CN104208108A - Method for effectively separating low-acid gingko leaf extract - Google Patents
Method for effectively separating low-acid gingko leaf extract Download PDFInfo
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Abstract
The invention relates to a method for effectively separating a low-acid gingko leaf extract and belongs to the technical field of extraction separation. According to the method, a gingko extract is subjected to continuous chromatographic separation by adopting mixed resin, namely mixed macroporous adsorption resin is used in the continuous chromatographic separation process to realize effective extraction separation of the gingko leaf extract; gingko leaf is subjected to proper extraction concentration to obtain gingko leaf extract liquid, and ginkgo flavones and lactone compounds are adsorbed by a continuous chromatographic system; and elution by using eluent, concentration and drying are carried out to obtain the low-acid gingko extract. According to the method, the gingko extract is separated by adopting continuous chromatography, so that the utilization of macroporous adsorption resin is improved, the production cycle is shortened, the production cost is reduced, the product quality is improved, and the discharge amount of waste water is reduced; and the aim of separation and deacidification can be achieved at one time by using the mixed resin. Through detection by an HPLC (high performance liquid chromatography) method, the product quality exceeds the quality standard specified in the Chinese pharmacopoeia, and the content of ginkgolic acid is less than 5 microgram.g<-1) and accords with the requirement of international standard.
Description
Technical field
The present invention relates to a kind of efficient separation method of extractive of low acidic gingko leaves, specifically a kind of efficient separation method of Folium Ginkgo extract, relate to the separation method of Folium Ginkgo extract, belong to extraction and separation technology field.
Background technology
Semen Ginkgo is one of the most ancient in the world seeds, and be described as " living fossil ", Semen Ginkgo is China's special product, and resource owning amount accounts for the world about 70%.Folium Ginkgo is a kind of important medicinal, healthy food material, its extract (GBE, Egb, Extract of Ginkgo, biloba) there is great antioxidation, free radical superfluous in organism can be removed, stop body lipid oxidation, improve body body immunity, the functions such as slow down aging.In recent years, the particularly chemical analysis of the country such as Germany, Japan, the U.S., France to Folium Ginkgo both at home and abroad, separation method has carried out large quantifier elimination, achieves greater advance.Large quantifier elimination has been carried out to solvent extraction and Flavonoids by Macroporous Adsorption Resin, domestic now based on Flavonoids by Macroporous Adsorption Resin.The GBE761 standard that Semen Ginkgo extrac adopts German Schwabe company to formulate: flavones content percent>=24%, lactone>=6%, ginkgoic acid phenol≤5 × 10
-6.
Folium Ginkgo extract is the basic material of each pharmacy corporation as various gingko leaf preparation.In recent years, the sale in the international market of natural medicinal plant extract day by day improves, and China's export medicinal plant extract Semen Ginkgo ranks first three, and international demand amount is very large.But, because preparation technology's level of Semen Ginkgo extrac respectively has the similarities and differences, cause its quality uneven, the region of domestic plantation Semen Ginkgo is extensive, plucking time is different, and the changes of contents in its leaf is comparatively large, and the extractive content causing same process conditions to obtain is unstable, sometimes do not reach state-promulgated pharmacopoeia standard, be difficult to meet domestic and international market demand.Therefore, a kind of preparation technology that is stable, Folium Ginkgo extract reliably is provided, also can reduces the waste of resource in the demand meeting market simultaneously, there is wide market prospect.In addition, the harmful components in Folium Ginkgo extract but limit further exploitation and the clinical practice of its preparation, and wherein gingkolic acid is the main matter causing clinical adverse, tool sensitization and mutagenic toxicity.Gingkolic acid is one of key index of Folium Ginkgo extract product quality, and German hygiene department specifies that the content of the gingkolic acid of ginkgo leaf extract preparation must be less than 5 μ gg
-1.
