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CN110384710B - Medicine and combination product for preventing and/or treating pain and application thereof - Google Patents

Medicine and combination product for preventing and/or treating pain and application thereof Download PDF

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CN110384710B
CN110384710B CN201910308683.XA CN201910308683A CN110384710B CN 110384710 B CN110384710 B CN 110384710B CN 201910308683 A CN201910308683 A CN 201910308683A CN 110384710 B CN110384710 B CN 110384710B
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pain
isovaleryl spiramycin
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isovalerylspiramycin
medicine
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CN110384710A (en
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姜勋雷
姜勋东
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Shenyang Fuyang Pharmaceutical Technology Co Ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
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    • A61K31/7048—Compounds having saccharide radicals and heterocyclic rings having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin, digitoxin or digoxin
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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Abstract

The invention discloses a medicament for preventing and/or treating pain, a combined product and application thereof, wherein the effective component of the medicament comprises one of colimycin, isovaleryl spiramycin III, isovaleryl spiramycin II and isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III. The medicine and the combined product have good treatment effect on pain, especially have obvious analgesic effect on cancer pain, have quick effect, short treatment period, small side effect and weak dependence, are not limited by the tolerance of patients, and have important economic benefit and social benefit.

Description

Medicine and combination product for preventing and/or treating pain and application thereof
Technical Field
The invention belongs to the field of medicine application, and particularly relates to a medicine for preventing and/or treating pain, a combination product and application thereof.
Background
The international association for pain defines pain as an unpleasant sensory and emotional feeling with or without tissue damage. In 1995, the american academy of pain suggested that pain was classified as the fifth leading vital sign, and was an important indicator of vital signs along with blood pressure, body temperature, respiration, and pulse.
Cancer pain is one of the most common and intolerable symptoms of cancer patients and one of the most difficult to control, about 30-50% of cancer patients are accompanied by different degrees of pain, and 60-90% of late-stage cancer patients suffer from moderate and severe pain, which seriously affects the quality of life of cancer patients.
In order to improve the quality of life of patients with chronic pain, various researchers have endeavored to find new ways of treating pain in an attempt to save patients from pain. Although many drugs are used for treating the pain at present, most drugs have slow efficacy, long treatment period, large side effect and strong dependence, and are limited by the tolerance of patients, so that the pain relieving effect of some patients is poor.
The colimycin (carriamycin), also known as bistspiramycin (Bitespiramycin) and shengmycin (Shengjimycin) is a novel antibiotic which is formed by cloning 4 ' -isovaleryl transferase (4 ' -o-acyl-transferase) of a carbomycin producing strain into a spiramycin producing strain through a transgenic technology by cooperating with the applicant of the institute of biotechnology of Chinese academy of medicine, directionally acylating 4 ' -OH of the spiramycin and adding an isovaleryl side chain to the 4 ' -position to form the 4 ' -position isovaleryl spiramycin serving as a main component.
Figure BDA0002030713200000021
The kelimycin is composed of a plurality of spiramycin derivatives, the total content of the main active ingredients isovaleryl spiramycin (I + II + III) is not less than 60%, the total content of acylated spiramycin is not less than 80%, and the kelimycin is an acceptable pharmaceutical composition in pharmacy. The central structure is a 16-membered lactone ring which is connected with a molecule of forosamine, a molecule of mycaminose and a molecule of mycaminose, and the main components of the isovaleryl spiramycin I, II and III are structurally different from spiramycin in that a group connected to the 4' position of the mycaminose is isovaleryl instead of hydroxyl. The chemical structure is shown as a formula (1), and the chemical structure comprises more than ten components. The composition standard of the prior finished product of the colimycin is that the content of isovaleryl spiramycin III is more than or equal to 30 percent, the sum of the proportions of isovaleryl spiramycin I, II and III is more than or equal to 60 percent, the content of total acylated spiramycin is more than or equal to 80 percent, and the sum of other unknown components is less than or equal to 5 percent.
Colimycin belongs to the class of 16-membered macrolide antibiotics, which has the active groups carboxyl, alkoxy, epoxy, keto and aldehyde groups and a pair of conjugated C = C, and has a molecular weight of about 884 to 982. Due to their similar chemical structures, colimycin has many commonalities with macrolide antibiotics: the compound is easily soluble in most organic solvents such as esters, acetone, chloroform, alcohols and the like, slightly soluble in petroleum ether and hardly soluble in water; the molecular structure contains two dimethylamino groups, is alkalescent and is easy to dissolve in an acidic aqueous solution; has the property of negative solubility that the solubility decreases with increasing temperature. The main component isovaleryl spiramycin of the colimycin has a longer carbon chain at the 4 'position and poor hydrophilicity, and has lower solubility in water than spiramycin and 4' -acetyl spiramycin.
