CN101502550B - Filifolium sibiricum effective component extract as well as preparation method and use thereof - Google Patents
Filifolium sibiricum effective component extract as well as preparation method and use thereof Download PDFInfo
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Abstract
The invention discloses a filifolium sibiricum effective part extract, a preparation method and an application thereof, belonging to the field of Chinese herbal medicine effective part extracts. The preparation method comprises the following steps: a. using water or ethanol for reflux extraction of filifolium sibiricum for 2-4 times, filtering and merging filtrate; b. recycling solvent from the filtrate till the volume of filtrate is equal to 3-6 times of medicinal materials and obtaining liquid medicine 1; c. adding 10-50 percent ethanol into the liquid medicine 1 for suspension and then standing; d. taking supernatant liquid, to recycle the solvent and obtaining the liquid medicine 2; e. sampling the liquid medicine 2 to the chromatogram of a macroporous resin column for absorption, washing by water till nearly colorless, using 20-60 percent ethanol for elution, merging eluate and concentrating for preparation. The filifolium sibiricum effective part extract contains one effective part group and can effectively treat bacterial infectious diseases. The preparation process of the filifolium sibiricum effective part extract can be applicable to the industrial production, thereby providing sufficient raw materials for the follow-up research of total flavonoids of filifolium sibiricum and broadening the application range of the total flavonoids of filifolium sibiricum.
Description
Technical field
The present invention relates to the Chinese herbal medicine effective part extract, relate in particular to Filifolium sibiricum (Linn.) Kitam. effective part extract and preparation method thereof, the invention still further relates to the purposes of this Filifolium sibiricum (Linn.) Kitam. effective part extract in treatment antibacterial and drug resistant bacterial infections disease, belong to Chinese herbal medicine effective part extract field.
Background technology
Filifolium sibiricum (Linn.) Kitam. [Filifolium sibiricum (L.) Kitam.] has another name called rabbit hair Artemisia, Tanacetum sibiricum L., Siberia tansy, Herba Violae dissectae, frightened grass, JINGCAO etc., is Compositae Filifolium sibiricum (Linn.) Kitam. platymiscium.Belong to non-pharmacopeia and include natural drug.This medicine is a kind of heat and toxic materials clearing away medicine, and herb is all pharmaceutically acceptable.Its nature and flavor bitter cold has heat-clearing and toxic substances removing, anti-inflammation, and it is relieving convulsion to calm the nerves, the function of regulating menstruation, hemostasis.With the treatment of diseases such as its treatment infectiousness hyperpyrexia, upper respiratory tract infection, bigger researching value is arranged clinically.
Up to now, few to Filifolium sibiricum (Linn.) Kitam. effective part extract research report both at home and abroad, to Filifolium sibiricum (Linn.) Kitam. treatment bacterial infection disease effective substance and indeterminate.Domestic its The Chemical Constituents is started from the end of the seventies in last century, therefrom separate and identified volatile oil, several flavonoid chemical constituents.Wait report to extract volatile oil in the Filifolium sibiricum (Linn.) Kitam. with steam distillation as Zhang Zhen outstanding person, yield is about 0.52%, with the gas chromatography-mass spectrography technical appraisement 25 kinds of compositions, account for 55.6% of volatile oil total amount.The antibacterial action of Filifolium sibiricum (Linn.) Kitam. decocting liquid has the research report in Medical Colleges of the Inner Mongol, Qiqihar Medical College.From Filifolium sibiricum (Linn.) Kitam., get three flavone compounds, be respectively filifolin, eriodictyol and quercetagetin 3, the 6-dimethyl ether.But all the antimicrobial effective site of Filifolium sibiricum (Linn.) Kitam. is not carried out more careful, deep research.The extraction process of Filifolium sibiricum (Linn.) Kitam. total flavones does not still have mature technique.
Summary of the invention
One of the object of the invention provides a kind of Filifolium sibiricum (Linn.) Kitam. effective part extract;
Two of the object of the invention provides a kind of method for preparing above-mentioned Filifolium sibiricum (Linn.) Kitam. effective part extract;
Three of the object of the invention provides a kind of pharmaceutical composition of inhibiting bacteria and diminishing inflammation;
Above-mentioned purpose of the present invention is achieved through the following technical solutions:
Filifolium sibiricum (Linn.) Kitam. effective part extract of the present invention contains 1 effective part group, comprise flavone compound, be mainly filifolin (filifolin), eriodictyol (eriodictyol) and quercetagetin 3,6-dimethyl ether (3,6-dimethoxyquercet-agetin), Quercetin (Quercetindihydrate), isoquercitrin (isoquercitrin) etc.The total flavonoid compounds content that comprises these 5 chromocor compounds in the extract of the present invention accounts for more than 60% of total extract weight;
Filifolium sibiricum (Linn.) Kitam. extractive of general flavone of the present invention is measured through high performance liquid chromatogram-uv detection method (UV-HPLC), and each composition of contained flavonoid occupies the percentage ratio of imitating the extractive part gross weight and is respectively: filifolin 10.26-20.26% is preferably 15.26%; Eriodictyol 3.6-7.02% is preferably 5.31%; Quercetagetin 3,6-dimethyl ether 2.43-5.89% is preferably 4.16%; Quercetin 6.87-13.47% is preferably 10.17%; Isoquercitrin 6.96-13.59% is preferably 10.29%.
