CN103432214A - Preparation method and application of effective components of polygonum capitatum - Google Patents
Preparation method and application of effective components of polygonum capitatum Download PDFInfo
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- CN103432214A CN103432214A CN2013104077436A CN201310407743A CN103432214A CN 103432214 A CN103432214 A CN 103432214A CN 2013104077436 A CN2013104077436 A CN 2013104077436A CN 201310407743 A CN201310407743 A CN 201310407743A CN 103432214 A CN103432214 A CN 103432214A
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
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Abstract
The invention relates to a preparation method and application of effective components of polygonum capitatum, and belongs to the field of extraction of effective components of traditional Chinese medicines. The preparation method comprises the following steps: a, taking polygonum capitatum medicinal raw material, adding water for extraction, so as to obtain water extracting solution; b, processing the water extracting solution by using a macroporous adsorption resin chromatographic column; c, firstly eluting the sampled macroporous adsorption resin chromatographic column by using water, then eluting by using 70-95wt% ethanol, and collecting eluent I; d, processing the eluent I by a polyamide chromatographic column; e, recovering effluent liquid from the sampled polyamide chromatographic column, eluting by using 40-95wt% ethanol, collecting eluent II; f, mixing the effluent liquid and the eluent II, recovering ethanol, concentrating and drying to finally obtain the effective components of polygonum capitatum. The preparation method has the beneficial effect that the effective components of the polygonum capitatum, which has strong antibacterial effect and can be used for preparing anti-infection drugs can be prepared by reasonable extracting and separating methods.
Description
Technical field
The invention belongs to the Chinese medicine active component and extract field, be specifically related to a kind of preparation method and purposes of Herba Polygoni Capitati active component.
Background technology
Herba Polygoni Capitati (Polygonum capitatum) has another name called Herba Polygoni Capitati, Herba Polygoni Capitati, the red acid bar, province's fourth grass, belong to Polygonaceae (Polygonaceae) Polygonum (Polygonum) herbaceos perennial, main product Guizhou, the Guizhou genuine medicinal materials, record in version " Guizhou Province's Chinese medicine in 2003, ethnic drug quality standard ", there is heat-clearing and toxic substances removing, dissipating blood stasis, the effect of inducing diuresis for treating stranguria syndrome, be mainly used in clinically urinary system infection, the treatment of the diseases such as urinary system calculus, to urgency, chronic urethritis, cystitis, pyelonephritis has good therapeutic effect, clinical safe to use (Miao Ethnomedicine-Herba Polygoni Capitati Review Study. the Chinese Hospitals medication is estimated and is analyzed 2005, 5 (6): 383-384).At present, built vertical normalized Herba Polygoni Capitati GAP planting base, Guizhou Province, take Herba Polygoni Capitati as crude drug a plurality of Chinese medicine preparation of having developed and gone on the market, relate to many enterprises, formed a larger Herba Polygoni Capitati industry, Herba Polygoni Capitati has become one of kind of Guizhou Province's " six large Miao Ethnomedicine " and Guizhou Province's modernization of Chinese medicine emphasis cultivation and development.
The Patents and the research report that now relate to Herba Polygoni Capitati are existing a plurality of, have document with 15 times of amount 55% ethanol to the extraction process of total flavones in Herba Polygoni Capitati studied (Herba Polygoni Capitati total flavone extracting process research. the time precious traditional Chinese medical science traditional Chinese medicines, 2009,20 (11): 2815-2816).Chinese patent application prospectus CN1481832A discloses preparation method and the purposes of Herba Polygoni Capitati water and alcohol extract; CN1570134A discloses Herba Polygoni Capitati and extract and goods microbial activity assay method; CN102441041 discloses the method that the Herba Polygoni Capitati water extract prepares oral formulations; CN102526219 discloses a kind of water extract and has prepared the method for Herba Polygoni Capitati extract with the 75-85% ethanol extraction; CN202772497 discloses a kind of method for preparing Herba Polygoni Capitati extract with the 50-90% alcohol reflux; CN102526218 discloses a kind of water extract and has prepared the method for Herba Polygoni Capitati extract with the 76-84% ethanol extraction.Yet, existing these documents just are studied or apply for the preparation method of Herba Polygoni Capitati extract, perhaps only water extract is wherein extracted with the high concentration alcoholic solution, and these extract ingredient complexity of Herba Polygoni Capitati, often effective and invalid components coexists, and is unfavorable for the performance of drug effect.CN101940625 discloses a kind of water or water-alcohol extraction is used the preparation method of the component of high concentration alcohol eluting acquisition after macroporous resin water or low concentration alcohol eluting, has certain progressive.But, from our research, this method not take and is removed the Galla Turcica (Galla Helepensis) acids and the tannin constituents is purpose, there is no relevant quality control method, obtained component is owing to containing tannins and 2% above gallic acid and analog thereof, be unfavorable for the performance of Herba Polygoni Capitati drug effect, the preparation technology of active component still needs and further optimizes.