The colimycin is a white non-crystalline powder, has slight hygroscopicity, the specific rotation degree is about minus 80.8 degrees, the maximum ultraviolet absorption wavelength is 231-232 nm, the colimycin has weak fluorescence chromophoric groups, and the colimycin has purple reaction when meeting concentrated sulfuric acid or hydrochloric acid to generate strong purple fluorescence, and the maximum light absorption value is 231-232 nm.
The medicine has good lipophilicity, strong tissue permeability, quick oral absorption, long in-vivo maintenance time and continuous antibiotic after-effect. According to the relation between the drug effect and the chemical conformation, after the 4 'position of the macrolide antibiotic is acylated, the lipophilicity and the in vivo activity are improved, the in vivo antibacterial activity and the clinical treatment effect are both obviously improved, and the stability of the antibiotic in vivo is enhanced along with the increase of the carbon chain of the 4' hydroxy ester, namely isovaleryl spiramycin > butyryl spiramycin > propionyl spiramycin > acetyl spiramycin.
Preliminary in vivo and in vitro pharmacodynamic tests show that the medicine is not only used for most G + The bacteria have good antibacterial activity, and can be used for treating part G - The bacteria also have certain effect, and each technical index is obviousThe antibacterial agent is superior to azithromycin, erythromycin, acetylspiramycin and midecamycin, has the strongest antibacterial activity on mycoplasma pneumoniae, has certain antibacterial activity on erythromycin resistant bacteria, gonococcus, pneumococcus, staphylococcus aureus, pseudomonas aeruginosa, bacillus influenzae, haemophilus influenzae, bacteroides fragilis, legionella, multirow bacillus and clostridium perfringens, and has only little cross-resistance on clinical erythromycin resistant staphylococcus aureus. The colimycin is mainly used for treating gram-positive bacteria infectious diseases, particularly upper respiratory tract infection, and possibly urinary system infection and the like.
The results of pharmacokinetic studies show that active effective components in the kelimycin are mainly isovaleryl spiramycin I, II and III. The kelimycin is metabolized into spiramycin soon after entering the body, and AUC of parent isovaleryl spiramycin I, II and III and active metabolite spiramycin I, II and III 0-t The oral absolute bioavailability was found to average 91.6% by summation. The structure of isovaleryl spiramycin is proved to obviously improve the bioavailability of the active ingredient spiramycin. A single dose of colimycin has a slower elimination, T 1/2β Between 23 and 27 hours.
The present invention has been made in view of this point.
Disclosure of Invention
The technical problem to be solved by the present invention is to overcome the disadvantages of the prior art and to provide a medicament, a combination product and the use thereof for the prevention and/or treatment of pain.
In order to solve the technical problems, the invention adopts the technical scheme that:
the invention provides a medicament for preventing and/or treating pain, wherein the effective component of the medicament comprises one of colimycin, isovaleryl spiramycin III, isovaleryl spiramycin II and isovaleryl spiramycin I;
or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
Further, the medicine is prepared into a pharmaceutically acceptable dosage form.
Further, the dosage of the medicine is 10-80 mg/kg.
The invention also provides a medicament for preventing and/or treating pain, wherein the effective component of the medicament comprises a derivative, a pharmaceutically acceptable salt, a solvate, a metabolite, a stereoisomer, a tautomer, a polymorph and a prodrug of the colimycin;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin III;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin II;
at least one of derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin I.
The invention also provides a combination product for preventing and/or treating pain, which comprises a first medicament, wherein the effective component of the first medicament comprises one of colimycin, isovaleryl spiramycin III, isovaleryl spiramycin II and isovaleryl spiramycin I;
or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
Further, the combination product further comprises a second medicament.
Further, the second agent includes at least one of the related drugs for preventing and/or treating pain.
Further, the related medicines for preventing and/or treating pain comprise narcotic analgesics and antipyretic analgesics.
The present invention also provides a combination for use in the prevention and/or treatment of pain, said combination comprising a first agent, the active ingredient of which comprises a derivative, pharmaceutically acceptable salt, solvate, metabolite, stereoisomer, tautomer, polymorph, prodrug of colimycin;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin III;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin II;
at least one of derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin I.
The invention also provides a medicament as defined in any one of the preceding claims or a combination product as defined in any one of the preceding claims for use in the prevention and/or treatment of pain.
The pain includes cancer pain. The pain includes nociceptive pain and neuropathic pain.
