CN108969555B - A Chinese medicinal composition comprising herba Epimedii with kidney invigorating, blood and essence replenishing, and immunity improving effects, and its preparation method - Google Patents
A Chinese medicinal composition comprising herba Epimedii with kidney invigorating, blood and essence replenishing, and immunity improving effects, and its preparation method Download PDFInfo
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- CN108969555B CN108969555B CN201810900857.7A CN201810900857A CN108969555B CN 108969555 B CN108969555 B CN 108969555B CN 201810900857 A CN201810900857 A CN 201810900857A CN 108969555 B CN108969555 B CN 108969555B
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Abstract
Disclosed are a herba Epimedii and radix Rhodiolae Chinese medicinal composition for invigorating kidney, replenishing vital essence and blood, and enhancing immunity, and a saussurea involucrate extraction device, wherein the extraction device has high efficiency, and can minimize the generation of by-products during the extraction process, so that the obtained extract has high pharmacological activity. The composition has effects of enhancing immunity, resisting inflammation, relieving fatigue, preventing radiation, invigorating kidney, and replenishing essence and blood.
Description
Technical Field
The invention relates to a preparation method of a herba epimedii and rhodiola rosea traditional Chinese medicine composition, in particular to a preparation method of a traditional Chinese medicine composition which is capable of improving immunity, resisting inflammation, fatigue and radiation, tonifying kidney and benefiting essence and blood, wherein the traditional Chinese medicine composition is especially a composition containing a snow lotus extract, an epimedium extract and a rhodiola rosea extract, namely the snow lotus, herba epimedii and rhodiola rosea traditional Chinese medicine composition.
Background
Saussurea involucrate is a rare and rare Chinese medicinal material in China, has long-term application history and rich folk medicine bases, contains flavone and glycoside thereof, phenylpropanoids, sesquiterpene lactone, coumarin, lignan, organic acid, amino acid, steroid, polysaccharide and other components in the saussurea involucrate, has biological activities of resisting rheumatism, easing pain, regulating cardiovascular system, resisting cancer, delaying aging, preventing radiation and the like, and has great medicinal development value. The effects of improving immunity, resisting inflammation and preventing radiation of the saussurea involucrate and the extract thereof are recorded in a plurality of documents. The research on the radiation damage resistance and mechanism of the saussurea involucrate extract indicates that the saussurea involucrate is well used as a traditional medicine for a long time, has definite pharmacological action and has good effects of resisting oxidation and removing free radicals, and systematically demonstrates the radiation damage resistance and the possible action mechanism of the saussurea involucrate extract through a whole animal experiment, a hydroxyl free radical removal experiment, Giemsa staining analysis of chromosome aberration and RT-PCR analysis of P21 gene expression for regulating and controlling the cell cycle. The research on the radiation protection effect of saussurea involucrata aqueous extract on mouse germ cells in radiation health of China, 4 th 2010, Na and the like researches the radiation protection effect of the saussurea involucrata aqueous extract on the mouse germ cells. The research on the effects of the snow lotus herb oral liquid on improving the immunity, promoting blood circulation, relieving pain and resisting inflammation is carried out by the methods of pharmacology and clinic of traditional Chinese medicines, Huanghua and the like, pharmacology and clinic of traditional Chinese medicines, 2002 and the research on the effects of the snow lotus herb oral liquid on promoting blood circulation, relieving pain and resisting inflammation.
The kidney-tonifying and essence-blood-benefiting effects and mechanisms of epimedium are also widely recognized and studied. In the fourth round of pharmaceutical planning teaching material of pharmaceutical products in higher medical colleges and universities in China (3 rd edition [ M ].2016), which is edited by Prunus mume, it is clearly indicated that Epimedium herb has the effects of invigorating kidney and tonifying yang, and the pharmacology of Epimedium herb is mainly to enhance the immune function, improve the gonadal function, delay senility, resist inflammation, resist allergy and the like. The kidney-warming and yang-supporting functions of epimedium and extracts thereof (mainly flavonoids and glycosides) are also indicated in the professional knowledge of traditional Chinese medicine (2015) organized by the qualification and certification center of medical practitioners in the State food and drug administration. The effect of tonifying kidney and replenishing essence and blood of epimedium and its extract is clearly indicated in the comparison of efficacy of traditional Chinese medicines (2013) compiled in Zhang Jinghu.
