CN114939141B - Traditional Chinese medicine composition for removing tear marks of pets and preparation method thereof - Google Patents
Traditional Chinese medicine composition for removing tear marks of pets and preparation method thereof Download PDFInfo
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- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
- A61K35/37—Digestive system
- A61K35/413—Gall bladder; Bile
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- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/28—Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
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- A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
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Abstract
The invention relates to the technical field of animal medicines, in particular to a traditional Chinese medicine composition for removing tear marks of pets and a preparation method thereof, wherein the traditional Chinese medicine composition is prepared from the following raw materials in parts by weight: 30-50 parts of animal bile, 5-10 parts of semen cassiae, 10-20 parts of mulberry fruit, 5-15 parts of marigold, 10-15 parts of medlar, 15-20 parts of eucommia ulmoides leaf, 1-5 parts of liquorice, 3-7 parts of herba euphorbiae lathyris and 8-12 parts of honeysuckle. The traditional Chinese medicine composition provided by the invention can continuously protect the liver and gall of a pet, so that the effects of continuously and efficiently preventing and relieving tear marks of the pet are achieved, the tear marks of the pet are radically treated, the traditional Chinese medicine composition has high utilization rate, one-time administration of the pet is realized, the long-term effective effect is realized, the administration period of the pet is effectively prolonged, and the effects of poor drug resistance and toxic and side effects of the drug of the pet can be avoided.
Description
Technical Field
The invention relates to the technical field of animal medicines, in particular to a traditional Chinese medicine composition for removing tear marks of pets and a preparation method thereof.
Background
With the gradual improvement of the living standard of people, the pet dogs are an important member in the family of keeping dogs, the welfare treatment is also more and more paid attention to, and the pet owners are also more and more concerned about the health of the self-family pet dogs, wherein the tear mark problem of the pet dogs is the most concerned. The tear is characterized by brown or pink tear spots, and is accompanied by bad smell, and even skin ulceration and other problems are caused when serious, and the main reason is caused by bacterial breeding around the eye orbit. At present, the domestic treatment method for tear marks is mainly drug treatment and operation treatment, wherein the operation treatment is carried out in a specific hospital, has high cost and has certain risk factors, so the treatment method is not easy to be accepted by a pet owner; the special convenience and low cost of the drug treatment become the primary choice for the pet owner to treat the tear marks.
For example, the chinese patent application No. CN202010268416.7 discloses a nutritional supplement formulation for improving tear marks of pet dogs and a preparation method thereof, which comprises purple sweet potato, perilla, inulin, oregano-phenol, magnolia flower, plantain seed, dodder seed, isomaltooligosaccharide, beta-glucan and mannooligosaccharide; according to the nutritional supplement, the raw materials are mixed in a specific proportion, the excessive tear phenomenon of the pet dogs can be improved to a certain extent after the pet dogs are fed, but the nutritional supplement does not have the effect of protecting the liver and the gallbladder, so that the liver and the gallbladder are heavily loaded when the bile is secreted to relieve the tear marks, the health of the liver and the gallbladder is not benefited, the tear marks of the pets cannot be treated fundamentally, the utilization rate of the nutritional supplement is low, only a small part of the nutritional supplement can play a role after the pets eat the nutritional supplement, and most of the nutritional supplement can be discharged from the body, so that the pets need to eat the pets in a regular and quantitative mode, the workload of the pets is greatly increased, and the phenomena of poor drug resistance and drug toxic and side effects of the pets are easy to occur.
Disclosure of Invention
The invention aims to provide a traditional Chinese medicine composition for removing tear marks of pets and a preparation method thereof, which can not only continuously protect liver and gall of the pets, thereby achieving the effects of continuously and efficiently preventing and relieving the tear marks of the pets, radically treating the tear marks of the pets, but also enabling the traditional Chinese medicine composition to have high utilization rate, realizing one-time administration of the pets, effectively prolonging the administration period of the pets for a long time and avoiding the drug resistance deterioration and the toxic and side effects of the drugs.
In order to achieve the above purpose, the present invention provides the following technical solutions:
the traditional Chinese medicine composition for removing the tear marks of the pets is prepared from the following raw materials in parts by weight: 30-50 parts of animal bile, 5-10 parts of semen cassiae, 10-20 parts of mulberry fruit, 5-15 parts of marigold, 10-15 parts of medlar, 15-20 parts of eucommia ulmoides leaf, 1-5 parts of liquorice, 3-7 parts of herba euphorbiae lathyris and 8-12 parts of honeysuckle.
As a further preferable scheme of the invention, the animal bile is at least one of chicken bile, duck bile and goose bile.
