WO2006017994A1 - An analgesic pharmaceutical composition - Google Patents

Abstract

An analgesic pharmaceutical composition consisting of at least two or three of benzoylmesaconine 0-600 parts, mesaconitine 0-6 parts, hypaconitine 0-3 parts as active components and at least two of them being not zero. The composition has synergism effect.

Description

 Pharmaceutical composition with analgesic effect

 The present invention relates to an analgesic pharmaceutical composition, in particular to an analgesic drug prepared by the aconitine component extracted from the aconite plant as an effective drug.

Background technique

Aconite is the genus Aconite (Aeonitam earm gae Debi). The lateral root of Aeonitam earm gae Debi is a traditional Chinese medicine. It is a commonly used anti-retroviral drug in ancient warming agents because it has obvious toxicity and dosage. and nearly toxic dose, poisoning and death frequently reported clinical phenomenon _s therefore has a large number of its students about drug prescription coverage for cooking up chemistry, pharmacology and toxicology, and clinical trials and other work .. such as: Zhang Minfeng Aconitum poisoning rhythm Disorder, "Shandong Journal of Traditional Chinese Medicine", Volume 1, May 1999, discloses that "orphanine 0.2 mg orally can cause human poisoning", so in order to reduce its toxicity, it increases the efficacy and constantly seeks high-efficiency and low toxicity with pharmacodynamic activity. Research on the composition of compounds is essential

Zhang Dihua et al., in Chinese herbal medicine 19S2:73(ll), l-4, reported that eight alkaloids including aconitine and benzoyl aconitine in aconitine were isolated from Baifu tablets. Ingredients _s and proved that it can have anti-inflammatory and anti-heart failure effects Wang Chaohong, extraction and separation of highly toxic alkaloids from Aconitum, Criminal Technology, No. 3, 2002, discloses the extraction of aconitine, Zhongwu from aconite Methods for the treatment of aconitine and aconitine with regard to the application of aconite in analgesics, Zhang Yongzhong et al., in Chinese herbal medicine 2002:33(2), 106-109, reported four traditional Chinese medicines such as Yuanhuwutou. The research on the composition of "analgesic soup" in the treatment of advanced pain in cancer has proved to have a good therapeutic effect. Studies on the extraction, separation, content and structure identification of single aconitine have also been reported. analgesic effects while also trying to be divided, and wherein the active ingredient to distinguish between "the toxic components of Aconitum compound compatibility, low toxicity purposes, there is no relevant reports _Θ

Summary of the invention

 SUMMARY OF THE INVENTION The technical solution of the present invention provides an analgesic pharmaceutical composition comprising an aconitine component isolated and extracted from an aconite plant as an effective pharmaceutical ingredient, which can achieve an effective analgesic effect of high efficiency and low toxicity.

 The invention provides an analgesic pharmaceutical composition, which is prepared by using aconitine compound benzoyl aconitine, aconitine, and aconitine as active ingredients. The dosage of the medicament is as follows: 0-600 parts of aconitine in benzoyl, 0-6 parts of hypoaconitine, 0-3 parts of metoconium, and at least two ratios of distribution ratio Can't be zero.

 The three alkaloids of Hypaconi «ne , Mesaconitine and Benzoylmesaconine are active ingredients in the pharmaceutical compositions of the present invention. The structure of the lower aconitine is as shown in the formula (Π), and the structure of the aconitine is as shown in the formula (ffl), and the structure of the aconitine in the benzoyl group is as shown in the formula (W). These components can be extracted and isolated from the medicinal plant aconite according to methods currently reported in the literature.

( Π ) (m) Confirmation

 (IV)

 The pharmaceutical composition is prepared by using a lower weight ratio of aconitine compound as an active ingredient: benzoyl aconitine 100-300 parts of aconitine 1-3 parts preferably: benzene Acetyl aconitine i is divided into 1 part of aconitine

 Or an agent prepared by the following weight-matching aconitine compound as an active ingredient: 2-6 parts of hypoaconitine, 1-3 parts of mesaconitine: 2 parts of hypoaconitine, Zhongwu Reduce the head by 3 copies.

 Or an agent prepared by the following weight-matching aconitine compound as an active ingredient: 200-600 parts of aconitine in benzoyl and 1-3 parts of aconitine in benzoyl: benzoyl Stereophylline 200 parts of aconitine 3 parts.

 Or an agent prepared by the following a weight-matched aconitine compound as an active ingredient: 100-600 parts of aconitine in benzoyl, 1-6 parts of aconitine, and aconitine 1- 3 parts of preference: 100 parts of benzoyl aconitine, 1 part of aconitine and 1 part of aconitine

 Wherein the agent is: an oral preparation, an external preparation, and an injection.

 The oral preparations are tablets, capsules, pills, granules.

 The external preparation is a lotion, an ointment, a suppository, or a medicinal dressing.

 The injection is an injection solution or a powder for injection.

The toxicity test results of the above effective pharmaceutical ingredients showed that the LD _S0 of the aconitine was 6.41 m _g / k _g , and the LD _{5 ( )} of the aconitine was 12.8 mg / kg. The toxicity of the base was relatively low, and the LD _{5 ()} value was not measured, and the maximum tolerated amount was 1000 mg/kg body weight.

 The amount of the three aconitine compounds in the present invention is within the above safe range, and the synergistic effect of the synergistic effect of the three compounds in combination or the simultaneous use of the three compounds can be achieved. Moreover, the drug composition after compatibility does not have a simple dose-effect linear relationship, that is, the more the dosage, the better the drug effect, but the drug is best at a specific ratio. Tests have shown that the best combination of pharmaceutical compositions in the best ratio can achieve the best efficacy and significantly reduced toxicity. In summary, the pharmaceutical composition of the present invention achieves the purpose of reducing efficacy and low toxicity, and provides a new choice for clinical use.

Specific implementation

 The above-mentioned aconitine alkaloids are used as effective pharmaceutical ingredients, mixed with other auxiliary and/or additional ingredients which are allowed and acceptable in the medicine, and processed according to the corresponding pharmaceutical method, so that they can be prepared for use. A corresponding analgesic formulation pharmaceutical composition. For example, it can be prepared into a tablet by mixing with a commonly used auxiliary component such as a disintegrator, an excipient, a lubricant, a binder, a filler, and the like which are acceptable in an oral preparation, and according to a corresponding conventional process. Oral drugs in the form of solid preparations such as pills, capsules, granules and corresponding sustained release agents, controlled release agents. The corresponding injectable drug can be prepared after the appropriate solvent and additive which are allowed to be used in the injectable pharmaceutical preparation and the corresponding process operation. Mixing with the corresponding external drug dispersing agent, curing agent, stabilizer, etc., can be prepared into a corresponding externally used drug including ointment, suppository, lotion and medicinal dressing.

Examples 1 to 7: Oral Capsule Drugs The composition of each drug in parts by weight is shown in Table 1.

 Preparation method: The effective pharmaceutical ingredient is pulverized and sieved according to the conventional preparation method of the capsule medicine, and then starch is added and mixed with an appropriate amount of medical ethanol to be granulated on a swing granulator, and after drying the whole granule, it is mixed with a proper amount of lubricant such as stearin and magnesium. The required dose of the drug is filled in the capsule.

 Examples 8~14: Oral tablet drugs

 The composition of each drug is the same as in Examples 1-7.

 Preparation method: According to the methods of Examples 1 to 7, granulation and drying, sieving and adding an appropriate amount of talc enamel compression tablets, and then using a conventional coating method with a combination of HPMC propylene glycol, titanium dioxide, medical ethanol and Tween-g0 Coating the coated tablets with the liquid

 Example 1S~21: Oral granule analgesic

 The composition of each drug in parts by weight is shown in Table 2.

Preparation: The pulverized active pharmaceutical ingredient in a conventional pharmaceutical formulation and manner sieved particles, praseodymium starch powder and then added an appropriate amount of ethanol medical mixed, granulated and dried sieved on _s swing granulator, the desired dose points Packing _beta

 Table 2 Composition of granule analgesics

 Example 15 Example 16 Case 17 Case 1 § Example W Example 20 Case 21 Aconitine 8.9 8.9 8.9 8.9 ～ Aconitine 7.8 7.8 A 7.8 7.8 Monobenzoyl aconitine 41.1 A 41.1 41.1 A 41.1 Starch (X 1000) 800 800 800 800 800 800 800 Sugar powder (1 00) 200 200 200 200 200 200 200

10 Examples 22~28: Analgesic injection drugs

 The composition of each drug in parts by weight is shown in Table 3.

 Composition of each injection drug

 Example 15 Case 16 Case 17 Case 18 Case 19 Case 20 Case 21 Aconitine 8.9 8.9 8.9 8.9 8.9 Aconitine 7.8 7.8 7.8 7.8 Benzyl acyl aconitine 41.1 41.1 41.1 One one 41.1 Tween-80 (X 1000) 5 5 5 5 5 5 5

EDTA ( X 1000) 3 3 3 3 3 3 3 Water for Injection CX 1000) to 1000 to 1000 to 1000 to 1000 to 1000 to 1000 to 1000 to 1000

, Preparation method: Add the effective pharmaceutical ingredients to the appropriate amount of water for injection, Tween-80, EDTA and CMC-Na in the conventional range, stir well and mix with ultrasonic wave, fully dissolve, dilute with water for injection, filter, and use appropriate amount of hydrochloric acid when needed. Adjust pH 4-9 and fill with ampoule.

 Example 29~35: topical analgesics

The composition and dosage of each effective drug are the same as in Examples 15-21. • L painkiller plaster

Preparation method: The effective pharmaceutical ingredients are dissolved by heating with medical ethanol, filtered, and then mixed with 1IMK X 10 parts (both weights) of cetostearyl alcohol and 150 (X 10 parts) of hard fat 3⁄4, and heated and stirred. After uniform heating, it is a spare oil phase material. Separately, 10 _s 000 parts of sodium laurylsulfate stone block ₁₅₀ parts Teng Dou stirred uniformly mixed with the high-speed standby oil phase of the composition sufficient to Canadian ethyl paraben 1000 (X 1000) parts of purified water was heated After mixing and mixing, cool the package.

 2. Preparation method of analgesic patch (adhesive plaster): According to the conventional preparation method of adhesive plaster, 4 leak (X leak) rubber matrix is pressed and dipped, and effective pharmaceutical ingredients and 100 (X 1000) parts of Vase #, 50 ( X 1000) parts of lanolin 000 ( X 1000) parts of rosin 30 ( X 1000 ) parts of fillers such as zinc oxide and dispersing agent, mixed with paste and filtered, and coated with paste on the carrier The recovered solvent is cut to size and packaged.

 Examples 36~42: Analgesic Aerosols

 The composition of each drug in parts by weight is shown in Table 4.

 Table 4 Composition of various analgesic aerosols

 'Example IS Case 1 β Case 17 Case IS Case 1 Case 21 Aconitine 89 89 89 One S9 One - Lower Aconitine 78 7§ _ 7S Monobenzoyl Aconitine 411 A 411 411 One 411 2⁄4 411 fluoromethane (X 1000 ) 7000 7000 7000 7000 7000 7000 7000 Medical Ethanol (X 1000) 2500 2500 2500 2500 2500 2500 2500 Amount of appropriate amount of flavoring agent Appropriate amount of appropriate amount of preparation method: Adding effective pharmaceutical ingredients to medical ethanol, according to It is also necessary to add an appropriate amount of flavoring agent to mix and filter. Quantitatively loaded into a pressure-resistant container, pressurized according to the prescribed standard and injected into the microporous filtered dichlorodifluoromethane, which becomes an analgesic aerosol drug.