The invention also provides a medicament as described in any one of the above or a combination product as described in any one of the above for inhibiting prostaglandin synthesis, inhibiting substance P release, inhibiting 5-hydroxytryptamine and norepinephrine reuptake.
The colimycin can be prepared by separation according to the method of the prior art, such as the method of the embodiment 1 of the Chinese patent application with the application number of CN 101785778A.
The invention also provides a medicament for preventing and/or treating fever, wherein the effective component of the medicament comprises one of colimycin, isovaleryl spiramycin I, isovaleryl spiramycin II and isovaleryl spiramycin III;
or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
Further, the medicament comprises a pharmaceutically acceptable carrier.
Further, the medicine is prepared into clinically acceptable tablets, capsules, pills, injections, sustained release agents and various microparticle administration systems.
The invention also provides a medicament for preventing and/or treating fever, wherein the effective component of the medicament comprises a derivative, a pharmaceutically acceptable salt, a solvate, a metabolite, a stereoisomer, a tautomer, a polymorph and a prodrug of the colimycin;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin III;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin II;
and at least one of derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin I.
The invention also provides a combination product for treating fever, which comprises a first medicament, wherein the effective component of the first medicament comprises one of colimycin, isovaleryl spiramycin I, isovaleryl spiramycin II and isovaleryl spiramycin III;
or the combination of two or three of isovaleryl spiramycin I, isovaleryl spiramycin II and isovaleryl spiramycin III.
Further, the combination product further comprises a second medicament, and the second medicament comprises at least one of related medicaments for preventing and/or treating common fever.
Further, the related medicines for preventing and/or treating common fever comprise antibiotic medicines, antipyretic analgesic medicines and antiviral medicines.
The invention also provides a combination for use in the treatment of fever, the combination comprising a first agent whose active ingredient comprises a derivative, pharmaceutically acceptable salt, solvate, metabolite, stereoisomer, tautomer, polymorph, prodrug of colimycin;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin III;
derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin II;
and at least one of derivatives, pharmaceutically acceptable salts, solvates, metabolites, stereoisomers, tautomers, polymorphs, prodrugs of isovaleryl spiramycin I.
The invention also provides application of any one of the medicines and the combination product in preventing and/or treating fever.
The invention also provides the use of any one of the medicaments or combination products described above to reduce or prevent prostaglandin synthesis by inhibiting cyclooxygenase enzyme activity.
In the scheme, as the mechanism of fever is that pathogens stimulate macrophages to release cytokines, epoxymycin is triggered to catalyze arachidonic acid to generate prostaglandin, and fever, pain and inflammation are caused.
After adopting the technical scheme, compared with the prior art, the invention has the following beneficial effects:
the medicine and the combined product have good treatment effect on pain, especially have obvious analgesic effect on cancer pain, have quick effect, short treatment period, small side effect and weak dependence, are not limited by the tolerance of patients, and have important economic benefit and social benefit.
The following describes embodiments of the present invention in further detail with reference to the accompanying drawings.
Drawings
The accompanying drawings, which are included to provide a further understanding of the invention, are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the invention without limiting the invention to the right. It is obvious that the drawings in the following description are only some embodiments, and that for a person skilled in the art, other drawings can be derived from them without inventive effort. In the drawings:
FIG. 1 is a graph showing the mechanical pain threshold of the vaccinated lateral hind limbs of rats of the invention.
FIG. 2 is the mechanical pain threshold of the hind limb of the rat of the present invention as opposed to the vaccinated side.
FIG. 3 is a graph showing the thermal pain threshold of vaccinated lateral hind limbs of rats of the invention.
FIG. 4 is the thermal pain threshold of the hind limb of the rat of the invention opposite the vaccinated side.
FIG. 5 is a line graph showing the body temperature changes of the drug and the combination product of the present invention on LPS fever model rats.
FIG. 6 is a line graph showing the body temperature changes of the drug and the combination product of the present invention on a dry yeast fever model rat.
It should be noted that the drawings and the description are not intended to limit the scope of the inventive concept in any way, but to illustrate it by a person skilled in the art with reference to specific embodiments.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and the following embodiments are used for illustrating the present invention and are not intended to limit the scope of the present invention.
