CN109851646A - A kind of glutinous rehmannia effective component extraction separation method and the Catalpol and polysaccharide of extraction - Google Patents
A kind of glutinous rehmannia effective component extraction separation method and the Catalpol and polysaccharide of extraction Download PDFInfo
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Abstract
The present invention provides a kind of glutinous rehmannia effective component extraction separation method and the Catalpols and polysaccharide of extraction, Multiple components in glutinous rehmannia are extracted respectively, it gets rid of simple target and extracts isolated limitation, the utilization rate for substantially increasing glutinous rehmannia is finally reached the multiple target coproduction of Catalpol and polysaccharide, easy to operate, equipment requirement is low, 3-5 times is improved using enzymatic hydrolysis yield, important source material is saved, is easy to industrialization mass production.The obtained Catalpol monomer of the extraction separation method and monomers and polysaccharide, wherein the purity of Catalpol monomer is more than or equal to 98%.
Description
Technical field
The present invention relates to technical field of Chinese medicine, in particular to a kind of glutinous rehmannia effective component extraction separation method and
The Catalpol and polysaccharide of extraction.
Background technique
Glutinous rehmannia Rehmannia glutinosa Libosch is Scrophulariaceae glutinous rehmannia platymiscium also known as ground marrow, fragrant plant etc., root tuber
It is used as medicine, artificial cultivation, large area planting area is distributed in Yuncheng, Shanxi, Linfen, Henan Jiaozuo, weinan.Beijing, Hebei,
Also there is plantation in Shandong, Gansu.Due to the difference of Processing methods mode, medicinal material is divided into fresh rehmannia root, radix rehmanniae recen and Rehmannia glutinosa, is Chinese medicine
It is processed processing and the changed typical case of pharmacological property, fresh rehmannia root cure mainly in wound, by blood-arthralgia, fill out marrow, long muscle.The chemistry of glutinous rehmannia
Ingredient is based on glycoside, wherein again based on the iridoid glycosides such as Catalpol.Catalpol have anticancer, neuroprotection, anti-inflammatory, diuresis,
A variety of pharmacological activity such as hypoglycemic, liver protection and anti-aging, studies have shown that Catalpol has significant in the case where ischemia
Neuroprotective function, and find that Catalpol can activate brain neuron, it can treat because of wound, dysbolism, cerebral ischemia and Parkinson
Cranial nerve disease caused by family name's disease.Animal experiment in vitro, which shows Catalpol also, has anti-inflammatory, the intracellular Ca of resistance2+, promote PC12 cell
The effects of differentiation and inhibition Apoptosis.Catalpol content is the important Testing index of glutinous rehmannia Processing methods technique.Also contain in glutinous rehmannia
There are many polysaccharide, polysaccharide can eliminate the intracorporal free radical of people in glutinous rehmannia, have anti-hepatoma, antiviral, anti-aging and other effects, can
To treat cardiovascular disease, diabetes, there is dual regulation to human immune system, can be low with epidemic prevention power, it mentions
High immune function.
Plant resources of the China containing Catalpol are quite abundant, especially high-content plant, if glutinous rehmannia is the large common medicine in China
Material, this provides Resource Guarantee for Catalpol new drug development.For this purpose, should be by the chemical conversion of Catalpol and its derivative and pharmacodynamics knot
Study altogether, for illustrate glutinous rehmannia by Processing methods inherent mechanism that pharmacological property changes, open up a feasible road,
It should reinforce studying the activated product that may be generated after Catalpol and its degradation, the therapeutic agent to develop new lays the foundation.
In the prior art, generally have to the extraction of Catalpol following several: (1) organic solvent extraction is (with 5 times of methanol eddies
It extracts 2 times, each 2h)；(2) ultrasonic extraction (68% ethyl alcohol, solid-liquid ratio 5:1, ultrasonic 36min)；(3) microwave loss mechanisms；
(4) Enzymatic Extraction；(5) methanol extracts (in methanol concentration 70%, 55 DEG C of Extracting temperature, solid-liquid ratio is in 1:18).Above several sides
Method yield is lower, and other active drug ingredient wastes are serious in glutinous rehmannia, and extraction cost is higher.
