Preparation method of bone hypertrophy eliminating medicine

Abstract

The invention provides an osteoproliferation eliminating preparation method, and belongs to the technical field of traditional Chinese medicine preparations. The bone hypertrophy eliminating medicine is prepared from the following raw materials: prepared rehmannia root, roasted herba epimedii, caulis spatholobi, rhizoma cibotii, rhizoma drynariae, cynomorium songaricum, radix dipsaci, fructus lycii The method comprises the following steps: heating and mixing choline salt, 1-octyl-3-methylimidazolium bromide and an acid hydrogen bond donor or an alcohol hydrogen bond donor to prepare a multi-element eutectic solvent A or a multi-element eutectic solvent B; the preparation method comprises the following steps: uniformly mixing prepared rehmannia root powder, a multi-element eutectic solvent A and water, and carrying out microwave heating to obtain a prepared rehmannia root extracting solution and prepared rehmannia root solid residues; mixing the dried radix rehmanniae praeparata solid residues, the residual raw material powder, the multi-element eutectic solvent B and an alcoholic solution, and carrying out ultrasonic-assisted extraction for multiple times in a short time to obtain an extracted supernatant; combining the prepared rehmannia root extracting solution and the extracted supernate, and concentrating to obtain an osteoproliferation eliminating stock solution; and preparing the osteoproliferation eliminating preparation. The method can be used for rapidly and efficiently extracting the effective active ingredients of the raw materials of the osteodynia, the conditions are mild, and the consumed time is short.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicine preparations, and particularly relates to a preparation method of bone augmentation and elimination. Background Art

[0002] Guzengxiao is a traditional Chinese medicine preparation used to treat bone hyperplasia such as pain in the waist and knee joints caused by liver and kidney deficiency. Its main ingredients are: Rehmannia root, roasted Epimedium, Millettia reticulata, Cibotium barometz, Drynaria fortunei, Cynomorium songaricum, Dipsacus asper, Lycium barbarum, and Radish seed.

[0003] At present, the traditional preparation process of bone augmentation and consumption includes the following methods: first, all the raw materials are crushed and decocted for multiple times, the filtrate is combined and concentrated, and then made into corresponding dosage forms, such as tablets or capsules; second, dog spine and roasted epimedium are crushed into fine powder; the remaining raw materials are decocted for multiple times, the filtrate is combined and concentrated, and then mixed with the fine powder to make corresponding dosage forms; third, different extraction methods are used to extract each raw material separately to obtain the extract concentrate, and then combined to make corresponding dosage forms.

[0004] Among the above methods, the first and second ones belong to traditional processes, and their processes are simple, but both have defects such as low utilization rate of effective active ingredients in raw materials and large amount of medicinal materials, especially Rehmannia root, which, as the main raw material, contains multiple effective active ingredients such as polysaccharides, saponin compounds, flavonoid compounds, etc.; the third method uses enzymatic hydrolysis, carbon dioxide supercritical extraction and other technical means to extract the active ingredients in medicinal materials respectively, but it has the problems of harsh extraction conditions, long time consumption, high cost, and large amount of waste liquid. Summary of the invention

[0005] In view of this, the object of the present invention is to provide a method for preparing bone augmentation and reduction, aiming to solve at least one technical problem in the background technology.

[0006] The present invention is achieved in that: A method for preparing bone augmentation and reduction, wherein the raw materials of the bone augmentation and reduction include prepared rehmannia root, roasted epimedium, millettia reticulata, cibotium barometz, drynariae, cynomorium songaricum, dipsacus root, wolfberry fruit and radish seed; The preparation method comprises the following steps: The choline salt, 1-octyl-3-methylimidazolium bromide and the acid hydrogen bond donor are heated and mixed according to a preset molar ratio to prepare a multi-element deep eutectic solvent A; The choline salt, 1-octyl-3-methylimidazolium bromide and the alcohol hydrogen bond donor are heated and mixed according to a preset molar ratio to prepare a multi-component deep eutectic solvent B; The solution of multi-element low eutectic solvent A is used as extractant A, and Rehmannia glutinosa powder is treated by microwave in a closed environment to obtain Rehmannia glutinosa extract and Rehmannia glutinosa solid residue; The dried Rehmannia glutinosa solid residue and the remaining raw material powder form a mixed material, and a solution of a multi-component low eutectic solvent B is used as an extractant B, and ultrasonic-assisted extraction is performed to obtain an extraction supernatant; The prepared Rehmannia root extract and the extracted supernatant are combined and concentrated to obtain the bone-increasing and detoxifying stock solution; The bone-increasing and consuming stock solution and adjuvants are used to prepare the bone-increasing and consuming preparation.

[0007] Preferably, the specific steps of obtaining the Rehmannia glutinosa extract and the Rehmannia glutinosa solid residue are: The Rehmannia root powder, multi-element low eutectic solvent A and water are uniformly mixed in a preset ratio, and the Rehmannia root powder is heated and extracted in a closed environment with microwave. After the extraction, it is cooled to room temperature and then centrifuged. After liquid phase distillation and impurity removal, the Rehmannia root extract is obtained, and the solid phase is the Rehmannia root solid residue.

[0008] Preferably, the specific steps of obtaining the extraction supernatant are: The dried Rehmannia glutinosa solid residue is mixed with the remaining raw material powder, and a mixed solution of a multi-component low eutectic solvent B and an alcohol solution is used as an extractant B. After sufficient stirring, ultrasonic-assisted extraction is performed, and then solid-liquid separation is performed. The obtained solid is added with the extractant B, and ultrasonic-assisted extraction and solid-liquid separation are repeated at least once. The filtrates after each solid-liquid separation are combined to obtain an extraction supernatant.

