile oils of Angelica sinensis and Chuanxiong, the content of ligustilide in the volatile oil of Angelica sinensis is not less than 30% w / w; the content of ligustilide A in the volatile oil of Chuanxiong is not less than 10% w / w, and the content of ligustilide is not less than 15% w / w; The preparation is a gel; the gel is a microemulsion gel, which is prepared from an oil phase, an aqueous phase, an emulsifier and an emulsifier to form a microemulsion, and is added to a gel matrix to form a microemulsion gel; wherein the oil phase is prepared from a tetrahydropalmatine oleic acid solution, angelica sinensis and chuanxiong volatile oil; the emulsifier is Tween-80; the emulsifier is polyethylene glycol-400; the mass ratio Km value of the emulsifier to the emulsifier is 1-3:1-3; the mass ratio of the oil phase to the emulsifier and the emulsifier is 1:1-9; the gel matrix is ​​carbomer 940; The preparation method of the microemulsion gel comprises the following steps: a. Dissolve tetrahydropalmatine in oleic acid by ultrasonication, and use it together with volatile oil as the oil phase; b. Evenly mix the emulsifier, the co-emulsifier and the oil phase, stir them at a speed of 100 r / min using a magnetic stirrer, and add deionized water while stirring to prepare a W / O type microemulsion; c. Continue to drop deionized water until the system forms a gel phase. Continue stirring and add deionized water to allow the system to quickly pass through the gel phase. The system changes from turbid to clear, forming an O / W microemulsion. d. Dissolve Carbomer 940 in water to prepare a 1.5% solution, and mix the microemulsion and the Carbomer solution in a ratio of 2:1 to form a microemulsion gel.

2. The external Chinese medicine composition for treating dysmenorrhea according to claim 1, characterized in that: The auxiliary emulsifier is polyethylene glycol-400; the mass ratio Km value of the emulsifier to the auxiliary emulsifier is 1:1; the mass ratio of the oil phase to the emulsifier and the auxiliary emulsifier is 1:

9.

3. A method for preparing the external Chinese medicine composition for treating dysmenorrhea according to claim 1 or 2, characterized in that: The preparation method of the microemulsion gel comprises the following steps: a. Dissolve tetrahydropalmatine in oleic acid by ultrasonication, and use it together with volatile oil as the oil phase; b. Evenly mix the emulsifier, the co-emulsifier and the oil phase, stir them at a speed of 100 r / min using a magnetic stirrer, and add deionized water while stirring to prepare a W / O type microemulsion; c. Continue to drop deionized water until the system forms a gel phase. Continue stirring and add deionized water to allow the system to quickly pass through the gel phase. The system changes from turbid to clear, forming an O / W microemulsion. d. Dissolve Carbomer 940 in water to prepare a 1.5% solution, and mix the microemulsion and the Carbomer solution in a ratio of 2:1 to form a microemulsion gel.

4. Use of the external Chinese medicine composition according to claim 1 or 2 in the preparation of a medicament for treating dysmenorrhea.

5. The use according to claim 4, characterized in that: The medicine is a medicine with analgesic effect.

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