In view of this, the present invention is specifically proposed.
Summary of the invention
The first object of the present invention is to provide a kind of glutinous rehmannia effective component extraction separation method, to solve the above problems,
The glutinous rehmannia effective component extraction separation method, extracts the Multiple components in glutinous rehmannia respectively, gets rid of simple target and extracts and divides
From limitation, substantially increase the utilization rate of glutinous rehmannia, be finally reached the multiple target coproduction of Catalpol and polysaccharide, easy to operate, equipment
It is required that it is low, 3-5 times is improved using enzymatic hydrolysis yield, important source material is saved, is easy to industrialization mass production.
The second object of the present invention is to provide the Catalpol of the extraction of glutinous rehmannia effective component extraction separation method described in one kind
Monomer and monomers and polysaccharide, wherein the purity of the Catalpol monomer is greater than 98%.
In order to realize above-mentioned purpose of the invention, the following technical scheme is adopted:
A kind of glutinous rehmannia effective component extraction separation method, comprising the following steps:
(1) enzyme will be added in glutinous rehmannia and water digests, successively use straight alcohol, volumetric concentration for the second of 65%-75%
Alcohol, the ethyl alcohol that volumetric concentration is 85%-95% and pure water extract, and merge all extracting solutions；
(2) extracting solution after the merging for obtaining step (1) through concentration and macroreticular resin after, then successively using pure water,
Volumetric concentration is the ethyl alcohol of 5%-15%, the ethyl alcohol of volumetric concentration 15%-25%, the ethyl alcohol of volumetric concentration 35%-45%, volume
The ethyl alcohol of concentration 55%-65% and the ethanol rinse chromatographic column of volumetric concentration 85%-95%；Volumetric concentration is 15%-25%'s
Catalpol extracting solution is obtained after ethanol rinse, obtains polysaccharide mixture extracting solution after the ethanol rinse that volumetric concentration is 85%-95%；
(3) Catalpol extracting solution and polysaccharide mixture extracting solution that step (2) obtains are concentrated and are dried respectively, obtained
The Catalpol monomer powders are further purified to obtain Catalpol monomer by Catalpol monomer powders and monomers and polysaccharide；
Preferably, the purifying is operated using silica column.
Preferably, in step (1), the enzyme is the 0.01%-0.1% of the glutinous rehmannia quality；It is furthermore preferred that enzyme is added
While suitable quantity of water is added, still more preferably, the quality of the water is 5-10 times of the glutinous rehmannia quality.
Preferably, the enzyme is selected from the group of one or more of pectase, cellulase, hemicellulase and protease
It closes；It is furthermore preferred that the temperature of the enzymatic hydrolysis is 30-70 DEG C；Temperature still more preferably is 40-50 DEG C；Still more preferably
, the time of the enzymatic hydrolysis is 24-48h.
Preferably, the enzyme is 1:(0.6-0.8 by mass ratio): (0.4-0.6): pectase, the cellulose of (0.3-0.7)
Enzyme, hemicellulase and protease composition；Preferred mass ratio is 1:0.7:0.5:0.5；Still more preferably, the egg
White enzyme is alkali protease.
Preferably, in step (1), the quality of the straight alcohol is 2.5-5 times of the glutinous rehmannia；The volumetric concentration is
The quality of the ethyl alcohol of 65%-75% is 5-10 times of the glutinous rehmannia；The quality of the ethyl alcohol of the 85%-95% is the glutinous rehmannia
5-10 times；The quality for the ethyl alcohol that the volumetric concentration is 85%-95% is 5-10 times of the glutinous rehmannia；The quality of the pure water is institute
5-10 times for stating glutinous rehmannia；
It is furthermore preferred that the temperature of the elution is 30-50 DEG C, it is still more preferably 40-50 DEG C；
It is furthermore preferred that the time of each elution is 30-60min, it is still more preferably 45-60min；
It is furthermore preferred that the temperature of the pure water is 90-100 DEG C.