Radix Rhodiolae has effects of invigorating qi, promoting blood circulation, and removing blood stasis. The research progress of pharmacological action of salidroside indicates that salidroside is one of the main active ingredients of traditional Tibetan medicine rhodiola rosea in China and has high medicinal value in 2015, and the research review of pharmacological action of salidroside on skeletal system, cardiovascular system, nervous system and liver from the protection action of salidroside on diabetic nephropathy, the treatment action of the salidroside, anti-inflammation, immunity improvement and the like provides theoretical basis for the research and clinical application of related diseases on medicament prevention and treatment. 98-99 of rhodiola root pharmacological action, has various pharmacological actions of resisting fatigue, resisting aging, regulating immunity, removing free radicals, resisting hepatic fibrosis and kidney damage, improving and protecting the functions of heart and brain, resisting tumor, resisting radiation and the like, and has wide application prospect.
In the development of saussurea involucrate traditional Chinese medicinal materials, the quality control is particularly critical, and particularly, the problem of controlling the content of impurities in the extraction of active ingredients of saussurea involucrate is a difficult problem. Rutin, chlorogenic acid, syringin, dehydrocostus lactone and other components are special expected components of non-snow lotus, and how to effectively separate the rutin, the chlorogenic acid, the syringin, the dehydrocostus lactone and the like from pharmacologically active components raises very high requirements and challenges for an extraction process.
CN101041653B discloses a method for extracting and enriching total flavone of saussurea involucrata, which comprises the steps of carrying out supercritical CO treatment on saussurea involucrata2Extracting to obtain volatile and heat labile components, and extracting with conventional solvent. The saussurea involucrate flavone after supercritical extraction is more easily released from the plant body.
CN102106890B discloses application and a preparation method of saussurea involucrate total sesquiterpene lactone and application of the saussurea involucrate total sesquiterpene lactone in preparation of an anti-hypoxic drug, wherein the total sesquiterpene lactone can be extracted by a solvent method, carbon dioxide supercritical extraction and petroleum ether reflux.
The extraction method of the CN104397683A jellyfish snow lotus essence comprises the following steps: (1) pre-treating saussurea medusa; (2) mixing pre-treated jellyfish herba Saussureae Involueratae, deionized water and bean curd, decocting for 2-10 hr, rinsing to remove bean curd,filtering to obtain a filtrate i, and drying residues obtained after filtering in the shade; (3) mixing the residue with rice vinegar, moistening for 6-10 min, parching with slow fire, taking out, and cooling; (4) grinding the cooled residue, sieving with 20-40 mesh sieve, placing in supercritical extraction kettle for extraction at 50-70 deg.C under 20-40Mpa with ethanol as entrainer and CO2Performing circulating extraction for 2-5 hours, and discharging at a discharge hole in the separation kettle after extraction is finished to obtain filtrate ii; (5) mixing the filtrate i and the filtrate ii, and filtering by using a microporous filter membrane with the pore diameter of 0.22 mu m to obtain a filtrate iii; (6) concentrating the filtrate iii at below 60 deg.C by rotary evaporation under reduced pressure until the solid content is 60-80%; (7) drying the concentrated solution to obtain jellyfish herba Saussureae Involueratae extract.
CN106511213A discloses a cosmetic with plant extract, which is prepared by mixing the following components in percentage by weight: 20-50% of aloe, 5-20% of chamomile, 5-15% of snow lotus herb, 10-30% of jasmine flower and 10-20% of tea; mixing the above components according to weight percentage, drying, pulverizing, performing carbon dioxide supercritical extraction on the medicinal powder, extracting with anhydrous ethanol as solvent, concentrating under reduced pressure, and recovering ethanol to obtain ethanol extraction concentrate; extracting the residue with 10-70% ethanol solution as extraction solvent, heating, concentrating, passing through macroporous resin column, eluting with 30-90% ethanol for several times, concentrating the eluate under reduced pressure, dissolving with 1.3-butanediol, filtering, and mixing with the solution obtained by supercritical carbon dioxide extraction to obtain plant extractive solution.
