CN116019848A - Method for extracting saikosaponin from bupleurum - Google Patents
Method for extracting saikosaponin from bupleurum Download PDFInfo
- Publication number
- CN116019848A CN116019848A CN202310191883.8A CN202310191883A CN116019848A CN 116019848 A CN116019848 A CN 116019848A CN 202310191883 A CN202310191883 A CN 202310191883A CN 116019848 A CN116019848 A CN 116019848A
- Authority
- CN
- China
- Prior art keywords
- bupleurum
- saikosaponin
- extracting
- powder
- alkyl glycoside
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 241000202726 Bupleurum Species 0.000 title claims abstract description 147
- KYWSCMDFVARMPN-LCSVLAELSA-N Saikosaponin D Chemical compound O([C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@]([C@H]3[C@]([C@@H]4[C@@]([C@@]5(C[C@@H](O)[C@]67CO[C@]5([C@@H]6CC(C)(C)CC7)C=C4)C)(C)CC3)(C)CC2)(C)CO)O[C@@H]([C@@H]1O)C)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O KYWSCMDFVARMPN-LCSVLAELSA-N 0.000 title claims abstract description 77
- 229930192014 saikosaponin Natural products 0.000 title claims abstract description 77
- 238000000034 method Methods 0.000 title claims abstract description 62
- 229930182470 glycoside Natural products 0.000 claims abstract description 63
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 62
- 239000000843 powder Substances 0.000 claims abstract description 58
- FCBUKWWQSZQDDI-UHFFFAOYSA-N rhamnolipid Chemical compound CCCCCCCC(CC(O)=O)OC(=O)CC(CCCCCCC)OC1OC(C)C(O)C(O)C1OC1C(O)C(O)C(O)C(C)O1 FCBUKWWQSZQDDI-UHFFFAOYSA-N 0.000 claims abstract description 48
- KWIUHFFTVRNATP-UHFFFAOYSA-N Betaine Natural products C[N+](C)(C)CC([O-])=O KWIUHFFTVRNATP-UHFFFAOYSA-N 0.000 claims abstract description 35
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 35
- KWIUHFFTVRNATP-UHFFFAOYSA-O N,N,N-trimethylglycinium Chemical compound C[N+](C)(C)CC(O)=O KWIUHFFTVRNATP-UHFFFAOYSA-O 0.000 claims abstract description 35
- 125000000217 alkyl group Chemical group 0.000 claims abstract description 35
- 229960003237 betaine Drugs 0.000 claims abstract description 35
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims abstract description 35
- 150000002338 glycosides Chemical class 0.000 claims abstract description 35
- 108090000790 Enzymes Proteins 0.000 claims abstract description 24
- 102000004190 Enzymes Human genes 0.000 claims abstract description 24
- 238000007710 freezing Methods 0.000 claims abstract description 19
- 230000008014 freezing Effects 0.000 claims abstract description 19
- 238000000605 extraction Methods 0.000 claims abstract description 18
- 239000011347 resin Substances 0.000 claims abstract description 17
- 229920005989 resin Polymers 0.000 claims abstract description 17
- 238000004108 freeze drying Methods 0.000 claims abstract description 15
- 239000003480 eluent Substances 0.000 claims abstract description 12
- 238000001179 sorption measurement Methods 0.000 claims abstract description 11
- 238000010992 reflux Methods 0.000 claims abstract description 7
- 239000007788 liquid Substances 0.000 claims abstract description 6
- 238000002791 soaking Methods 0.000 claims abstract description 6
- 238000005406 washing Methods 0.000 claims abstract description 6
- 230000000415 inactivating effect Effects 0.000 claims abstract description 5
- 101710121765 Endo-1,4-beta-xylanase Proteins 0.000 claims description 37
- 108010059892 Cellulase Proteins 0.000 claims description 32
- 229940106157 cellulase Drugs 0.000 claims description 32
- -1 alkyl glycoside Chemical class 0.000 claims description 28
- 229940088598 enzyme Drugs 0.000 claims description 23
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 20
- 239000000243 solution Substances 0.000 claims description 20
- 239000007864 aqueous solution Substances 0.000 claims description 5
- 239000003463 adsorbent Substances 0.000 claims 1
- 238000012546 transfer Methods 0.000 abstract description 26
- 230000001105 regulatory effect Effects 0.000 abstract 1
- 239000001397 quillaja saponaria molina bark Substances 0.000 description 31
- 229930182490 saponin Natural products 0.000 description 31
- 150000007949 saponins Chemical class 0.000 description 31