Containing multiple active component with pharmacological action such as flavonoid, terpenoid, phenols, alkaloids, polyisoprene, polysaccharide in Folium Ginkgo.Semen Ginkgo extrac has quite strong antioxidation, can eliminate free radical superfluous in organism, stops body lipid peroxidating, improves body immunity blood circulation promoting and brain metabolism, has defying age simultaneously, prevents the functions such as nervous system disease.Folium Ginkgo extracting method is different, but separation all adopts Flavonoids by Macroporous Adsorption Resin.The wherein preparation method macroporous adsorbent resin linear gradient elution method of Authorization Notice No. CN 100362011C Folium Ginkgo extract, eluant first time eluting concentration is 0%-30% eluant, and second time eluting concentration is 40%-95%.
This patented method separation of Silver Fructus Pruni extract need repeat upper prop, and suitability for industrialized production more complicated, is unfavorable for large-scale production.This purification mainly adopts fixed bed partition method, and needs to repeat secondary macroporous absorption post.This traditional separation method is separated the flavone that obtains and lactone and has that purity is low, eluant consumption is large, the cycle long, environmental protection pressure is large, the not high shortcomings of resin utilization rate.Therefore need to develop efficient process for separating and purifying, to improve the quality of products, advocate Green Chemistry.
The method removing gingkolic acid during Folium Ginkgo extract purification has two kinds, and a kind of is with the attached resin purification ginkgetin of general macroporous absorption and bilobalide, then removes gingkolic acid by DAD-1 macroporous adsorbent resin.Another kind is remove gingkolic acid with nonpolar organic solvent extraction.This technology takes hybrid resin one step to complete purification and the deacidification of ginkgetin and lactone, reaches good effect.
Summary of the invention
The object of the invention is to overcome above-mentioned weak point, provide a kind of efficient separation method of extractive of low acidic gingko leaves, utilize continuous chromatography to be separated Folium Ginkgo compound, flavone and lactone content are all higher.
According to technical scheme provided by the invention, a kind of efficient separation method of extractive of low acidic gingko leaves, step is as follows:
(1) preparation of ginkgo extract: get Folium Ginkgo, was ground into 15 ~ 25 object coarse powder; Carry out percolation after moistening, or obtain ginkgo extract through reflux, extract;
The process of carrying out percolation after moistening is: by coarse powder: ethanol weight ratio be 1:1 add mass concentration be 50% ~ 95% ethanol carry out moistening, after moistening 3 ~ 8 hours, continue to add ethanol again and carry out percolation, percolation 20 ~ 48 hours, the flow velocity of ethanol is 1 ~ 3mL/ minute/kg;
Or the process of reflux, extract, is: add coarse powder 5 ~ 8 times amount ethanol, 65 ~ 85 DEG C of reflux, extract, 2 ~ 3 hours, filter; Then add coarse powder 3 ~ 7 times amount ethanol and repeat reflux, extract, at the same terms, obtain ginkgo extract, after filtration, obtain ginkgo extract; The mass concentration of ethanol is 50% ~ 95%;
(2) thickening filtration: ginkgo extract step (1) obtained was 60 ~ 85 DEG C of vacuum concentration 2 ~ 8 hours, vacuum is-0.08MPa, be concentrated into every 2.5kg coarse powder to obtain concentrated solution and be less than or equal to 1L, then purified water is adopted to be settled to 1L, when being cooled to-1 DEG C ~ 20 DEG C, sedimentation 8 ~ 24 hours, is then filtered by quartz sand;
(3) adsorbing separation: adopt continuous chromatography piece-rate system to be separated the ginkgo extract after step (2) gained thickening filtration, the chromatographic column of employing is 20 ~ 30; Continuous chromatography piece-rate system is divided into adsorption zone, Xi Za district, resolves district and district of four, regeneration washing district;
Supernatant loading is got in hybrid resin continuous chromatography post in adsorption zone; Xi Za district first with purified water washing, with conductivity meter survey to confirm do not have inorganic salt to wash out, then by mass concentration be 5% ~ 30% washing with alcohol to cleaning mixture close to colourless; Resolving district is the ethanol elution of 60% ~ 80% with mass concentration; First renewing zone is the ethanol regeneration of 95% by mass concentration, then washes away ethanol by purified water;
(4) reclaim dry: by gained eluent at 60 ~ 75 DEG C of vacuum concentration to recycling design ethanol during proportion 1.08 ~ 1.35g/mL, vacuum is-0.08MPa; Then product extractive of low acidic gingko leaves is obtained by after the drying of gained concentrated solution.