Summary of the invention
The preparation method that the purpose of this invention is to provide a kind of Herba Polygoni Capitati active component.
Another object of the present invention is to provide the purposes of above-mentioned Herba Polygoni Capitati active component.
Herba Polygoni Capitati active component of the present invention, its preparation process comprises the following steps:
A, get medicinal material of polygonum capilalum, add water and extracted, obtain aqueous extract;
B, by macroporous adsorption resin chromatography post on described aqueous extract;
Macroporous adsorption resin chromatography post after c, loading, first wash with water, then use 70%～95% weight ratio ethanol elution, collects the eluent I;
D, by polyamide chromatography post on described eluent I;
Polyamide chromatography post after e, loading, reclaim effluent, then use 40%～95% weight ratio ethanol elution, collects the eluent II;
F, described effluent and eluent II are merged after, reclaim ethanol, concentrated, drying, obtain the Herba Polygoni Capitati active component.
As preferably, in described step a, the number of times of extracting in water is 1～4 time, and each time of extracting is 0.5～4 hour, and each amount of water extracted is 5～20 times of described medicinal material of polygonum capilalum weight.
As preferably, in described step b, macroporous adsorbent resin is a kind of in D101, D201, H-103, SIP-1300, D1, D2, Diaion HP20, HPD100, HPD400, AB-8, MD, DM130, CAD-40, SP820 or SP700.
As preferably, be effectively to remove the gallic acid constituents, wash the macroporous adsorption resin chromatography post after loading in described step c with water, be washed till water elution liquid FeCl
3reaction is aobvious negative; For effective ingredient such as effective eluting flavones ingredients, in described step c with 70%～95% weight ratio ethanol elution to the hydrochloric acid of eluent I-aobvious feminine gender of magnesium powder reaction.
As preferably, be effectively to remove tannins, in described steps d, polyamide is a kind of in chinlon 6, chinlon 66, chinlon 46, chinlon 11 or chinlon 1010.
As preferably, be the effective effective ingredient such as eluting flavones ingredient, with the polyamide chromatography post after 40%～95% weight ratio ethanol elution loading, be washed till the hydrochloric acid of eluent II-aobvious feminine gender of magnesium powder reaction in described step e.
As preferably, a kind of in concentrated of simmer down to concentrating under reduced pressure or normal pressure in described step f; In described step f, drying is a kind of in drying under reduced pressure, spray drying or lyophilization.
A kind of Herba Polygoni Capitati active component is the pharmaceutical formulation that active component is made, and the preparation method that described Herba Polygoni Capitati active component adds medical additive to allow according to pharmaceutics is made pharmaceutical formulation.
The application of a kind of Herba Polygoni Capitati active component in the preparation antibacterials.
Further, described antibiotic be that the urinary system infection common bacteria such as escherichia coli, staphylococcus aureus, proteus mirabilis and Pseudomonas aeruginosa are had to resistant function.
Prepared in the Herba Polygoni Capitati active component by the present invention, using and the most easily obtain also safe water as extracting solvent, by after twice column chromatography simple to operate, obtaining the Herba Polygoni Capitati active component.Wherein, at first the aqueous extract of Herba Polygoni Capitati by nonpolar or low pole macroporous adsorption resin chromatography post, in the enrichment flavones ingredient, has removed the gallic acid constituents; Go up again polyamide chromatography post, through 40%～95% weight ratio ethanol elution, obtain removing the active component that tannin contains a large amount of flavones ingredients.
In the present invention, FeCl
3it is complete whether reaction removes for the gallic acid constituents of qualitative detection water elution liquid, while showing feminine gender, illustrates fully and remove.Hydrochloric acid-magnesium powder reacts the flavones ingredient that is used for qualitative detection eluent II, and whether eluting is complete, while showing feminine gender, illustrates that eluting is complete containing flavones ingredient.
Herba Polygoni Capitati active component prepared by the present invention has stronger antibacterial action, can be used for treating the diseases such as urinary system infection.For the activity that proves the Herba Polygoni Capitati active component that the present invention is prepared and the necessity of preparation method, the applicant has carried out following experiment.
