Compound ant collateral activating capsule and preparation method thereof

Abstract

The invention relates to the technical field of capsules for treating rheumatoid diseases, and discloses a compound ant collateral activating capsule and a preparation method thereof.The preparation method comprises the following preparation steps that caulis sinomenii, kadsura pepper stem, garden balsam stem and long-noded pit viper are mixed and smashed into 25-30-mesh coarse powder, the coarse powder is cured and sterilized, then the coarse powder is smashed into 100-120-mesh fine powder, and a compound is obtained; and mixing the modified black ant particles, a compound, starch and ethanol to prepare particles of 0.1-0.2 mm, drying the particles, and filling the particles into hollow capsules to obtain the compound ant collateral activating capsules. The modified black ant granules are compounded with traditional Chinese medicinal materials such as caulis sinomenii, kadsura pepper stem, garden balsam stem and long-noded pit viper, the effects of resisting inflammation, regulating immunity, resisting oxidation, promoting blood circulation to remove meridian obstruction, inhibiting synovial hyperplasia and the like are achieved, the prepared compound ant collateral activating capsule can effectively relieve the symptoms of rheumatoid arthritis, and the situation that the overall curative effect is poor due to the fact that the expected effect is difficult to achieve through single medicine treatment is avoided.

Description

Technical Field

[0001] The present invention relates to the technical field of capsules for treating rheumatoid diseases, and specifically to a compound ant activating collaterals capsule and a preparation method thereof. Background Art

[0002] Rheumatoid arthritis is an autoimmune disease with synovitis as the pathological basis and may ultimately lead to joint deformity. Traditional Chinese medicine believes that this disease belongs to "bi syndrome", which is due to the patient's congenital endowment deficiency, wind-cold-damp pathogens, resulting in blockage of joint meridians, stagnation of qi and blood. Its pathological characteristics are multiple joint involvements, synovial hyperplasia and persistent recurrence of synovitis. The condition is lingering and difficult to cure, which can lead to destruction of intra-articular bone, joint dysfunction, and is extremely prone to disability, seriously affecting the quality of life of patients. Therefore, dredging meridians and strengthening the healthy qi to consolidate the root are the main principles for treating this disease. Currently, the drugs for treating rheumatoid arthritis clinically mainly include biological agents, glucocorticoids, antirheumatic drugs, non-steroidal anti-inflammatory drugs, etc. Long-term use will cause many adverse reactions, such as increasing the susceptibility to infection and causing peptic ulcers.

[0003] Black ants, as traditional Chinese medicines, are often used to treat joint pain, traumatic injuries, etc. The extract of black ants, as a drug ingredient, has the effects of promoting blood circulation to remove blood stasis, dispelling wind and dampness, and strengthening the healthy qi to consolidate the root, and can effectively relieve the symptoms of rheumatoid arthritis. However, the extract of black ants contains hydrophilic components such as polysaccharides and amino acids, which are easy to absorb moisture and react with oxygen or moisture in the environment, resulting in a decrease in the activity of the extract of black ants. Moreover, single-drug treatment is difficult to achieve the expected effect, resulting in poor overall efficacy. Summary of the Invention

[0004] The present invention provides a compound ant activating collaterals capsule and a preparation method thereof, which solves the problems that the extract of black ants, as a drug component, has poor stability and single-drug treatment is difficult to achieve the expected effect.

[0005] The technical solution of the present invention:

[0006] A preparation method of a compound ant activating collaterals capsule includes the following preparation steps:

[0007] S1. Mix caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus, and pulverize them into a coarse powder of 25-30 meshes. After the coarse powder is cooked and sterilized, it is pulverized into a fine powder of 100-120 meshes to obtain a composite.

[0008] S2. Mix the modified black ant particles, the composite, starch, and ethanol to form particles of 0.1-0.2 mm. After the particles are dried, they are filled into hollow capsules to obtain the compound ant activating collaterals capsule;

[0009] The modified black ant particles are obtained by mixing and reacting black ant particles, carboxyl-polyethylene glycol-carboxyl, and cyclic peptide RGD-polyethylene glycol-carboxyl;

[0010] The black ant particles are obtained by mixing lipid nanoparticles of black ant extract, lemon peel extract, sesbania leaf extract and silk fibroin as the core material, and β-cyclodextrin and gum arabic as the capsule material;

[0011] The lipid nanoparticles of black ant extract are obtained by mixing black ant extract, stearic acid and soy lecithin to form an oil phase, poloxamer 188 as the water phase, and reacting the oil phase and the water phase.

[0012] Further, in step S1, the mass ratio of caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae and agkistrodon acutus is (8 - 10):(5 - 7):(7 - 9):(10 - 12).

[0013] Further, in step S1, the ripening and sterilization temperature is 100 - 110 °C, and the ripening and sterilization time is 10 - 30 min.

[0014] Further, in step S2, the mass ratio of the modified black ant particles, the complex, starch and ethanol is (80 - 100):(30 - 40):(40 - 50):(20 - 25).

[0015] Further, the modified black ant particles are specifically prepared by the following steps:

[0016] A1. Crush black ants into powder with 80 - 110 meshes. After sterilization treatment, extract three times with ethanol at an extraction temperature of 75 - 85 °C. Mix the obtained extract a, extract b and extract c, and freeze-dry to obtain black ant extract;

[0017] A2. Add black ant extract, stearic acid and soy lecithin to ethanol, stir evenly to obtain an oil phase. Add poloxamer 188 to deionized water, stir evenly to obtain a water phase. Add the oil phase to the water phase, stir evenly, stir at 1 - 3 °C for 3 - 5 h, centrifuge and wash, add to mannitol solution, stir, filter, and freeze-dry to obtain lipid nanoparticles of black ant extract;

[0018] A3. Add lipid nanoparticles of black ant extract, lemon peel extract, sesbania leaf extract and silk fibroin to ethanol, stir evenly to obtain the core material. Add β-cyclodextrin and gum arabic to ethanol and deionized water, stir evenly, add the core material, stir and react at 55 - 65 °C for 4 - 6 h, filter with a vacuum pump to obtain a solid, and freeze-dry the solid to obtain black ant particles;

[0019] A4. Add black ant particles and carboxyl-polyethylene glycol-carboxyl to ethanol, stir evenly, cool to 20 - 30 °C, add cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 10 - 12 h, centrifuge to collect the particles to obtain modified black ant particles.

[0020] Further, in the above A2 reaction process, at 60°C, stearic acid, soy lecithin, and black ant extract are mixed to form an oil phase, and poloxamer 188 is dissolved in deionized water as the aqueous phase. The oil phase is added to the aqueous phase, the hydrophobic segments of poloxamer 188 extend into the oil phase, and the hydrophilic segments of poloxamer 188 extend into the aqueous phase, thereby forming lipid nanoparticles coated with stearic acid and soy lecithin on the black ant extract, and obtaining black ant extract lipid nanoparticles.