[0009] Preferably, in the multi-component deep eutectic solvent A, according to the molar ratio, choline salt: 1-octyl-3-methylimidazolium bromide: acid hydrogen bond donor = 1:1:2~4; the choline salt is choline chloride or choline bromide; the acid hydrogen bond donor is citric acid or acetic acid; the heating temperature when preparing the multi-component deep eutectic solvent A is 40°C~60°C.

[0010] Preferably, in the multi-component low eutectic solvent B, according to the molar ratio, choline salt: 1-octyl-3-methylimidazolium bromide: alcohol hydrogen bond donor = 1:1:2~4; the choline salt is choline chloride or choline bromide; the alcohol hydrogen bond donor is 1,4-butanediol, 1,2-propylene glycol or triethylene glycol; the heating temperature when preparing the multi-component low eutectic solvent B is 40°C~60°C.

[0011] Preferably, the Rehmannia root powder, the multi-element low eutectic solvent A, and water = 1 g: (10-15) g: (15-25) mL.

[0012] Preferably, the time for heating and extracting the Rehmannia glutinosa powder is 10 min to 20 min; the temperature for heating and extracting is 60° C. to 120° C.; and the power of the microwave is 150w to 250w.

[0013] Preferably, the step of preparing the extraction supernatant specifically comprises: Mix the dried Rehmannia glutinosa solid residue with the remaining raw material powder, and stir evenly to form a mixed powder; The mixed powder and the extractant B are mixed in a preset ratio, fully stirred, and then ultrasonic-assisted extraction is performed, and then solid-liquid separation is performed to obtain a first filtrate and a first solid residue; The first solid residue is dried and mixed with the extractant B according to a preset ratio, fully stirred and subjected to ultrasonic-assisted extraction, and then solid-liquid separation to obtain a second filtrate and a second solid residue; the second solid residue repeats this step to obtain a third filtrate and a third solid residue; The first filtrate, the second filtrate and the third filtrate are combined and centrifuged, and an extraction supernatant is obtained after impurities are removed by distillation; The power of the ultrasonic-assisted extraction is 200w-300w; the temperature is 40°C-60°C; and the time of a single ultrasonic-assisted extraction is 5min-10min.

[0014] Preferably, in a single ultrasonic-assisted extraction process, the mass ratio of solid material: multi-component low eutectic solvent B: alcohol solution is 1:1-2:30-60, and the solid material is a mixed powder, a first solid slag or a second solid slag.

[0015] Preferably, the alcohol solution is a methanol aqueous solution or an ethanol aqueous solution, and the mass concentration of alcohol in the alcohol solution is 40% to 80%.

[0016] Compared with the prior art, the present invention has the following beneficial effects: 1. The present invention adopts a ternary low eutectic solvent made of two different hydrogen bond donors, which can quickly and efficiently extract the effective active ingredients of various raw materials for bone augmentation and absorptivity, with mild conditions and short time consumption.

[0017] 2. The present invention uses choline salt + 1-octyl-3-methylimidazolium bromide + acid hydrogen bond donor to treat Rehmannia glutinosa, and cooperates with microwave treatment, so that multiple effective biologically active ingredients such as polysaccharides and saponin compounds in Rehmannia glutinosa can be fully extracted in a short time; the present invention uses a composite acidic ternary low eutectic solvent as an extractant, and during the mixing process of the composite acidic ternary low eutectic solvent with Rehmannia glutinosa powder, saponin compounds such as Rehmannia glutinosa polysaccharides and rehmannia glycoside D can be quickly dissolved, form hydrogen bonds with the multi-element low eutectic solvent A, and then dissolve in the solvent; at the same time, based on the thermal effect and electric field non-thermal effect generated by microwaves, electromagnetic waves are used to penetrate Rehmannia glutinosa to reach the inside and act on molecules, promote molecular motion to generate a large amount of heat energy, accelerate the molecular motion speed of biologically active ingredients, thereby accelerating their transfer to the solvent, and realizing efficient and rapid precipitation of biologically active ingredients.

[0018] 3. The present invention uses choline salt + 1-octyl-3-methylimidazolium bromide + alcohol hydrogen bond donor to treat Rehmannia glutinosa solid residue and other raw materials, and uses a composite alcohol ternary low eutectic solvent as an extractant. The hydroxyl groups in the alcohol can form intermolecular hydrogen bonds with the biologically active components in the raw materials, thereby increasing the solubility of components such as flavonoids, phenolic compounds, sinapines, betaine, etc.; and synergistic ultrasonic treatment, based on the thermal effect and mechanical effect of ultrasonic waves, enhances the penetration of the multi-component low eutectic solvent B, accelerates the transfer of target components, and thereby improves the extraction efficiency of the target components.

[0019] 4. The present invention is green and environmentally friendly, produces less waste liquid, and has a simple process. DETAILED DESCRIPTION

[0020] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the embodiments. It should be understood that the specific implementation cases described herein are only used to explain the present invention and are not used to limit the present invention.

[0021] A method for preparing bone augmentation and reduction, wherein the raw materials include the following components in parts by weight: 160 to 210 parts of Radix Rehmanniae Preparata; 100 to 150 parts of roasted Epimedium; 100 to 150 parts of Millettia Spatholobi; 100 to 150 parts of Cibotium Cibotium; 100 to 150 parts of Drynariae Rhizoma; 100 to 150 parts of Cynomorium songaricum; 50 to 80 parts of Dipsacus asper; 50 to 80 parts of Lycium barbarum; and 50 to 80 parts of Radish Seed. In specific implementation, the amount of each raw material can be adjusted according to actual conditions and is not specifically limited here.

[0022] The preparation method comprises steps S1 to S5.