Respectively component, tannin class component and Galla Turcica (Galla Helepensis) acids component and the active component of the present invention of only removing Galla Turcica (Galla Helepensis) acids component in water extract described in embodiment, water extract have been carried out to antibacterial research, the antibacterial activity of finding active component of the present invention is remarkable, can be used in the diseases such as treatment urinary system infection.
Beneficial effect of the present invention is:
(1) active component preparation method of the present invention, except last eluent concentrated all without concentrating, can greatly raise the efficiency, adopt present technique to obtain to take the active component that flavones ingredient is main component, can be for the relevant pharmacology of Herba Polygoni Capitati and drug efficacy study.
(2) preparation method of the present invention, utilize reasonably extraction, separation means, according to the construction features of gallic acid compounds, flavone compound and tannins and the difference on physicochemical property, remove Galla Turcica (Galla Helepensis) acids component and tannin class component in Herba Polygoni Capitati, improved widely drug effect.
The specific embodiment
For making those skilled in the art understand in detail production technology of the present invention and technique effect, below with concrete production instance, further introduce application of the present invention and technique effect.
Embodiment mono-:
The 2kg Herba Polygoni Capitati is pulverized, decocted respectively 2 times with 10 times of water gagings, each time decocted is 2 hours, merges extracted twice liquid, obtains aqueous extract;
By after D101 macroporous adsorption resin chromatography post on aqueous extract, wash with water, to water elution liquid FeCl
3reaction aobvious negative (TLC(chloroform/methanol/formic acid=9/1/0.1), gallic acid constituents and FeCl
3aobvious black-and-blue).Collect water elution liquid, add 60% weight ratio ethanol, polysaccharide and protein in precipitation water elution liquid, concentrated, drying, obtain the gallic acid constituents.
After water elution, on D101 macroporous adsorption resin chromatography post, remaining part is used 80% weight ratio ethanol elution again, to hydrochloric acid-magnesium powder reaction aobvious negative (it is aobvious red that flavones ingredient reacts with hydrochloric acid-magnesium powder), collects the eluent I.
By the 6 polyamide chromatography posts of chinlon on described eluent I, reclaim effluent, then use 90% weight ratio ethanol elution, to hydrochloric acid-magnesium powder reaction aobvious negative (it is aobvious red that flavones ingredient reacts with hydrochloric acid-magnesium powder), collect the eluent II; By after described effluent and the merging of eluent II, reclaim ethanol, concentrating under reduced pressure, drying under reduced pressure, obtain the Herba Polygoni Capitati active component.
Chinlon 6 polyamide chromatography posts 100% acetone eluting for remaining part, collect this eluent and reclaim acetone, is concentrated into extractum and obtains tannin class component.
Embodiment bis-:
The 2kg Herba Polygoni Capitati is pulverized, decocted respectively 3 times with 12 times of water gagings, each time decocted is 1.5 hours, merges extracting solution three times, obtains aqueous extract;
By after D201 macroporous adsorption resin chromatography post on aqueous extract, wash with water, to water elution liquid FeCl
3reaction aobvious negative (TLC(chloroform/methanol/formic acid=9/1/0.1), gallic acid constituents and FeCl
3aobvious black-and-blue).
After water elution, on D201 macroporous adsorption resin chromatography post, remaining part is used 70% weight ratio ethanol elution again, to hydrochloric acid-magnesium powder reaction aobvious negative (it is aobvious red that flavones ingredient reacts with hydrochloric acid-magnesium powder), collects the eluent I.
By the 66 polyamide chromatography posts of chinlon on described eluent I, reclaim effluent, then use 95% weight ratio ethanol elution, to hydrochloric acid-magnesium powder reaction aobvious negative (it is aobvious red that flavones ingredient reacts with hydrochloric acid-magnesium powder), collect the eluent II; By after described effluent and the merging of eluent II, reclaim ethanol, concentrating under reduced pressure, spray drying, obtain the Herba Polygoni Capitati active component.
Embodiment tri-:
The 2kg Herba Polygoni Capitati is pulverized, decocted respectively 3 times with 8 times of water gagings, each time decocted is 2 hours, merges extracting solution three times, obtains aqueous extract;
By after AB-8 macroporous adsorption resin chromatography post on aqueous extract, wash with water, to water elution liquid FeCl
3reaction aobvious negative (TLC(chloroform/methanol/formic acid=9/1/0.1), gallic acid constituents and FeCl
3aobvious black-and-blue).