Described hybrid resin is DM130 resin, one in D101 resin or SIP-1200 resin and DAD-1 macroporous resin mix.
The mixed proportion of a kind of and DAD-1 macroporous resin in described DM130 resin, D101 resin or SIP-1200 resin is 20 ~ 1:1.Described adsorption zone is made up of 3 ~ 8 unit; Xi Za district is made up of 5 ~ 11 unit; Resolve district to be made up of 5 ~ 12 unit, regeneration washing district is made up of 2 ~ 4 unit.
Step (4) described drying mode is vacuum drying or spraying dry, and wherein vacuum drying vacuum is-0.08MPa, and temperature is 60 ~ 85 DEG C; Spray-dired inlet temperature is 150 ~ 180 DEG C, and leaving air temp is 75 ~ 90 DEG C.
Described Folium Ginkgo ethanol extract, is the extracting solution obtained by Folium Ginkgo ethanol extraction, belongs to prior art, can conventionally method obtain, as the method for Chinese patent CN 100362011C.
Beneficial effect of the present invention: present invention improves over original fixed bed separation equipment, make the absorption of original fixed bed, wash assorted, eluting, the whole workshop sections such as regeneration are incorporated in disk conveying type adverse current continuous chromatography piece-rate system, whole section of resin in original fixed bed is divided into some sections by it, part resin before former process tradition district is positioned at one or several little resin column again, so just absorption can be reentered, eluting, regeneration waits in circulation, utilize original not by the part resin reinstated, resin utilization rate just substantially increases, chemical reagent can also be reduced simultaneously, the consumption of water etc.Disk conveying type continuous chromatography piece-rate system has a large amount of post (separation) unit, also makes them can very effectively be applied to series classification production process.According to the characteristic of composition each in Folium Ginkgo, the resin selected by the present invention is the macroporous adsorbent resin of mixed type, resin particle diameter at 30 ~ 80 orders, the uniformity more than 95%.
Concrete advantage is:
(1) institute of fixed-bed process is all integrated in a set of process system in steps, system is simplified, and reduces the layout of process pipe, system compact, Automated condtrol can be realized; Floor space saves 80%, and factory building height only needs 1/3 of fixed bed height, and the investment in fixed assets of same production capacity saves more than 30%.
(2) resin utilization rate is high, makes product design, purity and yield optimization; Present invention process compares with fixed-bed resin separating technology, and its resin demand is only original 30%, and can carry out forward and inverse stream than being easier in resin inside, and can loosen resin, prevents it from luming.
(3) adopt hybrid resin, by DM130 macroporous resin, D101 macroporous resin, SIP-1200 macroporous resin with except acid resin DAD-1 used in combination, once complete flavone, the separation of lactone and the removal of gingkolic acid.
(4) reduce the consumption of chemical reagent and water, reduce the discharge of waste water; Utilize this technique can carry out back cover to material to use, reach and recycle.
(5) system adopts self-con-tained unit, reduces work load.
(6) enhance productivity, improve production capacity, the production cycle decreased for 1/3 time relative to former fixed-bed resin separating technology.
Accompanying drawing explanation
Fig. 1 is embodiment 1 chromatographic isolation flow chart.
Fig. 2 is embodiment 2 chromatographic isolation flow chart.
Fig. 3 is embodiment 3 chromatographic isolation flow chart.
Detailed description of the invention
The following examples, for further illustrating and describing the present invention, but and do not mean that the present invention is only limitted to the content of this following Examples.In embodiment, value is the arbitrary concrete numerical value of scope of the present invention, is and can implements.