CN102106889A discloses an application of saussurea involucrate and saussurea involucrate extract in preparing anti-hypoxic drugs and a preparation method of saussurea involucrate extract, wherein the saussurea involucrate is selected from one of saussurea involucrate, saussurea medusa Maxim or saussurea bracteata, and the saussurea involucrate extract is one of saussurea involucrate extract, saussurea medusa Maxim extract or saussurea bracteata extract; the saussurea involucrate extract is water extract of saussurea involucrate or ethanol extract of saussurea involucrate and anti-anoxia effective part obtained by extraction with system solvent method and macroporous adsorption resin method.
CN101209274A A saussurea involucrate extract, which is characterized in that the saussurea involucrate extract contains 0.5 to 1.2 percent of syringin, 7 to 17 percent of chlorogenic acid and 8 to 25 percent of rutin according to weight percentage.
Research on extraction process of saussurea involucrata water-soluble polysaccharide, herba orostachyos and the like, food and biotechnology reports 2007, 26(3):37-40, compares the influence of factors such as material-liquid ratio, temperature, extraction time and the like on extraction rate of the snow lotus polysaccharide in the Sinkiang saussurea involucrata tissue culture, and provides the optimal extraction process of the saussurea involucrata water-soluble polysaccharide.
In summary, there are many defects in the prior art, especially the extraction method of saussurea involucrate in the prior art is still the traditional extraction method, although the report of extracting pharmacologically active substances of saussurea involucrate by using the supercritical method is available, the extraction method still uses the conventional supercritical extraction equipment and process, the content of the physiologically active ingredients in the extract is low, and the content of the undesirable impurities is high. In addition, no report exists for preparing a compound preparation by compounding the saussurea involucrate extract, the epimedium extract and the rhodiola rosea extract, or the three extracts containing complex components, namely crude extracts are usually mixed, so that the effect is poor and the synergistic effect is poor. There is a need in the art for an efficient extraction apparatus and method for obtaining saussurea involucrate extract with high pharmacological activity.
Disclosure of Invention
In order to solve the technical problems in the prior art at the same time, the invention provides a saussurea involucrate supercritical extraction device, which can effectively extract and obtain pharmacologically active ingredients with high content of the pharmacologically active ingredients and low content of undesirable impurities. The saussurea involucrate extract and the compound preparation containing the saussurea involucrate extract have high anti-inflammatory effect, immunity improving, radiation-proof, fatigue-resisting effect and the like, and have improved comprehensive pharmacological activity.
More specifically, the invention provides a composition for improving immunity, resisting inflammation and radiation, tonifying kidney and benefiting essence and blood, in particular to a traditional Chinese medicine composition containing a snow lotus extract, an epimedium extract and a rhodiola rosea extract, namely, the invention provides the snow lotus, epimedium and rhodiola rosea traditional Chinese medicine composition for improving immunity, resisting inflammation and radiation, tonifying kidney and benefiting essence and blood, so that the subject (name) of the invention can also be the snow lotus, epimedium and rhodiola rosea traditional Chinese medicine composition for improving immunity, resisting fatigue and preventing radiation and the preparation method thereof. Therefore, the invention provides the following technical scheme.
In one aspect of the invention, a saussurea involucrate supercritical extraction device is provided, which comprises: an inlet section, an extraction section, and an outlet section.
With the present invention, the inlet portion may comprise: a carbon dioxide supply tank (1), valves (2,4,7) connected to a pressure gauge (5), a compressor (3), a carbon dioxide storage vessel (6), a high pressure pump (8), a coil (9) and CO towards an extraction tank (11)2An inlet valve (10).
With the present invention, the extraction section may include: the device comprises an extraction pool (11), a pressure gauge (12) connected with the extraction pool, and a heating system consisting of a light-emitting bulb (13), a fan (14) for homogenizing the temperature and a precision thermometer connected with a temperature controller (15) for measuring the temperature.
With the present invention, the outlet portion may comprise: an outlet valve (16) of the extraction tank, an extract collector (17), a hot bath (18) and a flow meter (19).
Preferably, the extraction cell is made of a stainless steel alloy.