A preparation method of a traditional Chinese medicine composition for removing tear marks of pets comprises the following steps:
(1) Reflux-extracting animal bile to obtain bile extract, and treating the bile extract to obtain hydrophilic bile acid;
(2) Taking chitosan and hydrophilic bile acid as raw materials to synthesize a chitosan grafted bile acid derivative for standby, crushing semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaves, liquorice, millettia and honeysuckle to obtain traditional Chinese medicine powder, extracting by a supercritical carbon dioxide extraction technology to obtain traditional Chinese medicine extract, adding the chitosan grafted bile acid derivative into the traditional Chinese medicine extract, and uniformly mixing to obtain a medicament;
(3) Dissolving silk fibroin in a ternary solvent consisting of calcium chloride, ethanol and water, dialyzing in deionized water after complete dissolution to obtain silk fibroin solution, uniformly mixing the silk fibroin solution and sodium alginate solution to obtain a blending solution, adding a medicament into the blending solution, injecting the blending solution into a mold, performing ultrasonic treatment of 300-500W to gel to obtain composite hydrogel, and performing freeze drying and grinding into powder to obtain the medicine preparation.
As a further preferred embodiment of the present invention, in the step (1), the specific extraction method of the bile extract is as follows:
placing 20-30g animal bile into a container, adding 100-150mL petroleum ether, heating and refluxing at 50-60deg.C for 2-3h, suction filtering to obtain filter residue, repeating for 3-5 times, placing filtrate into a container, adding methanol with volume 8-10 times of that of the filtrate, heating and refluxing at 78-83deg.C for 1-2h, suction filtering to obtain filtrate, adding 1-5g montmorillonite, heating and refluxing at 78-83deg.C for 2-3h, suction filtering to obtain filtrate, repeating for 3-5 times, steaming the obtained filtrate until liquid-solid ratio is 3-4, and crystallizing at-4-0deg.C to obtain bile extract.
As a further preferable embodiment of the present invention, in the step (1), the method for treating the bile extract is as follows:
(1) Adding appropriate amount of bile extract into anhydrous methanol, heating to 50-55deg.C, stirring to dissolve completely, adding p-toluenesulfonic acid, refluxing for 2-5 hr, and rotary evaporating methanol to obtain product;
(2) Placing a proper amount of the standby product into a container, adding dichloromethane and triethylamine, dropwise adding acetic anhydride under stirring at room temperature, adding N, N-lutidine, reacting for 7-10h, rotationally steaming out dichloromethane to obtain an intermediate product, dissolving a proper amount of the intermediate product into tetrahydrofuran, adding triphenylphosphine and p-nitrobenzoic acid, slowly adding diisopropyl azodicarboxylate, stirring at room temperature for 20-25h under nitrogen, removing an organic solvent, and washing and drying to obtain a crude product;
(3) Dissolving the crude product in methanol, adding sodium hydroxide solution, reacting at room temperature for 1-3h, adding hydrochloric acid solution to adjust pH to 3-4, removing organic solvent, washing, and drying.
Still further, the bile extract, anhydrous methanol and p-toluenesulfonic acid were used in the ratio of (1-5) g: (50-100) mL: (0.1-0.3) g;
in the preparation process of the intermediate product, the dosage proportion of the standby product, dichloromethane, triethylamine, acetic anhydride and N, N-lutidine is (1-5) g: (20-30) mL: (3.5-4.3) mL: (1.2-1.8) g: (20-40) mg.
Further, in the preparation process of the crude product, the dosage ratio of the intermediate product, tetrahydrofuran, triphenylphosphine, p-nitrobenzoic acid and diisopropyl azodicarboxylate is (1-3) g: (70-100) mL: (1.1-1.5) g: (0.8-1.2) g: (0.2-0.5) g;
the dosage ratio of the crude product, methanol and sodium hydroxide solution is (1-3) g: (20-50) mL: (20-30) mL;
the concentration of the sodium hydroxide solution is 3-5wt%.
As a further preferable embodiment of the present invention, in the step (2), the synthesis method of the chitosan grafted bile acid derivative is as follows:
(1) 1-3g of chitosan is weighed and placed in a container, 20-30mL of acetic acid solution is added, stirring is carried out for 30-50min at normal temperature until the system is clear, 45-55mL of absolute ethyl alcohol is slowly added dropwise, 8-13g of hydrophilic bile acid is weighed and dissolved in 30-50mL of ethyl alcohol, then 0.6-0.8g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is slowly added dropwise into the clear system, then 0.6-10 mL of ethyl alcohol is weighed and dissolved, and stirring reaction is carried out for 20-30h at room temperature;
(2) After the reaction is finished, regulating the pH value of the reaction system to 7.5-8.5, adding enough ethanol, centrifuging at 3500-5000r/min for 3-5min, repeatedly washing the obtained product with ethanol, and freeze-drying for 40-50h to obtain the chitosan grafted bile acid derivative.
Further, the concentration of the acetic acid solution is 1-2wt%;
the volume ratio of the reaction system is adjusted to be (7-8): the ethanol/ammonia water mixed solution of (2- (3).