Example 1 coated tablet
Specification: 200mg/350mg
Tablet core prescription:
Figure BDA0002030713200000071
the prescription of the coating liquid is as follows:
Figure BDA0002030713200000072
the medicine A is one of colimycin, isovaleryl spiramycin III, isovaleryl spiramycin II and isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
The preparation process comprises the following steps:
preparation of the tablet core: the main drug and the auxiliary materials are respectively sieved by a 100-mesh sieve, the raw powder of the drug A, the microcrystalline cellulose and the carboxymethyl starch sodium with the prescription amount of 1/2 are evenly mixed, and then 5 percent of polyvidone K is added 30 Preparing soft material from water solution, granulating with 18 mesh sieve, and drying wet granules at 60 deg.C for 2 hr under ventilation condition; drying, granulating with 18 mesh sieve, adding 1/2 of carboxymethyl starch sodium and magnesium stearate, mixing, and tabletting with 11mm diameter shallow concave punch to obtain tablet core containing medicine with weight of 350mg and hardness of 6.5 kg.
Preparing a coating solution: weighing the required opadry II (white), adding the required amount of water into a liquid preparation container, adding the water in several times, reducing the stirring speed after all the water is added, eliminating the vortex, and continuing stirring for 30min to obtain the product.
Preparation of film-coated tablets: and (3) putting the tablet cores into a coating pot, determining coating conditions, coating at the speed of a main engine of 20r/min, the air inlet temperature of 40 ℃, the air outlet temperature of 30 ℃, the spraying pressure of 0.02Mpa and the spraying flow of 1ml/min, and continuously spraying for 1.5h after the constant speed until the surfaces of the tablet cores are smooth and uniform in color and luster, and the tablets are qualified according with the inspection standard of film coating. The weight of the coating is increased by about 5 percent.
Example 2 plain tablets (calculated as 10000 tablets)
Prescription:
medicine A1000 g
Low-substituted hydroxypropyl cellulose (5%) 92.5g
55.5g sodium carboxymethyl starch (3%)
Magnesium stearate (1%) 18.5g
Total weight of starch-weight of other raw and auxiliary materials
Total weight of 1850g
The drug A is one of the colimycin, the isovaleryl spiramycin III, the isovaleryl spiramycin II and the isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
The preparation process comprises the following steps: weighing a proper amount of starch, diluting to a concentration of 15%, and heating to paste to prepare an adhesive; respectively sieving raw powder of a main material medicament A, auxiliary material starch, low-substituted hydroxypropyl cellulose, sodium carboxymethyl starch and magnesium stearate with a 100-mesh sieve, and weighing the required main material and auxiliary material according to the prescription amount; mixing the raw powder of the medicine A, starch and low-substituted hydroxypropyl cellulose uniformly, preparing a soft material by starch paste with the concentration of 15% of starch, granulating by a 14-mesh sieve, drying at 50-60 ℃, controlling the water content to be 3-5%, granulating by the 14-mesh sieve, adding sodium carboxymethyl starch and magnesium stearate, mixing, and determining the content of granules; calculating the tablet weight according to the particle content, tabletting (a phi 9mm shallow concave punch), and detecting the tablet weight difference; and packaging after the inspection is qualified.
Example 3 Capsule (calculated as 10000 capsules)
Prescription:
medicine A1000 g
Starch 1080-drug A weight
Medicinal No. 3 capsule 1000 granules
Liquid paraffin 50ml
The drug A is one of the colimycin, the isovaleryl spiramycin III, the isovaleryl spiramycin II and the isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
The preparation process comprises the following steps: respectively weighing the raw powder of the main material medicament A and the medicinal starch of the auxiliary material according to the formula amount, and filling the raw powder and the medicinal starch into a mixer for fully mixing for 1.5 to 2 hours; the data obtained by sampling and detecting the content is basically consistent with theoretical data (the weight of each capsule is about 0.105 g), the qualified pharmaceutical No. 3 capsules and the mixed raw materials to be filled are respectively filled into a filling machine for filling according to the operation requirements of a full-automatic capsule machine, the filled capsules are subjected to difference detection (within +/-10 percent and less than 0.3 g), the dissolution rate meets the requirements, the capsules meeting the requirements after detection are put into a polishing machine, liquid paraffin is added for polishing for 15-20 minutes, and then the capsules are taken out for finished product packaging box detection.
Example 4 Dry syrup (calculated as 10000 bags)
Prescription:
drug A1250 g
Citric acid (0.5%) 15g
Total weight of sucrose-other raw and auxiliary materials
A total weight of about 500g
About 1g of pigment (curcumin)
The drug A is one of the colimycin, the isovaleryl spiramycin III, the isovaleryl spiramycin II and the isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
The preparation process comprises the following steps: the raw powder of the medicine A, citric acid and cane sugar are respectively crushed into particles with 85 percent passing 300 meshes and 15 percent passing 180 meshes by a high-speed airflow crusher, then the crushed fine powder is weighed according to the prescription amount and fully mixed for 1-1.5 hours, the content of the fine powder is measured, the filling amount (the theoretical filling amount is 500mg per bag) is calculated, then the mixture is filled into a bagging machine, aluminum foil paper is filled, the packaging is carried out according to the operation requirement of the packaging machine, the filling amount difference is within +/-5 percent, and the mixture is subjected to inspection and external packaging after being filled.