 Examples 43~49: Pain Relief / Lotion

 The effective drug composition of each case is the same as the above-mentioned analgesic aerosol.

 Preparation method: adding effective pharmaceutical ingredients to medical ethanol (300 (X 1000) parts for lotion; 100 (X 1000) parts for lotion), mixing and filtering, and adjusting to a total amount of 1000 (X 1000) parts with medical water , that is, the available analgesic/lust-fixing drugs respectively; after mixing and filtering, adding or not adding medical water as needed, quantitatively loading the pressure-resistant container and charging according to the prescribed standard, then becoming an analgesic aerosol Drugs.

 Examples 50-56: Analgesic Membrane Drugs

 The effective drug composition of each case is the same as the above-mentioned analgesic aerosol.

 Preparation method: After fully pulverizing the effective pharmaceutical ingredients, they are uniformly mixed with 100 (X 1000) parts of glycerin and about 5,000 parts of Tween-80, and then mixed with the solution prepared from polyvinyl alcohol (PVA). After defoaming, it is fully mixed with liquid paraffin to form a film, which can be dried.

 Example 57~63: Analgesic wet dressing

 The composition of each drug in parts by weight is shown in Table 5.

 Table 5 Composition of each wet dressing

Example 36 Case 37 Case 38 Case 39 Case 40 Case 41 Case 42 Aconitine 65 65 65 ― 65 65 Aconitine 65 65 One 65 One-one Benzopyrene Aconitine 130 130 130 One 130 Medical ethanol (X 1000) 200 200 200 200 200 200 200 Purified water (X 1000) 1000 1000 1000 1000 1000 1000 1000 Preparation method: Dissolve the effective pharmaceutical ingredient in medical ethanol, quantitatively dilute with water, and then apply medical gauze to the medicine. The gauze package with the drug solution is sufficiently immersed in the liquid. The beneficial effects of the present invention are demonstrated by pharmacodynamic test toxicological tests below. Test Example 1 Toxicological test of three aconitine compounds of the active ingredient of the present invention

 I. Acute toxicity test of lower aconitine

The test measured: Kunming mice were given a certain toxicity LD _{5 by} intragastric administration of the drug. The confidence limit of 12. 8 mg/kg, 953⁄4 is 10. 93~14. 99 mg/k _go

Determination of the median lethal dose (LD ₅ .)

 U test material

 1. 1 Test drug and preparation

 The medicinal aconitine white crystal granules, provided by the Chengdu Zhizhi Pharmaceutical Co., Ltd., the aconitine 20 mg, and the distilled water to 20 ml, diluted 1:0 to form a ratio of 0. 8C 0. 64, 0. 51, 0. 41 mg / ml - a series of concentrations of the drug solution.

 1. 2 test animals

Kunming mice, male and female, 18~22 g, provided by the Experimental Animal Center of Sichuan University, a ₃ -grade animal, certificate number: Chuan Shidong No. 67; Feed provider with animals, free Qin water breeding room The temperature is 21 ± 2 V and the relative humidity is: 50~60% ₀

 2. Test method

By pre-test to find the maximum lethal dose of 20 mg / kg, the minimum lethal dose of 5 mg / kg _ra water taken fasted for 14 hours 50 Kunming mice were randomly divided into 5 groups of 10 each, male and female 2 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 /10 The animal reaction was observed continuously for 7 days, and the toxic reaction and death of the mice were recorded day by day.

 3. Test results

 In the mice 10 minutes to 4 hours after gavage, most of them showed signs of fatigue, hypotonia, and vertical hair, and with increasing dose, there appeared an increasing degree of hiccup reaction and some or all animal death. The behavior, activity, diet, fur luster and weight gain of the undead mice returned to normal after one day. The dead mice were dissected, and no obvious abnormal changes were observed by visual observation of the major organs. After 7 days, the mice were sacrificed and dissected. No obvious abnormal changes were observed by the naked eyes. The death of mice in each dose group is shown in Table 6.

 Table 6 Number of mice killed in each dose of aconitine

 Group dose (mg/kg) log dose number of animals (only) number of deaths (only) mortality (%)

1 20. 00 1. 301 10 10 100

2 16. 00 1. 204 10 7 70

3 12. 80 1. 107 10 5 50

4 10. 24 1. 010 10 3 30

5 8. 19 0. 913 10 0 0

LD ₅ was calculated according to the Bliss method in the NDST software package. For a 12. 8 mg/kg, the 95% confidence limit is 10.93~; 14. 99 mg/kg.

 4 Conclusion

The drug aconitine has certain toxicity, Kunming mice are administered intragastrically, LD ₅ . For a 12. 8 mg/kg, the 95% confidence limit is 10.93~: 14. 99 mg/kgo

 Second, the acute toxicity test of aconitine

The test measured: Kunming mice were given a certain toxicity to the drug by intragastric administration of aconitine, 1^) ₅ . 6.41 mg/kg, The 95% confidence limit is 5. 19~7. 74 mg/kg _D

Determination of the median lethal dose (LD _M )

 experiment material

 1. 1 Test drug and preparation

 In the drug, aconitine, white crystalline granules, provided by Chengdu Zhizhi Pharmaceutical Co., Ltd. to take 15 mg of aconitine, add distilled water to 23. 4 ml (0. 64 rag/ml), take out 16 ml, ammonium 1: 0. 8 equal dilution was formulated into 0. 51 0. 41 , 0. 33 0. 26, 0. 21 mg / ml - a series of concentrations of the drug solution, spare.

 1. 2 test animals

 Kunming mice are male and female, 18~22 g, and the first-level animal certificate number is provided by the Experimental Animal Center of Sichuan University: Chuan Shidong Pipe Quality No. 67; Feeding provider is the same animal, free drinking water breeding room temperature is 21 Soil 2 V, relative humidity: 50~60%

 2, test method

Through pre-testing, the maximum lethal dose was 12.8 mg/kg, and the minimum lethal dose was 2.5 mg/kg. 60 Kunming mice were fasted and not for 14 hours, and were randomly divided into 6 groups. 2 ml / The volume of the aconitine administered to the mice in each group was 0. 2 ml / each of the two groups of mice was given a dose of 0. 2 ml / Immediately after the administration of 10 g _D , the response of the animals was observed. The toxic reaction and death of the mice were recorded on the 7th day.

 3. Test results

 In the mice 10 minutes to 4 hours after gavage, most of them showed fatigue and less movement, vertical hair, and increased dose-like hiccups with increasing doses and some or all animals died. Fur gloss and weight gain returned to normal. The dead mice were dissected, and no obvious abnormal changes were observed by visual observation of the major organs. Seven days later, the mice were sacrificed and the major organs were visually observed without obvious abnormal changes. The death of mice in each dose group is shown in Table 7.

 Table 7 Number of mice killed in each dose of aconitine in the drug

 Group dose (mg/kg) log dose number of animals (only) number of deaths (only) mortality (%)

1 12. 82 1. 108 10 10 100

- 2 10. 26 1. 011 10 8 80

 3 8. 21 0. 914 10 8 80

4 6. 56 0. 817 10 6 60

5 5. 25 0. 720 10 4 40

6 4. 20 0. 623 10 0 0 Calculate LD ₅ according to the Bliss method in NDST soft ί 牛. The preservative limit of 95% is 5.19~7. 74 mg/kg.

4 ^: , conclusion

Aconitine has certain toxicity in the drug, Kunming mice are administered once a day, LD ₅ . The preservative limit of 95% is 5.19~7. 74 mg/kg

 III. Acute toxicity test data of aconitine in benzoyl

 The test measured: The maximum tolerated dose of aconitine in the Kunming mice was 1 g/kg.

 Determination of maximum tolerated amount

 1. Test materials

 1. 1 test drug

The drug benzoyl aconitine, light yellow crystalline granules, has garlic odor, by Chengdu Zhizhi Pharmaceutical Co., Ltd. Offer

 1. 2 test animals

 Kunming mice are 18~22 g male and female, and the first-level animals are provided by the Experimental Animal Center of Sichuan University. Certificate No.: · Chuan Shi Dong Guan No. 67; Feeding Providers with Animals, Free Qin Water Breeding Room Temperature For 21 ± 2 3⁄4 pairs of humidity: δθ δθ^

 2, test method

 The drug benzoyl aconitine base granules were sonicated with a very small amount of ethanol, and then distilled water was added to prepare a concentration of 0.5 g/ml solution for 20 Kunming mice that were fasted for 14 hours. The stomach was given a solution of aconitine in the benzopyrene mill. 0. 2ml/10g, and the animal reaction was immediately observed. The behavioral activity of the mouse was recorded for 7 days and the skin weight and death of the skin were recorded.

 3, test results

In the mice 15 minutes to 4 hours after the gavage, most of the tired and less moving vertical hair, 4 hours after the behavioral active diet gradually returned to normal. During the observation period, no animals died. The animal's eating behavior was normal, and the normal hair color was normal, and the body weight was normal. (The body weight of the mice before the experiment was 19. 5 ± l. lg, and the weight was observed after 7 days. The weight was 25. 2 ± 1. 8g). In the future, the mice were sacrificed, and no obvious abnormalities were observed by naked eyes. Therefore, the maximum tolerated dose of benzoyl aconitine in a single gavage is 1 g/kg _e

 4 Conclusion

 The maximal tolerated dose of benzoyl aconitine in Kunming mice was 1 g/kg. The pharmacodynamic test was carried out to ensure the safety of the drug of the present invention. . Test Example 2 Analgesic effect test of three aconitine compounds of the active ingredient of the present invention

 1. Analgesia test of writhing mice

W ~ 22 g of body weight of the male and female Kunming mice as experimental animals were fed the same test under the conditions of test drugs _β: The following are aconitine (referred to as "component of Formula (Π)"), aconitine (referred to as "Formula (III) component"), benzoyl aconitine (referred to as "formula (IV) component") is a test drug. Wherein: The components of formula (II) are separately prepared into distilled liquids with a gradient content of 0.2, 0.1, 0.05 and 0.025 _mg / ml; the formula (ΙΠ) is formulated with distilled water to a gradient of 0.1, 0.05, 0.025 and 0.0125 mg, respectively. /ml solution: The compound of formula (IV) is formulated with distilled water to prepare a solution having a gradient content of 20, 10, 5 and 2.5 m _g /ml.

The control drug A was a drug solution in which the content of pethidine hydrochloride injection (Qinghai Pharmaceutical Factory, batch number 20010510) was diluted with physiological saline to a content of 2 m _g /ml.

 The control drug B was obtained by dissolving aspirated aspirin tablets (Nanjing Baijingyu Pharmaceutical Factory, batch number 031016) and dissolving in distilled water to obtain a drug solution of 10 mg/ml.