A kind of method for preparing above-mentioned Filifolium sibiricum (Linn.) Kitam. effective part extract may further comprise the steps:
A. with Filifolium sibiricum (Linn.) Kitam. water or alcohol reflux 2～4 times, filter merging filtrate;
B. filtrate is reclaimed the volume of solvent to 3～6 times of Filifolium sibiricum (Linn.) Kitam. medical materials, gets medicinal liquid 1;
C. after medicinal liquid 1 being added the suspension of 10～50% ethanol, leave standstill;
D. get supernatant and reclaim solvent, obtain medicinal liquid 2;
E. sample on the medicinal liquid 2 is adsorbed to macroporous adsorbent resin column chromatography, after the washing,, merge eluent, concentrate, promptly with 20%～60% ethanol elution.
In the above-mentioned preparation method, among the step a: extract preferably 5～20 times of Filifolium sibiricum (Linn.) Kitam. medical material weight of the water of Filifolium sibiricum (Linn.) Kitam. or alcoholic acid weight; Described reflux extracting time is preferably each reflux, extract, 1-3 hour; In order to reach better technique effect, more preferably be to extract solvent Filifolium sibiricum (Linn.) Kitam. is carried out reflux, extract, with ethanol, wherein, described concentration of alcohol is preferably 30～100wt%, more preferably 40～80wt%;
Time of repose described in the step c can be 12～48 hours etc.;
Macroporous adsorbent resin column chromatography described in the step e can be selected from D101, AB-8, HPD500, HPD300, macroporous adsorbent resin column chromatography such as N KA or H103; Described go up sample preferably flow velocity for the condition of 1.1～1.3 times of column volumes per hour under with medicinal liquid 2 on sample adsorb to macroporous adsorbent resin column chromatography;
Preferably carry out eluting among the step e, further preferably carry out eluting with 30% ethanol with 20～40% ethanol; The flow velocity of eluent is preferably per hour 0.5～0.6 times of column volume.
Filifolium sibiricum (Linn.) Kitam. of the present invention is the herb of Compositae Filifolium sibiricum (Linn.) Kitam. platymiscium Filifolium sibiricum (Linn.) Kitam. [Filifoliumsibiricum (L.) Kitam.].
Extracorporeal bacteria inhibitor test proves, Filifolium sibiricum (Linn.) Kitam. effective part extract of the present invention (total flavones) has highly antibacterial and bactericidal action to golden yellow and Staphylococcus albus, and penicillin drug resistance staphylococcus aureus is still had good inhibitory effect, Bacillus typhi, Bacillus proteus, dysentery bacterium, bacillus pyocyaneus are all had certain antibacterial and bactericidal action.Experimental animal models such as the bacillary keratoconjunctivitis of rabbit, rabbit peritonitis, experimental septicemia, mouse experiment staphy lococcus infection, local infection there is therapeutical effect, experimental results show that total flavones is a Filifolium sibiricum (Linn.) Kitam. anti-infective effective site.LD is not measured in this position oral administration administration
50, measure proof through maximum dosage-feeding, mice oral administration maximum dosage-feeding is 198.0g/kg, 69300 times of quite clinical adult's consumption.
Can obtain a kind of pharmaceutical composition for the treatment of bacterial infection disease after the Filifolium sibiricum (Linn.) Kitam. effective part extract of the present invention of effective dose and pharmaceutically acceptable carrier or adjuvant combined.
After Filifolium sibiricum (Linn.) Kitam. effective part extract of the present invention can add various adjuvants and pharmaceutically acceptable excipient or carrier required when preparing different dosage form, method of Chinese medicinal with routine is prepared into any suitable clinical preparation, for example can be injection, tablet, oral liquid, granule, capsule, soft capsule, drop pill etc., wherein, described adjuvant can be antioxygen chelating agent, filler, framework material etc.; Described pharmaceutically acceptable carrier is one or more in xylitol, mannitol, lactose, fructose, glucosan, glucose, polyvinylpyrrolidone, low molecular dextran, sodium chloride, calcium gluconate or the calcium phosphate, is preferably mannitol or lactose.