More preferably, the composition of the stainless steel alloy is, in weight percent: mn is more than or equal to 0.3 percent and less than or equal to 0.8 percent; cr is more than or equal to 3.0 percent and less than or equal to 6.0 percent; ni is more than or equal to 4.0 percent and less than or equal to 10.0 percent; al is more than or equal to 1.0 percent and less than or equal to 2 percent; ti is more than or equal to 1.0 percent and less than or equal to 1.5 percent, P is more than or equal to 0.001 percent and less than or equal to 0.02 percent, and W is more than or equal to 0.1 percent and less than or equal to 0.2 percent; s is more than or equal to 0.001% and less than or equal to 0.005%; n is more than or equal to 0.0001 percent and less than or equal to 0.0060 percent; c is more than or equal to 0.0001 percent and less than or equal to 0.025 percent; si is more than or equal to 0.0001% and less than or equal to 0.25%; o is more than or equal to 0.0001 percent and less than or equal to 0.0050 percent, and the balance is Fe.
The inventor researches and discovers that in the alloy, the addition of Ti can obviously improve the strength of the alloy, the fracture toughness of the alloy can be improved after the alloy is subjected to high-temperature deformation and aging refining of crystal grains, and the addition of Ni can improve the toughness of the alloy. The intermetallic compound formed by Al and Ni and Al and W can play a role of dispersion strengthening and antioxidation in the alloy, thereby improving the strength and the impact toughness of the alloy, for example, the strength is improved by 30 to 50 percent compared with the alloy without adding Ni and W.
The extraction cell of the invention can be made from the alloy by the following method:
(1) firstly, casting ingots by raw materials to prepare a semi-finished product;
(2) then carrying out isothermal heat treatment on the semi-finished product at 850-920 ℃, wherein the isothermal time is 3-6 hours, and then rapidly cooling to the temperature lower than or equal to-80 ℃ to carry out deep cooling treatment;
(3) the aging annealing treatment is carried out at the temperature of 400-480 ℃, and the aging treatment time is 10-20 hours.
The extraction cell obtained by the method has particularly good fracture resistance and toughness under extremely high pressure, and is not easy to generate CO under high pressure2Acid gas induced surface erosion thus enabling supercritical extraction at higher pressures and lower temperatures.
In another aspect of the present invention, there is provided a method for extracting effective components of saussurea involucrata using the above extraction apparatus, the method comprising: (1) bringing the device to the pressure conditions required for extraction at the inlet section; (2) extracting the extract under selected conditions of temperature and pressure in the extraction section; and (3) withdrawing the extract at the outlet portion.
In another aspect of the present invention, there is provided a saussurea involucrate extract obtained using the above apparatus or using the above method.
Preferably, the content of the volatile oil in the saussurea involucrate extract is less than 1.2 wt%. More preferably, the total content of rutin, chlorogenic acid, syringin and dehydrocostus lactone in the saussurea involucrate extract is lower than 2.5 wt%. Most preferably, the total content of lactone compounds having a parent nucleus represented by the following formula (a) in the saussurea involucrate extract is more than 65.0 wt%:
the inventor researches and discovers that when the components in the saussurea involucrate extract simultaneously meet the content requirement, the pharmacological activity of the saussurea involucrate extract can be particularly effectively ensured, and particularly the anti-inflammatory, immunity-improving and radiation-proof effects are achieved. Research shows that the lactone compound is a compound with better effects in the saussurea involucrate extract.
In the present invention, it is preferable that the extraction cell (11) has a diameter of 6.1 cm, a height of 18 cm, and an effective capacity of 0.50L; the power of the light-emitting bulb is 200-300W; the thermometer is an ASQ F200 precision thermometer, and the temperature controller is a temperature controller based on a single chip microcomputer. In the invention, because the extraction pool made of the alloy can bear higher pressure, the extraction can be carried out at lower temperature, and the required temperature can be obtained by heating with a bulb, thereby overcoming the problems of uneven heating and low heating control precision (such as temperature flying phenomenon) existing in the conventional heating means such as oil bath or water bath heating, effectively preventing the fluctuation of supercritical extraction conditions and effectively reducing the generation of extraction byproducts.
The inventor finds out through a large number of experiments that the optimal process conditions for extracting the saussurea involucrate are as follows: dried and pulverized (e.g., particle size of 400. + -. 25 μm) snow lotus (e.g., 25 g) is put into an ultrasonic extraction cell, the extract is extracted at 40 ℃ and 10.34MPa, each extraction is carried out for 120 minutes and the carbon dioxide discharge rate is 125.25. + -. 15.00m L/min, and the cell for collecting the extract is immersed in an ice water bath (8. + -. 0.5 ℃).