Embodiment 1
(1) preparation of ginkgo extract: get Folium Ginkgo, was ground into 15 object coarse powder; Percolation is carried out after moistening; By coarse powder: ethanol weight ratio be 1:1 add mass concentration be 95% ethanol carry out moistening, after moistening 3 hours, then continue add ethanol carry out percolation, percolation 20 hours, ethanol flow velocity is 2mL/ minute/kg, and percolation, to paler colour, obtains ginkgo extract;
(2) thickening filtration: ginkgo extract step (1) obtained was 60 DEG C of vacuum concentration 8 hours, vacuum is-0.08MPa, be concentrated into every 2.5kg coarse powder to obtain concentrated solution and be less than or equal to 1L, then purified water is adopted to be settled to 1L, when being cooled to 10 DEG C, sedimentation 8 hours, is then filtered by quartz sand;
(3) adsorbing separation: adopt continuous chromatography piece-rate system to be separated the ginkgo extract after step (2) gained thickening filtration, the chromatographic column of employing is 30; Continuous chromatography piece-rate system is divided into adsorption zone, Xi Za district, resolves district and district of four, regeneration washing district;
Select DM130 type macroporous adsorbent resin and DAD-1 macroporous adsorbent resin, its ratio is 9:1, and resin is 30 ~ 80 orders, and each resin column amount of fill is 0.005m
3, resin column is of a size of Ф 10 × 60mm, and actual filling ratio is about 90%.Resin column takes a step forward for every 3 hours.
Disk conveying type continuous chromatography piece-rate system divides following region:
A, adsorption zone: (Unit 6)
This region has 6 unit (25,26,27,28,29 and No. 30 chromatographic columns), by flow speed control, first raw material 6 enters the chromatographic column group by 25 and No. 26 chromatographic column parallel connections, then by all the other pole units of series connection, the effluent of No. 30 mouths is waste liquid, enters three wastes center processing.
B, Xi Za district: (9 unit)
This region has 9 unit (16 ~ No. 24 chromatographic columns), and after absorption, first No. 24 posts add water 5 and wash, and washing post is in parallel again with 25 and No. 26 parallel columns.No. 16 post resins need the ethanol 4 of 20% to clean, after being positioned at adsorption zone.Resin column rotates to after absorption 20% ethanol washes district, be entrained in the feed liquid (mainly clear liquor) between macroporous resin by 20% ethanol eject, wash away the feed liquid being mixed in resin gap and also take away impurity as far as possible, prevent feed liquid from carrying secretly and enter parsing district, improve the purity of stripping liquid, and its ethanol washing liquid of 20% is entered into No. 27 posts of adsorption zone, to determine clean result after being detected by No. 28 outlets, this outlet can not detect flavone or lactone compound.
C, parsing district (11 unit)
There are 10 unit (5 ~ No. 15 chromatographic columns) in this district, and in this parsing district, the ethanol 3 with 60% carries out eluting, resolve district and all take positive charging, in No. 5 posts ethanol 3 eluting of 60%, collect eluent in 15 post lower ends, be required ginkgetin and bilobalide.
D, regeneration washing district (4 unit);
There are 4 unit (1,2,3, No. 4 chromatographic column) in this district, the charging all separately of 4, this district unit, and is backward feed, and 3, No. 4 post parallel connections, the ethanol 2 with 95% regenerates.1, No. 2 posts, rinse by purified water 1, to forward loading district sample loading to.
Separation purity: the sample that No. 15 posts are collected is required containing more than 24%, and bilobalide is containing more than 6%, and gingkolic acid content is less than 5 μ gg
-1.What No. 30 posts were collected is the impurity such as 20% water soluble polysaccharide eluted.The part of pigment etc. 60% ethanol not easily eluting, elutes, as impurity completely with the ethanol of 95%.This part ethanol can be passed through reclaim under reduced pressure, can be used as 60% eluent use through regulating.Eluent collected by No. 15 posts.Detailed chromatographic isolation flow chart is as Fig. 1, and concrete active constituent content is as shown in table 1.
(4) reclaim dry: by gained eluent at 60 DEG C of vacuum concentration to recycling design ethanol during proportion 1.30 ~ 1.35g/mL, vacuum is-0.08MPa; Then gained concentrated solution is obtained product extractive of low acidic gingko leaves after 70 DEG C of vacuum dryings, vacuum is-0.08MPa.