The mice were randomly divided into groups of 10, one of which was a blank control group, which was given distilled water at a dose of 0.2 ml/10 g body weight; the positive control group A was 40 mg/kg body weight (equivalent to 20 times the clinical dose). The dose was administered intraperitoneally to the control drug A; the positive drug group B was administered with the control drug B at a dose of 200 m _g / kg body weight (equivalent to 20 times the clinical dose); the rest was administered by different dose gradients. The present invention corresponds to a test group of test drugs. The administration volume of each of the administration groups was 0.2 ml/10 g body weight. Except the control drug group A, after intraperitoneal injection of 0.6% glacial acetic acid solution 0.2ml/only after 15 minutes of administration, all the other groups were intraperitoneally injected with 0.2% glacial acetic acid solution 0.2ml/only after 30 minutes of administration. The number of writhing responses (abdominal hind leg extension, buttocks elevation) of each mouse within 5 to 20 minutes after acetic acid solution, and statistical analysis of the test results, calculating the writhing mean and standard deviation of each test group Pain inhibition rate, the difference between the groups of each administration group and the negative control group was compared by t test. The test results are shown in Table 8. Torsion test results of mice analgesia test

 Group. Dosage dose (mg/kg) Number of writhings (Ϊ ±§Β) Pain control rate (%) Blank control group - 39.3±13.S ―

 Control drug A 40 100.00

 Control drug B 200 3.1±3.2

 4 92.SS

2 23.6 ± 10.9 ^3⁄4

 . (Π) component

 1 · 37.66

 0.5 27.6±11.5 29.77

 2 50.89

1 21.§±12.6 ^ΔΔ 44.53

 Formula (no component

0.5 20.0 ± 9.6 ^ΔΔ

 0.25 33.6±11.5

400 20.4±12.9 ^ΔΔ Q.52

200 21.0±15.1 ^Δ 3S.78

 Formula (IV)

 100 27.4±1S.9 20.12

 50 + 1 20.70

Note: Compared with the blank group, *P<0.05, "*Ρ<0.01 ₉ ***Ρ<0.001; ^Λ Ρ<0.01;

<0.05 ₉ ^P<0.01 o

The above test results showed that the analgesic effects of the control drugs A and B were very significant, and the pain inhibition rates were 100% and 92.11%, respectively. In the formula (Π) of the aconitine component, except for the 0.5mg/kg dose group, the other groups can have different degrees of obvious analgesic effect; the formula (ΙΠ) of the aconitine component, except. In the 25mg/kg group, the other groups can also have obvious analgesic effect; the benzoyl aconitine til base in formula (IV) can be compared in the 400 mg/kg and 200 mg/k _g two dose groups. Significant analgesic effect. At the same time, the above results show that there is a dose-dependent inhibition rate of the three compounds on the pain.

 2: Hot plate method mouse analgesia test

 For the test animals and their groupings, the dose gradients, the doses and the administration methods of the test drugs were the same as those of the above-mentioned writhing test, and the control drug group only used pethidine hydrochloride as a control drug.

 The temperature of the constant temperature water bath was controlled to 55±0.5°C, and the mice were placed in a constant temperature aluminum drum, and the time from the placement to the hindfoot (pain threshold) was recorded, and the measurement was repeated twice, and the measurement interval was 5 times. Minutes, the average value is the pain threshold before the mice. Mice with a pain threshold of 5 to 30 seconds were grouped into groups of 10 per group. The pain thresholds of the mice were measured twice at 15, 30, 60, 90 and 120 minutes after administration, and the average pain threshold and pain threshold increase rate at each time point were calculated. Statistical analysis was performed on the test data, and the pain threshold increase rate and standard deviation of each group were calculated. The t test was used to compare the differences between the groups of the drug-administered group and the negative control group. During the test, the mice developed painful reactions such as lameness, kicking hind legs and jumping under thermal stimulation. Only the hind paws were selected as the pain response index. The test results are shown in Table 9.

'Rotary column - blank group twist = twisted times X let (the same below) Hot plate method mouse analgesia test results

 Dose 'pain B increase rate after administration (%) (X soil SD)

Group

 15 minutes 30 minutes 60 minutes 90 minutes 120 minutes Empty &3⁄4 photos A 12.1±45.0 -6.3±31.4 -0.5±3S.7 -11.3±3 .2 ll.l±3g.8

 40 284.4±229.5" 1S1.3±^.4'" 11§.9± 117.5** 0.7 soil 48.3

4 142.7± 139.4* 104.1± 137.4* 81.7+100.0* 34.4+59.3* S9.1 soil engraving) 2 72.4±57.6' 36.0 ±53.4 17.3 soil 56.1 1S.3±60.1 -13.0±33.1

 1 -16.6'±31.9 -31.9±24·2 -18.1±34.S -13.0±1 . 2§.3±S6.3

100 124.S± 16S.4 64.3 ±149.5 10δ.3±1ί)5·0 ^ΔΔ 49.0 ±52.9

50 21.4±42.9 14.4±34.7 7.9 ±37.3 30.4± β.1 ^Δ 36.0 ±42.6

25 39.7 ±76.6 59.2 ±146.9 37.2 ±88.5 61·3±32·1 ^Δ S0.7± 100.1

12.5 12.1±45.0 -6·3±31.4 -0.5±38.7 -11·3±34.2 11·1±3§.8

400 41.3±90.2 3S.2±57.2 ^A S3.7± 117.0

200 22,1±52.0 32.9士45·1 ^Δ 40.3 ±76.4 46.0±87.4 53.6±93.9

100 -0.4±33.8 13.7±52.3 64.6±76·7 ^Δ 71.1 ±104.5 106.2±116.5 Note: Compared with the blank group, *P<0.05, **P<0.01, ***Ρ<0.001; ^Δ Ρ<0.05 , ^ΔΑ Ρ<0.01

< 0.1) 5. Ρ <0.01

 It can be seen from the table that the active ingredients of the present invention have obvious analgesic effects and have a linear effect in a single dose. The pain threshold of the high dose group of the aconitine component of formula (II) was significantly increased at each time point measured, and the analgesic effect of the control drug meperidine hydrochloride was maintained for a long time, within 2 hours. Have obvious analgesic effect

 Test Example 3 Analgesic effect test of different ratios of aconitine in benzoyl and aconitine

The test showed that the drug benzoyl aconitine 100 mg / kg and the lower aconitine lmg / kg dose have an obvious analgesic effect, the pain inhibition rate was 33.7% and 46.9%. The 300:1, 100:1, 100:3 ratio of benzoyl aconitine and aconitine also significantly inhibited the writhing reaction in mice induced by acetic acid, and the pain inhibition rate was 57. 1%, 61.7% and 40.0%, of which 300: 1, 100: 1 ratio of the dose group is higher than 100mg/kg of benzoyl aconitine and 1mg/k _g of aconitine . It can be seen from the pain inhibition rate that when the ratio of aconitine and aconitine in benzoyl is 100:1, the analgesic effect is the strongest. It can be considered that the best ratio of analgesic effect of aconitine and aconitine in benzoyl is 100:1.

 First, the test materials

 .1, test drugs and preparation

 Benzoyl aconitine, light yellow powder; Hypaconitine, white crystalline granules, supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. Weigh 250 mg of aconitine in benzoyl, add 2.5 ml of absolute ethanol, heat in a water bath at 42 ° C and solubilize with ultrasound, add distilled water to 25 ml, and prepare benzoyl aconite. a solution of 10 mg/ml of the base; weigh 2. 5 mg of aconitine, add 2.5 ml of absolute ethanol, and after heating in a water bath at 42 ° C and ultrasonic solubilization, add distilled water to 25 ml to prepare a mixture. A solution of aconitine 0.1 mg/ml. According to Table 10, different volumes of benzoyl aconitine base V under aconitine solution were prepared and formulated into different proportions of benzoyl aconitine and aconitine solution.

Aspirin enteric-coated tablets, white, 25 mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312. Take 4 tablets of aspirin enteric-coated tablets, grind them, and use distilled water to 10 ml to prepare a liquid containing aspirin 10 mg/ml. Take 1ml of absolute ethanol, add distilled water to 10 ml of 103⁄4 ethanol

Kunming mice, male and female, 18~23⁄4, provided by the Experimental Animal Center of Sichuan University, the first-class animal certificate number: No. 10; Feed provider with animals, free drinking water, observation room temperature is 21±2Ό, relative humidity For: 5Q~60% _D

 3, reagents

 Glacial acetic acid: Analytically pure, Chengdu Chemical Reagent Factory Batch No.: 2005042

 Anhydrous ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory Batch No.: 20050313

 Second, the test method

70 Kunming mice were randomly divided into 7 groups, 10 rats, male and female weighing 18~22g _e Group 1 was the negative control group was given 10% ethanol 0.2 ml / 10g body weight; Group 2 positive The control group was given 200 mg/kg of escitalin (equivalent to 20 times of clinical dose); Groups 3, 4, 5, and 6. were the aconitine and aconitine in the drug benzamidine and its Three different proportions of the dose group were given benzoyl aconitine, aconitine and three different ratios, respectively. The dosages of each group were 0.2 ml/lOg, and 30 doses were administered. After a minute, intraperitoneal injection of 0.2% glacial acetic acid solution 0.2 ml / only, observe the writhing reaction of each mouse within 10 minutes after the injection of acetic acid solution for 5 minutes (abdominal hind leg extension, hip height) statistics on the test data The writhing mean, standard deviation and pain inhibition rate of each dose group were analyzed and calculated. The difference between the groups of the drug-administered group and the negative control group was compared by t test.

 Third, the test results

 Injection of acetic acid can cause longer-lasting painful stimulation in mice. The mice repeatedly have abdomen and hind leg extension, and the hips are high (twisted). In the aspirin-positive control group, the number of writhing in mice induced by glacial acetic acid was significantly decreased (P<0.001), and the analgesic effect was very obvious. The pain inhibition rate was 82.9%. The drug benzoyl aconitine 100mg/kg and lower aconitine lmg/kg doses significantly reduced the writhing response induced by acetic acid in mice, and the pain inhibition rates were 33.7% and 46.9%, respectively. 300:1, 100:1, 100:3 ratio of benzoyl aconitine and aconitine also inhibited the writhing reaction induced by acetic acid in different degrees. The number of writhing and negative control Compared with the group, there was a significant difference, showing obvious analgesic effect; the pain inhibition rate was 57.1%, 61.7% and 40.0%, respectively, of which 300:1, 100:1 ratio group was higher than 100mg/kg. Benzoic acid aconitine and lmg/kg of aconitine, the results are shown in Table 11. From the perspective of pain inhibition rate, the analgesic effect was the strongest when the ratio of aconitine and aconitine in benzoyl was 100:1. It can be considered that the best ratio of analgesic effect of aconitine and aconitine in benzoyl is 100:1.