[0021] Further, in the above A3 reaction process, lipid nanoparticles, lemon peel extract, sesbania leaf extract, and silk fibroin are mixed to form a complex as the core material, and β-cyclodextrin and gum arabic are used as the wall materials to achieve the encapsulation of the complex by β-cyclodextrin and gum arabic, forming microcapsules.

[0022] Further, in the above A4 reaction process, the carboxyl groups contained in carboxyl-PEG-carboxyl can chemically bond with the oxygen-containing functional groups on the surface of black ant particles, so that carboxyl-PEG-carboxyl is grafted onto the surface of black ant particles; the carboxyl-PEG-carboxyl on the surface of black ant particles has good compatibility with cyclic peptide RGD-PEG-carboxyl, and the molecular chains are bonded by hydrogen bonds, making cyclic peptide RGD-PEG-carboxyl more firmly grafted onto the surface of black ant particles, obtaining modified black ant particles.

[0023] Further, in step A1, the black ant extract is prepared by the following steps:

[0024] The black ants are crushed into a powder of 80 - 110 mesh, sterilized to obtain sterilized black ant powder. The sterilized black ant powder is added to ethanol, extracted at 75 - 85°C for 3 - 5 h, filtered, and the extract a and the residue a are collected. The residue a is added to ethanol and extracted continuously for 2 - 4 h, filtered to obtain extract b and residue b. The residue b is added to ethanol and extracted continuously for 1 - 3 h, filtered to obtain extract c. Extract a, extract b, and extract c are mixed and freeze-dried to obtain the black ant extract.

[0025] Further, the mass ratio of the sterilized black ant powder to ethanol is (8 - 12):(43.4 - 47.3 - 51.3).

[0026] Further, the mass ratio of the residue a to ethanol is (4 - 6):(10.3 - 11.8 - 13.4).

[0027] Further, the mass ratio of the residue b to ethanol is (2 - 4) g:(4 - 4.7 - 5.5).

[0028] Further, the sterilization temperature is 100 - 110°C, and the sterilization time is 10 - 30 min.

[0029] Further, in step A2, the dosage ratio of the black ant extract, stearic acid, soy lecithin, ethanol, poloxamer 188, deionized water, and mannitol solution is (1 - 2):(5 - 6):(1.6 - 2):(36 - 43):(1.3 - 1.5):(45 - 55):(45.8 - 56).

[0030] Further, in step A3, the dosage ratio of the black ant extract lipid nanoparticles, lemon peel extract, sesbania leaf extract, silk fibroin, and ethanol is (1.5 - 2.5):(0.4 - 0.6):(0.2 - 0.4):(0.4 - 0.6):(63.1 - 94.7).

[0031] Further, in step A3, the dosage ratio of β - cyclodextrin, arabic gum, ethanol, and deionized water is (4 - 6):(2 - 3):(11.8 - 19.7):(35 - 45).

[0032] Further, in step A4, the dosage ratio of the black ant particles, carboxyl - polyethylene glycol - carboxyl, ethanol, and cyclic peptide RGD - polyethylene glycol - carboxyl is (2 - 3):(1 - 1.2):(71 - 86.8):(0.1 - 0.2).

[0033] The present invention has the following beneficial effects:

[0034] (1) In the technical solution of the present invention, the black ant extract, stearic acid, and soy lecithin are mixed to form an oil phase, and poloxamer 188 is used as the water phase. The oil phase and the water phase are mixed and reacted to form black ant extract lipid nanoparticles. On the one hand, the functional groups of stearic acid, soy lecithin, and the black ant extract interact through hydrogen bonds to form stable lipid nanoparticles, improving the stability of the black ant extract and avoiding the functional groups such as polysaccharides and amino acids in the black ant extract from easily reacting with oxygen or moisture in the environment, resulting in a decrease in the activity of the black ant extract and affecting the treatment effect of rheumatoid arthritis. On the other hand, the drug for treating rheumatoid arthritis, the black ant extract, is coated in the lipid nanoparticles and can be targeted and delivered to the synovial fluid at the arthritis site, having an efficient effect on treating rheumatoid arthritis. In addition, the black ant extract has anti - inflammatory, sedative, analgesic, antibacterial and other effects, and can effectively relieve the symptoms of rheumatoid arthritis.

[0035] (2) In the technical solution of the present invention, the lipid nanoparticles of black ant extract, lemon peel extract, sesbania leaf extract and silk fibroin are mixed as the core material, and β-cyclodextrin and arabic gum are used as the wall material to prepare microcapsules, namely black ant particles. On the one hand, in terms of improving the stability of black ant extract, it has a good synergistic effect with the lipid nanoparticles of black ant extract, and the microcapsules have a certain sustained-release effect, which can achieve the sustained-release effect of the coated substance and increase the effect of black ant extract in treating rheumatoid arthritis. On the other hand, the hydrophilic groups contained in poloxamer 188 on the surface of the lipid nanoparticles can bind to silk fibroin through hydrogen bonds, and the polyphenols contained in lemon peel extract and sesbania leaf extract can also bind to silk fibroin through hydrogen bonds to form a cross-linked network structure, further enhancing the stability of the lipid nanoparticles of black ant extract, lemon peel extract and sesbania leaf extract and improving the treatment effect on rheumatoid arthritis.

[0036] (3) In the technical solution of the present invention, the lemon peel extract contains components such as pyrogallol, caffeic acid, eugenol, p-coumaric acid, p-hydroxybenzoic acid, resorcinol, salicylic acid, luteolin, quercetin, chrysin and luteolin-3-methoxy-7-rutinoside, and has the effects of free radical scavenging activity, antioxidant, antiviral, analgesic and antibacterial activities, etc. It can significantly reduce the levels of chondrocyte proliferation, reactive oxygen species, inflammatory cytokines and xanthine oxidase, and effectively relieve the symptoms of rheumatoid arthritis; the sesbania leaf extract contains a large number of phenolic compounds, has antioxidant, anti-inflammatory, immunomodulatory and cartilage protection effects, and has a synergistic effect with lemon peel extract in the treatment of rheumatoid arthritis as an extract with high anti-inflammatory effect.