- 230000000052 comparative effect Effects 0.000 description 22
- 229920002678 cellulose Polymers 0.000 description 20
- 239000001913 cellulose Substances 0.000 description 20
- 239000004094 surface-active agent Substances 0.000 description 19
- 230000000694 effects Effects 0.000 description 18
- 229920001277 pectin Polymers 0.000 description 15
- 229920002488 Hemicellulose Polymers 0.000 description 10
- AEMOLEFTQBMNLQ-BKBMJHBISA-N alpha-D-galacturonic acid Chemical compound O[C@H]1O[C@H](C(O)=O)[C@H](O)[C@H](O)[C@H]1O AEMOLEFTQBMNLQ-BKBMJHBISA-N 0.000 description 10
- 210000002421 cell wall Anatomy 0.000 description 10
- 239000013078 crystal Substances 0.000 description 8
- 150000001875 compounds Chemical class 0.000 description 6
- 238000004090 dissolution Methods 0.000 description 5
- 239000007791 liquid phase Substances 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 230000001737 promoting effect Effects 0.000 description 5
- 239000000758 substrate Substances 0.000 description 5
- 238000009736 wetting Methods 0.000 description 5
- 230000009849 deactivation Effects 0.000 description 4
- 239000007790 solid phase Substances 0.000 description 4
- 210000004185 liver Anatomy 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 239000002888 zwitterionic surfactant Substances 0.000 description 3
- 108010059820 Polygalacturonase Proteins 0.000 description 2
- 206010037660 Pyrexia Diseases 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 108010093305 exopolygalacturonase Proteins 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 230000006872 improvement Effects 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 1
- 206010008479 Chest Pain Diseases 0.000 description 1
- SHZGCJCMOBCMKK-UHFFFAOYSA-N D-mannomethylose Natural products CC1OC(O)C(O)C(O)C1O SHZGCJCMOBCMKK-UHFFFAOYSA-N 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 208000019255 Menstrual disease Diseases 0.000 description 1
- 206010038084 Rectocele Diseases 0.000 description 1
- 206010046814 Uterine prolapse Diseases 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 230000001914 calming effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000007071 enzymatic hydrolysis Effects 0.000 description 1
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 238000005470 impregnation Methods 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 201000004792 malaria Diseases 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 201000009240 nasopharyngitis Diseases 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 150000008265 rhamnosides Chemical class 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
- 238000003809 water extraction Methods 0.000 description 1
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses a method for extracting saikosaponin from bupleurum, which comprises the following steps: firstly, crushing bupleurum into bupleurum powder, soaking the bupleurum powder into water containing rhamnolipid and alkyl glycoside carboxyl betaine in an amount which is 3-5 times that of the bupleurum powder, freezing, crushing into ice residues by using a crusher, carrying out enzymolysis after the ice residues are melted, inactivating enzyme, and freeze-drying; step two, adding 6-8 times of ethanol solution with the volume fraction of 60-80% into the bupleurum processed in the step one, regulating the pH to be alkaline, carrying out reflux extraction for 2-3 times at 70-80 ℃ for 1-2 h each time, and combining the extracting solutions of the times; step three, the combined extracting solution is put on a macroporous adsorption resin column, then is washed by alkaline water and pure water in sequence, the washing liquid is discarded, then is eluted by ethanol solution with the volume fraction of 60-70%, and the eluent is collected; and step four, concentrating the eluent under reduced pressure, and freeze-drying. The invention can improve the transfer rate and the product purity of the saikosaponin.
Description
Technical Field
The invention relates to the field of bupleurum root extracts. More particularly, the present invention relates to a method for extracting saikosaponin from bupleurum.