[0023] S1. Raw material pretreatment The raw materials of bone augmentation and reduction are crushed and passed through a 80-100 mesh sieve to prepare powder; Weigh the materials according to the molar ratio of choline salt: 1-octyl-3-methylimidazolium bromide: acid hydrogen bond donor = 1:1:2-4, heat the three to 40°C-60°C and mix them to prepare a multi-element deep eutectic solvent A; Weigh the materials according to the molar ratio of choline salt: 1-octyl-3-methylimidazolium bromide: alcohol hydrogen bond donor = 1:1:2-4, heat the three to 40°C-60°C and mix them to prepare a multi-element deep eutectic solvent B; Among them, in the polyvalent deep eutectic solvent A and the polyvalent deep eutectic solvent B, the choline salt is choline chloride or choline bromide, the acid hydrogen bond donor is citric acid or acetic acid, and the alcohol hydrogen bond donor can be a polyol such as 1,4-butanediol, 1,2-propylene glycol or triethylene glycol.

[0024] S2, Rehmannia root powder, multi-element low eutectic solvent A, and water are uniformly mixed in a preset ratio, and the Rehmannia root powder is heated and extracted in a closed environment in coordination with microwave treatment; after the extraction, it is cooled to room temperature and then centrifuged, and the liquid phase is distilled to remove impurities to obtain Rehmannia root extract, and the solid phase is Rehmannia root solid residue; Since Rehmannia glutinosa contains multiple effective active ingredients such as polysaccharides and saponin compounds, single water extraction and alcohol extraction require several hours of processing and cannot effectively extract biologically active ingredients. In the present invention, a composite acidic ternary deep eutectic solvent is used as the extractant A. During the mixing process of the composite acidic ternary deep eutectic solvent with Rehmannia glutinosa powder, biologically active ingredients such as Rehmannia glutinosa polysaccharides and saponin compounds (mainly rehmannoside D) can be dissolved from Rehmannia glutinosa. The biologically active ingredients form hydrogen bonds with the multi-component deep eutectic solvent A and are then dissolved in the multi-component deep eutectic solvent A. In the above process, microwave treatment is used in coordination. Based on the thermal effect and electric field non-thermal effect generated by microwaves, electromagnetic waves are used to penetrate Rehmannia glutinosa and reach the inside and act on the molecules, thereby promoting molecular movement to generate a large amount of heat energy, speeding up the molecular movement speed of the bioactive components, thereby accelerating their transfer to the multi-electrolytic solvent A, and achieving efficient and rapid precipitation of the bioactive components. During the extraction process, the role of adding water as a solvent is to increase the wettability of Rehmannia glutinosa powder and reduce the viscosity of the reaction system, which is beneficial to the molecular transfer of bioactive components; however, if the amount of water is too high, it will reduce the effectiveness of the internal hydrogen bond structure of the multi-electrolytic solvent A when dissolving the bioactive components; By optimizing the ratio of raw materials to solvents, the extraction rate of bioactive components in Radix Rehmanniae Preparata can be effectively improved; in a specific implementation, Radix Rehmanniae Preparata powder, multi-element low eutectic solvent A, water = 1g: (10-15) g: (15-25) mL; for example, it can be 1g: 10g: 15mL, 1g: 12g: 15mL, 1g: 15g: 15m, 1g: 15g: 18mL, 1g: 15g: 20mL, 1g: 15g: 25mL; but it is not limited to the listed values, and other values ​​not listed within the numerical range are also applicable; Microwaves can promote the precipitation of active medicinal components in Radix Rehmanniae Preparata, but other components of Radix Rehmanniae Preparata will also be precipitated under high-power microwave conditions, resulting in an increase in impurities, so the present invention adopts low-power microwaves; the power of microwave treatment is 150w~250w, for example, it can be 150w, 170w, 190w, 210w, 230w, 250w; but it is not limited to the listed values, and other values ​​not listed in the numerical range are also applicable; The heating extraction time is 10 min to 20 min; for example, it can be 10 min, 12 min, 15 min, 18 min, 20 min; but it is not limited to the listed values, and other values ​​not listed in the numerical range are also applicable; The temperature during the extraction process is 60°C to 120°C, for example, 60°C, 80°C, 100°C, or 120°C; but not limited to the listed values, other values ​​not listed within the numerical range are also applicable.

[0025] S3, mixing the dried Rehmannia glutinosa solid residue with the remaining raw material powder, using a multi-element low eutectic solvent B and an alcohol solution as an extractant B, stirring thoroughly, performing ultrasonic-assisted extraction, and then performing solid-liquid separation. After drying the obtained solid, adding the extractant B, repeating ultrasonic-assisted extraction and solid-liquid separation at least once, combining the filtrate after each solid-liquid separation to obtain an extraction supernatant; In the process of extracting the biologically active components of Radix Rehmanniae Preparata in step S2, the extraction of the flavonoid compounds contained therein is limited. In order to further extract the effective components in Radix Rehmanniae Preparata, step S3 is to mix the solid residue of Radix Rehmanniae Preparata separated and treated in step S2 with the remaining raw material powders such as roasted Epimedium, Millettia Spatholobi, Cibotium Cibotium, Drynariae Rhizoma, Cynomorium Songaricum, Dipsacus Asper, Fructus Lycii, Semen Radishes and the like for extraction; The multi-electrolytic solvent B is composed of choline salt + 1-octyl-3-methylimidazolium bromide + alcohol hydrogen bond donor. The alcohol hydrogen bond donor is a polyol such as 1,4-butanediol, 1,2-propylene glycol or triethylene glycol. The hydroxyl groups in the polyol can form intermolecular hydrogen bonds with the bioactive components in the raw materials, thereby increasing the solubility of flavonoids, phenolic compounds, sinapines, betaine and other components. This step uses ultrasound assistance. Based on the thermal and mechanical effects of ultrasound, on the one hand, sound waves are converted into mechanical energy to generate cavitation, thereby enhancing the permeability of the multi-component low eutectic solvent B and accelerating the extraction of the target components. On the other hand, when ultrasound is transmitted through the powder and liquid media in the reaction system, part of the ultrasonic energy is converted into heat energy due to the mutual movement between the medium particles, the mutual friction on the interface, and the absorption of the medium, thereby accelerating the transfer of the bioactive components and increasing their precipitation efficiency.