[0037] (4) In the technical solution of the present invention, the cyclic peptide RGD-polyethylene glycol-carboxyl is grafted on the surface of the black ant particles through carboxyl-polyethylene glycol-carboxyl. On the one hand, carboxyl-polyethylene glycol-carboxyl serves as a bridge to graft the cyclic peptide RGD-polyethylene glycol-carboxyl more firmly on the surface of the microcapsules, enabling the microcapsules to have the function of targeted delivery to the arthritis site and improving the treatment effect on rheumatoid arthritis. On the other hand, at the site of rheumatoid arthritis inflammation, the neovascular endothelial cells highly express αvβ3 integrin, and the cyclic peptide RGD, as a specific ligand of αvβ3 integrin, enables the cyclic peptide RGD to actively target and bind to αvβ3 integrin, guiding the microcapsules to accumulate in the inflammation site and improving the treatment effect on rheumatoid arthritis. In addition, carboxyl-polyethylene glycol-carboxyl and cyclic peptide RGD-polyethylene glycol-carboxyl are coated on the surface of the microcapsules to form a protective layer on the surface of the microcapsules, delaying their recognition, interaction with opsonins and further absorption by the reticuloendothelial system, thereby prolonging the circulation time of the microcapsule particles in the body fluid until they reach the synovial tissue at the inflammation site and improving the treatment effect on rheumatoid arthritis.

[0038] (5) In the technical scheme of the present invention, the modified black ant granules are compounded with Chinese medicinal materials such as Caulis Agrimoniae, Caulis Piperis, Herba Polygoni Multiflori, and Agkistrodon acutus, which have the effects of anti-inflammatory, immunomodulatory, antioxidant, blood circulation and collateral activating, and inhibiting synovial hyperplasia. The prepared compound ant active capsules can effectively relieve the symptoms of rheumatoid arthritis, avoiding the problem that a single drug treatment is difficult to achieve the expected effect, resulting in poor overall efficacy. DETAILED DESCRIPTION

[0039] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0040] The raw materials used in the examples of the present invention are as follows, and all reagents used are of analytical grade.

[0041] Among them, black ants were purchased from Bozhou Renyi Chinese Medicine Sales Co., Ltd., Qingfengteng was purchased from Bozhou Huozhengtang Pharmaceutical Co., Ltd., Haifengteng was purchased from Bozhou Yongzheng Pharmaceutical Co., Ltd., Ligusticum chuanxiong was purchased from Bozhou Ru'antang Pharmaceutical Co., Ltd., and White Flower Snake was purchased from Bozhou Berlintang Pharmaceutical Co., Ltd.

[0042] The hollow capsule model was 1#, purchased from Shaoxing Kangke Capsule Co., Ltd.

[0043] Starch was corn starch purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0044] Silk fibroin product number xyh001 was purchased from Hubei Xinyuhong Biopharmaceutical Technology Co., Ltd.

[0045] Gum Arabic, food grade, was purchased from Tianjin Fengchuan Chemical Reagent Technology Co., Ltd.;

[0046] Carboxyl-polyethylene glycol-carboxyl (catalog number: P004001) was purchased from Shanghai Tuoyang Biotechnology Co., Ltd.

[0047] Poloxamer 188 model: CP2020, purchased from Shanxi Jinyang Pharmaceutical Excipients Co., Ltd.

[0048] Cyclic peptide RGD-polyethylene glycol-carboxyl, molecular weight 3400, was purchased from Chongqing Yusi Pharmaceutical Technology Co., Ltd.

[0049] The lemon peel extract with a particle size of 100 mesh was purchased from Lanzhou Waterless Biotechnology Co., Ltd.

[0050] The sesbania odorata leaf extract is specifically prepared by the following steps:

[0051] Take the leaves of Sesbania cannabina plants, dry them in an oven at 60 °C for 10 min, grind them into Sesbania cannabina leaf powder with a particle size of 1 mm, add 10 g of Sesbania cannabina leaf powder to 78.9 g of ethanol, heat up to 60 °C, stir evenly, perform ultrasonic extraction at 50 KHz for 2 h, and freeze-dry at -20 °C for 12 h to obtain Sesbania cannabina leaf extract.

[0052] Example 1

[0053] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0054] S1. Mix Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus, crush them into 25-mesh coarse powder, after aging and sterilizing the coarse powder, crush it into 100-mesh fine powder to obtain a compound;

[0055] S2. Mix the modified black ant granules, the compound, starch, and ethanol to make granules with a particle size of 0.1 mm, dry the granules at 60 °C for 20 min, and fill them into hollow capsules to obtain the compound ant activating collaterals capsule;

[0056] Among them, in step S1, the mass ratio of Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus is 8:5:7:10.

[0057] In step S1, the aging and sterilization temperature is 100 °C, and the aging and sterilization time is 10 min.

[0058] In step S2, the mass ratio of the modified black ant granules, the compound, starch, and ethanol is 80:30:40:20.

[0059] The modified black ant granules are specifically prepared by the following steps:

[0060] A1. Crush black ants into 80-mesh powder, sterilize it at 100 °C for 10 min to obtain sterilized black ant powder, add 8 g of sterilized black ant powder to 43.4 g of ethanol, extract at 75 °C for 3 h, filter, collect extract a and residue a, add 4 g of residue a to 10.3 g of ethanol, continue to extract for 2 h, filter to obtain extract b and residue b, add 2 g of residue b to 4 g of ethanol, continue to extract for 1 h, filter to obtain extract c, mix extract a, extract b, and extract c, and perform vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract;

[0061] A2. Add 1 g of black ant extract, 5 g of stearic acid, and 1.6 g of soy lecithin to 36 g of ethanol, stir at 55 °C for 20 min to obtain an oil phase. Add 1.3 g of poloxamer 188 to 45 g of deionized water, stir at 55 °C for 20 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 55 °C and 1800 r / min for 10 min, stir at 1 °C for 3 h to form particles, centrifuge at 17000 r / min to collect the particles, wash twice with deionized water, add to 45.8 g of a 5% mannitol solution, stir evenly, filter through a 0.45 μm membrane, and freeze-dry at -20 °C for 12 h to obtain black ant extract lipid nanoparticles;

[0062] A3. Add 1.5 g of black ant extract lipid nanoparticles, 0.4 g of lemon peel extract, 0.2 g of sesbania leaf extract, and 0.4 g of silk fibroin to 63.1 g of ethanol, stir at 1800 r / min for 10 min to obtain a core material. Add 4 g of β-cyclodextrin and 2 g of arabic gum to 11.8 g of ethanol and 35 g of deionized water, stir evenly, add the core material, stir and react at 55 °C and 800 r / min for 4 h, filter through a vacuum pump to obtain a solid, and freeze-dry the solid at -15 °C for 20 h to obtain microcapsules, namely black ant particles;

[0063] A4. Add 2 g of black ant particles and 1 g of carboxyl-polyethylene glycol-carboxyl to 71 g of ethanol, stir and mix at 60 °C for 20 min, cool to 20 °C, add 0.1 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 10 h, and centrifuge at 10000 r / min to collect the particles to obtain modified black ant particles.