Example 5 granules (calculated as 10000 bags)
Prescription:
drug A1250 g
20000g powdered sugar
Dextrin 9000g
5％PVP-K 30 Proper amount of
The drug A is one of the colimycin, the isovaleryl spiramycin III, the isovaleryl spiramycin II and the isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
The preparation process comprises the following steps: sieving the raw powder A, the powdered sugar and the dextrin with a 120-mesh sieve, weighing the raw powder A, the powdered sugar and the dextrin according to the prescription amount, uniformly mixing, and using 5 percent of PVP-K to uniformly mix the materials 30 Making the mucilage into soft material, granulating by swing type granule at 70 deg.C, drying, grading, inspecting, and packaging.
Example 6 lyophilized powder injection
Weighing 500mg of the drug A raw powder and equimolar adipic acid, uniformly mixing, and dissolving in 5ml of water to obtain a light yellow clear solution, wherein the pH value is 4.6-5.6. Adding mannitol 40mg as lyophilized proppant, rapidly freezing at low temperature for 9 hr, freeze drying to obtain yellowish loose block, and dissolving in 10ml sterile water before use.
The medicine A is one of colimycin, isovaleryl spiramycin III, isovaleryl spiramycin II and isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
Experimental example 1 establishment of tibial cancer pain model
Walker rat breast cancer cells were purchased from the shanghai institute of biomedical engineering. Collecting 1ml ascites tumor (about 2X 10) 7 One cell/ml) was inoculated into the abdominal cavity of 70-80g female Wistar rats, 5ml of ascites was extracted after 7-10 days and centrifuged for 3 minutes (1300 rpm), and after precipitation, washed with 10ml of PBS, centrifuged again and then counted in PBS, and the cell concentration was adjusted to 1X 10 7 Each cell/ml, put in ice box for use.
Rats were anesthetized with 10% chlorous acid hydrate (4 ml/kg body weight) by intraperitoneal injection, and the right knee joint was prepared with skin and the skin was sterilized with 75% alcohol. The left hand is used for fixing the knee joint to enable the surface skin to be tensed, and a No. 7 needle is used for drilling a hole from the inner side edge of patellar ligament of the knee joint to the far end of the tibia along the longitudinal axis of the tibia, and the depth is about 1cm. Then a microsyringe which is sucked with tumor cells is used for replacing (before injection, 1 mul of air, 2 mul of gelatin sponge aqueous solution, 1 mul of air, 4 mul of tumor cells or PBS are sucked in the microsyringe in sequence), and 4 mul of tumor cells are injected into the bone marrow cavity of the tibia; control rats were injected with an equal amount of PBS, and then sealed by pushing 2. Mu.l of gel sponge solution, and the needle was left for 1 minute after injection to prevent the injection from oozing out.
Rats were divided into two groups: PBS control group and Walker256 model group were injected. The radiation heat-shrinkable paw latency (PWL) was measured before molding as the basal pain threshold, and PWL was measured every other day after molding as the thermal hyperalgesia threshold, as shown in table 2, and the corresponding graphs are shown in fig. 3 and 4. Meanwhile, the mechanical pain caused by von Frey filaments is used for measuring, the mechanical pain threshold value is measured before the model is built and is used as the basic pain threshold value, the mechanical pain hypersensitivity threshold value is measured every other day after the model is built, and concretely, the table 1 shows that the corresponding graphs are shown in figures 1 and 2.
TABLE 1 Van Frey mechanical pain threshold (g) of post-molding rats: (
Figure BDA0002030713200000111
n＝8)
Figure BDA0002030713200000112
* p <0.05 to control, p <0.01 to control
TABLE 2 thermal pain threshold (sec) of the rats after molding: (
Figure BDA0002030713200000113
n＝8)
Figure BDA0002030713200000114
The results show a significant reduction in the mechanical pain threshold induced by Von Frey filaments in the vaccinated lateral hind limbs starting from day 2 after vaccination of Walker256 in rats compared to PBS rats (P < 0.05). Similar to the vaccinated side, the mechanical pain threshold of the lateral hind limb was similarly altered by rat vaccination with Walker 256. However, in the experiment of heat-sensitive pain, the two hind limbs of the rats after being inoculated with Walker256 have no significant difference to the threshold value of thermal hyperalgesia in the whole experiment process (P < 0.05).