Filifolium sibiricum (Linn.) Kitam. effective part extract of the present invention contains 1 effective part group, effectively treatment bacterial infection disease; The preparation technology of Filifolium sibiricum (Linn.) Kitam. effective part extract of the present invention can adapt to suitability for industrialized production, for the follow-up study of Filifolium sibiricum (Linn.) Kitam. total flavones furnishes ample material, has widened the range of application of Filifolium sibiricum (Linn.) Kitam. total flavones.
The specific embodiment
Further describe the present invention below in conjunction with specific embodiment, advantage of the present invention and characteristics will be more clear along with description.But these embodiment only are exemplary, scope of the present invention are not constituted any restriction.It will be understood by those skilled in the art that and down can make amendment or replace without departing from the spirit and scope of the present invention, but these modifications and replacing all fall within the scope of protection of the present invention the details of technical solution of the present invention and form.
Embodiment 1
Get Filifolium sibiricum (Linn.) Kitam. raw material 6000g, 50% ethanol extraction of 20 times of weight of first usefulness raw material 3 times, each 1.5 hours extraction times, merge said extracted liquid, decompression recycling ethanol obtains medicinal liquid 1 to being less than or equal to 180000ml;
Left standstill 1～2 day after medicinal liquid 1 stirred evenly, natural subsidence to interface no longer descends, and gets supernatant;
Supernatant is continued to reclaim solvent to there not being the alcohol flavor, add water and be settled to 6000ml, obtain medicinal liquid 2;
(resin weighs 80g to D101 type macroporous adsorbent resin column chromatography D101 with sample on the medicinal liquid 2, column volume 100ml, post footpath: post height=1: 2) adsorb, last sample flow velocity is 1.2 times of column volumes per hour, uses the distilled water flushing pillar behind the upward intact sample, and is closely colourless until water elution liquid, discard water elution liquid, contain 20% ethanol water eluting with 6 times of column volumes, eluent flow rate is controlled at per hour 0.5 times of column volume, uses Alcl in the elution process
3Reagent detects whether eluting is complete; Collect eluent, merge, concentrating under reduced pressure, drying under reduced pressure obtains Filifolium sibiricum (Linn.) Kitam. total flavones crude product 210g.
Above-mentioned Filifolium sibiricum (Linn.) Kitam. extractive of general flavone is measured through high performance liquid chromatogram-uv detection method (UV-HPLC), and the total flavonoid compounds content accounts for 66.27% of total extract gross weight; Wherein contained each composition of flavonoid occupies the percentage composition of imitating the extractive part gross weight and is respectively: filifolin (13.01%), eriodictyol (4.23%), quercetagetin 3,6-dimethyl ether (4.35%), Quercetin (9.16%), isoquercitrin (9.27%).
Embodiment 2
Line taking leaf Radix Chrysanthemi and rhizome coarse powder raw material 2500g, 50% ethanol extraction of 20 times of weight of first usefulness raw material 3 times, each 1.5 hours extraction times, merge said extracted liquid, decompression recycling ethanol obtains medicinal liquid 1 to being less than or equal to 75000ml;
Left standstill 1～2 day after medicinal liquid 1 stirred evenly, natural subsidence to interface no longer descends, and gets supernatant;
Supernatant is added water be settled to 2500ml, obtain medicinal liquid 2;
With (the heavy 80g of resin of AB-8 type macroporous adsorbent resin column chromatography on the medicinal liquid 2, column volume 100ml, post footpath: post height=1: 2), flow velocity is 1.3 times of column volumes per hour, uses the distilled water flushing pillar behind the upward intact sample, and is closely colourless until water elution liquid, discard water elution liquid, contain 30% ethanol elution with 6 times of column volumes, eluent flow rate is controlled at per hour 0.5 times of column volume, uses Alcl in the elution process
3Reagent detects whether eluting is complete; Collect eluent, merge, concentrating under reduced pressure, drying under reduced pressure obtains Filifolium sibiricum (Linn.) Kitam. total flavones crude product 175g.
Above-mentioned Filifolium sibiricum (Linn.) Kitam. extractive of general flavone is measured through high performance liquid chromatogram-uv detection method (UV-HPLC), and the total flavonoid compounds content accounts for 72.33% of total extract gross weight; Each composition of contained flavonoid occupies the percentage composition of imitating the extractive part gross weight and is respectively: filifolin (16.68%), eriodictyol (5.87%), quercetagetin 3,6-dimethyl ether (5.31%), Quercetin (11.65%), isoquercitrin (10.87%).