Embodiment 2
(1) preparation of ginkgo extract: get Folium Ginkgo, was ground into 25 object coarse powder; Ginkgo extract is obtained through reflux, extract; Reflux, extract, 2 times, first time coarse powder: ethanol weight ratio is 1:8,65 DEG C of reflux, extract, 3 hours, filters; Second time coarse powder: ethanol weight ratio is 1:7,65 DEG C of reflux, extract, 3 hours, filter;
(2) thickening filtration: ginkgo extract step (1) obtained was 80 DEG C of vacuum concentration 2 hours, vacuum is-0.08MPa, be concentrated into every 2.5kg coarse powder to obtain concentrated solution and be less than or equal to 1L, then purified water is adopted to be settled to 1L, when being cooled to 15 DEG C, sedimentation 24 hours, is then filtered by quartz sand;
(3) adsorbing separation: adopt continuous chromatography piece-rate system to be separated the ginkgo extract after step (2) gained thickening filtration, the chromatographic column of employing is 30; Continuous chromatography piece-rate system is divided into adsorption zone, Xi Za district, resolves district and district of four, regeneration washing district;
Select D101 type macroporous adsorbent resin and DAD-1 macroporous adsorbent resin, its ratio is 85:15, and resin is 30 ~ 80 orders, and each resin column amount of fill is 0.005m
3, resin column is of a size of Ф 10 × 60mm, and actual filling ratio is about 90%.Resin column takes a step forward for every 3 hours.
Disk conveying type continuous chromatography piece-rate system divides following region:
A, adsorption zone: (6 unit)
This region has 6 unit (25,26,27,28,29 and No. 30 chromatographic columns), by flow speed control, first raw material 6 enters the chromatographic column group by 25, No. 26 chromatographic column parallel connections, then by all the other pole units of series connection, the effluent of No. 30 mouths is waste liquid, enters three wastes center processing.
B, Xi Za district: (10 unit)
This region has 10 unit (15 ~ No. 24 chromatographic columns), and after absorption, No. 24 posts are washing post, add water 5.No. 15 post resins ethanol 4 of 20% is washed, and No. 16 posts are in parallel with No. 17 posts, and No. 18 posts are in parallel with No. 19 posts, and No. 20 posts are in parallel with No. 21 posts, and No. 22 posts are connected with No. 27 posts with after No. 23 post parallel connections.Resin column rotates to after absorption 20% ethanol washes district, be entrained in the feed liquid (mainly clear liquor) between macroporous resin by 20% ethanol eject, wash away the feed liquid being mixed in resin gap and also take away impurity as far as possible, prevent feed liquid from carrying secretly and enter parsing district, improve the purity of stripping liquid, and its ethanol washing liquid of 20% is entered into No. 27 posts of adsorption zone, the active component again in adsorbed water washing liquid, assorted elution profile before detecting after No. 28 posts.
C, parsing district (10 unit)
In this parsing district, No. 5 post ethanol 3 eluting of 65%, use parallel connection and series connection to connect, and resolve district and all take positive charging, collect eluent, be required ginkgetin and bilobalide in 14 post lower ends.Be specially No. 6 posts in parallel with No. 7 posts, No. 8 posts are in parallel with No. 9 posts, and No. 10 posts are in parallel with No. 11 posts, and No. 12 posts are in parallel with No. 13 posts, connecting with No. 14 posts.
D, regeneration washing district (4 unit);
The charging all separately of 4, this district unit, and be backward feed, 3, No. 4 post parallel connections, the ethanol 2 with 95% regenerates.1, No. 2 posts, rinse by purified water 1, to forward loading district sample loading to.
Separation purity: the sample that No. 14 posts are collected is required containing more than 24%, and bilobalide is containing more than 6%, gingkolic acid 5 μ gg
-1.What No. 30 posts were collected is the impurity such as 20% water soluble polysaccharide eluted.The part of pigment etc. 65% ethanol not easily eluting, elutes, as impurity completely with the ethanol of 95%.This part ethanol can be passed through reclaim under reduced pressure, can be used as 65% eluent use through regulating.No. 14 posts collect eluent, decompression recycling ethanol, and spray-dried or drying under reduced pressure may finished product.
Detailed separation process figure is as Fig. 2, and concrete active constituent content is as shown in table 1.