Experimental example 2 Effect of drug A on mechanical pain threshold of rat with tibial cancer pain
Rats were randomized into 6 groups: model group, 10, 20, 40 and 80mg/kg drug A administration group, positive drug (morphine hydrochloride) control group. After the tibial cancer pain model is successfully established, continuous gavage administration is carried out for 12 days, the model group is fed with physiological saline with the same volume through gavage, and the positive medicine group is fed with 15mg/kg of morphine hydrochloride on the last day of the experiment. The change in mechanical pain threshold was measured in rats using mechanical pain induced by van Frey fine needle as an index, as detailed in table 3. The effective component of the medicine A comprises one of colimycin, isovaleryl spiramycin III, isovaleryl spiramycin II and isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
TABLE 3 threshold for van Frey mechanical pain (g) in rats after drug administration: (C)
Figure BDA0002030713200000121
n＝8)
Figure BDA0002030713200000122
* P <0.01 compared to control, p <0.001 compared to control
The results show that the mechanical pain threshold of the postoperative limb of rats of 20, 40 and 80mg/kg drug A administration groups and positive drug control groups is remarkably increased (p is less than 0.01) compared with that of model control groups, the analgesic rate reaches 83.3%, 127.8%, 188.9% and 205.5% respectively, the influence of drug modeling on the mechanical pain threshold of the collateral postoperative limb is similar, and the analgesic rate reaches 40.7%, 88.9%, 125.9% and 133.3% respectively, and the results show that the drug A can remarkably improve the mechanical pain threshold of rats with cancer pain.
In conclusion, a rat tibia cancer pain model is established, and Walker256 breast cancer cells are injected into a rat tibia bone marrow cavity by using a microsyringe. With the growth of tumor cells, there was no significant thermal hyperalgesia in the rat hindlimb, but significant mechanical hyperalgesia was produced, suggesting successful establishment of the model. After the model is established and the continuous gavage administration is carried out for 12 days, the bilateral hindlimb mechanical pain threshold of the rat is tested, and the result shows that the bilateral hindlimb mechanical pain threshold of the rat in the drug A administration group of 20-80mg/kg is obviously increased. The administration of 20-80mg/kg of the drug A remarkably improves the mechanical pain threshold of cancer pain rats, and experimental results show that the drug A can be used for treating cancer pain.
Experimental example 3 analgesic experiment by writhing method in mouse
The mouse writhing method is a classical method for screening analgesics, the principle of which is to observe the tolerance degree of a drug-administered mouse to chemical pains, and the method is to inject 0.2 ml/mouse of 1.44% acetic acid into the abdominal cavity of the drug-administered mouse and record the writhing times of animals within 30min of injection. The twisting index is that the abdomen climbs forwards along the cylinder bottom, the hind limbs stretch or the hip is lifted and twisted to one side, and the sensitivity of the twisting experiment is high.
60 healthy Kunming mice are selected, male mice with the weight of 18-22g are selected, and after animal quarantine is qualified, the mice are randomly divided into 6 groups, namely a blank control group, a 10mg/kg, 20mg/kg, 40mg/kg and 80mg/kg drug A dose group and a positive drug morphine hydrochloride 15mg/kg control group, and 10 mice are selected in each group. Before the experiment, each group of mice is first subjected to intragastric administration, and a blank control group of mice is given physiological saline with the same volume every day and is continuously administered for 3 days; the positive drug morphine hydrochloride was administered only 1 time the day of the experiment. The effective component of the medicine A comprises one of the colimycin, the isovaleryl spiramycin III, the isovaleryl spiramycin II and the isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
After 1h of administration, 0.2ml/20g of 1.44% glacial acetic acid is injected into the abdominal cavity of each mouse, the writhing times of each mouse after 20min of acetic acid injection are recorded, differences among groups are compared, and then the percent of pain easing of the medicine is calculated, wherein the percent of pain easing is% = (the writhing times of a blank group-the writhing times of an administration group) ÷ the writhing times of the blank group multiplied by 100 percent, and the specific results are shown in a table 4.
TABLE 4 number of writhing times of mice: (
Figure BDA0002030713200000131
n＝10)
Figure BDA0002030713200000132
* p <0.05 to control group,. P <0.01 to control group
The results show that the times of writhing of the mice are reduced along with the increase of the dosage of the medicine A, and the analgesic rate of the dosage groups of 10mg/kg, 20mg/kg, 40mg/kg and 80mg/kg of the medicine A to the pain of the mice caused by the glacial acetic acid is 15.26%, 22.48%, 35.92% and 48.65% respectively; the analgesic rate of the positive morphine hydrochloride is 45.83 percent, so that the medicament A with the concentration of 20-80mg/kg has remarkable analgesic effect.