[0064] Example 2

[0065] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0066] S1. Mix Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus, crush them into a 28-mesh coarse powder. After the coarse powder is cooked and sterilized, crush it into a 110-mesh fine powder to obtain a composite;

[0067] S2. Mix the modified black ant particles, the composite, starch, and ethanol to make particles with a diameter of 0.15 mm. Dry the particles at 60 °C for 20 min, and fill them into hollow capsules to obtain a compound ant activating collaterals capsule;

[0068] Among them, in step S1, the mass ratio of Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus is 9:6:8:11.

[0069] In step S1, the cooking and sterilization temperature is 105 °C, and the cooking and sterilization time is 20 min.

[0070] In step S2, the mass ratio of the modified black ant particles, the complex, the starch and the ethanol is 90:35:45:23.

[0071] The modified black ant particles are specifically prepared by the following steps:

[0072] A1. Crush the black ants into 100-mesh powder, sterilize it at 105 °C for 20 min to obtain the sterilized black ant powder. Add 10 g of the sterilized black ant powder to 47.3 g of ethanol, extract at 80 °C for 4 h, filter, collect the extract a and the residue a. Add 5 g of the residue a to 11.8 g of ethanol, continue to extract for 3 h, filter to obtain the extract b and the residue b. Add 3 g of the residue b to 4.7 g of ethanol, continue to extract for 2 h, filter to obtain the extract c. Mix the extract a, the extract b and the extract c, and carry out vacuum freeze-drying at -20 °C for 12 h to obtain the black ant extract;

[0073] A2. Add 1.5 g of the black ant extract, 5.6 g of stearic acid and 1.8 g of soy lecithin to 39.5 g of ethanol, stir at 60 °C for 25 min to obtain the oil phase. Add 1.4 g of poloxamer 188 to 50 g of deionized water, stir at 60 °C for 25 min to obtain the water phase. Add the oil phase to the water phase, stir at 60 °C and 2000 r / min for 15 min, stir at 2 °C for 4 h to form particles, centrifuge at 17000 r / min to collect the particles, wash twice with deionized water, add to 50.9 g of a 5% mannitol solution, stir evenly, filter through a 0.45-μm membrane, and carry out freeze-drying at -20 °C for 12 h to obtain the black ant extract lipid nanoparticles;

[0074] A3. Add 2 g of the black ant extract lipid nanoparticles, 0.5 g of lemon peel extract, 0.3 g of sesbania leaf extract and 0.5 g of silk fibroin to 78.9 g of ethanol, stir at 2000 r / min for 15 min to obtain the core material. Add 5 g of β-cyclodextrin and 2.5 g of arabic gum to 15.8 g of ethanol and 40 g of deionized water, stir evenly, add the core material, stir and react at 60 °C and 1000 r / min for 5 h, filter with a vacuum pump to obtain a solid, and carry out freeze-drying of the solid at -15 °C for 20 h to obtain the microcapsules, that is, the black ant particles;

[0075] A4. Add 2.5 g of the black ant particles and 1.1 g of carboxyl-polyethylene glycol-carboxyl to 78.9 g of ethanol, stir and mix at 65 °C for 25 min, cool to 25 °C, add 0.15 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 11 h, centrifuge at 10000 r / min to collect the particles to obtain the modified black ant particles.

[0076] Example 3

[0077] A preparation method of compound ant and wind - activating capsule, comprising the following preparation steps:

[0078] S1. Mix caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus, and pulverize them into a 30 - mesh coarse powder. After the coarse powder is cooked and sterilized, it is further pulverized into a 120 - mesh fine powder to obtain a composite;

[0079] S2. Mix modified black ant granules, the composite, starch, and ethanol to make granules with a diameter of 0.2 mm. The granules are dried at 60 °C for 20 min and filled into hollow capsules to obtain compound ant and wind - activating capsules;

[0080] Among them, in step S1, the mass ratio of caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus is 10:7:9:12.

[0081] In step S1, the cooking and sterilization temperature is 110 °C, and the cooking and sterilization time is 30 min.

[0082] In step S2, the mass ratio of the modified black ant granules, the composite, starch, and ethanol is 100:40:50:25.

[0083] The modified black ant granules are specifically prepared by the following steps:

[0084] A1. Pulverize black ants into 110 - mesh powder, sterilize it at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b, and extract c, and perform vacuum freeze - drying at - 20 °C for 12 h to obtain black ant extract;

[0085] A2. Add 2 g of black ant extract, 6 g of stearic acid, and 2 g of soybean lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash them 2 times with deionized water, add them to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45 - μm membrane, and perform freeze - drying at - 20 °C for 12 h to obtain black ant extract - loaded lipid nanoparticles;

[0086] A3. Add 2.5 g of lipid nanoparticles of black ant extract, 0.6 g of lemon peel extract, 0.4 g of sesbania leaf extract, and 0.6 g of silk fibroin into 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain the core material. Add 6 g of β-cyclodextrin and 3 g of gum arabic into 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, and stir and react at 65 °C and 1200 r / min for 6 h. Filter through a vacuum pump to obtain a solid, and freeze-dry the solid at -15 °C for 20 h to obtain microcapsules, namely black ant particles;

[0087] A4. Add 3 g of black ant particles and 1.2 g of carboxyl-polyethylene glycol-carboxyl into 86.8 g of ethanol, stir at 70 °C for 30 min, cool to 30 °C, add 0.2 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 12 h, and collect the particles by centrifugation at 10000 r / min to obtain modified black ant particles.

[0088] Comparative Example 1

[0089] A preparation method of a compound ant activating collaterals capsule comprises the following preparation steps:

[0090] S1. Mix Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus, crush them into a 30-mesh coarse powder. After the coarse powder is cooked and sterilized, crush it into a 120-mesh fine powder to obtain a complex;

[0091] S2. Mix the modified black ant particles, the complex, starch, and ethanol to make particles with a size of 0.2 mm. Dry the particles at 60 °C for 20 min, and fill them into hollow capsules to obtain the compound ant activating collaterals capsule;

[0092] Among them, in step S1, the mass ratio of Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus is 10:7:9:12.

[0093] In step S1, the cooking and sterilization temperature is 110 °C, and the cooking and sterilization time is 30 min.

[0094] In step S2, the mass ratio of the modified black ant particles, the complex, starch, and ethanol is 100:40:50:25.

[0095] The modified black ant particles are specifically prepared by the following steps:

[0096] A1. Crush black ants into 110-mesh powder, sterilize it at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b and extract c, and carry out vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract;

[0097] A2. Add 2.5 g of black ant extract, 0.6 g of lemon peel extract, 0.4 g of sesbania leaf extract and 0.6 g of silk fibroin to 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain the core material. Add 6 g of β-cyclodextrin and 3 g of arabic gum to 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, stir and react at 65 °C and 1200 r / min for 6 h, and filter with a vacuum pump to obtain a solid. The solid is freeze-dried at -15 °C for 20 h to obtain microcapsules, namely black ant particles;

[0098] A3. Add 3 g of black ant particles and 1.2 g of carboxyl-polyethylene glycol-carboxyl to 86.8 g of ethanol, stir at 70 °C for 30 min, cool to 30 °C, add 0.2 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 12 h, and collect the particles by centrifugation at 10000 r / min to obtain modified black ant particles.