[0026] It has been found that as the extraction time increases, the amount of each component in the raw material increases, but as time increases, the structure of some compounds dissolved in the multi-element low eutectic solvent B will be destroyed. Therefore, the ultrasonic-assisted extraction in this step uses short-term and multiple extractions of effective components in the solid. The specific operation is as follows: (1) Mixing the dried Rehmannia glutinosa solid residue with the remaining raw material powder, and stirring evenly to form a mixed powder; (2) The mixed powder and the extractant B are mixed in a preset ratio, fully stirred, and then ultrasonic-assisted extraction is performed, followed by solid-liquid separation to obtain a first filtrate and a first solid residue; (3) The first solid residue is dried and mixed with the extractant B in a preset ratio, fully stirred and ultrasonic-assisted extraction is performed, and then solid-liquid separation is performed to obtain a second filtrate and a second solid residue; the second solid residue repeats this step to obtain a third filtrate and a third solid residue; (4) The first filtrate, the second filtrate and the third filtrate are combined and centrifuged, and the extracted supernatant is obtained after distillation and impurities are removed.

[0027] In a single ultrasonic-assisted extraction process, the mass ratio of the mixed powder, the first solid slag or the second solid slag and other solid materials: the multi-element low eutectic solvent B: the alcohol solution is 1:1~2:30~60, for example, it can be 1:1:30, 1:1.2:40, 1:1.5:45, 1:1.8:50, 1:2:60; but not limited to the listed values, other values ​​not listed in the numerical range are also applicable; when the number of ultrasonic-assisted extractions exceeds three times, the effective components of the subsequent extraction are limited and the effect is not obvious. Based on cost considerations, the number of ultrasonic-assisted extractions is controlled to three times; The alcohol solution is a methanol aqueous solution or an ethanol aqueous solution, and the mass concentration of alcohol in the alcohol solution is 40% to 80%; for example, it can be 40%, 50%, 60%, 70%, 80%; but it is not limited to the listed values, and other values ​​not listed within the numerical range are also applicable; In the extraction process, the role of adding the alcohol solution as a solvent to the multi-element low eutectic solvent B is: on the one hand, it improves the wettability of the raw material to be extracted and reduces the viscosity of the extraction reaction system, which is beneficial to the molecular transfer of the bioactive components; on the other hand, the bioactive components precipitated in this step are more easily soluble in methanol, ethanol, etc.; however, if the amount of alcohol solution is too high, the hydrogen bond structure effect will be reduced; The power of the ultrasonic assisted extraction is 200w~300w; for example, it can be 200w, 220w, 250w, 280w, 300w; but it is not limited to the listed values, and other values ​​not listed in the numerical range are also applicable; The temperature of the ultrasonic-assisted extraction is 40°C to 60°C; for example, it can be 40°C, 45°C, 50°C, 55°C, 60°C; but it is not limited to the listed values, and other values ​​not listed within the numerical range are also applicable; The time for a single ultrasound-assisted extraction is 5 minutes to 10 minutes; for example, it can be 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, or 10 minutes; but it is not limited to the listed values, and other values ​​not listed within the numerical range are also applicable.

[0028] S4, combining the Radix Rehmanniae Preparata extract and the supernatant to form a mixed liquid, and concentrating the mixed liquid to obtain a bone-increasing and eliminating stock solution; S5, preparing the bone-increasing and consuming preparation with the bone-increasing and consuming stock solution and the auxiliary agent; The bone-enhancing and detoxifying preparation can be in the form permitted in the art, such as tablets, capsules, pills, etc.; the adjuvants can be in the form permitted in the art, such as fillers, surfactants, lubricants, disintegrants, wetting agents, adhesives, etc., which are not specifically limited here. The following embodiments are described using tablets as an example, but are not limited to the dosage forms listed, and other dosage forms not listed in the art are also applicable.