Preferably, in step (2), the pure water volume is 1-2 times of the chromatography column volume；The volumetric concentration is
The ethyl alcohol of 5%-15% is 2-5 times of column volume of the chromatography；The ethyl alcohol that the volumetric concentration is 35%-45% is the chromatographic column
2-5 times of volume；The ethyl alcohol that the volumetric concentration is 55%-65% is 2-5 times of column volume of the chromatography；The volumetric concentration is
The ethyl alcohol of 85%-95% is 2-5 times of column volume of the chromatography.
Preferably, in step (1), the time of the extraction for the ethyl alcohol that the volumetric concentration is 65%-75% is 30-
60min, more preferably 45-60min.
Preferably, in step (1), the time of the extraction for the ethyl alcohol that the volumetric concentration is 85%-95% is 30-
60min, more preferably 45-60min.
Preferably, in step (2), the solvent of 70%-75% is fallen in the concentration for concentration.
Preferably, in step (2), the large aperture adsorption resin is selected from D101, AB-8, X-5, LX-60 or DM-130
One of.
The extracted Catalpol of glutinous rehmannia effective component extraction separation method and polysaccharide, it is preferred that the Catalpol it is pure
Degree is greater than 98%.
Compared with prior art, the invention has the benefit that
(1) glutinous rehmannia effective component extraction separation method provided herein, extracts the Multiple components in glutinous rehmannia respectively,
It gets rid of simple target and extracts isolated limitation, substantially increase the utilization rate of glutinous rehmannia, be finally reached the multiple target of Catalpol and polysaccharide
Coproduction.
(2) glutinous rehmannia effective component extraction separation method provided herein, easy to operate, equipment requirement is low, utilizes enzyme
It solves yield and improves 3-5 times, save important source material, be easy to industrialization mass production.
(3) purity for the Catalpol monomer that glutinous rehmannia effective component extraction separation method provided herein extracts is more than or equal to
98%.
Specific embodiment
Technical solution of the present invention is clearly and completely described below in conjunction with specific embodiment, but ability
Field technique personnel will be understood that following described embodiments are some of the embodiments of the present invention, instead of all the embodiments,
It is merely to illustrate the present invention, and is not construed as limiting the scope of the invention.Based on the embodiments of the present invention, the common skill in this field
Art personnel every other embodiment obtained without making creative work belongs to the model that the present invention protects
It encloses.The person that is not specified actual conditions in embodiment, carries out according to conventional conditions or manufacturer's recommended conditions.Agents useful for same or instrument
Production firm person is not specified, is the conventional products that can be obtained by commercially available purchase.
A kind of glutinous rehmannia effective component extraction separation method, comprising the following steps:
(1) enzyme will be added in glutinous rehmannia and water digests, successively use straight alcohol, volumetric concentration for the second of 65%-75%
Alcohol, the ethyl alcohol that volumetric concentration is 85%-95% and pure water extract, and merge all extracting solutions；
(2) extracting solution after the merging for obtaining step (1) through concentration and macroreticular resin after, then successively using pure water,
Volumetric concentration is the ethyl alcohol of 5%-15%, the ethyl alcohol of volumetric concentration 15%-25%, the ethyl alcohol of volumetric concentration 35%-45%, volume
The ethyl alcohol of concentration 55%-65% and the ethanol rinse chromatographic column of volumetric concentration 85%-95%；Volumetric concentration is 15%-25%'s
Catalpol extracting solution is obtained after ethanol rinse, obtains polysaccharide mixture extracting solution after the ethanol rinse that volumetric concentration is 85%-95%；
(3) Catalpol extracting solution and polysaccharide mixture extracting solution that step (2) obtains are concentrated and are dried respectively, obtained
The Catalpol monomer powders are further purified to obtain Catalpol monomer by Catalpol monomer powders and monomers and polysaccharide；
Preferably, the purifying is operated using silica column.