Embodiment 3
Line taking leaf Radix Chrysanthemi and rhizome coarse powder raw material 1000g, earlier with 95% ethanol extraction of 20 times of weight of raw material 3 times, each 1.5 hours extraction times, merge said extracted liquid, merge said extracted liquid, decompression recycling ethanol obtains medicinal liquid 1 to smaller or equal to 30000ml;
Left standstill 1～2 day after medicinal liquid 1 stirred evenly, natural subsidence to interface no longer descends, and gets supernatant;
Supernatant is continued to reclaim solvent to there not being the alcohol flavor, add water and be settled to 1000ml, obtain medicinal liquid 2;
With (the heavy 80g of resin of H103 type macroporous adsorbent resin column chromatography H103 on the medicinal liquid 2, column volume 100ml, post footpath: post height=1: 2), flow velocity is 1.1 times of column volumes per hour, uses the distilled water flushing pillar behind the upward intact sample, and is closely colourless until water elution liquid, discard water elution liquid, contain 60% ethanol elution with 6 times of column volumes, eluent flow rate is controlled at per hour 0.6 times of column volume, uses Alcl in the elution process
3Reagent detects whether eluting is complete; Collect eluent, merge, concentrating under reduced pressure, drying under reduced pressure obtains Filifolium sibiricum (Linn.) Kitam. total flavones crude product 58g.
Above-mentioned Filifolium sibiricum (Linn.) Kitam. extractive of general flavone is measured through high performance liquid chromatogram-uv detection method (UV-HPLC), and the total flavonoid compounds content accounts for 64.52% of total extract gross weight; Wherein contained each composition of flavonoid occupies the percentage composition of imitating the extractive part gross weight and is respectively: filifolin (15.03%), eriodictyol (5.32%), quercetagetin 3,6-dimethyl ether (4.89%), Quercetin (10.13%), isoquercitrin (10.56%).
Embodiment 4 makes injection
It is an amount of to get the prepared extract of embodiment 1, adds an amount of solubilizing agent, grinds, and adds a small amount of water for injection dilution again, mixing, and it is an amount of to add sodium chloride then, adds the injection water after the dissolving again to ormal weight, filter, embedding, sterilization, promptly.
Embodiment 5 makes oral liquid:
It is an amount of to get the prepared extract of embodiment 2, adds an amount of solubilizing agent, grinds, and adds the low amounts of water dilution again, and mixing adds correctives and antiseptic then, and mixing adds water to ormal weight.Mixing, packing, sterilization, promptly.
Embodiment 6 makes tablet:
It is an amount of to get the prepared extract of embodiment 3, adds right amount of auxiliary materials, and mixing is made granule, drying, compacting in flakes, promptly.
The investigation of sample flow velocity on the experimental example 1
Filifolium sibiricum (Linn.) Kitam. aqueous extract (the water reflux, extract, Filifolium sibiricum (Linn.) Kitam. with 5～20 times of weight of Filifolium sibiricum (Linn.) Kitam. medical material prepares for 2～4 times) (every 12.5ml extracting solution is equivalent to 1g extractum or 3g crude drug) is splined on AB-8 type macroporous resin column (the heavy 80g of resin, column volume 100ml, post footpath: in process post height=1: 2), adopt different flow velocitys respectively, detect with Liberman reagent, investigated the relation of last sample flow velocity and adsorbance, testing result sees Table 1.
Table 1
The result shows: sample solution flow velocity and applied sample amount are negative correlation, flow velocity is big more, adsorbance is more little, when considering flow velocity less than 1ml/min (1.1 times of column volumes), consuming time longer, and flow velocity is during greater than 3ml/min (1.3 times of column volumes), adsorbance has on the low side, the consumption amount of resin is big, adopt the flow velocity of 2ml/min during above sample, promptly go up the sample flow velocity and be 1.2 times of column volumes per hour.
The investigation of experimental example 2 eluting solvents
To exchange in AB-8 type macroporous adsorptive resins (the heavy 80g of resin, column volume 100ml, the post footpath: the Filifolium sibiricum (Linn.) Kitam. extracting solution post height=1: 2) (the described medicinal liquid 2 of embodiment 1-3), adopt following system to carry out eluting respectively: to contain Different concentrations of alcohol system: Alcl
3Reagent detects, and has investigated the elution efficiency of different elution systems, and elution flow rate adopts 2ml/min, and its result is as follows:
Elution system: respectively with containing 5%, 10%, 20%, 30%, 40%, 50% ethanol elution; 10%, 20% ethanol elution efficient does not have significant difference, has a brown colour band to be washed on the visible post of 20% ethanol, all total flavones can not be washed fully ALCL when being eluted to 10 times of column volumes
3Reacting positive, color of resin are still darker, are kermesinus.During 30% ethanol elution, total flavones can be washed fully Alcl during 10 times of column volumes
3Reagent reacting feminine gender, color of resin are still darker, are kermesinus.30% ethanol elution efficient height.