In another aspect of the present invention, there is provided a composition comprising the above extract. The composition also optionally comprises epimedium extract and rhodiola extract. The weight ratio of the saussurea involucrate extract, the epimedium extract and the rhodiola rosea extract is (10-5) to (1-3).
In another aspect of the invention, the use of the composition is provided, which is used for preparing medicines, health products or drinks capable of enhancing the immune function of human bodies.
Preferably, the purity of the glycosides in the epimedium composition is greater than 99.0 wt%. More preferably, the glycoside species has a purity of greater than 99.2 wt%. Most preferably, the glycoside species is greater than 99.7 wt% pure.
The glycoside is preferably or comprises a mixture of compounds of the following formulae (I) and (II):
the weight ratio of the compounds of the formula (I) to the compounds of the formula (II) is preferably from 1:5 to 5: 1.
Researches show that the two compounds are glycoside compounds, and compared with general flavonoid compounds such as icariin A and icariin B, the two compounds have particularly wide and relatively higher biological activities, such as anti-osteoporosis, anti-inflammatory and anti-cancer effects, kidney tonifying and essence and blood benefiting effects.
"influence of Epimedium general flavone on hypothalamus-pituitary-adrenal axis calmodulin gene expression in kidney yang deficiency rat", Jiang shujun et al, journal of Chinese applied physiology, 2004; 20(4), discussing the pathological changes of the kidney-yang deficiency syndrome and the action and the mechanism of the epimedium total flavonoids from the molecular level, and providing a theoretical basis for developing the new clinical application of the medicament. In many documents including the above document, the effects of tonifying kidney and benefiting essence and blood including the above epimedium flavone compounds (also glycoside compounds) are reported, but basically, the study is carried out as total flavone (i.e. total mixture).
The master thesis of the Jilin agriculture university, "research on chemical components of epimedium koreanum", the han bin, 20070601, and the master thesis of the Beijing chemical university, "research on chemical components and antibacterial activity of epimedium in Sichuan", Songhina, 20090601 all report that a plurality of flavone compounds (namely, the glycoside compounds) including the compounds of the formula (I) and the formula (II) have the effects of tonifying kidney and benefiting essence and blood, and specifically, the icariin can obviously promote basic secretion of rat interstitial cells TS, so that experimental basis is provided for the epimedium to treat male infertility with low TS level. The epimedium herb can obviously improve the yang deficiency syndrome caused by glucocorticoid, and all indexes can be recovered quickly. However, the above documents do not specifically indicate that the two specific compounds have superior effects.
The research on the chemical components and pharmacology of epimedium plants, the Xianjuan, the pharmaceutical of strait-Xian, No. 18 and No. 5 in 2006 reports the pharmacological effects of various flavone compounds including the compounds of the formula (I) and the formula (II) on benefiting essence, strengthening bones and muscles, tonifying waist and kidney and strengthening heart. Similarly, the two compounds have no better effects of tonifying kidney and benefiting essence and blood.
The inventor of the invention finds that the compounds of the formula (I) and the formula (II) have better efficacies relative to other compounds in the extract through screening, and extracts the compounds serving as target compounds by adopting a specific process to prepare the composition.