Glutinous rehmannia effective component extraction separation method provided by the present invention is first to raw material clean the surface crushed after being dried, so
Enzyme is added afterwards to be digested, carries out alcohol extracting three times in the 30-50 DEG C of ethyl alcohol using various concentration after enzymatic hydrolysis, is eventually adding water extraction
Once.It is gradually eluted after extraction, subsection receiing.After the solvent for extract three times being mixed and being concentrated 70%-75%, mistake
D101 or AB-8 macroreticular resin, respectively with water (1-2 times separates column volume) --- 5-15% ethyl alcohol (2-5V) --- 15-25%
Ethyl alcohol (2-5V) --- 35-45% ethyl alcohol (2-5V) --- 55-65% ethyl alcohol (2-5V) --- 85-95% ethyl alcohol (2-5V),
In obtain Catalpol after 20% ethanol rinse, obtain polysaccharide mixture after 35-45% ethyl alcohol -85-95% ethanol rinse.Finally incite somebody to action
To Catalpol solution and polysaccharide mixture solution concentration drying obtain corresponding monomer powders.
Preferably, in step (1), the enzyme is the 0.01%-0.1% of the glutinous rehmannia quality；It is furthermore preferred that enzyme is added
While suitable quantity of water is added, still more preferably, the quality of the water is 5-10 times of the glutinous rehmannia quality.
Preferably, the enzyme is selected from the group of one or more of pectase, cellulase, hemicellulase and protease
It closes；It is furthermore preferred that the temperature of the enzymatic hydrolysis is 30-70 DEG C；Temperature still more preferably is 40-50 DEG C；Still more preferably
, the time of the enzymatic hydrolysis is 24-48h.
Preferably, the enzyme is 1:(0.6-0.8 by mass ratio): (0.4-0.6): pectase, the cellulose of (0.3-0.7)
Enzyme, hemicellulase and protease composition；Preferred mass ratio is 1:0.7:0.5:0.5；Still more preferably, the egg
White enzyme is alkali protease.
Preferably, in step (1), the quality of the straight alcohol is 2.5-5 times of the glutinous rehmannia；The volumetric concentration is
The quality of the ethyl alcohol of 65%-75% is 5-10 times of the glutinous rehmannia；The quality of the ethyl alcohol of the 85%-95% is the glutinous rehmannia
5-10 times；The quality for the ethyl alcohol that the volumetric concentration is 85%-95% is 5-10 times of the glutinous rehmannia；The quality of the pure water is institute
5-10 times for stating glutinous rehmannia；
It is furthermore preferred that the temperature of the elution is 30-50 DEG C, it is still more preferably 40-50 DEG C；
It is furthermore preferred that the time of each elution is 30-60min, it is still more preferably 45-60min；
It is furthermore preferred that the temperature of the pure water is 90-100 DEG C.
Preferably, in step (2), the pure water volume is 1-2 times of the chromatography column volume；The volumetric concentration is
The ethyl alcohol of 5%-15% is 2-5 times of column volume of the chromatography；The ethyl alcohol that the volumetric concentration is 35%-45% is the chromatographic column
2-5 times of volume；The ethyl alcohol that the volumetric concentration is 55%-65% is 2-5 times of column volume of the chromatography；The volumetric concentration is
The ethyl alcohol of 85%-95% is 2-5 times of column volume of the chromatography.
Preferably, in step (1), the time of the extraction for the ethyl alcohol that the volumetric concentration is 65%-75% is 30-
60min, more preferably 45-60min.
Preferably, in step (1), the time of the extraction for the ethyl alcohol that the volumetric concentration is 85%-95% is 30-
60min, more preferably 45-60min.
Preferably, in step (2), the solvent of 70%-75% is fallen in the concentration for concentration.
Preferably, in step (2), the large aperture adsorption resin is selected from D101, AB-8, X-5, LX-60 or DM-130
One of.
The extracted Catalpol of glutinous rehmannia effective component extraction separation method and polysaccharide, it is preferred that the Catalpol it is pure
Degree is greater than 98%.
The structural formula of Catalpol is as follows:
Embodiment 1
Glutinous rehmannia effective component extraction separation method provided by the present embodiment, specifically includes the following steps:
1) fresh glutinous rehmannia and cauline leaf chopping are taken, the mixing of the pectase and cellulase of glutinous rehmannia quality 0.01% is added
5 times of glutinous rehmannia quality of water is added in object, and temperature control digests 48h at 30 DEG C.