 Table 11 Effect of different ratios of benzoyl aconitine and aconitine on pain response in mice induced by acetic acid. Group dose (mg/kg)

 Group animal number writhing number pain inhibition rate benzoyl aconitine aconitine - (only) (JC ± SD) (%) Negative control 11 - 10 17.5 ± 7.1 one aspirin - ― 10 3.0±2.9*** 82.9

 1 100 ― 10 11.6±5.2* 33.7

2 ― 1 10 9.3±7.8* 46.9

3 150 0.5 10 7.5 ±4.5** 57.1

4 100 1 10 6.7 ±5.2** 61.7

5 50 1.5 10 10.5 ±7.6* 40.0

:: t test, compared with the negative control group, *p<0.05, **p<0.01, * *p<0.001 Four test conclusions

The benzoyl aconitine 100mg/kg and the lower aconitine lm _g /k _g doses had obvious analgesic effects, and the pain inhibition rates were 33.7% and 46.93⁄4, respectively. 300:1 100:1, 100:3 ratio of aconitine in benzoyl aconitine also inhibited the writhing reaction of mice induced by acetic acid. The pain inhibition rate was 57.1%. 61.7% and 40.0% of the 300:1, 100:1 ratio of the dose group higher than 100mg/kg of benzoate in the aconitine and lmg/kg of the lower aconitine _D from the pain inhibition rate, to benzene The dosage and the efficacy of the combination of aconitine and aconitine in Jia 3⁄4 were not dose-dependent. When the ratio of the two was 100:1, the dosage was less and the analgesic effect was the strongest, achieving the goal of high efficiency and low toxicity.

 Test Example 4 Analgesic effect test of different ratios of aconitine and mesaconine

The test proved that the drug aconitine lmg / kg and aconitine 0.5mg / kg dose have obvious analgesic effect, the pain inhibition rate of 46.9% and 73.13⁄4 _D under aconitine, aconitine The 6:3U 2:1. 2:3 ratio drug also significantly inhibited the writhing response in mice induced by acetic acid, and the pain inhibition rates were 74.33⁄4 66.9% and 78.3%, respectively. From the pain inhibition rate, it can be seen that the following analgesic effect is the strongest when the ratio of aconitine and aconitine is 2:3. The best analgesic effect of hypocotyl and aconitine is the best. The ratio is 2:3 _Q

 First, the test materials

 1. Test drugs and preparation

Aconitine white crystalline granules; aconitine, near white powder, supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. Weigh 2.5 mg of aconitine, add 2.5 ml of absolute ethanol, heat in 42 Ό water bath and sonicate, add distilled water to 25 ml, and prepare a solution containing 0.1 mg/ml of aconitine; Head base 1.25 mg, add 2.5 ml of absolute ethanol, heated in 42 ° water bath and sonicated, add distilled water to 25 ml, and prepare a solution containing aconitine 0.05 mg / ml. According to Table 12, different volumes of hypoxazone and aconitine solution were prepared into different ratios of hypoxazone and aconitine solution, and spare _D

 Aspirin enteric-coated tablets, white, 25nig/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd. Batch number: 20050312. Take 4 tablets of aspirin enteric-coated tablets, grind them, and use distilled water to 10 ml to prepare a liquid containing aspirin 10 mg/ml.

 Take 1 ml of absolute ethanol and add distilled water to 10 ml to make 10% ethanol.

 Table 12 Lower aconitine / aconitine different ratios

 Group dose (mg/kg)

Group - Sample Volume (ml)

 Proportion of the group, under the sulphate, aconitine, distilled water, lower aconite, aconite

 2.5 ― 2.5 1

 2 one one 2.5 2.5 0.5

 3 3.75 1.25 1.5 0.25 6: 1

4 . 2.5 2.5 1 0.5 2: 1

5 1.25 3.75 0.5 0.75 2: 3

2. Test animals

 Kunming mice, male and female, 18~22 g, provided by Experimental Animal Center of Sichuan University, primary animal, certificate number: No. 10; vocabulary provider with animals, free drinking water, observation room temperature is 21± 2 ° C, relative humidity: 50~60%.

 3, reagents

 Glacial acetic acid.: Analytically pure, Chengdu Chemical Reagent Factory, batch number: 20050423.

 Anhydrous ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory, batch number: 20050313.

Second, the test method Seventy Kunming mice were randomly divided into 7 groups, 10 males and a half each, weighing 18-22 g. Group 1 was given a negative control group with 10% ethanol 0.2ml/10g 侔 weight; Group 2 was a positive control group, and aspirin was given 200 mg/kg by gavage (equivalent to 20 times the clinical dose); _3⁄4 5, 6. Group 7 was given aconitine, aconitine and its three different ratios in the treatment of aconitine and its three different ratios. The dosage volume of the group was 0.2 ml/lOg, and 0.2 ml/0.6 ml of glacial acetic acid solution was intraperitoneally injected 30 minutes after the administration, and the writhing reaction of each mouse within 10 minutes after the injection of the acetic acid solution for 5 minutes was observed. (The number of abdomen hind legs stretched up the hips). Statistical analysis was performed on the test data to calculate the mean, standard deviation and pain inhibition rate of the writhing body in each dose group. The difference between the groups of the drug-administered group and the negative control group was compared by t test.

 Three test results

Injection of acetic acid can cause longer-lasting pain and stimulation in mice, and the mice repeatedly appear in the abdomen and hind legs and the hips rise (twist #) ^: reaction. Aspirin yang. Sexual control group significantly decreased the number of writhing in mice induced by glacial acetic acid

(P < 0.001), the analgesic effect is very obvious, the pain inhibition rate of the drug aconitine Img / kg and aconitine 0.5mg / kg dose have significantly reduced the writhing reaction caused by acetic acid in mice The pain inhibition rates were 46.9% and 73.13⁄4, respectively. The 6:1, 2:1, 2:3 ratio of the lower aconitine and aconitine also inhibited the writhing reaction induced by acetic acid in different degrees. Compared with the negative control group, show significant differences obvious analgesic effect; their pain inhibition rate was 66.9% and 78.3% 74. respectively, results shown in Table ₁₃ Θ inhibiting amount of the composition from the ratio of pain, Bian aconitine aconitine proportions There is no dose-dependent effect on the drug; when the ratio is 2:3, the dosage is small, the analgesic effect is the strongest, and the purpose of high efficiency and low toxicity is achieved.

 Table 13 Effects of different ratios of aconitine and mesaconitine on pain response in mice induced by acetic acid Groups (mg/kg) Dynamic « Number of writhings (soil SD) Pain inhibition rate (%)

 Aconitine

 Negative control 1 - 10 17.5 ± 7.1 - - Aspirin - One 10 10 3.0 ± 2.9 *** 82.9

1 1 ― 10 9.3±7.8* 46.9

2 ― 0.5 10 4.7±6.8*** 73.1

3 1.5 0.25 10 4.5 ±4.1*** 74.3

4 1 0.5 10 5.8 ±4.3*** 66.9

5 0.5 0.75 10 3.8 ±3· 7*** 78.3 Note: t test, compared with the negative control group, <0.05, "p<0.01," *p<0.001

 Fourth, ΐ 验 test conclusion

 The drugs had a significant analgesic effect on the aconitine lmg/kg and the aconitine 0.5 mg/kg dose, and the pain inhibition rates were 46.9% and 73.1%, respectively. The 6:1, 2:1, and 2:3 ratio drugs of hypocotyl and aconitine also significantly inhibited the writhing reaction induced by acetic acid in mice, and the pain inhibition rates were 74.3%, 66.9%, and 78.3%. It can be seen from the pain inhibition rate that the following analgesic effect is the strongest when the ratio of aconitine and meconium is 2:3. It can be considered that the best ratio of the analgesic effect of the lower aconitine and the aconitine is 2:3.

 Test Example 5 Analgesic effect test of different ratios of aconitine and aconitine in benzoyl

The experiment showed that the drug benzoyl aconitine 100rag/kg and mesaconitine 0.5mg/kg dose had obvious analgesic effect, and the pain inhibition rates were 33.7% and 73.1%, respectively. The 600:1, 200:1, and 200:3 ratio of benzoyl aconitine and aconitine also significantly inhibited the writhing reaction in mice induced by acetic acid, and the pain inhibition rate was 51.4. %, 75.4% and 84.6%, of which the ratio of 200:1, 200:3 was higher than 100 mg/kg of benzoyl aconitine and 0.5 mg/kg of aconitine. It can be seen from the pain inhibition rate that as the proportion of aconitine increases, the analgesic effect gradually increases. When the ratio of aconitine and aconitine in benzoyl is 200:3, the town has the strongest effect. Can be considered as: benzoyl aconitine, aconitine analgesic as the strongest best match The ratio is 200:3.

 Test material

 K test drug and preparation

Benzoyl aconitine light yellow powder; Aconitine near white powder is supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. Weigh benzoyl aconitine 250 mg, add 2.5 ml of anhydrous acetamidine, After heating in 42 °C water bath and ultrasonic assisted solution, add steamed water to 25 ml to prepare a solution containing 10 mg/ml of benzoyl aconitine; weigh aconitine. 1.25 mg added 2.5 ml water, ethanol, 42 ° C water bath by heating and ultrasonic solubilization add distilled water to 25 ml, formulated in a solution containing the following table _β aconitine 0.05 mg / ml of benzoyl taken in different volumes of aconine Aconitine solution is formulated into different proportions of benzoyl aconitine aconitine solution, spare

Aspirin enteric-coated tablets, white 25mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312 ₀ Take 4 tablets of aspirin enteric-coated tablets, grind, use distilled water to 10 ml, and prepare liquid medicine containing aspirin 10 mg/ml spare"

Take 1 ml of anhydrous ethanol, add distilled water to 10 ml of 10% ethanol dubbed _Q

 Different proportions of benzoyl aconitine and aconitine

 Group • Sampling volume (ml) Group dose (rag/kg)

 Proportion of the group, benzoyl aconitine, aconitine, distilled water, benzoyl aconitine, aconitine

 1 2.5 —― 2.5 100 —― —―

2 —- 2.5 2.5 —― 0.5

 Kunming mice, male and female, 18 22 g, provided first-class animals by the Experimental Animal Center of Sichuan University, certificate number: No. 10; vocabulary provider with animals, free drinking water, observation room temperature is 21±2Ό , relative humidity: 50 60%

 3, reagents

 Glacial acetic acid: Analytically pure, Chengdu Chemical Reagent Factory, batch number:

 3⁄4 water ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory, batch number:

 Second, the test method

 Seventy Kunming mice were randomly divided into 7 groups, 10 in each group, half male and half female, weighing 18 22g. Group 1 was a negative control group, and 10% ethanol was administered orally with 0.2 ml/10 g body weight; Group 2 was a positive control group, and aspirin was given 200 mg/kg by gavage (equivalent to 20 times the clinical dose); 5 6 7 groups were the drug benzoyl aconitine, mesaconitine and its three different ratios. The benzoyl aconitine 3⁄43⁄4, aconitine and 3 of them were administered by gavage. Different ratios of the drug solution, each group of the drug volume was .2 ml / lOg, intraperitoneal injection of 0.6% glacial acetic acid solution 0.2 ml / only 30 minutes after administration, observed within 10 minutes after the injection of acetic acid solution for 5 minutes The number of times the writhing reaction of the mouse (the abdomen hind legs stretched and the buttocks rose). Statistical analysis was performed on the test data, and the mean, standard deviation and pain inhibition rate of the writhing body in each dose group were calculated, and the difference between the groups of the drug-administered group and the negative control group was compared by t test.