Claims (13)
1. method for preparing Filifolium sibiricum (Linn.) Kitam. [Filifolium sibiricum (L.) Kitam.] effective part extract may further comprise the steps:
A. water or alcohol reflux Filifolium sibiricum (Linn.) Kitam. are 2～4 times, filter merging filtrate;
B. filtrate is reclaimed the volume of solvent to 3～6 times of Filifolium sibiricum (Linn.) Kitam. medical materials, gets medicinal liquid 1;
C. after medicinal liquid 1 being added the suspension of 10～50% ethanol, leave standstill;
D. get supernatant and reclaim solvent, obtain medicinal liquid 2;
E. sample on the medicinal liquid 2 is adsorbed to macroporous adsorbent resin column chromatography, after the washing,, merge eluent, concentrate, promptly with 20～60% ethanol elutions.
2. it is characterized in that in accordance with the method for claim 1: the water of 5～20 times of weight of usefulness Filifolium sibiricum (Linn.) Kitam. medical material or alcohol reflux Filifolium sibiricum (Linn.) Kitam. are 2～4 times among the step a; Described reflux extracting time is each reflux, extract, 1-3 hour.
3. it is characterized in that in accordance with the method for claim 1: among the step a be to extract solvent Filifolium sibiricum (Linn.) Kitam. is carried out reflux, extract, with ethanol; Wherein, described concentration of alcohol is 30～100wt%.
4. in accordance with the method for claim 1, wherein, described concentration of alcohol is 40～80wt%.
5. in accordance with the method for claim 1, it is characterized in that: the macroporous adsorbent resin column chromatography described in the step e is selected from D101, AB-8, HPD500, HPD300, NKA or H103 macroporous adsorbent resin column chromatography.
6. it is characterized in that in accordance with the method for claim 1: the last sample described in the step e be flow velocity for the condition of 1.1～1.3 times of column volumes per hour under with medicinal liquid 2 on sample adsorb to macroporous adsorbent resin column chromatography.
7. it is characterized in that in accordance with the method for claim 1: carry out eluting with 30% ethanol among the step e; Wherein, the flow velocity of eluent is 0.5～0.6 times of column volume per hour.
8. Filifolium sibiricum (Linn.) Kitam. effective part extract according to the preparation of the described method of claim 1, it is characterized in that: the total flavonoid compounds content accounts for more than 60% of total extract gross weight in the described effective part extract; Wherein, described total flavonoid chemical compound comprises: filifolin, eriodictyol, quercetagetin 3,6-dimethyl ether, Quercetin and isoquercitrin.
9. according to the described Filifolium sibiricum (Linn.) Kitam. effective part extract of claim 8, it is characterized in that: filifolin occupies the 10.26-20.26% of imitating the extractive part gross weight; Eriodictyol occupies the 3.6-7.02% of imitating the extractive part gross weight; Quercetagetin 3,6-dimethyl ether occupy the 2.43-5.89% of imitating the extractive part gross weight; Quercetin occupies the 6.87-13.47% of imitating the extractive part gross weight; Isoquercitrin occupies the 6.96-13.59% of imitating the extractive part gross weight.
10. according to the described Filifolium sibiricum (Linn.) Kitam. effective part extract of claim 9, it is characterized in that: filifolin occupies imitates 15.26% of extractive part gross weight; Eriodictyol occupies imitates 5.31% of extractive part gross weight; Quercetagetin 3,6-dimethyl ether occupy imitates 4.16% of extractive part gross weight; Quercetin occupies imitates 10.17% of extractive part gross weight; Isoquercitrin occupies imitates 10.29% of extractive part gross weight.
11. claim 8, the purposes of 9 or 10 described Filifolium sibiricum (Linn.) Kitam. effective part extracts in the medicine of preparation treatment bacterial infection disease.
12. a pharmaceutical composition for the treatment of bacterial infection disease is made up of with pharmaceutically acceptable carrier or adjuvant claim 8, the 9 or 10 described Filifolium sibiricum (Linn.) Kitam. effective part extracts of effective dose.
13., it is characterized in that: be prepared into capsule, tablet, granule, drop pill, oral liquid or injection liquor according to method of Chinese medicinal according to the described pharmaceutical composition of claim 12.
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