(4) reclaim dry: by gained eluent at 60 DEG C of vacuum concentration recycling design ethanol to proportion 1.35g/mL, vacuum is-0.08MPa; Then gained concentrated solution is obtained product extractive of low acidic gingko leaves through vacuum drying.Vacuum drying vacuum is-0.08MPa, and temperature is 70 DEG C.
Embodiment 3
(1) preparation of ginkgo extract: get Folium Ginkgo, was ground into 20 object coarse powder; Percolation is carried out after moistening; By coarse powder: ethanol weight ratio is the ethanol that 1:1 adds that mass concentration is 95%, then continuation interpolation ethanol carries out percolation, after moistening 5 hours, and percolation 20 hours, the flow velocity of ethanol is 2mL/ minute/kg, and percolation is to paler colour;
(2) thickening filtration: ginkgo extract step (1) obtained was 60 DEG C of vacuum concentration 8 hours, vacuum is-0.08MPa, be concentrated into every 2.5kg coarse powder to obtain concentrated solution and be less than or equal to 1L, then purified water is adopted to be settled to 1L, when being cooled to 20 DEG C, sedimentation 10 hours, is then filtered by quartz sand;
(3) adsorbing separation: adopt continuous chromatography piece-rate system to be separated the ginkgo extract after step (2) gained thickening filtration, the chromatographic column of employing is 30; Continuous chromatography piece-rate system is divided into adsorption zone, Xi Za district, resolves district and district of four, regeneration washing district;
Select D101 type macroporous adsorbent resin and DAD-1 macroporous adsorbent resin, its ratio is 9:1, and resin is 30 ~ 80 orders, and each resin column amount of fill is 0.005m
3, resin column is of a size of Ф 10 × 60mm, and actual filling ratio is about 90%.Resin column takes a step forward for every 2.5 hours.
Disk conveying type continuous chromatography piece-rate system divides following region:
A, adsorption zone: (Unit 6)
This region has 6 unit (25,26,27,28,29 and No. 30 chromatographic columns), by flow speed control, first raw material 6 enters the chromatographic column group by 25 and No. 26 chromatographic column parallel connections, then by all the other pole units of series connection, the effluent of No. 30 mouths is waste liquid, enters three wastes center processing.
B, Xi Za district: (8 unit)
This region has 8 unit (17 ~ No. 24 chromatographic columns), and after absorption, first No. 24 posts add water 5, washing, and washing post is in parallel again with 25 and No. 26 parallel columns.No. 17 post resins need the ethanol 4 of 25% to wash, after being positioned at adsorption zone.Resin column rotates to after absorption 25% ethanol washes district, be entrained in the feed liquid (mainly clear liquor) between macroporous resin by 25% ethanol eject, wash away the feed liquid being mixed in resin gap and also take away impurity as far as possible, prevent feed liquid from carrying secretly and enter parsing district, improve the purity of stripping liquid, and its ethanol washing liquid of 25% is entered into No. 27 posts of adsorption zone, detected after (measuring flavone, lactone with efficient liquid phase) by No. 28 outlets to determine clean result.
C, parsing district (12 unit)
There are 12 unit (5 ~ No. 16 chromatographic columns) in this district, and in this parsing district, the ethanol 3 with 70% carries out eluting, resolve district and all take positive charging, in No. 5 posts ethanol 3 eluting of 70%, collect eluent in 16 post lower ends, be required ginkgetin and bilobalide.
D, regeneration washing district (4 unit);
There are 4 unit (1,2,3, No. 4 chromatographic column) in this district, the charging all separately of 4, this district unit, and is backward feed, and 3, No. 4 post parallel connections, the ethanol 2 with 95% regenerates.1, No. 2 posts, rinse by purified water 1, to forward loading district sample loading to.
Separation purity: the sample that No. 16 posts are collected is required containing more than 24%, and bilobalide is containing more than 6%.What No. 30 posts were collected is the impurity such as 25% water soluble polysaccharide eluted.The part of pigment etc. 70% ethanol not easily eluting, elutes, as impurity completely with the ethanol of 95%.This part ethanol can be passed through reclaim under reduced pressure, and can be used as 70% eluent use through regulating, No. 16 posts collect eluent, decompression recycling ethanol.