 Third, the test results

Injection of acetic acid can cause longer-lasting painful stimulation in mice. The mice have repeated abdominal and hind leg extension, and the buttocks rise (the writhing reaction. Aspirin-positive control group, the number of writhing in mice caused by glacial acetic acid decreased significantly (P< 0.001), the analgesic effect is very obvious, the pain inhibition rate is 82.9%. The drug benzoyl aconitine lOOmg/kg and mesaconitine 0.5mg/kg dose have significantly reduced mice induced by acetic acid Twisting reaction, its pain Pain inhibition rates were 33.7% and 73.1¾ "in aconine benzoyl aconitine 600: 1,200: 1,200: 3 ratio of drugs have varying degrees of inhibition of writhing in mice caused by acetic acid _s The number of writhings was significantly different from that of the negative control group, showing a significant 12345 difference, showing a significant analgesic effect; the pain inhibition rates were 51.43⁄4, 75.43⁄4 and 84.6%, respectively, of which 200:1, 200:3 ratio In the dose group, the aconitine and the aconitine of 0.5 mg/kg were higher than 100 m _g /kg. The results are shown in Table 15. From the inhibition rate of pain, the aconitine in aconitine in benzal milling stoichiometric amount of the composition and the pharmaceutical efficacy and dose not depend Lennon; ratio of 200: 3-benzoyl pharmaceutical strongest in table 15 with less aconine _s analgesic effects, with different aconitine Comparison of the effects of acetic acid on pain response in mice

 Group dose (mg/kg)

 Negative control 10 17.5±7.1

 Aspirin 10 3.0±2- 82.9

 10 11.6±5.2' 33.7

0.5 10 4.7 ±6.8**' 73.1 0.25 10 8.5±6.4** 51.4 0.5 10 4.3 ±3.9** ^f 75.4 0.75 10 2.7±3.3 ⁱ 14 Note: The t test is <0.05, p<0.01 compared with the negative control group. , <0.001

 Fourth, the test conclusion '

The drug benzoyl aconitine 100mg/kg and mesaconitine 0.5m _g / kg dose have obvious analgesic effect, the pain inhibition rate is 33.7% and 73.1%. The 600:1, 200:1, 200:3 ratio of benzoyl aconitine and aconitine also significantly inhibited the writhing reaction in mice induced by acetic acid, and the pain inhibition rate was 51.4. %, 75.4% and 84.6%, of which the ratio of 200:1, 200:3 in the dose group was higher than 100 mg/kg of benzoyl aconitine and 0.5 mg/kg of aconitine. It can be seen that as the proportion of aconitine increases, the analgesic effect gradually increases. When the ratio of aconitine and aconitine in benzoyl is 200:3, the analgesic effect is the strongest. It can be considered that the best ratio of analgesic effect of aconitine and aconitine in benzoyl is 200:3.

 Test Example 6 Analgesic effect test of benzoyl aconitine, aconitine and mesaconine in different proportions.

 Tests have shown that the combination of benzoyl aconitine, aconitine and lower aconitine has a significant analgesic effect. In benzoyl aconitine, Zhongwutou The ratio of alkali to hypocotamine is 200:1:2, 400:1:2, 100:1:1, and 200:1:4. The ratio of 100:1:1 is the strongest analgesic effect. Therefore, the best ratio of benzoyl aconitine, aconitine and hypoaconitine is 100:1:1.

 , experiment material

 1. Test drugs and preparation

Benzoyl aconitine, light yellow powder; aconitine, near white powder; hypocotyl, white crystalline granules, supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. Weigh 250 mg of acetoyl aconitine, add 2.5 ml of anhydrous alcohol, heat in 42 °C water bath and sonicate, add distilled water to 25 ml, and prepare benzoyl aconitine 10 m _g /ml solution; weigh 1.25 m _g of aconitine, add 2.5 ml of absolute ethanol, after heating in 42 ° C water bath and ultrasonic solubilization, add distilled water to 25 ml, and prepare aconitine a solution of 0.05 mg/ml; weigh 2.5 mg of aconitine, add 2.5 ml of absolute ethanol, heat it in a water bath at 42 ° C and solubilize with ultrasound, add distilled water to 25 ml, and formulate aconitine 0.1 mg. /ml solution. According to Table 16, take different volumes of benzoyl aconitine, aconitine, and aconitine solution, and prepare different proportions of benzoyl aconitine, aconitine, and lower aconite. Alkaline solution, spare. Table 16 Different ratios of aconitine and aconitine in aconitine in benzamidine

 Take volume (ml) group dose (mg/kg)

 Group

 Among the benzoyl groups, benzoyl group, the proportion of the mixture, aconitine, aconitine, aconite, lower aconite, 3⁄4

 Aconitine aconitine

 1 1.67 1.67 1.67 33 0.167 0.33 200:1:2

2 1.25 1.25 50 0.125 0.25 400:1:2

1.25 2.5 1 · 25 25 0.25 0.25 100:1:1

4 1.25 1.25 2_ 5 25 0.125 0.5 200:1:4 Aspirin enteric-coated tablets, white, 25 mg/tablet Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312. Taking Aspirin 4 with distilled water to 10 ml, formulated with a chemical called aspirin 10 mg / ml backup _Θ

Take 1 ml of anhydrous ethanol, add distilled water to 10 ml of 10% ethanol dubbed _s

2 _3⁄4 test animals

 Kunming Mingming mice, male and female, 18~22 g, provided by Experimental Animal Center of Sichuan University

Certificate No.: No. 10; Feed provider with animal, free drinking water observation room temperature is 21±2Ό:

Degree is: 50 ~ hidden.

 3, reagent

 Glacial acetic acid: Analytical purity Chengdu Chemical Reagent Factory, batch number: 20050423

 Anhydrous ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory, batch number: 20050313

 Second test method

Kunming mice were randomly divided into 6 groups, 10 per group, male and female weighing 18 ~ 22g _Q Group 1 was the negative control group, 10% of intragastric administration of ethanol 0.2ml / 10g body weight; Group 2 For the positive control group, aspirin was given 200 mg/kg by gavage (equivalent to 20 times of the clinical dose); the third group was the drug of benzoyl aconitine, aconitine and hypoaconitine. Four different proportions of the dose group were administered with 4 different ratios of benzoyl aconitine and aconitine and aconitine. The dosages of each group were 0.2ml/10g. After 30 minutes of administration, a 0.6% glacial acetic acid solution (0.2 ml/mouse) was intraperitoneally injected, and the number of writhing responses (abdominal hind leg extension, buttocks height) of each mouse within 10 minutes after the injection of the acetic acid solution for 5 minutes was observed. Statistical analysis was performed on the test data, and the mean, standard deviation and pain inhibition rate of the writhing body in each dose group were calculated. The difference between the groups of the drug-administered group and the negative control group was compared by t test.

 Third, the test results

 Injection of acetic acid can cause longer-lasting painful stimulation in mice. The mice repeatedly have abdomen and hind leg extension, and the hips are high (twisted). In the aspirin-positive control group, the number of writhing in mice induced by glacial acetic acid was significantly decreased (P<0.001), and the analgesic effect was very obvious. The pain inhibition rate was 88.0%. The compound benzoyl aconitine, aconitine and hypoaconitine have a ratio of 200:1:2, 400:1:2, 100:1:1 and 200:1:4. The writhing response of mice induced by acetic acid was reduced to some extent. The number of writhing reactions in mice was significantly different from that of the negative control group, showing obvious analgesic effect. The pain inhibition rate was 85.0%. 65.3%, 90.1%, and 86.1%. The results are shown in Table 17. From the perspective of pain inhibition rate, the ratio of aconitine, aconitine and hypoaconitine in benzoyl is not dose-dependent; the ratio is 100:1:1, the dosage is small. The analgesic effect is the strongest, achieving the goal of high efficiency and low toxicity.

 Table 17 drugs benzoyl aconitine, aconitine, hypoaconitine different ratio

 Effect of acetic acid on pain response in mice

 Group dose (mg/kg) number of animals writhing number pain suppression group

 Aconitine aconitine in benzoyl aconitine (only) (X soil SD) rate (%) Negative control ― ― 10 27.4±10.1 ― Aspirin one ― 10 3.3 ±3.2*** 88.0

1 33 0.167 0.33 10 4· 1±4· 0*** 85.0

2 50 0.125 0.25 10 9.5 ±6.7*** 65,3

3 25 0.25 0.25 10 2.7 ±3.3"* 90.1

4 25 0.125 0.5 10 3.8 ±3· 7*** 86.1 Note: The t test is 3⁄4<0.05, *p<0.01 *3⁄4<0.001 compared with the negative control group.

 Fourth, the conclusion of the test

 The compound benzoyl aconitine, aconitine and lower aconitine have different analgesic effects. The aconitine and wuwu in the benzoyl aconitine The ratio of head to base ratio is 200:1:2, 5 400:1:2, 100:1:1 and 200:1:4, the analgesic effect is the strongest at a ratio of 100:1:1, so benzoyl The best analgesic effect of aconitine and aconitine in Zhongwutou base is 100:1:1.

 Test Example 7 Analgesic effect test of intramuscular injection of aconitine in benzoyl

The test proved that the intramuscular injection of benzoyl aconitine 100 and 25 mg / kg doses have obvious 10 pain effects, and the analgesic effect is significantly stronger than 100m _g / kg. 98.6% and 93.5%, and the pain inhibition rate of intragastric administration of 100mg/kg was 37.2, but the same dose of toxic reaction was also more important than intragastric administration, indicating that benzoyl aconite was used in commercial applications. when alkali intramuscular dose oral administration should be much lower than to avoid the occurrence of toxicity rabbit _β

 First, the test materials

 15 1. Test drugs and preparation

 Benzoyl aconitine, light yellow powder was supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. Weighed 125 mg of benzoyl aconitine, added 1.25 ml of absolute ethanol, heated by 42 Ό water bath and ultrasonically assisted. Serve to 25 ml., prepare a solution containing 5 mg/ml of aconitine in benzoyl, take 2.5 ml, add normal saline to 10 ml,

, formulated into a 1.25mg/inl solution, ready for use. '

 '20 Aspirin enteric-coated tablets white, 25mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312. Take 4 tablets of aspirin enteric-coated tablets, grind, use distilled water to 10 ml, and prepare medicine containing aspirin 10fflg/ml 0.5 ml of absolute ethanol was added to the solution, and physiological saline was added to 10 ml to prepare 5% ethanol.

 , 2, test animals

 Kunming mice, male and female, 18~22 g, provided by Experimental Animal Center of Sichuan University, first-class animals, certificate number: 笫10; vocabulary provider with animals, free drinking water, observation room temperature is 21± 2 ° C, relative humidity: 50~60%.

 3, reagents

 , Glacial acetic acid: Analytically pure, Chengdu Chemical Reagent Factory, Lot No.: 20050423

 0 absolute ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory, batch number: 20050313

 , experiment method

 Fifty Kunming mice were randomly divided into 5 groups, 10 in each group, half male and half female, weighing 18~22g. Group 1 was a negative control group, and 5% ethanol was given to the leg muscles by 0.2 ml/10 g body weight; the second group was a positive pair.

