CN103159820B - A kind of method utilizing microwave counter current technology to extract kincup lactone in kincup - Google Patents
A kind of method utilizing microwave counter current technology to extract kincup lactone in kincup Download PDFInfo
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- CN103159820B CN103159820B CN201210595585.7A CN201210595585A CN103159820B CN 103159820 B CN103159820 B CN 103159820B CN 201210595585 A CN201210595585 A CN 201210595585A CN 103159820 B CN103159820 B CN 103159820B
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Abstract
The invention belongs to field of natural organic chemistry, adopt microwave counter current extract, there is homogeneous heating, process be easy to control, without the need to pre-treatment such as dryings, simplify technique, decrease investment, substantially increase extraction efficiency; Utilize macroporous resin adsorption-column chromatography technology separation and purification kincup lactone, good to the adsorption selectivity of kincup lactone, absorption is fast, parsing is also fast, loading capacity is larger, extract convenient and swift, low production cost, separating effect is obvious, DNA purity is high, kincup lactone can be adsorbed selectively from solution by physical adsorption, obtain the kincup lactone work in-process of content more than 20% and the kincup lactone finished product of content more than 70%, overcome the shortcoming that traditional extraction rate is relatively low, DNA purity is low.
Description
Technical field:
The invention belongs to field of natural organic chemistry, relate to a kind of purification process of kincup lactone, particularly relate to a kind of method utilizing kincup lactone in microwave counter current extraction-macroporous resin adsorption-column chromatography coupling technology enriching and purifying kincup.
Background technology:
Kincup: English name Herb of Membranaceous Marshmarigold, another name: donkey hoof dish, marsh marigold.. perennial herb, high 20-48cm, without hair, fibrous root meat.Stem is upright, solid, the thin longitudinal furrow of tool, middle part or middle part with upper branch, rare not branch.Basal leaf 3-7, herbaceous stem, has long handle; The long 7-24cm of handle; Blade is circular, circle kidney shape or with shape, long 2.5-5cm, wide 3-9cm, tip circle, base portion innermost being shape, the close raw denticle in edge; Stem leaf is less, tool short handle or stockless.Single over-expense cyme is born in stem or branch top, usually has 2 flowers; The long 2-10cm of bennet; Flower both sexes, diameter 1.6-3.2cm; Sepal 5, petal-shaped, yellow, obovate or narrow obovate, long 1-1.8cm, wide 0.6-1.2cm, tip circle; Petal without; Stamen is most, long 4.5-7mm, and filigree is narrow linear, flower pesticide Long Circle, long 1-1.6mm; Carpel 7-12, closely isometric with stamen, stockless, style Duan ， Follicle radish fruit, be about 1cm, wide about 3mm, have transverse vein line, beak is about 1mm.Seed is most, and narrow ovoid, long 1.5-2mm, black is glossy, and tool minority indulges wrinkle.The florescence 5-9 month, the fruit phase 6-10 month.Chemical composition: herb is containing corytuberine (corytuberine), magnoflorine (magnoflorine), nicotine (nicotine), kincup lactone (caltholide), the compositions such as table kincup lactone (epiacltholide).
Kincup lactone is a kind of natural active matter human health being had to remarkable health-care effect, has caused the common concern of international community.Prove through scientific research and clinical application, kincup lactone has clearing heat and promoting diuresis, removing toxic substances.For heatstroke, urinary tract infections; Burn and scald is controlled in external application, venomous snake bite.In recent years, it is found that containing a large amount of kincup lactones in natural phant kincup, therefrom extract kincup lactone and achieve encouraging progress, raw material kincup source is wider.
Summary of the invention:
The object of the invention is to overcome the shortcomings such as routine techniques extraction yield is relatively low, DNA purity is low, a kind of method utilizing kincup lactone in microwave counter current extraction-macroporous resin adsorption-column chromatography coupling technology enriching and purifying kincup is provided.