[0099] Comparative Example 2

[0100] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0101] S1. Mix Sinomenium acutum, Piper kadsura, Speranskia tuberculata and Agkistrodon acutus, crush them into 30-mesh coarse powder. After the coarse powder is cooked and sterilized, it is further crushed into 120-mesh fine powder to obtain a compound;

[0102] S2. Mix the modified black ant particles, the compound, starch and ethanol to make 0.2-mm particles. The particles are dried at 60 °C for 20 min and filled into hollow capsules to obtain the compound ant activating collaterals capsule;

[0103] Among them, in step S1, the mass ratio of Sinomenium acutum, Piper kadsura, Speranskia tuberculata and Agkistrodon acutus is 10:7:9:12.

[0104] In step S1, the cooking and sterilization temperature is 110 °C and the cooking and sterilization time is 30 min.

[0105] In step S2, the mass ratio of the modified black ant particles, the complex, the starch, and the ethanol is 100:40:50:25.

[0106] The modified black ant particles are specifically prepared by the following steps:

[0107] A1. Crush black ants into 110-mesh powder, sterilize it at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b, and extract c, and carry out vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract;

[0108] A2. Add 2 g of black ant extract, 6 g of stearic acid, and 2 g of soy lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash them twice with deionized water, add them to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45-μm membrane, and carry out freeze-drying at -20 °C for 12 h to obtain black ant extract lipid nanoparticles;

[0109] A3. Add 2.5 g of black ant extract lipid nanoparticles, 0.4 g of sesbania leaf extract, and 0.6 g of silk fibroin to 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain a core material. Add 6 g of β-cyclodextrin and 3 g of arabic gum to 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, stir and react at 65 °C and 1200 r / min for 6 h, filter with a vacuum pump to obtain a solid, and carry out freeze-drying of the solid at -15 °C for 20 h to obtain microcapsules, that is, black ant particles;

[0110] A4. Add 3 g of black ant particles and 1.2 g of carboxyl-polyethylene glycol-carboxyl to 86.8 g of ethanol, stir at 70 °C for 30 min, cool to 30 °C, add 0.2 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 12 h, centrifuge at 10000 r / min to collect the particles to obtain modified black ant particles.

[0111] Comparative Example 3

[0112] A preparation method of a compound ant and activating collaterals capsule, comprising the following preparation steps:

[0113] S1. Mix caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus, and pulverize them into a 30-mesh coarse powder. After the coarse powder is cooked and sterilized, it is pulverized into a 120-mesh fine powder to obtain a composite;

[0114] S2. Mix the modified black ant granules, the composite, starch, and ethanol to make granules with a diameter of 0.2 mm. The granules are dried at 60 °C for 20 min and filled into hollow capsules to obtain the compound ant and activating collaterals capsule;

[0115] Among them, in step S1, the mass ratio of caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus is 10:7:9:12.

[0116] In step S1, the cooking and sterilization temperature is 110 °C, and the cooking and sterilization time is 30 min.

[0117] In step S2, the mass ratio of the modified black ant granules, the composite, starch, and ethanol is 100:40:50:25.

[0118] The modified black ant granules are specifically prepared by the following steps:

[0119] A1. Pulverize black ants into a 110-mesh powder, sterilize it at 110 °C for 30 min to obtain the sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b, and extract c, and perform vacuum freeze-drying at -20 °C for 12 h to obtain the black ant extract;

[0120] A2. Add 2 g of the black ant extract, 6 g of stearic acid, and 2 g of soybean lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash them twice with deionized water, add them to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45-μm membrane, and perform freeze-drying at -20 °C for 12 h to obtain the black ant extract lipid nanoparticles;

[0121] A3. Add 2.5 g of black ant extract lipid nanoparticles, 0.6 g of lemon peel extract, and 0.6 g of silk fibroin into 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain the core material. Add 6 g of β-cyclodextrin and 3 g of gum arabic into 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, and stir and react at 65 °C and 1200 r / min for 6 h. Filter through a vacuum pump to obtain a solid, and freeze-dry the solid at -15 °C for 20 h to obtain microcapsules, namely black ant particles;

[0122] A4. Add 3 g of black ant particles and 1.2 g of carboxyl-polyethylene glycol-carboxyl into 86.8 g of ethanol, stir at 70 °C for 30 min, cool to 30 °C, add 0.2 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 12 h, and collect the particles by centrifugation at 10000 r / min to obtain modified black ant particles.

[0123] Comparative Example 4

[0124] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0125] S1. Mix caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus, crush them into a 30-mesh coarse powder. After aging and sterilizing the coarse powder, crush it into a 120-mesh fine powder to obtain a composite;

[0126] S2. Mix the modified black ant particles, the composite, starch, and ethanol to make particles with a size of 0.2 mm. Dry the particles at 60 °C for 20 min, and fill them into hollow capsules to obtain the compound ant activating collaterals capsule;

[0127] Among them, in step S1, the mass ratio of caulis sinomenii, caulis piperis kadsurae, herba speranskiae tuberculatae, and agkistrodon acutus is 10:7:9:12.

[0128] In step S1, the aging and sterilizing temperature is 110 °C, and the aging and sterilizing time is 30 min.

[0129] In step S2, the mass ratio of the modified black ant particles, the composite, starch, and ethanol is 100:40:50:25.

[0130] The modified black ant particles are specifically prepared by the following steps:

[0131] A1. Crush black ants into 110-mesh powder, sterilize it at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b and extract c, and conduct vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract;

[0132] A2. Add 2 g of black ant extract, 6 g of stearic acid and 2 g of soy lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash twice with deionized water, add to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45-μm membrane, and conduct freeze-drying at -20 °C for 12 h to obtain black ant extract lipid nanoparticles;

[0133] A3. Add 2.5 g of black ant extract lipid nanoparticles, 0.6 g of lemon peel extract and 0.4 g of sesbania leaf extract to 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain a core material. Add 6 g of β-cyclodextrin and 3 g of arabic gum to 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, stir and react at 65 °C and 1200 r / min for 6 h, conduct vacuum filtration with a vacuum pump to obtain a solid, and conduct freeze-drying of the solid at -15 °C for 20 h to obtain a microcapsule, namely black ant particles;

[0134] A4. Add 3 g of black ant particles and 1.2 g of carboxyl-polyethylene glycol-carboxyl to 86.8 g of ethanol, stir at 70 °C for 30 min, cool to 30 °C, add 0.2 g of cyclic peptide RGD-polyethylene glycol-carboxyl, continue to stir for 12 h, centrifuge at 10000 r / min to collect the particles to obtain modified black ant particles.