[0029] Example 1 Embodiment 1 of the present invention provides a method for preparing bone augmentation and reduction, the method comprising steps 1 to 5: 1. Pretreatment of bone augmentation and digestion raw materials Weigh the following raw materials for bone enhancement and reduction according to weight: 200 parts of Radix Rehmanniae Preparata; 120 parts of roasted Epimedium; 120 parts of Millettia Spatholobi; 120 parts of Drynariae Rhizoma; 120 parts of Cynomorium songaricum; 60 parts of Dipsacus asper; 60 parts of Lycium barbarum; 60 parts of Radish Seed; grind the raw materials respectively and pass through 80-100 mesh sieve to make powder; 2. Extract the active ingredients from Rehmannia glutinosa Choline bromide: 1-octyl-3-methylimidazolium bromide: citric acid are mixed in a molar ratio of 1:1:2, and the three are heated to 40°C to 60°C to prepare a multi-component deep eutectic solvent A; Prepared Rehmannia root powder, multi-element low eutectic solvent A, and water are uniformly mixed in a ratio of 1g:10g:15mL, and the prepared Rehmannia root powder is heated to 60°C-70°C in a closed environment with a 150W microwave, and the extraction process is performed for 20 minutes; after the extraction is completed, the mixture is cooled to room temperature and then centrifuged at a speed of 5000rmp, and the centrifugation process not only separates the solid and liquid, but also dissociates the multi-element low eutectic solvent A from the extract; the liquid phase (i.e., the supernatant) is subjected to reduced pressure distillation to remove impurities such as acid hydrogen bond donors to obtain a prepared Rehmannia root extract, and the solid phase is prepared Rehmannia root solid residue; 3. Preparation of other extracts According to the molar ratio of choline bromide salt choline bromide: 1-octyl-3-methylimidazolium bromide: alcohol hydrogen bond donor 1,4-butanediol = 1:1:2, the three are heated to 40°C to 60°C and mixed to prepare a multi-element deep eutectic solvent B; The dried Rehmannia glutinosa solid residue is mixed with the remaining raw material powder, and stirred evenly to form a mixed powder; After being fully mixed and stirred in a mass ratio of mixed powder: multi-element low eutectic solvent B: 70% ethanol solution = 1:1:30, a first ultrasonic-assisted extraction was performed at 40°C for 10 minutes with an ultrasonic power of 200w, and then solid-liquid separation was performed to obtain a first filtrate and a first solid residue; After being fully mixed and stirred in a mass ratio of 1:1:30, the first solid residue after drying: multi-element low eutectic solvent B: 70% ethanol solution were subjected to a second ultrasonic-assisted extraction at 40°C for 10 minutes with an ultrasonic power of 200w, and then solid-liquid separation was performed to obtain a second filtrate and a second solid residue; After being mixed and stirred in a mass ratio of 1:1:30, the dried second solid residue: multi-electrolytic solvent B: 70% ethanol solution were subjected to a third ultrasonic-assisted extraction at 40°C for 10 minutes with an ultrasonic power of 200w, and then solid-liquid separation was performed to obtain a third filtrate and a third solid residue; The first filtrate, the second filtrate and the third filtrate are combined and centrifuged, and an extraction supernatant is obtained after impurities are removed by distillation; 4. The Radix Rehmanniae Preparata extract and the supernatant are combined into a mixed liquid, and the mixed liquid is concentrated to obtain a bone-enhancing and eliminating stock solution; 5. Add compressible starch and 70% ethanol to the bone-dense and detoxifying stock solution, dry and compress to make bone-dense and detoxifying tablets.

[0030] Example 2 On the basis of Example 1, only the component ratio of the multi-component low eutectic solvent A in step 2 was changed, and other steps and reaction parameters remained unchanged. The changes in the extraction amounts of rehmannia glutinosa glycoside D and rehmannia polysaccharides in the prepared rehmannia extract prepared in step 2 were shown in Table 1 below.

[0031] The content of rehmannia glutinosa D in the raw material of Rehmannia glutinosa was measured by high performance liquid chromatography, and the content of Rehmannia glutinosa polysaccharide was measured by phenol-sulfuric acid method. The extraction rate is the ratio of the content of the extracted component to the content of the corresponding component in the raw material of Rehmannia glutinosa.

[0032] Table 1

[0033] In Table 1, A1 refers to choline bromide, A1´ refers to choline chloride; A2 refers to 1-octyl-3-methylimidazolium bromide; A3 refers to citric acid, A3´ refers to acetic acid; It can be seen from the data in Table 1 that in the present invention, the use of multi-element low eutectic solvent A as extractant A can simultaneously, efficiently and quickly extract effective glycosides and rehmannia polysaccharides in Rehmannia glutinosa. As the amount of acid hydrogen bond donor increases, the extraction amount of rehmannia glutinosa glycoside D and rehmannia polysaccharides will increase. However, when it increases to a certain value, the effect of increasing the extraction amount of rehmannia glutinosa glycoside D and rehmannia polysaccharides is not obvious. Based on cost considerations, in the multi-element low eutectic solvent A, the molar ratio of choline salt: 1-octyl-3-methylimidazolium bromide: alcohol hydrogen bond donor is 1:1:2~4.

[0034] Example 3 On the basis of Example 1, only the ratio of Rehmannia glutinosa powder, multi-electrolytic solvent A and water in step 2 was changed, and other steps and reaction parameters remained unchanged. The changes in the extraction amounts of Rehmannia glutinosa glycoside D and Rehmannia glutinosa polysaccharides in the Rehmannia glutinosa extract prepared in step 2 were shown in Table 2 below.

[0035] Table 2

[0036] From the data in Table 3, it can be seen that with the increase in the dosage of multi-electrolyte A, the extraction amount of rehmannia glycoside D and rehmannia polysaccharide will increase. However, when the multi-electrolyte A increases to a certain value, the extraction amount of rehmannia glycoside D and rehmannia polysaccharide decreases. The reason is that with the increase in the dosage of multi-electrolyte A, the amount of water remains unchanged, resulting in an increase in the viscosity of the reaction system, which reduces the movement and transfer of molecules, thereby reducing the extraction amount. After further increasing the amount of water, the extraction amount of rehmannia glycoside D and rehmannia polysaccharide increases. Therefore, Rehmannia glutinosa powder, multi-electrolyte A, water = 1g: (10~15) g: (15~25) mL.

[0037] Example 4 On the basis of Example 1, only the parameters of the microwave in step 2 are changed, and the change of the microwave power will affect the reaction temperature of the system. Under the premise that other steps and reaction parameters remain unchanged, the changes in the extraction amount of rehmannia glutinosa glycoside D and rehmannia polysaccharide in the prepared rehmannia extract prepared in step 2 are shown in Table 3 below.

[0038] Table 3

[0039] From the data in Table 3, it can be seen that with the increase of microwave power and system reaction temperature, the extraction amount of rehmannia glutinosa glycoside D and rehmannia glutinosa polysaccharide will increase significantly. The reason is that the increase of microwave parameters will promote the destruction of rehmannia glutinosa cells and accelerate the analysis of components, and microwaves will accelerate the movement and transfer of molecules; however, when the microwave power and system reaction temperature increase to a certain value, the extraction amount of rehmannia glutinosa glycoside D and rehmannia glutinosa polysaccharide will decrease. The reason is that the increase of microwave power and system reaction temperature causes the water to gradually change from liquid to gas, and the viscosity of the reaction system increases, thereby reducing the movement and transfer of molecules, resulting in a decrease in the extraction amount of rehmannia glutinosa glycoside D and rehmannia glutinosa polysaccharide. Based on cost considerations, the temperature for heating and extracting rehmannia glutinosa powder is set at 60℃~120℃; the microwave power is set at 150w~250w.