For achieving the above object, the technical solution used in the present invention is:
Utilize a method for kincup lactone in microwave counter current extraction-macroporous resin adsorption-column chromatography coupling technique enriching and purifying kincup, its step is as follows:
(1) after dry kincup being pulverized, add pure water mixing, carry out microwave counter current extraction, forward feeding, oppositely to intake, the flowing of material and water is reverse continuous dynamic flow, and microwave power is at 400 ~ 800W, and extraction time is 60min ~ 120min; Extraction temperature is 50 ~ 60 DEG C, extracts three times, and filter, merging filtrate, obtains extracting solution;
(2) the kincup extracting solution suction filtration that will process through step (1), obtains extracting solution and concentrating under reduced pressure obtains kincup primary extract;
(3) by obtained primary extract water heating for dissolving, add extraction into ethyl acetate, standing, suction filtration, concentrating under reduced pressure obtain kincup lactone crude extract;
(4) gained kincup lactone crude extract macroporous adsorbent resin is adsorbed;
(5) with the ethanol of different concns, wash-out is carried out to macroporous adsorbent resin, use tlc tracing detection, collect each stepwise elution liquid of kincup lactone;
(6) by each for kincup lactone stepwise elution liquid concentration and recovery ethanol, obtain kincup lactone work in-process, its content is more than 20%;
(7) gained kincup lactone work in-process are dissolved in ethyl acetate and add proper silica gel stirring, dried 60-80 mesh sieve, dry method loading, upper technical grade post 200-300 order silica gel column chromatography, and carry out wash-out with ethanol, ether with different volumes ratio, tlc tracing detection, collects each stepwise elution liquid of kincup lactone;
(8) by collected kincup lactone elutriant, adopt activated carbon decolorizing, recrystallization method to refine to obtain kincup lactone, its content is more than 70%;
In described step (1), kincup crosses 60 ~ 80 mesh sieves after pulverizing;
In described step (1), the volume adding purified water is 8 ~ 10 times that kincup pulverizes volume;
In described step (3), the amount adding ethyl acetate be by volume calculated as extract dissolve after 10 ~ 20 times;
One in NKA-9, AB-8H and NKA-II type resin that macroporous adsorbent resin used in described step (4) produces for Tianjin Nankai university, is adsorbed as Static Adsorption or dynamic adsorption;
The described wash-out described in step (5) be with 10 ~ 90% ethanol carry out wash-out;
In described step (5) and (7), the detection method taked is tlc tracing detection;
The ethanol taked in described step (7) and the volume of ether are 1 ~ 5: 1.
The present invention adopts microwave counter current to extract, there is homogeneous heating, process be easy to control, without the need to pre-treatment such as dryings, simplify technique, decrease investment, substantially increase extraction efficiency; Macroporous adsorption resin technology and column chromatographic isolation and purification kincup lactone, good to the adsorption selectivity of kincup lactone, absorption is fast resolves also soon, and loading capacity is comparatively large, extracts convenient and swift, abundant raw material source, low production cost, separating effect is obvious, and DNA purity is high, the kincup lactone work in-process of content more than 20% and the kincup lactone finished product of content more than 70% can be obtained, overcome the shortcoming that traditional extraction rate is relatively low, DNA purity is low.
The present invention selects that stable in physicochemical property, surface-area are comparatively large, exchange velocity is very fast, physical strength is high, contamination resistance is strong, the macroporous adsorbent resin of Heat stability is good, be insoluble to acid, alkali and organic matchmaker, better to organism selectivity, not by the impact that inorganic salts and strong ion low molecular compound exist, kincup lactone can be adsorbed by physical adsorption selectively from solution, absorption is fast, resolve also fast, loading capacity is larger.
Embodiment:
Embodiment 1: a kind of method utilizing kincup lactone in microwave counter current extraction-macroporous adsorbent resin-column chromatography coupling technology enriching and purifying kincup, its step is as follows:
(1) cross 60 mesh sieves after dry for 500g kincup being pulverized, add the pure water that kincup pulverizes volume 10 times and carry out microwave counter current extraction, forward feeding, oppositely to intake, the flowing of material and water is reverse continuous dynamic flow.Microwave power is at 400W, and extraction time is 60min; Extraction temperature is 50 DEG C, extracts three times, and filter, merging filtrate, obtains extracting solution;
(2) the kincup extracting solution suction filtration that will process through step (1), obtains extracting solution and concentrating under reduced pressure obtains kincup primary extract;
(3) by obtained primary extract water heating for dissolving, adding volume is that the ethyl acetate of solution of extract volume 10 times extracts, and leaves standstill, suction filtration, concentrating under reduced pressure obtain kincup lactone crude extract;
(4) gained kincup lactone extract macroporous adsorbent resin is adsorbed, the NKA-II type resin that macroporous resin used is produced for Tianjin Nankai university;
(5) in macroporous adsorbent resin layer post, gradient elution is carried out with the ethanol of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% different concns successively, use tlc tracing detection, collect each stepwise elution liquid of kincup lactone;
(6) by each for kincup lactone stepwise elution liquid concentration and recovery ethanol, obtain kincup lactone work in-process, its content is 23.1%;
(7) gained kincup lactone work in-process are dissolved in ethyl acetate and add proper silica gel stirring, dried 60 mesh sieves, dry method loading, upper technical grade post 200-300 order silica gel column chromatography, and carry out gradient elution with the volume ratio of 1: 1,2: 1,3: 1,4: 1,5: 1 successively with ethanol, ether, tlc tracing detection, collects each stepwise elution liquid of kincup lactone;
(8) by collected kincup lactone elutriant, adopt activated carbon decolorizing, recrystallization method to refine and obtain kincup lactone, its content is 72.3%.