2) 2.5 times of straight alcohols of glutinous rehmannia quality are added, 30min is extracted at 30 DEG C, filter, filter cake continues plus 5 times of 70% second
Alcohol extracting 30min, filtering, filter cake continues plus 5 times of 90% ethyl alcohol extracts 30min, filtering, and filter cake adds 5 times of 90 DEG C of Aqua pure extracts
30min.Filtering obtains filter residue and filtrate, can be used as plant bacterial manure after filter residue drying, mix finally by the extracting solution of all steps
Extracting solution.
3) 70% solvent is fallen into the concentration of final extracting solution, then the ethyl alcohol that solvent recovery is 70% or so crosses macroporous absorption
Resin D101 successively chromatographs pure water, 2 times of 10% ethyl alcohol of chromatography column volume, 2 times of chromatography 20% second of column volume of column volume with 1 times
Alcohol, 2 times of 40% ethyl alcohol of chromatography column volume, 2 times of 60% ethyl alcohol of chromatography column volume, 2 times of chromatography 90% ethanol rinse of column volume chromatographies
Column；
Wherein, Catalpol extracting solution is obtained after 20% ethanol rinse, and polysaccharide mixture extracting solution is obtained after 90% ethanol rinse.
4) drying is concentrated in Catalpol extracting solution and polysaccharide mixture extracting solution respectively, obtains Catalpol monomer powders and polysaccharide list
Catalpol monomer powders are further purified to obtain the Catalpol monomer of high-purity through silica column for body.
Embodiment 2
Glutinous rehmannia effective component extraction separation method provided by the present embodiment, specifically includes the following steps:
1) fresh glutinous rehmannia and cauline leaf chopping are taken, pectase, cellulase and the hemicellulose of glutinous rehmannia quality 0.1% is added
10 times of glutinous rehmannia quality of water is added in enzyme, and temperature control is at 50 DEG C, and enzymatic hydrolysis is for 24 hours.
2) 5 times of straight alcohols of glutinous rehmannia quality are added, 60min is extracted at 50 DEG C, filter, filter cake continues plus 10 times of 70% ethyl alcohol
60min, filtering are extracted, filter cake continues plus 10 times of 90% ethyl alcohol extracts 60min, filtering, and filter cake adds 10 times of 100 DEG C of Aqua pure extracts
60min.Filtering obtains filter residue and filtrate, can be used as plant bacterial manure after filter residue drying, mix finally by the extracting solution of all steps
Extracting solution.
3) 75% solvent is fallen into the concentration of final extracting solution, then the ethyl alcohol that solvent recovery is 70% or so crosses macroporous absorption
Resin A B-8 successively chromatographs pure water, 5 times of 10% ethyl alcohol of chromatography column volume, 5 times of chromatography 20% second of column volume of column volume with 2 times
Alcohol, 5 times of 40% ethyl alcohol of chromatography column volume, 5 times of 60% ethyl alcohol of chromatography column volume, 5 times of 90% ethanol rinse chromatographic columns of volume；
Wherein, Catalpol extracting solution is obtained after 20% ethanol rinse, and polysaccharide mixture extracting solution is obtained after 90% ethanol rinse.
4) drying is concentrated in Catalpol extracting solution and polysaccharide mixture extracting solution respectively, obtains Catalpol monomer powders and polysaccharide list
Catalpol monomer powders are further purified to obtain the Catalpol monomer of high-purity through silica column for body.
Embodiment 3
Glutinous rehmannia effective component extraction separation method provided by the present embodiment, specifically includes the following steps:
1) fresh glutinous rehmannia and cauline leaf chopping are taken, the pectase of glutinous rehmannia quality 0.05% is added, is added 7 times of glutinous rehmannia quality
Water, temperature control digest 36h at 40 DEG C.
2) 3 times of straight alcohols of glutinous rehmannia quality are added, 40min is extracted at 40 DEG C, filter, filter cake continues plus 7 times of 70% ethyl alcohol
40min, filtering are extracted, filter cake continues plus 7 times of 90% ethyl alcohol extracts 40min, filtering, and filter cake adds 7 times 95 DEG C of pure water, extracts
4min.Filtering obtains filter residue and filtrate, can be used as plant bacterial manure after filter residue drying, mix finally by the extracting solution of all steps
Extracting solution.