As a further preferable mode of the invention, in the step (3), the ratio of the amount of the silk fibroin to the ternary solvent is (10-20) g: (150-200) mL;
in the ternary solvent, the dosage proportion of calcium chloride, ethanol and water is (5-10) g: (30-50) mL: (100-150) mL;
the mass fraction of the sodium alginate solution is 0.5-1.0%;
in the blend liquid, the mass ratio of silk fibroin to sodium alginate is (60-90): (10-40);
the medicament accounts for 20-30% of the volume of the blend.
Compared with the prior art, the invention has the beneficial effects that:
the traditional Chinese medicine composition provided by the invention comprises animal bile, semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaf, liquorice, millet grass and honeysuckle; the method comprises the steps of taking animal bile as a raw material, carrying out reflux extraction to obtain a bile extract, taking the bile extract as a raw material, taking p-toluenesulfonic acid as a catalyst, and sequentially carrying out esterification, acetylation and alkali hydrolysis to obtain hydrophilic bile acid, wherein the bile acid has good water solubility, is beneficial to quick absorption of animals, can directly reduce the task of synthesizing bile acid by livers, plays a role in helping the livers to reduce load and make the livers rest more, and can also effectively promote the livers to secrete a large amount of rarefaction bile, help the livers to discharge harmful bacteria, heavy metals, self metabolic wastes and the like which are considered more along with the bile, provide safe and healthy internal environment for liver cells, and directly protect the liver cells from being affected by toxins, and play a role in protecting the livers and the gall; meanwhile, bile acid is supplemented to the animal body, so that bile secretion of the animal body can be promoted, and excretion of bilirubin is promoted, and tear marks of pets are fundamentally relieved; meanwhile, the supercritical carbon dioxide extraction technology is adopted to extract semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaves, liquorice, millettia and honeysuckle, and plant extracts extracted from semen cassiae, mulberry fruit, marigold and medlar can protect liver cells by protecting cell membranes, scavenging oxygen free radicals and inhibiting lipid peroxidation, so that the health degree of liver and gall of a pet is improved, and the liver and gall can not be subjected to heavy load when the pet secretes bile to relieve tear marks; meanwhile, the extract obtained by extracting eucommia ulmoides leaves, liquorice, herba euphorbiae lathyris and honeysuckle has good antibacterial and anti-inflammatory effects, and can reduce the occurrence of heavy pet eye inflammation, so that the outflow of tears is reduced, the periphery of the eye socket of a pet is not excessively moist, the breeding of bacteria on the periphery of the eye socket is reduced, and the effect of preventing and relieving the tear marks of the pet is further achieved.
In the invention, in order to further improve the protection effect of the traditional Chinese medicine composition on liver and gall, chitosan is used as a raw material, hydrophilic bile acid is connected with a chitosan main chain through an amide bond, and polysaccharide is connected to the bile acid, so that the generation of AST and ALT can be effectively inhibited, the antioxidation effect is enhanced, and simultaneously, the liver function can be activated, and liver cells are repaired, so that the liver protection effect is further enhanced; meanwhile, after the polysaccharide is accessed, the chitosan grafted bile acid derivative can be endowed with good antibacterial performance by utilizing the antibacterial effect of the chitosan, and the occurrence of heavy pet eye inflammation can be reduced, so that the effect of the traditional Chinese medicine composition in preventing and reducing the tear marks of pets is obviously improved.
Meanwhile, in order to improve the utilization efficiency of the traditional Chinese medicine composition, the high-porosity carrier is prepared by taking the silk fibroin and the sodium alginate as raw materials, and the carrier is used for loading the medicament, so that the carrier has good biocompatibility, can realize the sustained release of the medicament with a certain concentration, and can continuously protect the liver and gall of the pet, thereby achieving the effects of continuously and efficiently preventing and relieving tear marks of the pet, simultaneously, the traditional Chinese medicine composition has high utilization rate, realizes the once administration of the pet, effectively prolongs the administration period of the pet for a long time, and can avoid the effects of poor drug resistance and toxic and side effects of the medicament.
Detailed Description
The following description of the technical solutions in the embodiments of the present invention will be clear and complete, and it is obvious that the described embodiments are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Example 1
The traditional Chinese medicine composition for removing the tear marks of the pets is prepared from the following raw materials in parts by weight: 30 parts of chicken bile, 5 parts of semen cassiae, 10 parts of mulberry fruit, 5 parts of marigold, 10 parts of medlar, 15 parts of eucommia ulmoides leaf, 1 part of liquorice, 3 parts of herba euphorbiae lathyris and 8 parts of honeysuckle.