Case study on implementation 2: a kind of method utilizing kincup lactone in microwave counter current extraction-macroporous adsorbent resin-column chromatography coupling technology enriching and purifying kincup, its step is as follows:
(1) cross 70 mesh sieves after dry for 500g kincup being pulverized, add the pure water that kincup pulverizes volume 15 times and carry out microwave counter current extraction, forward feeding, oppositely to intake, the flowing of material and water is reverse continuous dynamic flow.Microwave power is at 600W, and extraction time is 90min; Extraction temperature is 55 DEG C, extracts three times, and filter, merging filtrate, obtains extracting solution;
(2) the kincup extracting solution suction filtration that will process through step (1), obtains extracting solution and concentrating under reduced pressure obtains kincup primary extract;
(3) by obtained primary extract water heating for dissolving, adding volume is that the ethyl acetate of solution of extract volume 15 times extracts, and leaves standstill, suction filtration, concentrating under reduced pressure obtain kincup lactone crude extract;
(4) gained kincup lactone extract macroporous adsorbent resin is adsorbed, the AB-8H type resin that macroporous resin used is produced for Tianjin Nankai university;
(5) in macroporous adsorbent resin layer post, gradient elution is carried out with the ethanol of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% different concns successively, use tlc tracing detection, collect each stepwise elution liquid of kincup lactone;
(6) by each for kincup lactone stepwise elution liquid concentration and recovery ethanol, obtain kincup lactone work in-process, its content is 23.7%;
(7) gained kincup lactone work in-process are dissolved in ethyl acetate and add proper silica gel stirring, dried 80 mesh sieves, dry method loading, upper technical grade post 200-300 order silica gel column chromatography, and carry out gradient elution with the volume ratio of 1: 1,2: 1,3: 1,4: 1,5: 1 successively with ethanol, ether, tlc tracing detection, collects each stepwise elution liquid of kincup lactone;
(8) by collected kincup lactone elutriant, adopt activated carbon decolorizing, recrystallization method to refine and obtain kincup lactone, its content is 74.5%.
Case study on implementation 3: a kind of method utilizing kincup lactone in microwave counter current extraction-macroporous adsorbent resin-column chromatography coupling technology enriching and purifying kincup, its step is as follows:
(1) cross 80 mesh sieves after dry for 500g kincup being pulverized, add the pure water that kincup pulverizes volume 20 times and carry out microwave counter current extraction, forward feeding, oppositely to intake, the flowing of material and water is reverse continuous dynamic flow.Microwave power is at 800W, and extraction time is 120min; Extraction temperature is 60 DEG C, extracts three times, and filter, merging filtrate, obtains extracting solution;
(2) the kincup extracting solution suction filtration that will process through step (1), obtains extracting solution and concentrating under reduced pressure obtains kincup primary extract;
(3) by obtained primary extract water heating for dissolving, adding volume is that the ethyl acetate of solution of extract volume 20 times extracts, and leaves standstill, suction filtration, concentrating under reduced pressure obtain kincup lactone crude extract;
(4) gained kincup lactone extract macroporous adsorbent resin is adsorbed, the NKA-9 type resin that macroporous resin used is produced for Tianjin Nankai university;
(5) in macroporous adsorbent resin layer post, gradient elution is carried out with the ethanol of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% different concns successively, use tlc tracing detection, collect each stepwise elution liquid of kincup lactone;
(6) by each for kincup lactone stepwise elution liquid concentration and recovery ethanol, obtain kincup lactone work in-process, its content is 24.2%;
(7) gained kincup lactone work in-process are dissolved in ethyl acetate and add proper silica gel stirring, dried 80 mesh sieves, dry method loading, upper technical grade post 200-300 order silica gel column chromatography, and carry out gradient elution with the volume ratio of 1: 1,2: 1,3: 1,4: 1,5: 1 successively with ethanol, ether, tlc tracing detection, collects each stepwise elution liquid of kincup lactone;
(8) by collected kincup lactone elutriant, adopt activated carbon decolorizing, recrystallization method to refine and obtain kincup lactone, its content is 75.3%.