The compounds of formula (I) and formula (II) are preferably obtained by extraction by the following method:
(1) extracting herba Epimedii herbal powder (preferably 1.18kg) with MeOH (preferably 3X 6L, each soaking time is about 20h) at room temperature, and evaporating the combined extracts under reduced pressure to obtain brown residue (162.5g) as methanol extract;
(2) using a reverse phase silica gel column (e.g., RP-18 silica gel column), MeOH-H is first used2Subjecting the residue to gradient elution with a mixed solution of O to perform column chromatography, preferably with elution steps of 0:1(1L), 1:9(1L), 2:8(1L), 3:7(1L), 4:6(1L), 5:5(2L), 6:4(2L), 7:3(1L), 1:0(2L), followed by acetone (2L) to obtain 10 elution fractions in sequence, denoted as elution fractions A-J;
(3) the eluted fraction B (e.g. 6.5g) is chromatographed on a column of silica gel (preferably 60 cm. times.5 cm) using CHCl3-MeOH-H2Gradient elution is carried out on the O mixed solution (the elution gradient is preferably 20:1:0(1L), 10:1:0(2L), 30:8:1(2L) and 15:6:1(2.5L)), so that 4 elution components are obtained and are marked as elution components B1-B4;
(4) elute B2 (e.g., 300.2mg) with MeOH-H2Separating O (1:1) on Sephadex LH-20 (preferably 100cm × 3cm) to obtain 7 secondary elution components, which are marked as elution components B2.1-B2.7;
(5) a mixture of the eluting components B2.5 and B2.6 (e.g. 210.5mg total weight) is passed through a CC column on silica gel (preferably 80 cm. times.2.5 cm) eluting with EtOAc-CHCl3-MeOH-H2O (12:6:3:1), and concentrating the eluate under reduced pressure to dryness to obtain pale yellow powder (e.g. 11.2mg), which is a mixture of compounds represented by formula (I) and formula (II).
The method can effectively eliminate interference of protein and other impurities, and other low-titer polysaccharides can be effectively separated. The applicant obtained for the first time the compounds of formula (I) and formula (II) in extremely high purity by the above 5-stage column chromatography separation, and it is important that the 5-stage separation system have close hierarchical and coordination relationships, such as sequential selection.
In the context of the present invention, the rhodiola rosea extract is preferably salidroside. Salidroside can be extracted by conventional method in the art. More preferably, the rhodiola rosea extract is a modified rhodiola rosea extract, and most preferably, the rhodiola rosea extract is a compound represented by the following formula (II) or (III) obtained by modifying salidroside:
compared with general natural glycosides, the compound shown in the formula (II) or (III) has more obvious immunoregulation effect on normal mice, enhances the capacity of scavenging free radicals due to the introduction of electron-withdrawing groups Cl and F, can obviously increase the number of specific antibody secretory cells of the mice, can enhance the delayed hypersensitivity reaction intensity at the dosage of 50 mg/(kg. d), also shows the mixed lymphocyte reaction on heterotypic mice and the phagocytic function of macrophages, and can reduce the activity of interleukins to a certain extent, thereby indicating that the two are promising immunomodulators.
The substances having the structural formulae (II) to (III) can be synthesized by the following methods: adding MCM-22 molecular sieve as selective catalyst into a reaction container, adding chloroform solvent, adding natural salidroside and chloro-or fluoro-acetyl chloride into the reaction solvent in a molar ratio of 1:1-1.3 (the natural salidroside and the chloro-or fluoro-acetyl chloride are preferably added in the form of chloroform solution), reacting at 20-30 ℃ for 10-30min, separating and drying to obtain the compound. The method has the advantages of high yield and less impurities. Especially, the occurrence of side reaction can be obviously reduced due to the shape-selective action of the MCM-22 molecular sieve.
In another aspect of the invention, the use of the composition is provided, which is used for preparing medicines, health products or drinks with anti-inflammatory effect, anti-fatigue effect and radiation protection effect, which can enhance the immune function of human body.
Preferably, the medicine is a compound traditional Chinese medicine preparation. Preferably, the composition is used for preparing oral capsules, granules and the like. The health product comprises oral products such as chewable tablets or buccal tablets.
Drawings
Fig. 1 is a schematic view of an extraction device according to the invention.