A preparation method of a traditional Chinese medicine composition for removing tear marks of pets comprises the following steps:
(1) Reflux extracting chicken bile to obtain bile extract, and treating the bile extract to obtain hydrophilic bile acid;
(2) Taking chitosan and hydrophilic bile acid as raw materials to synthesize a chitosan grafted bile acid derivative for standby, crushing semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaves, liquorice, millettia and honeysuckle to obtain traditional Chinese medicine powder, extracting by a supercritical carbon dioxide extraction technology to obtain traditional Chinese medicine extract, adding the chitosan grafted bile acid derivative into the traditional Chinese medicine extract, and uniformly mixing to obtain a medicament;
(3) 10g of silk fibroin is dissolved in a ternary solvent consisting of 150mL of calcium chloride, ethanol and water, wherein the dosage ratio of the calcium chloride to the ethanol to the water is 5g:30mL:100mL, after complete dissolution, dialyzing in deionized water to obtain a silk fibroin solution, uniformly mixing the silk fibroin solution and a sodium alginate solution with the mass fraction of 0.5%, and obtaining a blend solution, wherein the mass ratio of silk fibroin to sodium alginate is controlled to be 60:40, adding the medicament into the blending liquid, controlling the medicament to occupy 20 percent of the volume of the blending liquid, injecting the mixture into a mould, performing ultrasonic treatment at 300W for gelation to obtain composite hydrogel, and grinding the composite hydrogel into powder after freeze drying to finish the preparation of the medicament.
The specific extraction method of the bile extract comprises the following steps:
placing 20g of chicken bile in a container, adding 100mL of petroleum ether, heating and refluxing for 2h at 50 ℃, suction-filtering to obtain filter residues, repeating for 3 times, placing the filtrate in the container, adding methanol which is 8 times of the volume of the filtrate, heating and refluxing for 1h at 78 ℃, suction-filtering to obtain the filtrate, adding 1g of montmorillonite, heating and refluxing for 2h at 78 ℃, suction-filtering to obtain the filtrate, repeating for 3 times, steaming the obtained filtrate until the liquid-solid ratio is 3, and crystallizing at-4 ℃ to obtain the bile extract.
The treatment method of the bile extract comprises the following steps:
(1) Adding 1g of bile extract into 50mL of absolute methanol, heating to 50 ℃, stirring to dissolve the bile extract completely, then adding 0.1g of p-toluenesulfonic acid, refluxing for 2 hours, and then steaming out the methanol in a rotary manner to obtain a product for later use;
(2) 1g of the standby product is weighed and placed in a container, 20mL of dichloromethane and 3.5mL of triethylamine are added, 1.2g of acetic anhydride is added dropwise under stirring at room temperature, 20mg of N, N-lutidine is added for reaction for 7h, dichloromethane is distilled off in a rotary way to obtain an intermediate product, 1g of the intermediate product is dissolved in 70mL of tetrahydrofuran, 1.1g of triphenylphosphine and 0.8g of p-nitrobenzoic acid are added, 0.2g of diisopropyl azodicarboxylate is slowly added, stirring is carried out at room temperature for 20h under the condition of nitrogen, an organic solvent is distilled off in a rotary way, the organic phase is washed by saturated sodium chloride solution, anhydrous sodium sulfate is dried, and a crude product is obtained after concentration;
(3) 1g of the crude product is dissolved in 20mL of methanol, then 20mL of sodium hydroxide solution with the concentration of 3wt% is added for reaction for 1h at room temperature, hydrochloric acid solution with the concentration of 5wt% is added for regulating the pH value to 3, and the crude product is obtained through rotary evaporation of methanol, extraction of ethyl acetate, washing with saturated sodium chloride solution and drying with anhydrous sodium sulfate.
The preparation method of the chitosan grafted bile acid derivative comprises the following steps:
(1) 1g of chitosan is weighed and placed in a container, 20mL of acetic acid solution with the concentration of 1wt% is added, stirring is carried out for 30min at normal temperature until the system is clear, 45mL of absolute ethyl alcohol is slowly added dropwise, 8g of hydrophilic bile acid is weighed and dissolved in 30mL of ethyl alcohol, then 0.6g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is slowly added dropwise into the clear system, 0.6g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is weighed and dissolved in 6mL of ethyl alcohol, and stirring is carried out for 20h at room temperature;
(2) After the reaction is finished, the volume ratio is 7:3, regulating the pH value of the reaction system to 7.5 by using an ethanol/ammonia water mixed solution, adding enough ethanol, centrifuging at 3500r/min for 3min, repeatedly washing the obtained product by using ethanol, and freeze-drying for 40h to obtain the chitosan grafted bile acid derivative.
Example 2
The traditional Chinese medicine composition for removing the tear marks of the pets is prepared from the following raw materials in parts by weight: 40 parts of duck bile, 7 parts of semen cassiae, 15 parts of mulberry fruit, 10 parts of marigold, 13 parts of medlar, 18 parts of eucommia ulmoides leaf, 3 parts of liquorice, 5 parts of herba euphorbiae lathyris and 10 parts of honeysuckle.