Claims (6)
1. a method for kincup lactone in microwave counter current extraction-macroporous resin adsorption-column chromatography coupling technique enriching and purifying kincup, its step is as follows:
(1) after dry kincup being pulverized, add pure water mixing, carry out microwave counter current extraction, forward feeding, oppositely to intake, the flowing of material and water is reverse continuous dynamic flow, and microwave power is at 400 ~ 800W, and extraction time is 60min ~ 120min; Extraction temperature is 50 ~ 60 DEG C, extracts three times, and filter, merging filtrate, obtains extracting solution;
(2) the kincup extracting solution suction filtration that will process through step (1), obtains extracting solution and concentrating under reduced pressure obtains kincup primary extract;
(3) by obtained primary extract water heating for dissolving, add extraction into ethyl acetate, standing, suction filtration, concentrating under reduced pressure obtain kincup lactone crude extract;
(4) gained kincup lactone crude extract macroporous adsorbent resin is adsorbed;
(5) with the ethanol of different concns, wash-out is carried out to macroporous adsorbent resin, use tlc tracing detection, collect each stepwise elution liquid of kincup lactone;
(6) by each for kincup lactone stepwise elution liquid concentration and recovery ethanol, obtain kincup lactone work in-process, its content is more than 20%;
(7) gained kincup lactone work in-process are dissolved in ethyl acetate and add proper silica gel stirring, dried 60-80 mesh sieve, dry method loading, upper technical grade post 200-300 order silica gel column chromatography, and carry out wash-out with ethanol, ether with different volumes ratio, tlc tracing detection, collects each stepwise elution liquid of kincup lactone;
(8) by collected kincup lactone elutriant, adopt activated carbon decolorizing, recrystallization method to refine to obtain kincup lactone, its content is more than 70%;
One in NKA-9, AB-8H and NKA-II type resin that macroporous adsorbent resin used in described step (4) produces for Tianjin Nankai university, is adsorbed as Static Adsorption or dynamic adsorption;
In described step (5) and (7), the detection method taked is tlc tracing detection.
2. the method for kincup lactone in a kind of microwave counter current extraction-macroporous resin adsorption according to claim 1-column chromatography coupling technique enriching and purifying kincup, its speciality is: in described step (1), and kincup crosses 60 ~ 80 mesh sieves after pulverizing.
3. the method for kincup lactone in a kind of microwave counter current extraction-macroporous resin adsorption according to claim 2-column chromatography coupling technique enriching and purifying kincup, it is characterized in that: in described step (1), the volume adding purified water is 10 ~ 20 times that kincup pulverizes volume.
4. in a kind of microwave counter current extraction-macroporous resin adsorption according to claim 3-column chromatography coupling technique enriching and purifying kincup, the method for kincup lactone, is characterized in that; In described step (3), the amount adding ethyl acetate is 10 ~ 20 times after extract dissolving by volume.
5. the method for kincup lactone in a kind of microwave counter current extraction-macroporous resin adsorption according to claim 4-column chromatography coupling technique enriching and purifying kincup, it is characterized in that: in described step (5), the described elutriant aqueous ethanolic solution of 10 ~ 90% carries out wash-out.
6. the method for kincup lactone in a kind of microwave counter current extraction-macroporous resin adsorption according to claim 5-column chromatography coupling technique enriching and purifying kincup, it is characterized in that: in described step (7), adopt the volume ratio of ethanol, ether to be 1 ~ 5: 1.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101062069A (en) * | 2007-06-18 | 2007-10-31 | 石任兵 | Whole coumarins extract from root of dahuriae angelica and the preparing method thereof |
| CN101073590A (en) * | 2007-06-18 | 2007-11-21 | 石任兵 | Muxiang total lactone extract and its production |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101062069A (en) * | 2007-06-18 | 2007-10-31 | 石任兵 | Whole coumarins extract from root of dahuriae angelica and the preparing method thereof |
| CN101073590A (en) * | 2007-06-18 | 2007-11-21 | 石任兵 | Muxiang total lactone extract and its production |
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