, according to the group, intragastric administration of aspirin 200 mg / kg (equivalent to 20 times the clinical dose); group 3 by intragastric administration of 5mg / ml

:) 5 benzoyl aconitine solution, group 4 intramuscular injection of 5 mg / ml benzoyl aconitine solution, group 5 intramuscular injection of 1.25 mg / ml of benzoyl zhongwu The cephalospergic acid solution was administered at a volume of 0.2 ml/lOg in each group. 0.2 ml of glacial acetic acid solution was intraperitoneally injected 30 minutes after administration, and each small 10 minutes after the injection of the acetic acid solution for 5 minutes was observed. The number of times the rat's writhing reaction (the abdomen's hind legs stretched and the buttocks rose). Correct

According to the statistical analysis, the writhing mean, standard deviation and pain inhibition rate of each dose group were calculated, using t test,

Λ0 is different from the group of each administration group and the negative control group.

 Third, the test results

Injection of acetic acid can cause longer-lasting painful stimulation in mice, and the mice repeatedly have abdomen hind legs stretched and hips twitched (twisted body). In the aspirin-positive control group, the number of writhing in mice induced by glacial acetic acid decreased significantly (Ρ<0.001), and the analgesic effect was very obvious. The pain inhibition rate was 81.6%. The drug benzoyl aconitine tendon 5 meat injection and intragastric administration of 100m _g / kg can significantly reduce the writhing response induced by acetic acid in mice, the pain inhibition rate was 98.6% and ' 37.2%. However, the analgesic effect of intramuscular injection was significantly stronger than that of intragastric administration. Even if the intramuscular injection of 25 mg/kg was more effective than 100 mg/kg, the pain inhibition rate was 93.5%. 18. We also found that mice injected intramuscularly with 100 mg/k _{g of} benzoyl aconitine showed atrophy and decreased movement. The toxicity of the drug is more obvious. In practice, the dosage of benzoyl aconitine in intramuscular injection should be much lower than that of intragastric administration to avoid the occurrence of toxicity.

 Effect of benzoyl aconitine on pain response in mice induced by acetic acid

 Group 3⁄4Jfi(mg/kg) Route of administration Animal convergence (only) Number of writhing (soil SD) 3⁄43⁄4 window » System rate (%) Negative control One by one im. 10 27. 7± 12· 4 ―

 Aspirin 200 ig. 10 5. 1±6. 0*** 81. 6 Benzoyl 100 ig. 10 17. 4±8· 6* 37. 2 Aconit original monument 100 im. 10 0· 4±0 · 8*** 98. 6

25 10 1 8 + 3⁄4 {3⁄4*** 93. 5 Note: t test _s compared with the negative control group: 'ρ<0. 05» * ρ<0. 01, ***ρ<0. 001

 Four test conclusions

 The muscle-stimulating drugs benzoyl aconitine 100 and 25 mg/kg had significant analgesic effects, and the analgesic effect was significantly stronger than that of 100 mg/kg. The pain inhibition rate was 98.63⁄4. And 93.5%, and the pain inhibition rate of lOOmg/kg by intragastric administration is 37.2% „but the same dose of toxic reaction is also more important than intragastric administration, indicating that in practical application, benzoyl Intramuscular injection of aconitine should be administered at a much lower dose than intragastric administration in order to avoid the occurrence of toxic reactions. Tests have shown that benzoyl aconitine is compatible with mesaconitine or hypoaconitine. The analgesic effect can obviously achieve high efficiency and low toxicity. It is a new drug route test. § Analgesic effect test of benzoyl aconitine skin administration

The test proved that the drug benzoyl aconitine skin external application of about SOOmg / kg dose can significantly reduce the writhing reaction caused by acetic acid in mice, the pain inhibition rate of 33. 6%; 400mg / kg dose group analgesia is not obvious, "the results showed that the benzoyl aconine for skin external analgesic effect, the doses showed high skin absorption efficiency benzoyl _Θ in aconine not high in this experiment, if the future Improved addition of excipients promotes skin absorption of aconitine in benzoyl, which will help reduce dosage and reduce cost.

 First, the test materials

 1-Test drug and preparation

 _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ In the Vaseline, mix well, and evaporate the ether in a 37 °C oven overnight to prepare a Vaseline cream containing about 16 mg/g of benzoyl aconitine.

 Aspirin enteric-coated tablets, white, 25 mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312» Take 4 tablets of aspirin enteric-coated tablets, grind, use distilled water to 10 ml, and mix with aspirin 10 mg/ml Liquid, spare.

 Take 2g of Vaseline, add 2. 5 ml of ether, mix well, and evaporate the ether in a 37-inch oven overnight to prepare a negative control vaseline paste.

 2. Test animals

 Kunming mice, male and female, 18~22 g, provided by Experimental Animal Center of Sichuan University, primary animal, certificate number: No. 10; vocabulary provider with animals, free drinking water, observation room temperature is 21 ± 2 ° C, relative humidity: 50~60%.

 3, reagents

 Ether, analytical grade, Chengdu Kelon Chemical Reagent Factory, batch number: 20050407.

 Sodium sulfide: Analytically pure, Chengdu Chemical Reagent Factory, batch number: 20040921.

 Glacial acetic acid: Analytically pure, Chengdu Chemical Reagent Factory, batch number: 20050423.

 Medicine Vaseline: Production at Wuhan Petrochemical Plant.

 Second, the test method

Forty Kunming mice were randomly divided into 4 groups, 10 in each group, half male and half female, weighing 18~22 g. The day before the administration, the back of the mouse was roughly cut with a pair of scissors, and the hair was removed with an 8% sodium sulfide solution, and the area was 1. 5 cmX 1. 5 cm. The first group was a negative control group, and the skin of the back was uniformly coated with a negative control of the Vaseline cream 0. Ig/ Only ^ then gently covered with a layer of gauze (l. 5emX 1. 5cm); the second group was given aspirin 200 mg/kg (equivalent to 20 times the clinical dose) for the positive control group; The ointment containing the 16 g / g benzoyl aconitine 0. lg / only; Group 4 coated with 16 mg / g benzoyl aconitine ointment 0. 05 ₈ / only „ two coated ointment group is also covered with a layer of gauze (1. 5cmX 1. 5cm) after application. In addition to the second group 30 minutes after the administration of intraperitoneal injection of 0. 63⁄4 glacial acetic acid solution, the rest Each group was given an intraperitoneal injection of 0. 63⁄4 glacial acetic acid solution 0. 2 ml / only, and the writhing response of each mouse within 10 minutes after 5 minutes of injection of vinegar vinegar solution was observed. The number of times of leg extension and hip height was statistically analyzed. The standard deviation of the mean writhing body and the pain inhibition rate of each dose group were calculated by t test, and the differences between the groups of the drug group and the negative control group were compared.

 Third, the test results

 Injection of acetic acid can cause longer-lasting pain in mice. Repetition of abdominal hind leg stretch, sacral height (twisted body) response Aspirin-positive control group, the number of writhing in mice caused by glacial acetic acid decreased significantly

(P<0. 001), the analgesic effect is very obvious, and the pain inhibition rate is 81.6%. _{Θ The} drug 荜formyl zhongtou yoshioji is used as an external application for skin, which can be significantly reduced at a dose of about 800 mg/kg. The inhibition rate of writhing reaction in mice induced by acetic acid was 33.6%; there was no significant difference in the number of writhing reactions between the 400m _g /kg group and the negative control group. The results showed that the analgesic effect was not obvious. 19. The results showed that benzoyl aconitine played an analgesic effect on the skin for external use, and the dosage was higher. It is indicated that the skin absorption efficiency of aconitine in benzoyl is not high in this experiment. If the addition of excipients can be improved in the future, promoting the skin absorption of aconitine in benzoyl will help to reduce the dose and reduce the dosage. cost.

 Effect of topical uracilyl aconitine on pain response in mice induced by acetic acid Group dose (mg/kg) Number of animals administered (only) Number of writhing (X ± SD) Pain inhibition rate (%) Negative control for external use 10 27. 7±12. 4 ― Aspirin 200 ig. 10 5. 1 ±6. 0*** 81. 6 benzoyl in about 800 external use 10 18. 4±6. 5* 33. 6 Oral base approximately 400 10 24· 5±7· 4 11. 6 Note: t test, compared with the negative control group, 0.05., "p<0. 01, "*p<0. 001

 Fourth, the conclusion of the test

 The drug benzoyl aconitine can significantly reduce the writhing reaction induced by acetic acid when the skin is applied at a dose of about 800 mg/kg. The pain inhibition rate is 33.6%; the analgesic effect is not obvious in the 400 mg/kg dose group. . The results showed that acetoyl aconitine played an analgesic effect on the skin for external use, and the dose used was higher. It is indicated that in this experiment, the skin absorption efficiency of aconitine in benzoyl is not high, and if the addition of excipients can be improved in the future, the skin absorption of aconitine in benzoyl can be promoted, which will help to reduce the dose. , cut costs. Tests have shown that benzoyl aconitine can be synergistically synergistic when used in combination with mesaconine or hypoaconitine, achieving high efficiency and low toxicity, and is a new route of administration.

 Test Example 9 Analgesic effect test of intramuscular injection of aconitine

 Tests have shown that the intramuscular injection of aconitine 100 and 25 rag / kg doses have a very significant analgesic effect, the inhibition rate of sputum is 100%. The analgesic effect of intramuscular injection of the same dose was significantly stronger than that of intragastric administration, but the toxicity was also heavier than that of intragastric administration. It indicates that in practical application, the dose of aconite should be administered intramuscularly. The medicine is much lower, which not only maintains the efficacy but also avoids the occurrence of toxic reactions.

 First, the test materials

 1. Test drugs and preparation

 Lower aconitine, white crystalline granules, supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. The sulphate containing 0. 05 mg / 5% of aconitine 0. 05 mg / The solution of ml; take 2.5 ml, add physiological saline to 10 ml, and prepare a solution of 0. 0125 mg / ml, spare

Aspirin enteric-coated tablets, white, 25 mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312. Take 4 tablets of aspirin enteric-coated tablets, grind with distilled water to lO niL to prepare a liquid containing aspirin and weigh 10 mg/ml.

Take 0.5 ml of absolute ethanol, add physiological saline to 10 ml and prepare 5% ethanol _.

 2, test animals

 Kunming mice. Male and female 18~22 g, provided by the Experimental Animal Center of Sichuan University, the first-class animal combination certificate number: 'No. 10; Feed provider with the animal free drinking water, the observation room temperature is 21 soil relative humidity : 50~60% o

 3 reagents

 Glacial acetic acid: Analytically pure, Chengdu Chemical Reagent Factory, batch number: 20050423.

 Anhydrous ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory Batch No.: 20050313

 Second test method

50 ml of Kunming mice were randomly divided into 5 groups, each group of 10 males and females, body weight 18 _~ 22 _gffl, group 1 was negative control group, ₅ leg muscles were injected intramuscularly with 52⁄4 ethanol 0. 2 ml/lOg body weight; 2 groups were positive control group, aspirin 200 mg/kg was administered by gavage (corresponding to 20 times of clinical dose); group 3 was given 0. 05 mg/ml of hypotocaline solution, and group 4 was given intramuscular injection of 0. 5毫升/10g, intraperitoneal injection 30 minutes after administration, the lower aconitine solution of the 05 mg / ml group, the intramuscular injection of 0. 0125mg / ml of the lower aconitine solution is 0. 2 ml / 10g, 30 minutes after administration 0.6% glacial acetic acid number of 0. 2 ml / mouse, injected acetate solution was observed and recorded for each mouse writhing within 10 minutes after 5 minutes the reaction (abdominal stretching the hind legs, buttocks raised) _a test data done Statistical analysis was performed to calculate the mean, standard deviation and pain inhibition rate of the writhing body in each dose group. The difference between the groups of the drug-administered group and the negative control group was compared by t test.