[0040] Example 5 On the basis of Example 1, only the component ratio of the multi-component deep eutectic solvent B in step 3 was changed, and the other steps and reaction parameters remained unchanged. The changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 4 below.

[0041] HPLC was used to determine the content of flavonoids in the supernatant. Since the raw materials of bone-increasing and decomposing preparations such as Herba Epimedii, Millettia Spatholobi, and Rhizoma Drynariae had the highest content of flavonoids, other raw materials also contained effective ingredients such as phenols, sinapines, betaine, and protocatechuic acid, but the content was not high, flavonoids were used as reference amounts in step 3 to reflect the influence of different parameters on the extraction amount of effective ingredients in the raw materials. The following flavonoid content refers to the content of flavonoids in the supernatant.

[0042] Table 4

[0043] In Table 4, B1 refers to choline bromide, B1´ refers to choline chloride; B2 refers to 1-octyl-3-methylimidazolium bromide; B3 refers to 1,4-butanediol, B3´ refers to 1,2-propylene glycol, and B3´´ refers to triethylene glycol; As can be seen from the data in Table 4, in the present invention, the use of multi-element low eutectic solvent B as extractant B can simultaneously and efficiently and quickly extract the effective components in each raw material. As the amount of alcohol hydrogen bond donor increases, the extraction amount of flavonoid compounds will increase, but when it increases to a certain value, its effect on increasing the extraction amount of flavonoid compounds is not obvious. In addition, the comparison of the extraction effects of different alcohol hydrogen bond donors and choline salts is: triethylene glycol>1,2-propylene glycol>1,4-butanediol; choline bromide>choline chloride. Therefore, in the multi-element low eutectic solvent B, the molar ratio of choline salt: 1-octyl-3-methylimidazolium bromide: acid hydrogen bond donor = 1:1:2~4.

[0044] Example 6 On the basis of Example 1, only the ratio of the solid material, the multi-element deep eutectic solvent B and water in step 3 was changed, and the other steps and reaction parameters remained unchanged. The changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 5 below.

[0045] Table 5

[0046] From the data in Table 5, it can be seen that with the increase in the dosage of multi-electrolyte B, the extraction amount of flavonoids will increase. However, when the multi-electrolyte B increases to a certain value, the increase in the extraction amount of flavonoids is not obvious. The reason is that with the increase in the dosage of multi-electrolyte B, the alcohol solution remains unchanged, resulting in an increase in the viscosity of the reaction system, which reduces the movement and transfer of molecules, thereby reducing the extraction amount. After further increasing the dosage of alcohol solution, the extraction amount of flavonoids gradually increases. Therefore, in the single ultrasonic-assisted extraction process, the mass ratio of solid material: multi-electrolyte B: alcohol solution is set to 1:1~2:30~60.

[0047] Example 7 On the basis of Example 1, only the ultrasonic parameters in step 3 and the temperature of the reaction system during extraction were changed, and the other steps and reaction parameters remained unchanged. The changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 6 below.

[0048] Table 6

[0049] From the data in Table 6, it can be seen that with the increase of ultrasonic power and system reaction temperature, the extraction amount of flavonoid compounds will increase significantly. The reason is that the increase of ultrasonic parameters will strengthen its cavitation effect, enhance the penetration of multi-electrolytic solvent B, accelerate the extraction of target components, and microwaves will accelerate molecular movement and transmission; however, when the ultrasonic power increases to a certain value, the increase in the extraction amount of the components is not obvious. The reason is that: with the increase of ultrasonic power, its cavitation effect is too strong, which will cause the structure of biologically active components to be destroyed. In addition, the increase in temperature will cause the alcohol solution to volatilize, resulting in an increase in the viscosity of the reaction system, thereby reducing the movement and transmission of biologically active components and limiting the increase in the extraction amount. Therefore, based on cost considerations, the power of ultrasonic-assisted extraction is 200w~300w; the temperature is 40℃~60℃.

[0050] Comparative Example 1 In this comparative example, based on Example 1, only the component of the multi-component low eutectic solvent A in step 2 was changed, and other steps and reaction parameters remained unchanged. The changes in the extraction amounts of rehmannia glutinosa glycoside D and rehmannia polysaccharides in the prepared rehmannia extract prepared in step 2 are shown in Table 7 below.

[0051] In this comparative example, the multi-component deep eutectic solvent A does not contain choline bromide. Specifically, 1-octyl-3-methylimidazolium bromide and citric acid are mixed in a molar ratio of 1:1, and the two are heated to 40° C. to 60° C. to prepare the multi-component deep eutectic solvent A.

[0052] Comparative Example 2 In this comparative example, based on Example 1, only the component of the multi-component low eutectic solvent A in step 2 was changed, and other steps and reaction parameters remained unchanged. The changes in the extraction amounts of rehmannia glutinosa glycoside D and rehmannia polysaccharides in the prepared rehmannia extract prepared in step 2 are shown in Table 7 below.

[0053] In this comparative example, the multi-component low eutectic solvent A does not contain 1-octyl-3-methylimidazolium bromide. Specifically, choline bromide and citric acid are mixed in a molar ratio of 1:1, and the two are heated to 40° C. to 60° C. to prepare the multi-component low eutectic solvent A.

[0054] Comparative Example 3 In this comparative example, based on Example 1, the multi-electrolytic solvent A in step 2 was replaced with 75% ethanol solution, and the other steps and reaction parameters were kept unchanged. The changes in the extraction amounts of rehmannia glutinosa glycoside D and rehmannia polysaccharides in the prepared rehmannia extract obtained in step 2 were shown in Table 7 below.