Experimental example 4 Hot plate analgesia experiment in mice
The mouse hot plate method is a classical method for screening analgesic, and the principle is that a mouse is placed on a constant-temperature hot plate which is heated to 55 +/-0.1 ℃ in advance, the time required for adding the pain when the foot of the mouse is stimulated by the hot plate to generate the pain is used as a pain threshold value, the pain threshold value is used as an observation index, and the change of the threshold value before and after administration is compared to evaluate the effect of the analgesic.
60 healthy Kunming female mice are selected, the weight is 18-22g, after animal quarantine is qualified, the healthy Kunming female mice are randomly divided into 6 groups, namely a blank control group, a 10mg/kg control group, a 20mg/kg control group, a 40mg/kg control group, an 80mg/kg drug A dose group and a positive drug morphine hydrochloride 15mg/kg control group, and 10 mice are selected in each group. Before the experiment, each group of mice is first subjected to intragastric administration, and a blank control group of mice is given physiological saline with the same volume every day and is continuously administered for 3 days; the positive drug morphine hydrochloride was administered 1 time before the end of the experiment. The effective component of the medicine A comprises one of the colimycin, the isovaleryl spiramycin III, the isovaleryl spiramycin II and the isovaleryl spiramycin I; or a combination of two or three of isovalerylspiramycin I, isovalerylspiramycin II and isovalerylspiramycin III.
The mice were placed on a hot plate pain tester pre-heated to 55 ℃ and the time from the time the mice were thrown on the hot plate to the time the licking occurred was recorded by a stopwatch as the pain threshold for the mice. The pain threshold of each mouse was determined prior to dosing, and was found to be acceptable if the duration of the administration did not exceed 30 seconds. The test is carried out once at 30, 60, 90 and 120min after the administration, the test is finished, and the pain threshold increase percentage is calculated according to the average value of the measured pain threshold, wherein the pain threshold increase percentage is% = (average pain threshold after the administration-average pain threshold before the administration) ÷ average pain threshold before the administration is multiplied by 100 percent, which is shown in tables 5 and 6.
TABLE 5 pain threshold of mouse by Hot plate method: (
Figure BDA0002030713200000141
n＝10)
Figure BDA0002030713200000142
* p <0.05 to control group, p <0.01 to control group, p <0.001 to control group
TABLE 6 percentage increase in pain threshold by mouse hot plate method (n = 10)
Figure BDA0002030713200000143
The results show that the pain threshold increase percentages of 30min,40mg/kg and 80mg/kg of the drug A group and the positive drug group after administration are 21.21%, 49.38% and 68.11% respectively, the pain threshold increase percentages of the drug A group and the positive drug group after administration are 35.77%, 49.33% and 54.01% respectively, and the pain threshold increase percentages of the drug A group and the positive drug group after administration are 40.29%, 42.37% and 51.90% respectively after administration are 90min,40mg/kg and 80mg/kg respectively. Therefore, 40-80mg/kg of the drug A has a remarkable analgesic effect.
In conclusion, the mouse writhing method analgesia experiment shows that the writhing frequency of a mouse is reduced along with the increase of the administration dosage of the medicine A, and the analgesia rates of the medicine A dosage groups of 10mg/kg, 20mg/kg, 40mg/kg and 80mg/kg on the pain of the mouse caused by glacial acetic acid are 15.26%, 22.48%, 35.92% and 48.65% respectively; the analgesic rate of the positive morphine hydrochloride is 45.83 percent. A mouse hot plate method analgesia experiment shows that after the drug A is given, the time of the mouse licking the foot is prolonged to different degrees, wherein, the pain threshold values of the 40mg/kg and 80mg/kg dose groups at 30min and 60min are obviously improved, so the drug A has obvious analgesia effect and has important economic benefit and social benefit. The analgesic experiment of the mouse writhing method proves that the drug A with the dosage of 20-80mg/kg shows obvious analgesic effect. A mouse hot plate analgesic experiment shows that the drug A with the dosage of 40mg/kg and 80mg/kg produces a remarkable analgesic effect 30min after administration, and an experimental result shows that the drug A can be used for analgesia.
Experimental example 5 Effect of drug A on LPS-induced common fever model in rats
Experimental animals:
male healthy Sprague Dawley rats (grade SPF), weighing 220-260g, purchased from Liaoning Biotechnology Ltd, license number: SCXK (Liao) 2015-0001.