Detailed Description
Example 1
The saussurea involucrate used for extraction is saussurea involucrate (purchased from Wulu wood Qi North Yuan Chun agricultural and sideline products business Co., Ltd.). The extraction device adopted comprises: an inlet portion, an extraction portion, and an outlet portion, the inlet portion may include: a carbon dioxide supply tank (1), valves (2,4,7) connected to a pressure gauge (5), a compressor (3), a carbon dioxide storage vessel (6), a high pressure pump (8), a coil (9) and CO towards an extraction tank (11)2An inlet valve (10); the extraction section may include: an extraction cell (11), a pressure gauge (12) connected to the extraction cell, a heating system consisting of a light emitting bulb (13), a fan (14) for homogenizing the temperature, and a precision thermometer connected to a temperature controller (15) for measuring the temperature, the outlet part may include: an outlet valve (16) of the extraction tank, an extract collector (17), a hot bath (18) and a flow meter (19); the extraction cell is made of the stainless steel alloy described above; the process conditions are as follows: 25 g of dried and crushed (400 +/-25 mu m) snow lotus herb is put into an ultrasonic extraction tank, the extract is extracted at 40 ℃ and 10.34MPa, each batch of extraction is carried out for 180 minutes, the carbon dioxide discharge flow rate is 125.25 +/-17.22 mL/min, and the tank for collecting the extract is soaked in ice water bath (8 +/-0.5 ℃). The detection shows that the content of the volatile oil in the saussurea involucrate extract1.0 wt%, and the total content of rutin, chlorogenic acid, syringin and dehydrocostuslactone is 1.1 wt%. Most preferably, the total content of lactone compounds having a parent nucleus represented by formula (a) in the saussurea involucrate extract is 81.1 wt%.
Comparative example 1
The supercritical extraction was carried out according to the method described in the example of CN 102106890B. Through detection, the content of the volatile oil in the saussurea involucrate extract is 2.3 wt%, and the total content of rutin, chlorogenic acid, syringin and dehydrocostuslactone is 4.3 wt%. Most preferably, the total content of lactone compounds having a parent nucleus represented by formula (a) in the saussurea involucrate extract is 77.3 wt%.
As can be seen from the above examples and comparative examples, the extraction apparatus of the present invention can significantly reduce the content of undesired components with a slight increase in the total content of lactone compounds.
Example 2
The saussurea involucrate extract of example 1 was tested for its inhibitory effect on secondary damage to adjuvant-induced arthritis.
Selecting 60 Wistar rats with half male and half female, body weight of 150-200 g, half male and half female, randomly dividing into 3 groups, namely normal control group, and conventional extract group (0.05g kg. per cent-1Control group, in which saussurea involucrate extract was obtained by the method described in CN 101041653B), example 1 extract group (0.05 g/kg)-1) (ii) a The volume of the left hind paw of each group of rats was measured as a baseline by toe volume measurement. Except for the normal control group, Freund's complete adjuvant is injected subcutaneously to the right hind foot sole of each group by 0.1mL, the administration groups of 12d after inflammation begin to administrate the stomach, and the normal control group and the model group administrate the same amount of distilled water for 8d continuously. The volume of the left hind sole (non-injected toe) was measured to 20d on alternate days in each group after dosing using a toe volumeter. The toe swelling degree of each group of animals was calculated.
Compared with a model group and a control group, the saussurea involucrate extract in example 1 has significant difference on the inhibition effect of adjuvant arthritis secondary damage, which shows that the compound saussurea involucrate capsule of the invention can effectively reduce the adjuvant arthritis secondary damage and has significant anti-inflammatory effect, even under the condition of low dose, the compound saussurea involucrate capsule has significant inhibition effect along with the prolonging of the administration time, and the compound saussurea involucrate capsule is shown in the following table 1:
table 1: saussurea involucrata extract for inhibiting adjuvant-induced arthritis secondary damage
As is clear from the above examples, the saussurea involucrate extract of the present invention has a good inhibitory effect, particularly a significantly better anti-inflammatory effect, than the extract obtained by the conventional supercritical extraction method.
Example 3
Preparation of capsules containing saussurea involucrate extract, epimedium extract and rhodiola rosea extract, wherein the saussurea involucrate extract described in example 1, the epimedium extract and rhodiola rosea extract described above are taken, the weight ratio of the saussurea involucrate extract, the epimedium extract and the rhodiola rosea extract is 8:1:1, and each capsule contains 0.1g of content. The capsules were tested for inhibition of secondary damage to adjuvant arthritis according to the method of example 2, and the degree of swelling (ml) of toes on day 3 after inflammation was 0.86 ± 0.11 at the same dose and experimental conditions.
It can be seen from the same results in examples 2 and 3 that the addition of the epimedium extract and the rhodiola root extract enhances the anti-inflammatory effect of the saussurea involucrate extract, and further has better effects of improving immunity and resisting fatigue. The reason is presumed that, besides the epimedium extract and the rhodiola root extract have certain anti-inflammatory effect, the three may generate certain mutual synergistic and promoting effect.