[0055] Comparative Example 4 In this comparative example, based on Example 1, microwave treatment is not used in step 2, and other steps and reaction parameters remain unchanged. The changes in the extraction amounts of rehmannia glutinosa glycoside D and rehmannia polysaccharides in the prepared rehmannia extract prepared in step 2 are shown in Table 7 below.

[0056] In this comparative example, step 2 is specifically as follows: Choline bromide: 1-octyl-3-methylimidazolium bromide: citric acid are mixed in a molar ratio of 1:1:2, and the three are heated to 40°C~60°C to prepare a multi-element low eutectic solvent A; Rehmannia glutinosa powder, multi-element low eutectic solvent A, and water are evenly mixed in a ratio of 1g:10g:15mL, and Rehmannia glutinosa is heated to 60°C~70°C in a closed environment, and the extraction treatment is carried out for 20 minutes; after the extraction is completed, it is cooled to room temperature and then centrifuged at a speed of 5000rmp, and the centrifugation treatment not only separates the solid and liquid, but also dissociates the multi-element low eutectic solvent A from the extract; the liquid phase (i.e., the supernatant) is subjected to reduced pressure distillation to remove impurities such as acid hydrogen bond donors to obtain a Rehmannia glutinosa extract, and the solid phase is Rehmannia glutinosa solid residue.

[0057] Comparative Example 5 In this comparative example, based on Example 1, the multi-component deep eutectic solvent A in step 2 was replaced with the multi-component deep eutectic solvent B, and the other steps and reaction parameters remained unchanged. The changes in the extraction amounts of rehmannia glutinosa glycoside D and rehmannia polysaccharides in the prepared rehmannia extract obtained in step 2 are shown in Table 7 below.

[0058] Table 7

[0059] It can be seen from the data in Table 7 that the low eutectic solvent of 1-octyl-3-methylimidazolium bromide + hydrogen bond donor or choline salt + hydrogen bond donor, ethanol solution or multi-component low eutectic solvent B is used as the extractant, and the effect of extracting the active ingredients from Rehmannia glutinosa is lower than that of the multi-component low eutectic solvent A of the present invention; in the extraction process, the effect of microwave synergistic operation is better than that of non-microwave operation.

[0060] Comparative Example 6 In this comparative example, based on Example 1, only the component of the multi-component deep eutectic solvent B in step 3 was changed, and the other steps and reaction parameters remained unchanged. The changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 8 below.

[0061] In this comparative example, the multi-component low eutectic solvent B does not contain choline bromide. Specifically, 1-octyl-3-methylimidazolium bromide and 1,4-butanediol are mixed in a molar ratio of 1:1, and the two are heated to 40° C. to 60° C. to prepare the multi-component low eutectic solvent B.

[0062] Comparative Example 7 In this comparative example, based on Example 1, only the component of the multi-component deep eutectic solvent B in step 3 was changed, and the other steps and reaction parameters remained unchanged. The changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 8 below.

[0063] In this comparative example, the multi-component low eutectic solvent B does not contain 1-octyl-3-methylimidazolium bromide. Specifically, choline bromide: 1,4-butanediol are mixed in a molar ratio of 1:1, and the two are heated to 40° C. to 60° C. to prepare the multi-component low eutectic solvent B.

[0064] Comparative Example 8 In this comparative example, based on Example 1, ultrasonic co-treatment is not used in step 3, and under the premise that other steps and reaction parameters remain unchanged, the changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 8 below.

[0065] Comparative Example 9 In this comparative example, based on Example 1, a one-time ultrasonic synergistic treatment was used in step 3, and under the premise that other steps and reaction parameters remained unchanged, the changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 8 below.

[0066] The specific step 3 of this comparative example is: According to the molar ratio of choline bromide salt choline bromide: 1-octyl-3-methylimidazolium bromide: alcohol hydrogen bond donor 1,4-butanediol = 1:1:2, the three are heated to 40°C to 60°C and mixed to prepare a multi-element deep eutectic solvent B; The dried Rehmannia glutinosa solid residue is mixed with the remaining raw material powder, and stirred evenly to form a mixed powder; After being fully mixed and stirred in a mass ratio of mixed powder: multi-component low eutectic solvent B: 70% ethanol solution = 1:3:90, ultrasonic-assisted extraction was carried out at 40°C for 30 minutes with an ultrasonic power of 200w, and then solid-liquid separation was carried out to obtain a filtrate and a solid residue; the filtrate was centrifuged and the extraction supernatant was obtained after distillation and impurity removal.

[0067] Comparative Example 10 In this comparative example, based on Example 1, the mixed powder in step 3 only contains the remaining raw material powder except Rehmannia glutinosa, and the other steps and reaction parameters remain unchanged. The changes in the extraction amount of each effective ingredient (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 8 below.

[0068] Comparative Example 11 In this comparative example, based on Example 1, only the multi-component deep eutectic solvent B in step 3 was replaced with the multi-component deep eutectic solvent A, and the other steps and reaction parameters remained unchanged. The changes in the extraction amount of each effective component (taking flavonoid compounds as an example) in the extraction supernatant obtained in step 3 are shown in Table 8 below.

[0069] Table 8

[0070] It can be seen from the data in Table 8 that the low eutectic solvent or multi-component low eutectic solvent A of 1-octyl-3-methylimidazolium bromide + hydrogen bond donor or choline salt + hydrogen bond donor is used as the extractant, and its effect of extracting active ingredients in the drug is lower than that of the multi-component low eutectic solvent B of the present invention; during the extraction process, the comparison results of the effects of different ultrasonic treatments are: multiple ultrasonic treatment > one-time ultrasonic treatment > no ultrasonic treatment; after removing the Rehmannia glutinosa solid residue in step 2, the extraction amount of flavonoid compounds is slightly reduced.