After water elution, on AB-8 macroporous adsorption resin chromatography post, remaining part is used 95% weight ratio ethanol elution again, to hydrochloric acid-magnesium powder reaction aobvious negative (it is aobvious red that flavones ingredient reacts with hydrochloric acid-magnesium powder), collects the eluent I.
By the 11 polyamide chromatography posts of chinlon on described eluent I, reclaim effluent, then use 60% weight ratio ethanol elution, to hydrochloric acid-magnesium powder reaction aobvious negative (it is aobvious red that flavones ingredient reacts with hydrochloric acid-magnesium powder), collect the eluent II; By after described effluent and the merging of eluent II, reclaim ethanol, concentrating under reduced pressure, spray drying, obtain the Herba Polygoni Capitati active component.
For only removing the antibacterial action of component, tannin class component and Galla Turcica (Galla Helepensis) acids component and the active component of the present invention of Galla Turcica (Galla Helepensis) acids component in research water extract, water extract, this experiment adopts agar diffusion method of the paper to detect its extract and the antibacterial action of each component to the urinary system infection common bacteria.
Experiment material
1, bacterium is used in experiment: staphylococcus aureus, escherichia coli, Pseudomonas aeruginosa, proteus mirabilis.
2, the preparation of test organisms liquid: on the MHA plate, picking colony is seeded in the 5mLMH broth bouillon, cultivate 4～5 hours through 36 ℃, than turbid to 0.5 Maxwell unit (approximately containing 1～2 * 10
8cFU/mL).
3, the preparation of the pastille scraps of paper: medicinal liquid to be checked is added respectively in 30 aseptic roundlet filter papers (diameter 6mm) to the 0.3mL/ ware.
Experimental technique and result
Experimental technique:
Agar diffusion method of the paper.From the MHA plate, picking staphylococcus aureus, escherichia coli, Pseudomonas aeruginosa, proteus mirabilis bacterium colony are inoculated in respectively in the 5mLMH broth bouillon in 36 ℃ and cultivate 4～5h, than turbid to 0.5 Maxwell unit (approximately containing 1～2 * 10
8cFU/mL).To be inoculated on the MHA culture medium that is placed in plate than turbid good bacterium liquid three plates of each inoculation.Postvaccinal plate is put room temperature for a moment, after slightly dry, with aseptic ophthalmic tweezers, the pastille scraps of paper is affixed on to agar surface.Each scraps of paper centre distance > 24mm, the scraps of paper and plate inner edge distance > 15mm, plate is placed in to 36 ℃ and cultivates 18～24h, measure the size of inhibition zone.
Result:
The results are shown in Table 1.The antibacterial activity of Galla Turcica (Galla Helepensis) acids component the most weak (there is no activity under the survey condition).And, in water extract, only remove component after the Galla Turcica (Galla Helepensis) acids to the antibacterial activity of staphylococcus aureus, escherichia coli, Pseudomonas aeruginosa and proteus mirabilis strong than water extract; The antibacterial activity of active component of the present invention is only removed the antibacterial activity of component after the Galla Turcica (Galla Helepensis) acids and tannin class component apparently higher than water extract, illustrate that tannin class component and Galla Turcica (Galla Helepensis) acids component are mixed in the water extraction species, are unfavorable for the antibacterial action of Herba Polygoni Capitati.Compared to the tannin component, the antibacterial action of active component of the present invention is stronger.
The antibacterial circle diameter (mm) of prepared each component of table 1 the present invention to strain subject
In experiment, pastille scraps of paper diameter is 7mm; The concentration of gentamycin is 1.3mgmL
-1, the concentration of all the other medicinal liquids is 30mgmL
-1.Water extract is that described in embodiment mono-, aqueous extract is through concentrating under reduced pressure, and drying makes; Only removing component after the Galla Turcica (Galla Helepensis) acids in water extract is that described in embodiment mono-, the eluent I is through concentrating under reduced pressure, and drying makes; Tannin class component is the tannin class component described in embodiment mono-; Active component is the Herba Polygoni Capitati active component described in embodiment mono-; The gallic acid constituents is the gallic acid constituents described in embodiment mono-.
Finally it should be noted that, above embodiment is the unrestricted technical scheme of the present invention in order to explanation only, although with reference to above-described embodiment, the present invention is had been described in detail, those skilled in the art are to be understood that, still can modify or be equal to replacement the present invention, and not breaking away from any modification or partial replacement of the spirit and scope of the present invention, it all should be encompassed in claim scope of the present invention.