[0135] Control Example 5

[0136] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0137] S1. Mix sinomenium acutum, piper kadsura, herba speranskiae tuberculatae and agkistrodon acutus, crush them into 30-mesh coarse powder, after the coarse powder is cooked and sterilized, crush it into 120-mesh fine powder to obtain a complex;

[0138] S2. Mix the modified black ant granules, the complex, starch and ethanol to make granules with a size of 0.2 mm. Dry the granules at 60 °C for 20 min, and then fill them into hollow capsules to obtain the compound ant and lumbricus capsules.

[0139] Among them, in step S1, the mass ratio of sinomenium acutum, caulis piperis kadsurae, herba speranskiae tuberculatae and agkistrodon acutus is 10:7:9:12.

[0140] In step S1, the ripening and sterilization temperature is 110 °C and the ripening and sterilization time is 30 min.

[0141] In step S2, the mass ratio of the modified black ant granules, the complex, starch and ethanol is 100:40:50:25.

[0142] The modified black ant granules are specifically prepared by the following steps:

[0143] A1. Crush black ants into 110-mesh powder, sterilize the powder at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b and extract c, and carry out vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract.

[0144] A2. Add 2 g of black ant extract, 6 g of stearic acid and 2 g of soy lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash them twice with deionized water, add them to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45-μm membrane, and carry out freeze-drying at -20 °C for 12 h to obtain black ant extract lipid nanoparticles.

[0145] A3. Add 2.5 g of black ant extract lipid nanoparticles, 0.6 g of lemon peel extract, 0.4 g of sesbania leaf extract and 0.6 g of silk fibroin to 94.7 g of ethanol, stir at 12200 r / min for 20 min, and carry out freeze-drying at -15 °C for 20 h to obtain black ant granules.

[0146] A4. Add 3 g of black ant particles and 1.2 g of carboxyl-polyethylene glycol-carboxyl to 86.8 g of ethanol, stir at 70 °C for 30 min, cool to 30 °C, add 0.2 g of cyclic peptide RGD-polyethylene glycol-carboxyl, and continue to stir for 12 h. Centrifuge to collect the particles at 10,000 r / min to obtain modified black ant particles.

[0147] Comparative Example 6

[0148] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0149] S1. Mix Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus, crush them into a 30-mesh coarse powder. After the coarse powder is cooked and sterilized, crush it into a 120-mesh fine powder to obtain a complex.

[0150] S2. Mix the modified black ant particles, the complex, starch, and ethanol to make particles with a diameter of 0.2 mm. Dry the particles at 60 °C for 20 min, and fill them into hollow capsules to obtain the compound ant activating collaterals capsule.

[0151] Among them, in step S1, the mass ratio of Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus is 10:7:9:12.

[0152] In step S1, the cooking and sterilization temperature is 110 °C, and the cooking and sterilization time is 30 min.

[0153] In step S2, the mass ratio of the modified black ant particles, the complex, starch, and ethanol is 100:40:50:25.

[0154] The modified black ant particles are specifically prepared by the following steps:

[0155] A1. Crush black ants into a 110-mesh powder, sterilize it at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b, and extract c, and carry out vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract.

[0156] A2. Add 2 g of black ant extract, 6 g of stearic acid, and 2 g of soy lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash twice with deionized water, add to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45 μm membrane, and freeze-dry at -20 °C for 12 h to obtain black ant extract lipid nanoparticles;

[0157] A3. Add 2.5 g of black ant extract lipid nanoparticles, 0.6 g of lemon peel extract, 0.4 g of sesbania leaf extract, and 0.6 g of silk fibroin to 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain a core material. Add 6 g of β-cyclodextrin and 3 g of arabic gum to 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, stir and react at 65 °C and 1200 r / min for 6 h, filter through a vacuum pump to obtain a solid, and freeze-dry the solid at -15 °C for 20 h to obtain microcapsules, namely black ant particles;

[0158] A4. Add 3 g of black ant particles to 86.8 g of ethanol, stir at 30 °C for 12 h, centrifuge at 10000 r / min to collect the particles to obtain modified black ant particles.

[0159] Control Example 7

[0160] A preparation method of a compound ant activating collaterals capsule, comprising the following preparation steps:

[0161] S1. Mix Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus, pulverize them into a 30-mesh coarse powder, after aging and sterilizing the coarse powder, pulverize it into a 120-mesh fine powder to obtain a composite;

[0162] S2. Mix the modified black ant particles, the composite, starch, and ethanol to make 0.2 mm particles, dry the particles at 60 °C for 20 min, and fill them into hollow capsules to obtain the compound ant activating collaterals capsule;

[0163] Among them, in step S1, the mass ratio of Sinomenium acutum, Piper kadsura, Speranskia tuberculata, and Agkistrodon acutus is 10:7:9:12.

[0164] In step S1, the aging and sterilizing temperature is 110 °C, and the aging and sterilizing time is 30 min.

[0165] In step S2, the mass ratio of the modified black ant particles, the composite, starch, and ethanol is 100:40:50:25.

[0166] The modified black ant granules are specifically prepared by the following steps:

[0167] A1. Crush black ants into 110-mesh powder, sterilize at 110 °C for 30 min to obtain sterilized black ant powder. Add 12 g of the sterilized black ant powder to 51.3 g of ethanol, extract at 85 °C for 5 h, filter, collect extract a and residue a. Add 6 g of residue a to 13.4 g of ethanol, continue to extract for 4 h, filter to obtain extract b and residue b. Add 4 g of residue b to 5.5 g of ethanol, continue to extract for 3 h, filter to obtain extract c. Mix extract a, extract b and extract c, and carry out vacuum freeze-drying at -20 °C for 12 h to obtain black ant extract;

[0168] A2. Add 2 g of black ant extract, 6 g of stearic acid and 2 g of soy lecithin to 43 g of ethanol, stir at 65 °C for 30 min to obtain an oil phase. Add 1.5 g of poloxamer 188 to 45 g of deionized water, stir at 65 °C for 30 min to obtain an aqueous phase. Add the oil phase to the aqueous phase, stir at 65 °C and 2200 r / min for 20 min, stir at 3 °C for 5 h to form particles, centrifuge at 17000 r / min to collect the particles, wash twice with deionized water, add to 56 g of a 5% mannitol solution, stir evenly, filter through a 0.45-μm membrane, and carry out freeze-drying at -20 °C for 12 h to obtain black ant extract lipid nanoparticles;

[0169] A3. Add 2.5 g of black ant extract lipid nanoparticles, 0.6 g of lemon peel extract, 0.4 g of sesbania leaf extract and 0.6 g of silk fibroin to 94.7 g of ethanol, stir at 12200 r / min for 20 min to obtain a core material. Add 6 g of β-cyclodextrin and 3 g of arabic gum to 19.7 g of ethanol and 45 g of deionized water, stir evenly, add the core material, stir and react at 65 °C and 1200 r / min for 6 h, filter with a vacuum pump to obtain a solid, and carry out freeze-drying of the solid at -15 °C for 20 h to obtain microcapsules, namely black ant granules;

[0170] A4. Add 3 g of black ant granules and 1.2 g of carboxyl-polyethylene glycol-carboxyl to 86.8 g of ethanol, stir at 70 °C for 30 min, filter to obtain modified black ant granules.