3) 70% solvent is fallen into the concentration of final extracting solution, then the ethyl alcohol that solvent recovery is 70% or so crosses macroporous absorption
Resin (D101, AB-8, X-5, LX-60 or DM-130) successively chromatographs pure water, the 3 times of chromatography column volumes 10% of column volume with 2 times
Ethyl alcohol, 3 times of 20% ethyl alcohol of chromatography column volume, 3 times of 40% ethyl alcohol of chromatography column volume, 3 times of 60% ethyl alcohol of chromatography column volume, 3 times of layers
Analyse 90% ethanol rinse chromatographic column of column volume.
Wherein, Catalpol extracting solution is obtained after 20% ethanol rinse, and polysaccharide mixture extracting solution is obtained after 90% ethanol rinse.
4) drying is concentrated in Catalpol extracting solution and polysaccharide mixture extracting solution respectively, obtains Catalpol monomer powders and polysaccharide list
Catalpol monomer powders are further purified to obtain the Catalpol monomer of high-purity through silica column for body.
Embodiment 4
Glutinous rehmannia effective component extraction separation method provided by the present embodiment, specifically includes the following steps:
1) fresh glutinous rehmannia and cauline leaf chopping are taken, pectase, cellulase, the hemicellulose of glutinous rehmannia quality 0.08% is added
8 times of glutinous rehmannia quality of water is added in enzyme and protease, and temperature control digests 24-48h at 45 DEG C.
2) 2.5-5 times of straight alcohol of glutinous rehmannia quality is added, 45min is extracted at 40 DEG C, filters, filter cake continues to add 8 times 70%
Ethyl alcohol extracts 45min, filtering, and filter cake continues plus 8 times of 90% ethyl alcohol extracts 45min, filtering, and filter cake adds 8 times 90 DEG C of pure water, mentions
Take 45min.Filtering obtains filter residue and filtrate, can be used as plant bacterial manure after filter residue drying, mix most by the extracting solution of all steps
Whole extracting solution.
3) 70% solvent is fallen into the concentration of final extracting solution, then the ethyl alcohol that solvent recovery is 70% or so crosses macroporous absorption
Resin DM-130 successively chromatographs pure water, 4 times of 10% ethyl alcohol of volume, 4 times of 20% ethyl alcohol of volume, 4 times of volumes of column volume with 1 times
40% ethyl alcohol, 4 times of 60% ethyl alcohol of volume, 4 times of 90% ethanol rinse chromatographic columns of volume.
Wherein, Catalpol extracting solution is obtained after 20% ethanol rinse, and polysaccharide mixture extracting solution is obtained after 90% ethanol rinse.
4) drying is concentrated in Catalpol extracting solution and polysaccharide mixture extracting solution respectively, obtains Catalpol monomer powders and polysaccharide list
Catalpol monomer powders are further purified to obtain the Catalpol monomer of high-purity through silica column for body.
Embodiment 5 is substantially the same manner as Example 4, unlike, in the composition of enzyme, use mass ratio for 1:0.7:0.5:
0.5 pectase, cellulase, hemicellulase and alkali protease composition.
Embodiment 6 is substantially the same manner as Example 4, unlike, in the composition of enzyme, use mass ratio for 1:0.6:0.4:
0.3 pectase, cellulase, hemicellulase and alkali protease composition.
Embodiment 7 is substantially the same manner as Example 4, unlike, in the composition of enzyme, use mass ratio for 1:0.8:0.6:
0.7 pectase, cellulase, hemicellulase and alkali protease composition.
Comparative example 1 is substantially the same manner as Example 4, unlike, without enzymatic hydrolysis, directly extract step.
Comparative example 2 is substantially the same manner as Example 4, unlike, in step (3), the volumetric concentration of the ethyl alcohol of extraction according to
It is secondary are as follows: 4%, 10%, 30%, 50% and 80%.