A preparation method of a traditional Chinese medicine composition for removing tear marks of pets comprises the following steps:
(1) Reflux-extracting duck bile to obtain bile extract, and treating the bile extract to obtain hydrophilic bile acid;
(2) Taking chitosan and hydrophilic bile acid as raw materials to synthesize a chitosan grafted bile acid derivative for standby, crushing semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaves, liquorice, millettia and honeysuckle to obtain traditional Chinese medicine powder, extracting by a supercritical carbon dioxide extraction technology to obtain traditional Chinese medicine extract, adding the chitosan grafted bile acid derivative into the traditional Chinese medicine extract, and uniformly mixing to obtain a medicament;
(3) 15g of silk fibroin is dissolved in a ternary solvent consisting of 180mL of calcium chloride, ethanol and water, wherein the dosage ratio of the calcium chloride to the ethanol to the water is 8g:40mL:130mL, after complete dissolution, dialyzing in deionized water to obtain a silk fibroin solution, uniformly mixing the silk fibroin solution and a sodium alginate solution with the mass fraction of 0.8%, and obtaining a blend solution, wherein the mass ratio of silk fibroin to sodium alginate is controlled to be 70:30, adding the medicament into the blend, controlling the medicament to account for 25 percent of the volume of the blend, injecting the blend into a mold, performing 400W ultrasonic treatment to gel to obtain composite hydrogel, and grinding the composite hydrogel into powder after freeze drying to finish the preparation of the medicament.
The specific extraction method of the bile extract comprises the following steps:
placing 25g of animal bile into a container, adding 120mL of petroleum ether, heating and refluxing for 2.5h at 55 ℃, suction-filtering to obtain filter residues, repeating for 4 times, placing the filtrate into the container, adding methanol which is 9 times of the volume of the filtrate, heating and refluxing for 1.5h at 80 ℃, suction-filtering to obtain the filtrate, adding 3g of montmorillonite, heating and refluxing for 2.5h at 80 ℃, suction-filtering to obtain the filtrate, repeating for 4 times, steaming the obtained filtrate until the liquid-solid ratio is 3.5, and crystallizing at-2 ℃ to obtain the bile extract.
The treatment method of the bile extract comprises the following steps:
(1) Adding 3g of bile extract into 70mL of absolute methanol, heating to 52 ℃, stirring to dissolve the bile extract completely, then adding 0.2g of p-toluenesulfonic acid, refluxing for 3 hours, and then steaming out the methanol in a rotary manner to obtain a product for later use;
(2) Weighing 3g of the standby product, placing the standby product into a container, adding 25mL of dichloromethane and 3.9mL of triethylamine, dropwise adding 1.5g of acetic anhydride under stirring at room temperature, adding 30mg of N, N-lutidine, reacting for 8 hours, rotationally evaporating dichloromethane to obtain an intermediate product, dissolving 2g of the intermediate product into 80mL of tetrahydrofuran, adding 1.3g of triphenylphosphine and 1g of p-nitrobenzoic acid, slowly adding 0.3g of diisopropyl azodicarboxylate, stirring at room temperature for 23 hours under nitrogen, rotationally evaporating an organic solvent, washing an organic phase with saturated sodium chloride solution, drying with anhydrous sodium sulfate, and concentrating to obtain a crude product;
(3) 2g of the crude product is dissolved in 35mL of methanol, 25mL of sodium hydroxide solution with the concentration of 4wt% is added for reaction for 2h at room temperature, hydrochloric acid solution with the concentration of 7wt% is added for regulating the pH value to 3.5, and the crude product is obtained through rotary evaporation of methanol, extraction of ethyl acetate, washing with saturated sodium chloride solution and drying with anhydrous sodium sulfate.
The preparation method of the chitosan grafted bile acid derivative comprises the following steps:
(1) 2g of chitosan is weighed and placed in a container, 25mL of acetic acid solution with the concentration of 1.5wt% is added, stirring is carried out for 40min at normal temperature until the system is clear, 50mL of absolute ethyl alcohol is slowly added dropwise, 10g of hydrophilic bile acid is weighed and dissolved in 40mL of ethyl alcohol, then 0.7g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is slowly added dropwise into the clear system, then 0.7g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is weighed and dissolved in 8mL of ethyl alcohol, and stirring is carried out for 25h at room temperature;
(2) After the reaction is finished, the volume ratio is 8:2, regulating the pH value of the reaction system to 8 by using an ethanol/ammonia water mixed solution, adding enough ethanol, centrifuging for 5min at 4000r/min, repeatedly washing the obtained product by using ethanol, and freeze-drying for 45h to obtain the chitosan grafted bile acid derivative.
Example 3
The traditional Chinese medicine composition for removing the tear marks of the pets is prepared from the following raw materials in parts by weight: 50 parts of goose bile, 10 parts of semen cassiae, 20 parts of mulberry fruit, 15 parts of marigold, 15 parts of medlar, 20 parts of eucommia ulmoides leaf, 5 parts of liquorice, 7 parts of herba euphorbiae lathyris and 12 parts of honeysuckle.