 Third, the test results

 Injection of acetic acid can cause longer-lasting painful stimulation in mice, and the mice repeatedly have abdomen hind legs and hips high (twisted body) response. In the aspirin-positive control group, the number of writhing in the mice induced by glacial acetic acid was significantly decreased (P < 0.001), and the analgesic effect was very obvious. The pain inhibition rate was 81.6%. Both intramuscular injection of aconitine and intragastric administration of lmg/kg significantly reduced the writhing response induced by acetic acid in mice, and the pain inhibition rates were 47.7% and 100%, respectively. However, the analgesic effect of intramuscular injection was significantly stronger than that of intragastric administration. Even if the analgesic effect of intramuscular injection of 0.25 mg/kg was significantly stronger than that of 100 mg/kg, the pain inhibition rate was also 1003⁄4. See Table 20. We also found that mice with intramuscular injection of lmg/kg lower aconitine group showed atrophy, less movement, and hiccup-like toxicity. It is indicated that, in practical application, when intramuscular injection of hypoxazone is administered, the dose should be It is much lower than the intragastric administration to avoid the occurrence of toxic reactions.

 Table 20 Effect of drug aconitine on pain response in mice induced by acetic acid. Dose (mg/kg) Number of animals in the route of administration (only) Number of writhing (X ± SD) Pain inhibition rate (%) Negative Control. im. 10 27. 7±12. 4

 Aspirin 200 ig- 10 5. 1±6. 0"* 81. 6 Lower aconitine 1 ig- 10 14, 5 ± 7. 6* 47. 7

 1 ― im. 10 0±0*** 100

0. 25 10 0 soil ( * 100 Note: t test, compared with the negative control group, *ρ<0· 05, **ρ<0· 01, ***ρ<0. 001

 Fourth, the conclusion of the test

 The doses of aconitine 100 and 25 mg/kg in the filth-injected drugs have very obvious analgesic effects, and the pain inhibition rate is 100%. The analgesic effect of intramuscular injection of the same dose was significantly stronger than that of intragastric administration, but the toxicity was also heavier than that of intragastric administration. It indicated that in practical application, the dose of aconite should be administered intramuscularly. The medicine is much lower, which not only maintains the efficacy but also avoids the occurrence of toxic reactions. Tests have shown that when aconitine is used in combination with mesaconitine or benzoyl aconitine, it can synergistically increase efficiency and achieve high efficiency and low toxicity. It is a new route of administration.

 Test Example 10 Analgesic effect test of subcutaneous aconitine skin administration

3⁄4 to verify that the drug can significantly reduce the small amount caused by acetic acid when the topical skin of aconitine is about 8mg/kg. The writhing response of the rats, the pain inhibition rate was 37.2%; although the pain inhibition rate was 31.8 in the 4 mg/kg dose group, there was no significant difference in the number of writhing responses in the mice compared with the negative control group, indicating analgesic effect. The results are not obvious. The results show that the lower aconitine plays an analgesic effect on the skin for external use, and the dosage is higher. It is indicated that the skin absorption efficiency of aconitine is not high in this experiment. If the addition of excipients can promote the skin absorption of aconitine, it will help to reduce the dosage and reduce the cost.

 -. experiment material

 '1 test drug and preparation

Under aconitine, white crystalline granule _s Pharmaceutical Co. provided by Chengdu Zhizhi. The sulphate containing 1. 6 mg / g of the lower aconitine was added to the mixture. Vaseline cream is available.

 Aspirin enteric-coated tablets white, 25 mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd. Batch number: 20050312 Take 4 pieces of aspirin enteric-coated tablets, use distilled water to 10 ml, and prepare a medicine containing aspirin # 10 mg/nil Liquid spare.

 Take 3⁄4 medicine Vaseline Add 2.5 ml of ether, mix well 37 Ό for the overnight evaporation of ether to make a negative control Vaseline cream, spare.

 2. Test animals

Kunming mice _s male and female, 18~22 g, provided the first-class animal certificate number by the Experimental Animal Center of Sichuan University: No. 10; Feed provider with animals, free drinking water, observation room temperature is 21 ± 2 °C , the relative humidity is: 50~60%.

 3, reagents

 Ether: Analytically pure, Chengdu Kelon Chemical Reagent Factory, batch number: 20050407.

 Sodium sulfide: Analytical purity Chengdu Chemical Reagent Factory, batch number: 20040921.

 Glacial acetic acid: Analytical purity Chengdu Chemical Reagent Factory, batch number: 20050423.

 Medicine Vaseline: Production at Wuhan Petrochemical Plant.

 Second, the test method

40 Kunming mice were randomly divided into 4 groups, 10 in each group, half male and half female, weighing 18~22 g. One day before the administration, the back hair of the mice was roughly cut with scissors, and the hair was removed with 8% sodium sulfide solution. The area was 1. 5 cmX 1. 5cm _{0 The} first group was a negative control group, and the skin of the back was depilated with a negative control of Vaseline cream 0. lg / only, then a layer of gauze ( 1. 5cmX 1 5cm) lightly covered; the second group was a positive control group, and the aspirin was given 200 mg/kg by gavage (equivalent to 20 times the clinical dose); the third group was coated with a skin at the back of the hair removal treatment. The medicinal ointment containing 1. 6mg / g of aconitine 0. 05g / only. The two-coated ointment group was also lightly covered with a layer of gauze (1.5 cm X 1. 5 cm) after application. The 5% glacial acetic acid solution 0. 2 ml/only, observed, after the administration of 0.5% glacial acetic acid solution. The number of writhing responses (abdominal hind leg extension, buttocks height) of each mouse within 10 minutes after injection of the acetic acid solution for 5 minutes was recorded. Statistical analysis was performed on the test data, and the mean, standard deviation and pain inhibition rate of the writhing body in each dose group were calculated. The difference between the groups of the drug-administered group and the negative control group was compared by t test.

 Third, the test results

Injection of acetic acid can cause longer-lasting painful stimulation in mice, and the mice repeatedly have abdominal and hind leg extensions, and the buttocks rise (twisted body) response. In the aspirin-positive control group, the number of writhing in the mice induced by glacial acetic acid was significantly decreased (P < 0.001), and the analgesic effect was very significant, and the pain inhibition rate was 81.6%. The drug aconitine as an attempt to externally use the skin at a dose of about 8 mg/kg significantly reduced the writhing response in mice caused by acetic acid, and the pain inhibition rate was 37.2%; the 1⁄2 g/kg dose group despite the pain inhibition rate The index was 31.8%, but the number of writhing reactions in mice was not significantly different from that in the negative control group, indicating that the analgesic effect was not obvious. The results are shown in Table 21. The results showed that the lower aconitine exerted an analgesic effect on the external use of the skin, and the dosage used was higher. It is indicated that in this experiment, the skin absorption efficiency of the lower aconitine is not high, and if the addition of excipients can be improved in the future, the skin absorption of the aconitine can be promoted, which will help to reduce the dosage and reduce the cost. Table 21 Effect of topical aconitine on pain response in mice induced by acetic acid

 Group dose (mg/kg) Route of administration Number of animals (only) Torsion 3⁄4 (X soil SD) Pain inhibition rate (%) Negative control ― External use 10 27. 7±12. 4

 Aspirin 200 ί »3T· 10 5. 1±β. 0*** 81. 6 Lower aconitine 8 External use 10 17. 4±9. 0* 37. 2

 4 10 18. 9±9. 3 31. 8 Note: t test, compared with the negative control group p<0.05, **p<0. 01, <0. 001

 Four test conclusions

 The drug used in the treatment of aconitine skin at a dose of about 8mg / kg can significantly reduce the mouse torsion reaction caused by ester acid. 5. Pain inhibition rate is 37.2%; 4mg / kg dose group despite the pain inhibition rate of 31 8%, but the number of writhing reactions in mice was not significantly different from that in the negative control group, indicating that the analgesic effect was not obvious. The results showed that the higher dose of aconitine for external analgesia was indicated in this experiment. The skin absorption efficiency of middle and lower aconitine is not high. If the addition of excipients can be improved in the future, promoting the skin absorption of lower aconitine will help to reduce the dose reduction. Tests have shown that the combination of aconitine and mesaconine or benzoyl aconitine can synergistically increase the efficacy of 0 to achieve high efficiency and low toxicity is a new route of administration.

 Test Example 11 Analgesic effect test of intramuscular injection of aconitine

 5 mg/kg. The effect of the analgesic effect is significantly stronger than that of the 0. 5mg / 0. 5mg / kg of aconitine. The intragastric administration of kg, the pain inhibition rate was 99.6% and 77.6%, respectively, and the pain inhibition rate of 0.5 mg/kg of intragastric administration was 56. 7% „ intramuscular injection 0. 125 mg/kg The mice in the aconitine group also showed atrophy and less sputum-like toxicity. The toxicity was only slightly milder. In practical application, when the intramuscular injection of aconitine was administered, the dose should be much lower than that of the intragastric administration. Avoid the occurrence of toxic reactions in rabbits.

1. Test materials

 θ 1, test drug and preparation

Aconitine, near white powder, supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. 5 025 mg, the aconitine is added to the mixture. After the solution is heated in a water bath at 42 ° C and sonicated, the physiological saline is added to 50 ml. /ml的溶液; Take 2. 5 ml, add physiological saline to 10ml _{s to} form a solution of 0. 00625mg / ml, and then take 0. 0625mg / ml of the solution 2. 5 ml, add physiological saline to 10ml, with 0. 00156 mg/ml 15 solution, ready for use.

Aspirin enteric-coated tablets, white, 25 _mg / tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312. Take 4 tablets of aspirin enteric-coated tablets, grind them, use distilled water to 10 ml, and prepare a liquid containing aspirin 10 mg/ml.

 Take 0.5 ml of absolute ethanol and add normal saline to 10 ml to make 5% ethanol.

 ')0 2, test animals

 Kunming mice, male and female, 18~22 g, provided by Experimental Animal Center of Sichuan University, primary animal, certificate number: No. 10; vocabulary provider with animals, free drinking water, observation room temperature is 21 ± 2 ° C, relative humidity: 50 ~ 603⁄4.

, 3, reagent

 ■35 Glacial acetic acid: Analytically pure, Chengdu Chemical Reagent Factory, batch number: 20050423.

 Anhydrous ethanol, analytical grade, Chengdu Kelon Chemical Reagent Factory, batch number: 20050313.