Comparative example 3 is substantially the same manner as Example 4, unlike, in the composition of enzyme, use mass ratio for 1:0.5:0.3:
0.3 pectase, cellulase, hemicellulase and alkali protease composition.
Experimental example Catalpol and polyoses content test
(1) experimental material: fresh rehmannia root (originates from Henan Jiaozuo)；Acetonitrile (chromatographically pure)；Pure water.
(2) laboratory apparatus, as shown in table 1.
1 laboratory apparatus of table
(3) method provided by the embodiment of the present application 1-7 and comparative example 1-3 is respectively adopted and extracts experiment.Experiment knot
Fruit is as shown in table 2.
2 row of table extracts experimental results
The experimental results showed that glutinous rehmannia effective component extraction separation method provided herein, Catalpol and polysaccharide yield are high,
The utilization rate for substantially increasing glutinous rehmannia is finally reached the multiple target coproduction of Catalpol and polysaccharide, and easy to operate, equipment requirement is low, benefit
3-5 times is improved with enzymatic hydrolysis yield, important source material is saved, is easy to industrialization mass production.Also, obtained Catalpol purity is high,
Greater than 98%.
Wherein, embodiment 5,6 and 7 is digested using the combination of specific enzyme, can effectively improve the purity of Catalpol,
Effect is best under the proportion of middle embodiment 5, and the purity of Catalpol reaches maximum value.Compared with comparative example 3, the proportion of enzyme is not preferred
In the case where range, the purity of obtained Catalpol is lower than the purity of the obtained Catalpol under preferred scope.
And in comparative example 1, enzymatic hydrolysis operation is not carried out using any enzyme, will be greatly reduced the purity for extracting obtained Catalpol.
And pass through the comparison of comparative example 2, it is not eluted using the ethyl alcohol of ethyl alcohol volume concentration range provided herein, can also be made
It is reduced at the purity of the Catalpol of extraction.Thus illustrating, extracting method provided herein effectively increases the purity of Catalpol,
Moreover, polyoses extract can also be further obtained, the multiple target coproduction of Catalpol and polysaccharide is realized, substantially increases glutinous rehmannia
Utilization rate, and method is easy to operate, and equipment requirement is low, saves important source material, is easy to industrialization mass production.
Although illustrate and describing the present invention with specific embodiment, it will be appreciated that the above various embodiments is only used
To illustrate technical solution of the present invention, rather than its limitations；Those skilled in the art should understand that: without departing substantially from this hair
It in the case where bright spirit and scope, is possible to modify the technical solutions described in the foregoing embodiments, or to wherein
Some or all of technical characteristic is equivalently replaced；And these are modified or replaceed, and do not make the essence of corresponding technical solution
It departs from the scope of the technical solutions of the embodiments of the present invention；It is, therefore, intended that in the following claims including belonging to the present invention
All these substitutions and modifications in range.
Claims (10)
1. a kind of glutinous rehmannia effective component extraction separation method, which comprises the following steps:
(1) enzyme will be added in glutinous rehmannia and water digests, successively use straight alcohol, volumetric concentration for the ethyl alcohol of 65%-75%, body
Product concentration is that the ethyl alcohol of 85%-95% and pure water extract, and merges all extracting solutions；
(2) extracting solution after the merging for obtaining step (1) is after concentration and macroreticular resin, then successively uses pure water, volume
Concentration is the ethyl alcohol of 5%-15%, the ethyl alcohol of volumetric concentration 15%-25%, the ethyl alcohol of volumetric concentration 35%-45%, volumetric concentration
The ethyl alcohol of 55%-65% and the ethanol rinse chromatographic column of volumetric concentration 85%-95%；Volumetric concentration is the ethyl alcohol of 15%-25%
Catalpol extracting solution is obtained after elution, obtains polysaccharide mixture extracting solution after the ethanol rinse that volumetric concentration is 85%-95%；
Preferably, ethyl alcohol, the ethyl alcohol of volumetric concentration 20%, volumetric concentration 40% of pure water, volumetric concentration for 10% are successively used
Ethyl alcohol, the ethyl alcohol of volumetric concentration 60% and the ethanol rinse chromatographic column of volumetric concentration 90%, volumetric concentration be 20% ethyl alcohol
Catalpol extracting solution is obtained after elution, obtains polysaccharide mixture extracting solution after the ethanol rinse that volumetric concentration is 90%；
(3) Catalpol extracting solution and polysaccharide mixture extracting solution that step (2) obtains are concentrated and are dried respectively, obtain Catalpol
The Catalpol monomer powders are further purified to obtain Catalpol monomer by monomer powders and monomers and polysaccharide；
Preferably, the purifying is operated using silica column.