[0171] Now, performance tests are carried out on the compound ant activating collaterals capsules prepared in Examples 1-3 and Comparative Examples 1-7.

[0172] Effect detection of the compound ant activating collaterals capsule in the treatment of rheumatoid arthritis:

[0173] Patients: 70 males and 50 females; Ages: 30 cases aged ≤ 30 years, 41 cases aged 31 - 40 years, 35 cases aged 41 - 50 years, 8 cases aged 51 - 60 years, 6 cases aged over 60 years; Course of disease: 20 cases with a course of less than 1 year, 78 cases with a course of 1 - 5 years, 22 cases with a course of over 5 years;

[0174] Criteria: (1) Western medicine diagnosis conforms to the diagnostic criteria for rheumatoid arthritis in the "Combined Diagnosis and Treatment Guidelines for Rheumatoid Arthritis and Traditional Chinese Medicine Syndromes"; (2) Traditional Chinese medicine diagnosis conforms to the dialectical criteria in the "International Clinical Practice Guidelines for Traditional Chinese Medicine: Rheumatoid Arthritis", syndrome type: cold-dampness obstruction syndrome; Main symptoms: joint pain, aggravated by cold, relieved by heat, not warm to the touch, skin color not red; Secondary symptoms: joint contracture, limited flexion and extension, afraid of cold and preferring warmth, cold limbs and numbness; Tongue and pulse: pale tongue, enlarged tongue body, white or greasy tongue coating, string-taut and rapid pulse.

[0175] Treatment: 2 capsules each time, three times a day, orally, and all patients were treated continuously for 4 weeks;

[0176] Detection of patients' physical symptoms: Record the average value after 4 weeks of treatment. Patients' joint swelling index (no swelling is grade 0; mild swelling with skin texture becoming shallow is grade 1; moderate swelling with skin texture disappearing is grade 2; severe swelling with skin being tight is grade 3), joint tenderness index (no tenderness is grade 0; mild tenderness is grade 1; moderate tenderness is grade 2; severe tenderness, pain is unbearable to touch is grade 3); Before treatment, the patients' joint swelling index was 2.35 ± 0.56 grades, and the joint tenderness index was 2.38 ± 0.33 grades.

[0177] Detection of patients' bone and joint metabolite levels: Record the average value after 4 weeks of treatment. Collect 5 mL of venous blood from the patient's elbow, centrifuge at 3000 r / min for 5 min, take the upper serum, and detect the serum bone morphogenetic protein (BMPs) level by enzyme-linked immunosorbent assay, and detect the levels of serum matrix metalloproteinase-1 (MMP-1) and matrix metalloproteinase-3 (MMP-3) by electrochemiluminescence method; Before treatment, the patients' BMPs was 8.21 ± 1.51 pg / mL, MMP-1 was 22.4 ± 2.18 pg / mL, and MMP-3 was 63.26 ± 7.75 pg / mL.

[0178] The test results are shown in Tables 1 and 2 below.

[0179] Table 1 Performance Detection of Compound Ant Ant-Activating Collagen Capsules Prepared in Examples 1 - 3 and Comparative Examples 1 - 7

[0180]

[0181] Table 2 Performance Detection of Compound Ant Ant-Activating Collagen Capsules Prepared in Examples 1 - 3 and Comparative Examples 1 - 7

[0182]

[0183] As can be seen from the data in Table 1 and Table 2, the compound ant and activating collaterals capsules prepared in Examples 1-3 have significant therapeutic effects on rheumatoid arthritis.

[0184] In Comparative Example 1, the modified black ant particles prepared by replacing the lipid nanoparticles of black ant extract with black ant extract were added to the compound ant and activating collaterals capsules, and the therapeutic effect decreased. This proves that the black ant extract forms lipid nanoparticles, which improves the stability of the black ant extract, avoids the functional groups such as polysaccharides and amino acids in the black ant extract, and is easy to react with oxygen or moisture in the environment, resulting in a decrease in the activity of the black ant extract and affecting the therapeutic effect of rheumatoid arthritis. Moreover, the formed lipid nanoparticles can be targeted and delivered to the synovial fluid at the arthritis site, having an efficient therapeutic effect on rheumatoid arthritis.

[0185] In Comparative Example 2, the modified black ant particles prepared without adding lemon peel extract were added to the compound ant and activating collaterals capsules, and the therapeutic effect decreased. This proves that lemon peel extract contains components such as pyrogallol, caffeic acid, eugenol, p-coumaric acid, p-hydroxybenzoic acid, resorcinol, salicylic acid, luteolin, quercetin, chrysin and luteolin-3-methoxy-7-rutinoside, which significantly reduce the levels of chondrocyte proliferation, reactive oxygen species, inflammatory cytokines and xanthine oxidase, and effectively relieve the symptoms of rheumatoid joints.

[0186] In Comparative Example 3, the modified black ant particles prepared without adding sesbania leaf extract were added to the compound ant and activating collaterals capsules, and the therapeutic effect decreased. This proves that sesbania leaf extract contains a large number of phenolic compounds, having antioxidant, anti-inflammatory, immunomodulatory and cartilage protection effects. As an extract with high anti-inflammatory effect, it has a synergistic effect with lemon peel extract in the treatment of rheumatoid arthritis.

[0187] In Comparative Example 4, the modified black ant particles prepared without adding silk fibroin were added to the compound ant and activating collaterals capsules, and the therapeutic effect decreased. This proves that the hydrophilic groups contained in poloxamer 188 on the surface of the lipid nanoparticles can bind to silk fibroin through hydrogen bonds, and the polyphenols contained in lemon peel extract and sesbania leaf extract can also bind to silk fibroin through hydrogen bonds, forming a cross-linked network structure, further enhancing the stability of the black ant extract lipid nanoparticles, lemon peel extract and sesbania leaf extract, and improving the therapeutic effect on rheumatoid arthritis.