A preparation method of a traditional Chinese medicine composition for removing tear marks of pets comprises the following steps:
(1) Reflux-extracting goose bile to obtain bile extract, and treating the bile extract to obtain hydrophilic bile acid;
(2) Taking chitosan and hydrophilic bile acid as raw materials to synthesize a chitosan grafted bile acid derivative for standby, crushing semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaves, liquorice, millettia and honeysuckle to obtain traditional Chinese medicine powder, extracting by a supercritical carbon dioxide extraction technology to obtain traditional Chinese medicine extract, adding the chitosan grafted bile acid derivative into the traditional Chinese medicine extract, and uniformly mixing to obtain a medicament;
(3) 20g of silk fibroin is dissolved in a ternary solvent consisting of 200mL of calcium chloride, ethanol and water, wherein the dosage ratio of the calcium chloride to the ethanol to the water is 10g:50mL:150mL, completely dissolving, dialyzing in deionized water to obtain a silk fibroin solution, uniformly mixing the silk fibroin solution and a sodium alginate solution with the mass fraction of 1.0%, and obtaining a blend solution, wherein the mass ratio of silk fibroin to sodium alginate is controlled to be 90:10, adding the medicament into the blend, controlling the medicament to account for 30 percent of the volume of the blend, injecting the blend into a mould, carrying out 500W ultrasonic treatment for gelation to obtain composite hydrogel, and grinding the composite hydrogel into powder after freeze drying to finish the preparation of the medicament.
The specific extraction method of the bile extract comprises the following steps:
placing 30g of animal bile in a container, adding 150mL of petroleum ether, heating and refluxing for 3h at 60 ℃, suction-filtering to obtain filter residues, repeating for 5 times, placing the filtrate in the container, adding methanol which is 10 times of the volume of the filtrate, heating and refluxing for 2h at 83 ℃, suction-filtering to obtain the filtrate, adding 5g of montmorillonite, heating and refluxing for 3h at 83 ℃, suction-filtering to obtain the filtrate, repeating for 5 times, steaming the obtained filtrate until the liquid-solid ratio is 4, and crystallizing at 0 ℃ to obtain the bile extract.
The treatment method of the bile extract comprises the following steps:
(1) Adding 5g of bile extract into 100mL of absolute methanol, heating to 55 ℃, stirring to dissolve completely, then adding 0.3g of p-toluenesulfonic acid, refluxing for 5h, and then steaming out methanol in a rotary manner to obtain a product for later use;
(2) Weighing 5g of the standby product, placing the standby product into a container, adding 30mL of dichloromethane and 4.3mL of triethylamine, dropwise adding 1.8g of acetic anhydride under stirring at room temperature, adding 40mgN, N-lutidine, reacting for 10 hours, rotationally evaporating dichloromethane to obtain an intermediate product, dissolving 3g of the intermediate product into 100mL of tetrahydrofuran, adding 1.5g of triphenylphosphine and 1.2g of p-nitrobenzoic acid, slowly adding 0.5g of diisopropyl azodicarboxylate, stirring at room temperature for 25 hours under nitrogen, rotationally evaporating an organic solvent, washing an organic phase with saturated sodium chloride solution, drying with anhydrous sodium sulfate, and concentrating to obtain a crude product;
(3) 3g of the crude product is dissolved in 50mL of methanol, 30mL of 5wt% sodium hydroxide solution is added for reaction at room temperature for 3h, 8wt% hydrochloric acid solution is added for regulating the pH value to be 4, and the crude product is obtained through rotary evaporation of methanol, extraction of ethyl acetate, washing with saturated sodium chloride solution and drying with anhydrous sodium sulfate.
The preparation method of the chitosan grafted bile acid derivative comprises the following steps:
(1) 3g of chitosan is weighed and placed in a container, 30mL of acetic acid solution with the concentration of 2wt% is added, stirring is carried out for 50min at normal temperature until the system is clear, then 55mL of absolute ethyl alcohol is slowly added dropwise, 13g of hydrophilic bile acid is weighed and dissolved in 50mL of ethyl alcohol, then 0.8g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is slowly added dropwise into the clear system, then 0.8g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is weighed and dissolved in 10mL of ethyl alcohol, and stirring is carried out for 30h at room temperature;
(2) After the reaction is finished, the volume ratio is 8:2, regulating the pH value of the reaction system to 8.5 by using an ethanol/ammonia water mixed solution, adding enough ethanol, centrifuging for 5min at 5000r/min, repeatedly washing the obtained product by using ethanol, and freeze-drying for 50h to obtain the chitosan grafted bile acid derivative.
Comparative example 1: this comparative example is substantially the same as example 1 except that the bile extract is not treated in the preparation step (1) of the Chinese medicinal composition.