2. glutinous rehmannia effective component extraction separation method according to claim 1, which is characterized in that described in step (1)
Enzyme is the 0.01%-0.1% of the glutinous rehmannia quality；
Preferably, suitable quantity of water is added while enzyme is added, it is furthermore preferred that the quality of the water is the 5-10 of the glutinous rehmannia quality
Times；
Preferably, the enzyme is selected from the combination of one or more of pectase, cellulase, hemicellulase and protease；
Preferably, the temperature of the enzymatic hydrolysis is 30-70 DEG C；Preferred temperature is 40-50 DEG C；It is furthermore preferred that the enzymatic hydrolysis
Time is 24-48h.
3. glutinous rehmannia effective component extraction separation method according to claim 2, which is characterized in that the enzyme is by mass ratio
1:(0.6-0.8): (0.4-0.6): pectase, cellulase, hemicellulase and the protease composition of (0.3-0.7)；It is preferred that
Mass ratio be 1:0.7:0.5:0.5；It is furthermore preferred that the protease is alkali protease.
4. glutinous rehmannia effective component extraction separation method according to claim 1, which is characterized in that described in step (1)
The quality of straight alcohol is 2.5-5 times of the glutinous rehmannia；The quality for the ethyl alcohol that the volumetric concentration is 65%-75% is the glutinous rehmannia
5-10 times；The quality of the ethyl alcohol of the 85%-95% is 5-10 times of the glutinous rehmannia；The volumetric concentration is 85%-95%
Ethyl alcohol quality be 5-10 times of the glutinous rehmannia；The quality of the pure water is 5-10 times of the glutinous rehmannia；
Preferably, the temperature of the elution is 30-50 DEG C, more preferably 40-50 DEG C；
Preferably, the time of the elution is 30-60min, more preferably 45-60min every time；
Preferably, the temperature of the pure water is 90-100 DEG C.
5. glutinous rehmannia effective component extraction separation method according to claim 1, which is characterized in that described in step (2)
Pure water volume is 1-2 times of the chromatography column volume；The ethyl alcohol that the volumetric concentration is 5%-15% is the chromatography column volume
2-5 times；The ethyl alcohol that the volumetric concentration is 35%-45% is 2-5 times of column volume of the chromatography；The volumetric concentration is 55%-
65% ethyl alcohol is 2-5 times of column volume of the chromatography；The ethyl alcohol that the volumetric concentration is 85%-95% is the chromatography column volume
2-5 times.
6. glutinous rehmannia effective component extraction separation method according to claim 1, which is characterized in that described in step (1)
The time of the extraction for the ethyl alcohol that volumetric concentration is 65%-75% is 30-60min, preferably 45-60min.
7. glutinous rehmannia effective component extraction separation method according to claim 1, which is characterized in that described in step (1)
The time of the extraction for the ethyl alcohol that volumetric concentration is 85%-95% is 30-60min, preferably 45-60min.
8. glutinous rehmannia effective component extraction separation method according to claim 1, which is characterized in that described in step (2)
The solvent of 70%-75% is fallen in concentration for concentration.
9. glutinous rehmannia effective component extraction separation method according to claim 1-8, which is characterized in that in step
(2) in, the large aperture adsorption resin is selected from one of D101, AB-8, X-5, LX-60 or DM-130.
10. the extracted Catalpol of -9 described in any item glutinous rehmannia effective component extraction separation methods and polysaccharide according to claim 1,
Preferably, the purity of the Catalpol is greater than 98%.
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