Experimental drugs and reagents:
LPS (100 mg): (Sigma company), lot number: l2880;
sodium chloride injection (0.45g, 500ml): (Shijiazhuang four drugs Co., ltd.) batch number: 101217406; dissolving and diluting 0.9% sodium chloride injection to 1mg/ml, subpackaging with EP tube, and storing at-80 deg.C; the solution was diluted with 0.9% sodium chloride injection to 10. Mu.g/ml (100. Mu.g/kg) of sterile injection just before use.
An experimental instrument:
electron body temperature type MC-347: produced by Ohlong company Limited (Dalian), the temperature error is +/-0.1 ℃;
TE2101-L electronic balance: beijing Sadolis Instrument systems, inc.;
PB602-N electronic balance: mettler-toledo instruments ltd (shanghai);
the operation steps are as follows:
grouping:
male SD rats, weighing 220-260g, were acclimatized for 3 days, fed with free water and food, and cycled for 12h day and night. The groups were randomized into 6 groups, blank, model, drugs A25, 50 and 100mg/kg or the positive drug ibuprofen 30mg/kg, 10 per group. Each group was administered by intragastric administration at a volume of 1ml/100g body weight. The normal group and the model group are given equal volume of distilled water, the drug A group and the positive drug group are given corresponding doses of drugs, the drug administration of each group is carried out for 2 days and 3 days continuously, after all mice are fasted for 6 hours, the normal group and the model group are given equal volume of distilled water, and the drug A group and the positive drug group are given last time. The rats in the other groups except the normal group after the last administration were molded by intraperitoneal (ip) injection of LPS (100. Mu.g/kg) and subcutaneous (sc) injection of 20% dry yeast suspension (1 ml/100 g) at the back. The anal temperature was measured 1h, 2h, 4h, 6h, 8h, and 10h after the administration in the manner described above, and the maximum body temperature rise Δ T (Δ T = body temperature at fever — basal body temperature) was calculated at each time point.
The results of the experiment are shown in table 7:
TABLE 7 Effect of drug A on LPS fever model rat body temperature changes (means. + -. S, n = 10)
Figure BDA0002030713200000161
According to the experimental results, compared with the blank group, the LPS model group generates heat obviously within 2-10 h, wherein 6h reaches the peak period and then gradually decreases. Compared with the model group, the drug A administration group can significantly reduce the body temperature of rats. At 2h and 10h, 50mg/kg and 100mg/kg were significant, and at 4h, 6h and 8h time points, the respective dose groups were significant. Therefore, it could be shown that drug A could improve LPS-induced common fever in rats.
Experimental example 6 Effect of drug A on common fever model in rats induced by Dry Yeast
Experimental drugs and reagents: in the experiment, LPS in the experimental example 1 is replaced by dry yeast which is a product of Angel yeast of Hubei, and is prepared into 20 percent uniform suspension by using distilled water at 0-4 ℃ before use.
Other experimental drugs and reagents were the same as in experimental example 1.
An experimental instrument: same as in experiment 1.
The method comprises the following operation steps: in this experiment, except that LPS in experiment 1 was replaced with dry yeast, the procedure and dose selection were the same as in experiment example 1, and finally the maximum body temperature rise Δ T (Δ T = body temperature at fever-basal body temperature) was calculated at each time point based on the anal temperatures measured 1h, 2h, 4h, 6h, 8h, and 10h after administration.
The results of the experiment are shown in table 8:
TABLE 8 Effect of drug A on changes in body temperature in rats in the Dry Yeast fever model (means. + -. S, n = 10)
Figure BDA0002030713200000171
The above experimental results show that, compared with the blank group, the dry yeast model group generates heat obviously within 2-10 h, wherein 6h reaches the peak period and then gradually decreases. Compared with the model group, the drug A administration group can significantly reduce the body temperature of rats. At 2h and 10h, 50mg/kg and 100mg/kg were significant, and at 4h, 6h and 8h time points, the respective dose groups were significant. Therefore, the medicine A can improve the common fever of rats caused by dry yeast.
Although the present invention has been described with reference to the preferred embodiments, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof without departing from the scope of the present invention.

Claims (5)

1. Use of a kelimycin for the manufacture of a medicament for the treatment of pain.
2. Use according to claim 1, characterized in that: the medicine is prepared into a pharmaceutically acceptable dosage form.
3. Use according to claim 1, characterized in that: the dosage of the medicine is 10-80 mg/kg.
4. Use according to claim 1, characterized in that: the pain includes cancer pain.
5. The use of any one of claims 1-4, wherein the medicament inhibits prostaglandin synthesis, inhibits substance P release, and inhibits reuptake of 5-hydroxytryptamine and norepinephrine.
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