As shown in Figure 3, concrete active constituent content is as shown in table 1 for detailed chromatographic isolation flow chart.
(4) reclaim dry: by gained eluent at 60 DEG C of vacuum concentration recycling design ethanol to proportion 1.08 ~ 1.15g/mL, vacuum is-0.08MPa; Then by spray-dried for gained concentrated solution, inlet temperature is 170 DEG C, and leaving air temp is 80 DEG C, obtains product extractive of low acidic gingko leaves.
In Folium Ginkgo, the content assaying method of ginkgetin, bilobalide is shown in " Chinese Pharmacopoeia " (one) 2010 editions.
Total flavonoids take octadecylsilane chemically bonded silica as filler; With methanol-0.4% phosphoric acid solution (50:50) for mobile phase; Determined wavelength is 360nm.Number of theoretical plate calculates should be not less than 2500 by quercetin peak.With Quercetin reference substance, kaempferide reference substance, isorhamnetin reference substance in right amount, accurately weighed, add methanol and make every 1mL respectively containing the mixed solution of 30 μ g, 30 μ g, 20 μ g, product solution in contrast.
Total flavonoids content=(quercetin content+kaempferide content+isorhamnetin content) * 2.5L
Terpene lactone take octadecylsilane chemically bonded silica as filler; With normal propyl alcohol-oxolane-water (1:15:84) for mobile phase; Detect with evaporative light scattering detector.Number of theoretical plate calculates should be not less than 2500 by bilobalide peak.
The preparation of reference substance solution: get bilobalide reference substance, ginkalide A reference substance, ginkalide B reference substance and ginkalide C comparing device respectively appropriate, accurately weighed, add methanol and make the mixed solution of 1mg containing 2mg, 1mg, 1mg, 1mg, product solution in contrast.Calculate the content of bilobalide, ginkalide A, ginkalide B and ginkalide C with external standard two-point method logarithmic equation respectively, to obtain final product.
The assay method of gingkolic acid is as follows: measure with efficient liquid phase, chromatographic condition: C18 chromatographic column (4.6mm × 250mm, 5 μm), mobile phase methanol-water-phosphoric acid (289.5:10.3:0.2), determined wavelength 310nm, flow velocity 1mlmin
-1, column temperature 25 DEG C.Be reference substance with gingkolic acid, measure by external standard method, to obtain final product
Above-mentioned three embodiment test results are as shown in table 1:
Table 1
| Batch | Embodiment 1 | Embodiment 2 | Embodiment 3 |
| Total flavonoid glycoside content (%) | 30.8 | 28.2 | 29.6 |
| Bilobalide content (%) | 11.2 | 10.2 | 10.8 |
| Gingkolic acid content (ppm) | 2.7 | 0.9 | 1.1 |
Thermopnore continuous chromatography separation method of the present invention, and the single macroporous resin Technical comparing of existing fixed bed, the comparison data of beneficial effect are as shown in table 2.