This written description uses examples to disclose the invention, including the best mode, and also to enable any person skilled in the art to make and use the invention. The patentable scope of the invention is defined by the claims, and may include other examples that occur to those skilled in the art. Such other examples are intended to be within the scope of the claims if they have structural elements that do not differ from the literal language of the claims, or if they include equivalent structural elements with insubstantial differences from the literal languages of the claims. All citations referred to herein are incorporated herein by reference to the extent that no inconsistency is made.
Claims (1)
1. A preparation method of a saussurea involucrate epimedium and rhodiola rosea composition capable of improving immunity, resisting inflammation and preventing radiation comprises saussurea involucrate extract and rhodiola rosea extract, wherein the weight ratio of the saussurea involucrate extract to the epimedium extract to the rhodiola rosea extract is (10-5) to (1-3);
the saussurea involucrate extract is obtained by using an extraction apparatus comprising: an inlet portion, an extraction portion, and an outlet portion, the inlet portion comprising: carbon dioxide supply tank, valve connected to pressure meter, compressor, carbon dioxide storage vessel, high pressure pump, coil and CO towards extraction tank2An inlet valve; the technological conditions for extracting saussurea involucrate are as follows: placing dried and pulverized herba Saussureae Involueratae in an ultrasonic extraction tank, extracting at 40 deg.C and 10.34MPa for 120-180 min each time, with carbon dioxide discharge rate of 125.25 + -15.00 m L/min, and soaking the tank in ice-water bath at 8 + -0.5 deg.C; the content of volatile oil in the snow lotus extract is lower than 1.2 wt%, the total content of rutin, chlorogenic acid, syringin and dehydrocostuslactone in the snow lotus extract is lower than 2.5 wt%, and the total content of lactone compounds of parent nucleus shown in the following formula (a) in the snow lotus extract is higher than 65.0 wt%:
the purity of glucoside substances in the epimedium extract is more than 99.0 wt%;
the glucoside substances are a mixture of compounds shown in formulas (I) and (II):
the weight ratio of the compound shown in the formula (I) to the compound shown in the formula (II) is 1:5-5: 1;
the rhodiola rosea extract is a compound which is obtained by modifying salidroside and is shown as the following formula (III):
the compound shown in the formula (III) is synthesized by the following method: adding an MCM-22 molecular sieve as a selective catalyst into a reaction vessel, then adding a chloroform solvent, then adding natural salidroside and chlorinated acetyl chloride into the reaction solvent according to the molar ratio of 1:1-1.3, wherein the reaction temperature is 20-30 ℃, the reaction time is 10-30min, and separating and drying to obtain the compound shown in the formula (III);
wherein the compounds of formula (I) and formula (II) are extracted by the following method:
(1) extracting the powder of Epimedium herb with MeOH at room temperature, and evaporating the combined extracts under reduced pressure to obtain a brown residue, i.e., a MeOH extract;
(2) using a reverse phase silica gel column, first MeOH-H was used2Performing gradient elution on the residue by using an O mixed solution to perform column chromatography, wherein the elution steps comprise 0:1 to 1L, 1:9 to 1L, 2:8 to 1L, 3:7 to 1L, 4:6 to 1L, 5:5 to 2L, 6:4 to 2L, 7:3 to 1L and 1:0 to 2L, and then using acetone 2L to sequentially obtain 10 elution components which are marked as elution components A-J;
(3) chromatographing the eluted fraction B on a column of silica gel using CHCl3-MeOH-H2Carrying out gradient elution on the O mixed solution, wherein the elution gradient is 20:1:0 to 1L, 10:1:0 to 2L, 30:8:1 to 2L and 15:6:1 to 2.5L, so as to obtain 4 elution components marked as elution components B1-B4;
(4) elute fraction B2 with 1:1 MeOH-H2Separating O on Sephadex LH-20 to obtain 7 secondary elution groupsDivide, and mark as elution component B2.1-B2.7;
(5) the mixture of eluted fractions B2.5 and B2.6 was passed through a silica gel CC column eluting with 12:6:3:1 EtOAc-CHCl3-MeOH-H2And O, concentrating the eluent under reduced pressure to dryness to obtain light yellow powder, namely the mixture of the compounds shown in the formula (I) and the formula (II).
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