Comparative example 2: this comparative example is substantially the same as example 1 except that in the preparation step (2) of the Chinese medicinal composition, the hydrophilic bile acid is directly added to the Chinese medicinal extract.
Comparative example 3: this comparative example is substantially the same as example 1 except that the preparation step (3) of the Chinese medicinal composition is omitted.
Performance detection and effect evaluation:
by linking a local pet hospital and a pet store, 20 dogs with weight of about 4kg and white ratio Xiong Quan with certain tear mark problem are selected and randomly divided into 4 groups, and each group is tested by 5 dogs. In the experimental process, the pet dogs can eat and drink water freely, and are fed with common commercial dog foods respectively, and meanwhile, the traditional Chinese medicine compositions provided in the examples 1-3 and the comparative examples 1-3 are used for feeding the pet dogs once a day, and the using amount of each time is 10g/kg. Schirmer Tear Test (STT) was performed once a week and observed for tear stain change for a period of 4 weeks.
Table 1 comparison of tear levels in test period of test dogs
Table 2 comparison of tear stain sensory changes during the test period of the test dogs
As can be seen from tables 1 and 2, after 4 weeks of feeding of the traditional Chinese medicine composition, the tear level of the test dogs is gradually restored to the normal level (the normal value of the tear level of the dogs is 15+/-5 mm/min), wet hair around the eyesockets is gradually dried, the necessary conditions for microorganism breeding are reduced, so that the unpleasant smell generated by tear marks is gradually reduced until the tear marks disappear, and the traditional Chinese medicine composition provided by the invention has good effects of preventing, relieving and treating the tear marks of pets.
The preferred embodiments of the invention disclosed above are intended only to assist in the explanation of the invention. The preferred embodiments are not exhaustive or to limit the invention to the precise form disclosed. Obviously, many modifications and variations are possible in light of the above teaching. The embodiments were chosen and described in order to best explain the principles of the invention and the practical application, to thereby enable others skilled in the art to best understand and utilize the invention. The invention is limited only by the claims and the full scope and equivalents thereof.
Claims (7)
1. The traditional Chinese medicine composition for removing the tear marks of the pets is characterized by being prepared from the following raw materials in parts by weight: 30-50 parts of animal bile, 5-10 parts of semen cassiae, 10-20 parts of mulberry fruit, 5-15 parts of marigold, 10-15 parts of medlar, 15-20 parts of eucommia ulmoides leaf, 1-5 parts of liquorice, 3-7 parts of herba euphorbiae lathyris and 8-12 parts of honeysuckle;
the preparation method of the traditional Chinese medicine composition for removing tear marks of pets comprises the following steps:
(1) Reflux-extracting animal bile to obtain bile extract, and treating the bile extract to obtain hydrophilic bile acid;
(2) Taking chitosan and hydrophilic bile acid as raw materials to synthesize a chitosan grafted bile acid derivative for standby, crushing semen cassiae, mulberry fruit, marigold, medlar, eucommia ulmoides leaves, liquorice, millettia and honeysuckle to obtain traditional Chinese medicine powder, extracting by a supercritical carbon dioxide extraction technology to obtain traditional Chinese medicine extract, adding the chitosan grafted bile acid derivative into the traditional Chinese medicine extract, and uniformly mixing to obtain a medicament;
(3) Dissolving silk fibroin in a ternary solvent consisting of calcium chloride, ethanol and water, dialyzing in deionized water after complete dissolution to obtain silk fibroin solution, uniformly mixing the silk fibroin solution and sodium alginate solution to obtain a blending solution, adding a medicament into the blending solution, injecting the blending solution into a mold, performing ultrasonic treatment of 300-500W to gel to obtain composite hydrogel, and performing freeze drying and grinding into powder to obtain the medicine preparation;
in the step (1), the specific extraction method of the bile extract is as follows:
placing 20-30g of animal bile into a container, adding 100-150mL of petroleum ether, heating and refluxing for 2-3h at 50-60 ℃, suction filtering to obtain filter residues, repeating for 3-5 times, placing the filtrate into the container, adding methanol with the volume being 8-10 times that of the filtrate, heating and refluxing for 1-2h at 78-83 ℃, suction filtering to obtain the filtrate, adding 1-5g of montmorillonite, heating and refluxing for 2-3h at 78-83 ℃, suction filtering to obtain the filtrate, repeating for 3-5 times, steaming the obtained filtrate until the liquid-solid ratio is 3-4, and crystallizing at-4-0 ℃ to obtain a bile extract;
in the step (1), the treatment method of the bile extract comprises the following steps:
(1) Adding appropriate amount of bile extract into anhydrous methanol, heating to 50-55deg.C, stirring to dissolve completely, adding p-toluenesulfonic acid, refluxing for 2-5 hr, and rotary evaporating methanol to obtain product;
(2) Placing a proper amount of the standby product into a container, adding dichloromethane and triethylamine, dropwise adding acetic anhydride under stirring at room temperature, adding N, N-lutidine, reacting for 7-10h, rotationally steaming out dichloromethane to obtain an intermediate product, dissolving a proper amount of the intermediate product into tetrahydrofuran, adding triphenylphosphine and p-nitrobenzoic acid, slowly adding diisopropyl azodicarboxylate, stirring at room temperature for 20-25h under nitrogen, removing an organic solvent, and washing and drying to obtain a crude product;
(3) Dissolving the crude product in methanol, adding sodium hydroxide solution, reacting at room temperature for 1-3h, adding hydrochloric acid solution to adjust pH to 3-4, removing organic solvent, washing, and drying.