Table 2
Claims (5)
1. an efficient separation method for extractive of low acidic gingko leaves, is characterized in that step is as follows:
(1) preparation of ginkgo extract: get Folium Ginkgo, was ground into 15 ~ 25 object coarse powder; Carry out percolation after moistening, or obtain ginkgo extract through reflux, extract;
The process of carrying out percolation after moistening is: by coarse powder: ethanol weight ratio be 1:1 add mass concentration be 50% ~ 95% ethanol carry out moistening, after moistening 3 ~ 8 hours, the ethanol continuing interpolation 50% ~ 95% again carries out percolation, percolation 20 ~ 48 hours, and the flow velocity of ethanol is 1 ~ 3mL/ minute/kg;
Or the process of reflux, extract, is: add coarse powder 5 ~ 8 times amount ethanol, 65 ~ 85 DEG C of reflux, extract, 2 ~ 3 hours, filter; Then add coarse powder 3 ~ 7 times amount ethanol and repeat reflux, extract, at the same terms, obtain ginkgo extract, after filtration, obtain ginkgo extract; The mass concentration of ethanol is 50% ~ 95%;
(2) thickening filtration: ginkgo extract step (1) obtained was 60 ~ 85 DEG C of vacuum concentration 2 ~ 8 hours, vacuum is-0.08MPa, be concentrated into every 2.5kg coarse powder to obtain concentrated solution and be less than or equal to 1L, then purified water is adopted to be settled to 1L, when being cooled to-1 DEG C ~ 20 DEG C, sedimentation 8 ~ 24 hours, is then filtered by quartz sand;
(3) adsorbing separation: adopt continuous chromatography piece-rate system to be separated the ginkgo extract after step (2) gained thickening filtration, the chromatographic column of employing is 20 ~ 30; Continuous chromatography piece-rate system is divided into adsorption zone, Xi Za district, resolves district and district of four, regeneration washing district;
Supernatant loading is got in hybrid resin continuous chromatography post in adsorption zone; Xi Za district first with purified water washing, with conductivity meter survey to confirm do not have inorganic salt to wash out, then by mass concentration be 5% ~ 30% washing with alcohol to cleaning mixture close to colourless; Resolving district is the ethanol elution of 60% ~ 80% with mass concentration; First renewing zone is the ethanol regeneration of 95% by mass concentration, then washes away ethanol by purified water;
(4) reclaim dry: by gained eluent at 60 ~ 75 DEG C of vacuum concentration to recycling design ethanol during proportion 1.08 ~ 1.35g/mL, vacuum is-0.08MPa; Then product extractive of low acidic gingko leaves is obtained by after the drying of gained concentrated solution.
2. the efficient separation method of extractive of low acidic gingko leaves as claimed in claim 1, is characterized in that: described hybrid resin is DM130 resin, one in D101 resin or SIP-1200 resin and DAD-1 macroporous resin mix.
3. the efficient separation method of extractive of low acidic gingko leaves as claimed in claim 2, is characterized in that: the mixed proportion of a kind of and DAD-1 macroporous resin in described DM130 resin, D101 resin or SIP-1200 resin is 20 ~ 1:1.
4. the efficient separation method of extractive of low acidic gingko leaves as claimed in claim 1, is characterized in that: described adsorption zone is made up of 3 ~ 8 unit; Xi Za district is made up of 5 ~ 11 unit; Resolve district to be made up of 5 ~ 12 unit, regeneration washing district is made up of 2 ~ 4 unit.
5. the efficient separation method of extractive of low acidic gingko leaves as claimed in claim 1, it is characterized in that: step (4) described drying mode is vacuum drying or spraying dry, wherein vacuum drying vacuum is-0.08MPa, and temperature is 60 ~ 85 DEG C; Spray-dired inlet temperature is 150 ~ 180 DEG C, and leaving air temp is 75 ~ 90 DEG C.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107698691A (en) * | 2017-11-02 | 2018-02-16 | 厦门福美科技有限公司 | A kind of separating and purifying flavone from oldenlandia diffusa, the system and method for polysaccharide |
| CN110152353A (en) * | 2019-06-18 | 2019-08-23 | 大连博迈科技发展有限公司 | A kind of continuous chromatography device and arasaponin production method |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1515582A (en) * | 2003-01-08 | 2004-07-28 | 浙江康恩贝制药股份有限公司 | Preparation method of low-acid ginkgo leaf extract |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1515582A (en) * | 2003-01-08 | 2004-07-28 | 浙江康恩贝制药股份有限公司 | Preparation method of low-acid ginkgo leaf extract |
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| 庄玲华DENG: "银杏叶活性成分的提取分离研究概况", 《华西药学杂志》, vol. 17, no. 6, 31 December 2002 (2002-12-31), pages 437 - 439 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107698691A (en) * | 2017-11-02 | 2018-02-16 | 厦门福美科技有限公司 | A kind of separating and purifying flavone from oldenlandia diffusa, the system and method for polysaccharide |
| CN110152353A (en) * | 2019-06-18 | 2019-08-23 | 大连博迈科技发展有限公司 | A kind of continuous chromatography device and arasaponin production method |
| CN110152353B (en) * | 2019-06-18 | 2024-05-28 | 大连博迈科技发展有限公司 | Continuous chromatographic device and production method of total saponins of panax notoginseng |
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