[0071] The above-mentioned embodiments only express several implementation methods of the present invention, and the description thereof is relatively specific and detailed, but it cannot be understood as limiting the scope of the patent of the present invention. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. A method for preparing bone augmentation and reduction, characterized in that: The raw materials for bone enhancement and reduction include prepared rehmannia root, roasted epimedium, millettia reticulata, cibotium barometz, drynariae, cynomorium songaricum, dipsacus root, wolfberry fruit and radish seed; The preparation method comprises the following steps: The choline salt, 1-octyl-3-methylimidazolium bromide and the acid hydrogen bond donor are heated and mixed according to a preset molar ratio to prepare a multi-element deep eutectic solvent A; The choline salt, 1-octyl-3-methylimidazolium bromide and the alcohol hydrogen bond donor are heated and mixed according to a preset molar ratio to prepare a multi-component deep eutectic solvent B; The solution of multi-element low eutectic solvent A is used as extractant A, and Rehmannia glutinosa powder is treated by microwave in a closed environment to obtain Rehmannia glutinosa extract and Rehmannia glutinosa solid residue; The dried Rehmannia glutinosa solid residue and the remaining raw material powder form a mixed material, and a solution of a multi-component low eutectic solvent B is used as an extractant B, and ultrasonic-assisted extraction is performed to obtain an extraction supernatant; The prepared Rehmannia root extract and the extracted supernatant are combined and concentrated to obtain the bone-increasing and detoxifying stock solution; The bone-increasing and consuming stock solution and adjuvants are used to prepare the bone-increasing and consuming preparation.

2. A method for preparing bone augmentation and reduction according to claim 1, characterized in that: The specific steps of obtaining the Radix Rehmanniae Preparata extract and the Radix Rehmanniae Preparata solid residue are: The Rehmannia root powder, multi-element low eutectic solvent A and water are uniformly mixed in a preset ratio, and the Rehmannia root powder is heated and extracted in a closed environment with microwave. After the extraction, it is cooled to room temperature and then centrifuged. After liquid phase distillation and impurity removal, the Rehmannia root extract is obtained, and the solid phase is the Rehmannia root solid residue.

3. The method for preparing bone augmentation and reduction according to claim 1, characterized in that: The specific steps of obtaining the extraction supernatant are: The dried Rehmannia glutinosa solid residue is mixed with the remaining raw material powder, and a mixed solution of a multi-component low eutectic solvent B and an alcohol solution is used as an extractant B. After sufficient stirring, ultrasonic-assisted extraction is performed, and then solid-liquid separation is performed. The obtained solid is added with the extractant B, and ultrasonic-assisted extraction and solid-liquid separation are repeated at least once. The filtrates after each solid-liquid separation are combined to obtain an extraction supernatant.

4. The method for preparing bone augmentation and reduction according to claim 1, characterized in that: According to the molar ratio, in the multi-component low eutectic solvent A, choline salt: 1-octyl-3-methylimidazolium bromide: acid hydrogen bond donor = 1:1:2~4; the choline salt is choline chloride or choline bromide; the acid hydrogen bond donor is citric acid or acetic acid; the heating temperature when preparing the multi-component low eutectic solvent A is 40°C~60°C.

5. The method for preparing bone augmentation and reduction according to claim 1, characterized in that: According to the molar ratio, in the multi-component low eutectic solvent B, choline salt: 1-octyl-3-methylimidazolium bromide: alcohol hydrogen bond donor = 1:1:2~4; the choline salt is choline chloride or choline bromide; the alcohol hydrogen bond donor is 1,4-butanediol, 1,2-propylene glycol or triethylene glycol; the heating temperature when preparing the multi-component low eutectic solvent B is 40°C~60°C.

6. The method for preparing bone augmentation and reduction according to claim 2, characterized in that: The prepared Rehmannia root powder, multi-element low eutectic solvent A, and water = 1 g: (10-15) g: (15-25) mL.

7. A method for preparing bone augmentation and reduction according to claim 2, characterized in that: The time for heating and extracting the Radix Rehmanniae Radix Powder is 10-20 minutes; the temperature for heating and extracting is 60-120° C.; and the power of the microwave is 150-250W.

8. The method for preparing bone augmentation and reduction according to claim 3, characterized in that: The steps of preparing the extraction supernatant specifically include: The dried Rehmannia glutinosa solid residue is mixed with the remaining raw material powder, and stirred evenly to form a mixed powder; The mixed powder and the extractant B are mixed in a preset ratio, fully stirred, and then ultrasonic-assisted extraction is performed, and then solid-liquid separation is performed to obtain a first filtrate and a first solid residue; The first solid residue is dried and mixed with the extractant B according to a preset ratio, fully stirred and subjected to ultrasonic-assisted extraction, and then solid-liquid separation to obtain a second filtrate and a second solid residue; the second solid residue repeats this step to obtain a third filtrate and a third solid residue; The first filtrate, the second filtrate and the third filtrate are combined and centrifuged, and an extraction supernatant is obtained after impurities are removed by distillation; The power of the ultrasonic-assisted extraction is 200w-300w; the temperature is 40°C-60°C; and the time of a single ultrasonic-assisted extraction is 5min-10min.

9. A method for preparing bone augmentation and reduction according to claim 8, characterized in that: In a single ultrasonic-assisted extraction process, the mass ratio of solid material: multi-element low eutectic solvent B: alcohol solution is 1:1-2:30-60, and the solid material is a mixed powder, a first solid residue or a second solid residue.

10. The method for preparing bone augmentation and reduction according to claim 3, characterized in that: The alcohol solution is a methanol aqueous solution or an ethanol aqueous solution, and the mass concentration of alcohol in the alcohol solution is 40% to 80%.

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