[0188] The modified black ant granules prepared without adding β-cyclodextrin and arabic gum in Comparative Example 5 were added to the Compound Black Ant Huoluo Capsules, and the therapeutic effect decreased. This proved that β-cyclodextrin and arabic gum, as capsule materials, coated the lipid nanoparticles of black ant extract, lemon peel extract, sesbania leaf extract and silk fibroin to form microcapsules, that is, black ant granules, which improved the stability of black ant extract. Moreover, the microcapsules had a certain sustained-release effect, which could achieve the sustained-release effect on the coated substances and increase the therapeutic effect of black ant extract on rheumatoid arthritis.

[0189] The modified black ant granules prepared without adding carboxyl-polyethylene glycol-carboxyl in Comparative Example 6 were added to the Compound Black Ant Huoluo Capsules, and the therapeutic effect decreased. This proved that carboxyl-polyethylene glycol-carboxyl, as a bridge, could graft cyclic peptide RGD-polyethylene glycol-carboxyl on the surface of the microcapsules, enabling the microcapsules to have the function of targeted delivery to the arthritis site, improving the therapeutic effect on rheumatoid arthritis. Moreover, carboxyl-polyethylene glycol-carboxyl and cyclic peptide RGD-polyethylene glycol-carboxyl coated on the surface of the microcapsules formed a protective layer on the surface of the microcapsules, delaying their recognition, interaction with opsonin and further absorption by the reticuloendothelial system, thereby prolonging the circulation time of the microcapsule particles in the body fluid until they reached the synovial tissue at the inflammatory site, improving the therapeutic effect on rheumatoid arthritis.

[0190] The modified black ant granules prepared without adding cyclic peptide RGD-polyethylene glycol-carboxyl in Comparative Example 7 were added to the Compound Black Ant Huoluo Capsules, and the therapeutic effect decreased. This proved that neovascular endothelial cells highly expressed αvβ3 integrin, and cyclic peptide RGD, as a specific ligand of αvβ3 integrin, enabled cyclic peptide RGD to actively target and bind to αvβ3 integrin, guiding the microcapsules to accumulate in the inflammatory site and improving the therapeutic effect on rheumatoid arthritis.

[0191] In the description of the specification, the descriptions referring to terms such as "one embodiment", "example", "specific example", etc. mean that the specific features, structures, materials or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in a suitable manner in any one or more embodiments or examples.

[0192] The above content is only an example and illustration of the present invention. Those skilled in the art of this technology can make various modifications or supplements to the specific embodiments described or use similar methods to substitute, as long as they do not deviate from the invention or exceed the scope defined by the claims of the present invention, they should all fall within the protection scope of the present invention.

Claims

1. A method for preparing a compound ant active capsule, characterized in that: The method comprises the following preparation steps: S1. The green wind vine, sea wind vine, bone grass, white flower snake mixed, crushed into 25-30 mesh coarse powder, the coarse powder after maturation and sterilization, and then crushed into 100-120 mesh fine powder to obtain a composite; S2. The modified black ant particles, the complex, starch and ethanol were mixed to form particles of 0.1-0.2 mm, and the particles were dried and filled into hollow capsules to obtain compound ant active capsules; The modified black ant particles are obtained by a mixed reaction of black ant particles, carboxyl-polyethylene glycol-carboxyl and cyclic peptide RGD-polyethylene glycol-carboxyl; The black ant particles are prepared by mixing black ant extract lipid nanoparticles, lemon peel extract, sesbania leaf extract and silk fibroin as core materials, and β-cyclodextrin and gum arabic as capsule materials; The black ant extract lipid nanoparticles are obtained by mixing black ant extract, stearic acid and soybean lecithin to form an oil phase, using poloxamer 188 as a water phase, and reacting the oil phase with the water phase.

2. The method for preparing the compound ant active capsule according to claim 1, characterized in that: The modified black ant particles are specifically prepared by the following steps: A1. The black ants were crushed into 80-110 mesh powder, sterilized, extracted three times with ethanol, the extraction temperature was 75-85 ° C, extract a, extract b and extract c were mixed, freeze-dried to obtain a black ant extract; A2. Add black ant extract, stearic acid and soybean lecithin to ethanol, stir evenly to obtain an oil phase, add poloxamer 188 to deionized water, stir evenly to obtain an aqueous phase, add the oil phase to the aqueous phase, stir evenly, stir at 1-3°C for 3-5h, centrifuge, wash, add to a mannitol solution, stir, filter, and lyophilize to obtain black ant extract lipid nanoparticles; A3. Add black ant extract lipid nanoparticles, lemon peel extract, sesbania leaf extract and silk fibroin to ethanol, stir evenly to obtain a core material, add β-cyclodextrin and gum arabic to ethanol and deionized water, stir evenly, add the core material, stir and react at 55-65° C. for 4-6 hours, filter with a vacuum pump to obtain a solid, freeze-dry the solid to obtain black ant particles; A4. Add black ant particles and carboxyl-polyethylene glycol-carboxyl to ethanol, stir at 60-70°C for 20-30 minutes, cool to 20-30°C, add cyclic peptide RGD-polyethylene glycol-carboxyl, continue stirring for 10-12 hours, collect particles by centrifugation, and obtain modified black ant particles.

3. The method for preparing the compound ant active capsule according to claim 2, characterized in that: In step A1, the sterilization temperature is 100-110°C, and the sterilization time is 10-30 minutes.

4. The method for preparing the compound ant active capsule according to claim 2, characterized in that: In step A2, the black ant extract, stearic acid, soy lecithin, ethanol, poloxamer 188, deionized water and mannitol solution are used in a ratio of (1-2):(5-6):(1.6-2):(36-43):(1.3-1.5):(45-55):(45.8-56).

5. The method for preparing the compound ant active capsule according to claim 2, characterized in that: In step A3, the ratio of the black ant extract lipid nanoparticles, lemon peel extract, sesbania leaf extract, silk fibroin and ethanol is (1.5-2.5):(0.4-0.6):(0.2-0.4):(0.4-0.6):(63.1-94.7); The dosage ratio of the β-cyclodextrin, gum arabic, ethanol and deionized water is (4-6):(2-3):(11.8-19.7):(35-45).

6. The method for preparing the compound ant active capsule according to claim 2, characterized in that: In step A4, the ratio of the black ant particles, carboxyl-polyethylene glycol-carboxyl, ethanol and cyclic peptide RGD-polyethylene glycol-carboxyl is (2-3):(1-1.2):(71-86.8):(0.1-0.2).

7. The method for preparing the compound ant active capsule according to claim 1, characterized in that: In step S1, the mass ratio of Caulis Sinomenii, Caulis Piperis, Herba Polygoni Multiflori and Agkistrodon is (8-10):(5-7):(7-9):(10-12).

8. The method for preparing the compound ant active capsule according to claim 1, characterized in that: In step S2, the mass ratio of the modified black ant particles, the complex, starch and ethanol is (80-100):(30-40):(40-50):(20-25).

9. A compound ant active capsule prepared by the preparation method of the compound ant active capsule according to any one of claims 1 to 8.

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