Claims (10)
1. the preparation method of a Herba Polygoni Capitati active component, is characterized in that, comprises the following steps:
A, get medicinal material of polygonum capilalum, add water and extracted, obtain aqueous extract;
B, by macroporous adsorption resin chromatography post on described aqueous extract;
Macroporous adsorption resin chromatography post after c, loading, first wash with water, then use 70%～95% weight ratio ethanol elution, collects the eluent I;
D, by polyamide chromatography post on described eluent I;
Polyamide chromatography post after e, loading, reclaim effluent, then use 40%～95% weight ratio ethanol elution, collects the eluent II;
F, described effluent and eluent II are merged after, reclaim ethanol, concentrated, drying, obtain the Herba Polygoni Capitati active component.
2. the preparation method of Herba Polygoni Capitati active component according to claim 1, it is characterized in that: in described step a, the number of times of extracting in water is 1～4 time, each time of extracting is 0.5～4 hour, and each amount of water extracted is 5～20 times of described medicinal material of polygonum capilalum weight.
3. the preparation method of Herba Polygoni Capitati active component according to claim 1 is characterized in that: in described step b, macroporous adsorbent resin is a kind of in D101, D201, H-103, SIP-1300, D1, D2, Diaion HP20, HPD100, HPD400, AB-8, MD, DM130, CAD-40, SP820 or SP700.
4. the preparation method of Herba Polygoni Capitati active component according to claim 1 is characterized in that: wash the macroporous adsorption resin chromatography post after loading in described step c with water, be washed till water elution liquid FeCl
3reaction is aobvious negative; Aobvious negative to the hydrochloric acid of eluent I-magnesium powder reaction with 70%～95% weight ratio ethanol elution in described step c.
5. the preparation method of Herba Polygoni Capitati active component according to claim 1 is characterized in that: in described steps d, polyamide is a kind of in chinlon 6, chinlon 66, chinlon 46, chinlon 11 or chinlon 1010.
6. the preparation method of Herba Polygoni Capitati active component according to claim 1 is characterized in that: in described step e with the polyamide chromatography post after 40%～95% weight ratio ethanol elution loading, be washed till the eluent II hydrochloric acid-the magnesium powder reaction is aobvious negative.
7. the preparation method of Herba Polygoni Capitati active component according to claim 1 is characterized in that: a kind of in concentrated of simmer down to concentrating under reduced pressure or normal pressure in described step f; In described step f, drying is a kind of in drying under reduced pressure, spray drying or lyophilization.
8. one kind is utilized the pharmaceutical formulation that in claim 1-7, the described Herba Polygoni Capitati active component of any one is made for active component, it is characterized in that: the preparation method that described Herba Polygoni Capitati active component adds medical additive to allow according to pharmaceutics is made pharmaceutical formulation.
9. one kind is utilized the application of the described Herba Polygoni Capitati active component of any one in the preparation antibacterials in claim 1-7.
10. Herba Polygoni Capitati active component application in antibacterials in preparation according to claim 9 is characterized in that: described antibiotic be that escherichia coli, staphylococcus aureus, proteus mirabilis and Pseudomonas aeruginosa are had to resistant function.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104447720A (en) * | 2014-12-12 | 2015-03-25 | 东莞广州中医药大学中医药数理工程研究院 | Method for separating vicenin-2 from linearstripe rabdosia herb |
| CN104614480A (en) * | 2015-02-04 | 2015-05-13 | 东莞广州中医药大学中医药数理工程研究院 | Water-soluble general flavones of rabdosia lophanthide and fingerprint chromatographic detection method of water-soluble general flavones of rabdosia lophanthide |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104447720A (en) * | 2014-12-12 | 2015-03-25 | 东莞广州中医药大学中医药数理工程研究院 | Method for separating vicenin-2 from linearstripe rabdosia herb |
| CN104447720B (en) * | 2014-12-12 | 2017-01-18 | 东莞广州中医药大学中医药数理工程研究院 | Method for separating vicenin-2 from linearstripe rabdosia herb |
| CN104614480A (en) * | 2015-02-04 | 2015-05-13 | 东莞广州中医药大学中医药数理工程研究院 | Water-soluble general flavones of rabdosia lophanthide and fingerprint chromatographic detection method of water-soluble general flavones of rabdosia lophanthide |
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|---|---|
| CN103432214B (en) | 2015-07-01 |
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