 Second, the test method

Sixty Kunming mice were randomly divided into 6 groups, 10 in each group, half male and half female, weighing 18~22 g. The first group was a negative control group, and the intramuscular injection of 5% ethanol was 0. 2 ml/lOg body weight; the second group was positive for the 40 group, and the aspirin was given 200 mg/kg by gavage (equivalent to 20 clinical doses). 00625rag/ml。 The first group of intramuscular injection of 0. 025mg / ml of aconitine solution, the fourth group of intramuscular injection of 0. 025mg / ml of aconitine solution, the fifth group of intramuscular injection of 0. 00625rag / ml _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ Ice Acetic acid solution 0.2 ml/only observe the number of times the writhing reaction (abdominal hind leg extension hip height) of each mouse within 10 minutes after 5 minutes of injection of the acetic acid solution. Statistical analysis of the experimental data, was calculated for each dose group mean writhing inhibition rate of pain and the standard deviation, t-test comparison between each picking-administered group and negative control group of group differences _Θ

 Three test results

 Injection of acetic acid can cause longer-lasting painful stimulation in mice, and the mice repeatedly have abdomen hind legs stretching and hips high (twisting) response. The aspirin-positive control group had a significant decrease in the number of writhing in mice induced by glacial acetic acid (Ρ<0. 001), and the analgesic effect was very obvious. The pain inhibition rate was 81.63⁄4. In this experiment, due to the toxicity of intramuscular injection of 0.5 mg/kg of aconitine, 10 mice showed obvious atrophy and less dyspnea after intramuscular injection. In 30 minutes, 8 died, although The remaining two did not show a torsion reaction in the experiment. The concentration was further diluted and the dose was lowered. The results showed that the analgesic effect of Wutou 橼0. 125 and 0.031 mg/kg in the intramuscular injection was significantly stronger than that of the 0.5 mg/kg intragastric administration. The rate of pain inhibition was 56.7%. The results are shown in Table 22. We also found that mice injected intramuscularly with 0. 125mg/kg of aconitine also showed atrophy and less mitigation. It is indicated that in practical application, when intramuscular injection of aconitine is administered, the dose should be much lower than that of intragastric administration to avoid the occurrence of toxic reactions.

 Effect of different routes of administration of aconitine on pain response in mice induced by acetic acid Group dose (nig/kg) Number of animals in the route of administration (only) Number of writhing soil SD) Pain inhibition rate (%) Negative control - An im. 10 27. 7±12. 4 —1

 Aspirin 200 ig. 10 5. 1±6. 0*** 81. 6 Aconitine 0. 5 ig. 10 12. 0 ±8. 2*" 56. 7

0. 5 im. 10 0±(T* ^SS 100

0. 125 10 0. 1 + 0. 3***

 0. 031 10 6. 2±6. 4 * 77. 6 Note: t test, compared with the negative control group, <0. 05, * <0. 01, ***p<0. 001

 0 mice were intramuscularly injected with 0.5 mg/kg of aconitine 30 min later, and 8 died. This data is the observation result of the remaining two.

 Fourth, the conclusion of the test

 Intramuscular injection of 0.5 mg/kg of aconitine is too toxic. The analgesic effect of the analgesic effect of the aconitine was significantly higher than that of the 0. 5 mg/kg intragastric administration, and the pain inhibition rate was 99. 6 7%。 The pain inhibition rate of 56. 7%. Muscle injection 0. 125 mg / kg in the aconitine group also showed atrophy, less movement, hiccup-like toxicity, only a lesser degree. It is indicated that in practical application, when intramuscular injection of aconitine is administered, the dose should be much lower than that of intragastric administration to avoid the occurrence of toxicity.

 Test Example 12 Analgesic effect test of aconitine skin administration

 First, 3⁄4 test materials

 1. Test drugs and preparation

 Aconitine, near white powder, supplied by Chengdu Zhizhi Pharmaceutical Co., Ltd. 5 mg, containing aconitine, about 0. 8 mg, containing a solution of aconitine, and then adding a mixture of the mixture. /g of Vaseline cream, spare.

 Aspirin enteric-coated tablets, white, 25 mg/tablet, Xuzhou Enhua Pharmaceutical Group Co., Ltd., batch number: 20050312. Take 4 tablets of aspirin enteric-coated tablets, grind them, and use distilled water to 10 ml to prepare a liquid containing aspirin 10 mg/ml.

 Take 2g of Vaseline, add 2. 5 ml of ether, mix well, and evaporate the ether in a 37 °C oven overnight to prepare a negative control Vaseline paste.

 2. Test animals

Kunming mice, male and female, 18~22 g, provided by Experimental Animal Center of Sichuan University, first-class animal, pile number: No. 10; feeding provider with animals, free drinking water, observation room temperature is 21 ±2°C, relatively wet The degree is: 50~603⁄4.

 3. Reagent

I : Analytical pure, Chengdu Kelon Chemical Reagent Factory Lot No.: 20050407 _β

 Sodium Sulfide: Analytical Pure Chengdu Chemical Reagent Factory, Lot Number: 20040921

Ice vinegar: Analytically pure, Chengdu Chemical Reagent Factory Lot Number: 20050423 _β

 Medicine Vaseline: Production at Wuhan Petrochemical Plant.

 Second, the test method

40 Kunming mice were randomly divided into 4 groups, 10 rats in each group, half male and half female, weighing 18~22 g _ffl . The day before the administration, scissors were used to roughly cut off the back hair of the mice, and the area was removed with 8% sodium sulfide solution. 1. 5 cmX 1. 5cm. The first group was a negative control group, and the skin was evenly coated with a negative control in the back of the hair removal treatment. 0. Ig / only then with a layer of gauze (1. 5cmX 1. 5cm 8m _g /g。 The second group of the positive control group was given aspirin 200 mg / kg (equivalent to 20 times the clinical dose); Oral aconitine ointment O. lg / only; Group 4 coated with 0. 8m _g / g of aconitine ointment two coated ointment group also used a layer of gauze after application (1.5 cm X 1. 5 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 。 Ml/only, observe the record of the writhing reaction (abdominal hind leg extension, hip height) of each mouse within 10 minutes after injection of acetic acid solution for 5 minutes. Writhing dose group mean, standard deviation and pain suppression rate, t-test, comparison between drug-treated groups and the negative control group of a difference

 Third, the test results

Injection of acetic acid can cause longer-lasting painful stimulation in mice, and the mice repeatedly have a high back (twisted body) reaction in the hind legs. In the aspirin-positive control group, the number of writhing in mice induced by glacial acetic acid was significantly decreased (P < 0.001), and the analgesic effect was very obvious. The pain inhibition rate was 81.6% _D. Aconitine was used as skin for external use. At the dose of about 4 mg/kg, the writhing response of mice induced by acetic acid was significantly reduced, and the pain inhibition rate was 34.3%; the number of writhing reactions in the 2 mg/kg dose group was compared with the negative control group. There was no significant difference, indicating that the analgesic effect was not obvious. The results are shown in Table 23. The results showed that aconitine exerted an analgesic effect on external skin for a high dose. It is indicated that in this experiment, the skin absorption efficiency of aconitine is not high, and if the addition of excipients can be improved in the future, the skin absorption of aconitine can be promoted, which will help to reduce the dosage and reduce the cost.

 Table 23 Effect of topical aconitine on pain response in mice induced by acetic acid Group dose (mg/kg) Number of animals administered (only) Number of writhing (X soil SD) Pain inhibition rate (%) Negative control ― ―— 10 27. 7± 12. 4 one by one

 Aspirin 200 ig. 10 5. 1 ±6. 0*** 81. 6 aconitine about 4 external use 10 18. 2±7. 34. 3

 Approx. 2 10 20· 2± 14· 1 27. 1 Note: t test, compared with the negative control group, p<0.05, **p<0. 01, p<0. 001

 Fourth, the conclusion of the test

Drug skin external aconitine about 4m _g / mouse can significantly reduce the acetic acid induced writhing when k _g dose, pain inhibition rate was 34.3%; the number of reaction 2mg / kg dose group and writhing Compared with the negative control group, there was no significant difference, indicating that the analgesic effect was not obvious. The results showed that aconitine played an analgesic effect on the external use of the skin, and the dosage used was relatively high. It is indicated that in this experiment, the skin absorption efficiency of mesaconitine is not high, and if the addition of excipients can be improved in the future, the skin absorption of mesaconitine can be promoted, which will help to reduce the dosage and reduce the cost. Tests have shown that aconitine can be synergistically synergistic when combined with aconitine or benzoyl aconitine, which is a new route of administration.

 The above test results show that the amount of a single compound is dose-effect-dependent, and the pharmaceutical composition after compatibility does not have a simple dose-effect linear relationship, but is used in a specific ratio at a specific ratio. In the case of the best efficacy, high-efficiency, low-toxic analgesic pharmaceutical compositions provide a clinical choice.

Claims

 An analgesic pharmaceutical composition characterized in that: it is prepared from aconitine compound benzamidine in which aconitine or aconitine or two or three of aconitine are prepared as active ingredients. The weight ratio of the obtained medicament is:

 0-600 parts of aconitine in benzoyl, 0-6 parts of aconitine 0-3 · 汾, at least two of which have a distribution ratio of not zero.

 The pharmaceutical composition having an analgesic action according to claim 1, which is characterized in that it is an agent prepared by using a weight-matching aconitine compound as an active ingredient:

 : benzoyl acyl aconitine 100-300 parts, hypoaconitine 1-3 parts

 The pharmaceutical composition having an analgesic effect according to Claim 2, which is characterized in that it is an agent prepared by using an aconitine compound having the following weight ratio as an active ingredient:

 J _

 Benzoyl aconitine 100 parts of aconitine 1 part.

 The pharmaceutical composition having an analgesic effect according to Claim 1, which is prepared by the following a weight ratio of an aconitine compound as an active ingredient:

 2-6 parts of lower aconitine and 1-3 parts of mesaconitine.

 The pharmaceutical composition having an analgesic effect according to Claim 4, which is an agent prepared by the following a weight-matching aconitine compound as an active ingredient:

 2 parts of lower aconitine and 3 parts of aconitine.

 The pharmaceutical composition having an analgesic effect according to Claim 1, which is characterized in that it is an agent prepared by using the a weight-matching aconitine compound as an active ingredient:

 200-600 parts of aconitine in benzoyl and 1-3 parts of aconitine.

 The pharmaceutical composition having an analgesic effect according to claim 6, which is an agent prepared by the following a weight ratio of an aconitine compound as an active ingredient:

 200 parts of aconitine in benzoyl and 3 parts of aconitine.

 The pharmaceutical composition having an analgesic effect according to Claim 1, which is prepared by the following a weight ratio of an aconitine compound as an active ingredient:

 Benzoyl aconitine 100-600 parts, hypoaconitine 1-6 parts, and aconitine 1-3 parts.

 The pharmaceutical composition having an analgesic effect according to claim 8, which is an agent prepared by the following a weight-matching aconitine compound as an active ingredient:

 100 parts of aconitine in benzoyl, 1 part of hypoaconitine, and 1 part of aconitine.

 The pharmaceutical composition having an analgesic effect according to any one of claims 1 to 9, wherein the pharmaceutical agent is an oral preparation, an external preparation, or an injection.

 An analgesic pharmaceutical composition according to claim 10, wherein the oral preparation is a tablet, a capsule, a pill, or a granule.

 The pharmaceutical composition having an analgesic effect according to claim 10, wherein the external preparation is a lotion, an ointment, a suppository, or a medicinal dressing.

 The pharmaceutical composition having an analgesic effect according to claim 10, wherein the injection is an injection solution or a powder for injection.