2. The composition of claim 1, wherein the animal bile is at least one of chicken bile, duck bile, and goose bile.
3. The traditional Chinese medicine composition for removing tear marks of pets according to claim 1, wherein the dosage ratio of the bile extract, the absolute methanol and the p-toluenesulfonic acid is (1-5) g: (50-100) mL: (0.1-0.3) g;
in the preparation process of the intermediate product, the dosage proportion of the standby product, dichloromethane, triethylamine, acetic anhydride and N, N-lutidine is (1-5) g: (20-30) mL: (3.5-4.3) mL: (1.2-1.8) g: (20-40) mg.
4. The composition of claim 1, wherein the ratio of the amounts of the intermediate product, tetrahydrofuran, triphenylphosphine, p-nitrobenzoic acid and diisopropyl azodicarboxylate in the crude product is (1-3) g: (70-100) mL: (1.1-1.5) g: (0.8-1.2) g: (0.2-0.5) g;
the dosage ratio of the crude product, methanol and sodium hydroxide solution is (1-3) g: (20-50) mL: (20-30) mL;
the concentration of the sodium hydroxide solution is 3-5wt%.
5. The traditional Chinese medicine composition for removing tear marks of pets according to claim 1, wherein in the step (2), the synthesis method of the chitosan grafted bile acid derivative is as follows:
(1) 1-3g of chitosan is weighed and placed in a container, 20-30mL of acetic acid solution is added, stirring is carried out for 30-50min at normal temperature until the system is clear, 45-55mL of absolute ethyl alcohol is slowly added dropwise, 8-13g of hydrophilic bile acid is weighed and dissolved in 30-50mL of ethyl alcohol, then 0.6-0.8g of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride is slowly added dropwise into the clear system, then 0.6-10 mL of ethyl alcohol is weighed and dissolved, and stirring reaction is carried out for 20-30h at room temperature;
(2) After the reaction is finished, regulating the pH value of the reaction system to 7.5-8.5, adding enough ethanol, centrifuging at 3500-5000r/min for 3-5min, repeatedly washing the obtained product with ethanol, and freeze-drying for 40-50h to obtain the chitosan grafted bile acid derivative.
6. The composition of claim 5, wherein the concentration of the acetic acid solution is 1-2wt%;
the volume ratio of the reaction system is adjusted to be (7-8): the ethanol/ammonia water mixed solution of (2-3).
7. The traditional Chinese medicine composition for removing tear marks of pets according to claim 1, wherein in the step (3), the dosage ratio of the silk fibroin to the ternary solvent is (10-20) g: (150-200) mL;
in the ternary solvent, the dosage proportion of calcium chloride, ethanol and water is (5-10) g: (30-50) mL: (100-150) mL;
the mass fraction of the sodium alginate solution is 0.5-1.0%;
in the blend liquid, the mass ratio of silk fibroin to sodium alginate is (60-90): (10-40);
the medicament accounts for 20-30% of the volume of the blend.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102754756A (en) * | 2012-08-08 | 2012-10-31 | 上海比瑞吉宠物用品有限公司 | Natural plant feed additive for improving tear stains of poodles and preparation method thereof |
| CN113143971A (en) * | 2021-02-03 | 2021-07-23 | 山东龙昌动物保健品有限公司 | Application of bile acid complex microbial inoculum in preparation of preparation for preventing and treating pet tear stains |
| CN114009587A (en) * | 2021-11-03 | 2022-02-08 | 华兴宠物食品有限公司 | Composition for improving tear stains of dogs and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102754756A (en) * | 2012-08-08 | 2012-10-31 | 上海比瑞吉宠物用品有限公司 | Natural plant feed additive for improving tear stains of poodles and preparation method thereof |
| CN113143971A (en) * | 2021-02-03 | 2021-07-23 | 山东龙昌动物保健品有限公司 | Application of bile acid complex microbial inoculum in preparation of preparation for preventing and treating pet tear stains |
| CN114009587A (en) * | 2021-11-03 | 2022-02-08 | 华兴宠物食品有限公司 | Composition for improving tear stains of dogs and preparation method thereof |
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| 王晓玲 ; 王成永 ; .正交试验法优选六味茶口服液中菊花、金银花的提取工艺.中医药临床杂志.-,2012,第24卷(第07期),679-681. * |
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