Background
Bupleuri radix is a common traditional Chinese medicine, and has effects of relieving exterior and interior, soothing liver and invigorating yang. Can be used for treating common cold, fever, cold and heat, malaria, liver depression, qi stagnation, chest pain, rectocele, uterine prolapse, and menoxenia. The bupleurum root mainly contains saponin, volatile oil, polysaccharide and other components, wherein the saponin component is the main active component. Pharmacological experiments prove that the saikosaponin has the effects of relieving fever, resisting bacteria, resisting inflammation, calming, resisting viruses, protecting liver and the like. The existing extraction method of the traditional Chinese medicine saponin comprises an alcohol extraction method, a water extraction method, a macroporous adsorption resin method, an impregnation method, a reflux method and the like, and the alcohol extraction method is a method which is more used for extracting the saikosaponin, but has the problems of lower saikosaponin transfer rate and lower product purity.
Disclosure of Invention
The invention aims to provide a method for extracting saikosaponin from bupleurum, which comprises the steps of firstly mixing bupleurum powder with water containing rhamnolipid and alkyl glycoside carboxyl betaine, freezing and crushing, then carrying out enzymolysis, damaging the structure of cellulose through freezing and crushing, and in the enzymolysis process, the compound surfactant consisting of rhamnolipid and alkyl glycoside carboxyl betaine can improve the activity of cellulase, xylanase and pectinase, thereby improving the enzymolysis efficiency, further promoting the rupture of the cell wall of bupleurum rootstock, fully releasing saponin, improving the transfer rate of the saponin, carrying out alcohol extraction for multiple times, extracting the saponin in bupleurum, purifying the saikosaponin through macroporous resin in the later stage, and improving the purity of the saikosaponin.
To achieve these objects and other advantages and in accordance with the purpose of the invention, there is provided a method for extracting saikosaponin from bupleurum, comprising:
step one, crushing bupleurum into bupleurum powder, soaking the bupleurum powder in water containing rhamnolipid and alkyl glycoside carboxyl betaine, freezing the bupleurum powder, crushing the bupleurum powder into ice residues by using a crusher, adding cellulase, xylanase and pectase for enzymolysis after the ice residues are melted at room temperature, inactivating the enzyme, and freeze-drying;
step two, adding ethanol solution with the volume fraction of 60-80% which is 6-8 times of the weight of the bupleurum processed in the step one, adjusting the pH value to be alkaline, carrying out reflux extraction for 2-3 times at 70-80 ℃ for 1-2 h each time, and combining the extracting solutions of each time;
step three, the combined extracting solution is put on a macroporous adsorption resin column, then is washed by alkaline water and pure water in sequence, the washing liquid is discarded, then is eluted by ethanol solution with the volume fraction of 60-70%, and the eluent is collected;
and step four, concentrating the eluent under reduced pressure, and freeze-drying to obtain the saikosaponin powder.
Preferably, in the method for extracting saikosaponin from bupleurum, the mass fractions of rhamnolipid and alkyl glycoside carboxyl betaine in water containing rhamnolipid and alkyl glycoside carboxyl betaine are respectively 0.005-0.015% and 0.03-0.05%.
Preferably, in the method for extracting saikosaponin from bupleurum, cellulase is added according to 15-20 FPU/g bupleurum powder, xylanase is added according to 200-250U/g bupleurum powder, and pectase is added according to 300-600U/g bupleurum powder.
Preferably, in the method for extracting saikosaponin from bupleurum, the freezing temperature in the first step is-30 to-20 ℃ and the freezing time is 20-30 hours.
Preferably, in the method for extracting saikosaponin from bupleurum, naOH and Na are adopted in the second step 2 CO 3 Or KOH to adjust the pH to 9-14.
Preferably, in the method for extracting saikosaponin from bupleurum, the macroporous adsorption resin column is D101.
Preferably, in the method for extracting saikosaponin from bupleurum, the alkaline water is 2% NaOH aqueous solution by mass fraction.
Preferably, in the method for extracting saikosaponin from bupleurum, the bupleurum powder is crushed into 40-60 meshes.
Preferably, in the method for extracting saikosaponin from bupleurum, the enzymolysis temperature in the first step is 30-50 ℃ and the enzymolysis time is 2-3 h.
Preferably, in the method for extracting saikosaponin from bupleurum, the mass fractions of rhamnolipid and alkyl glycoside carboxyl betaine in water containing rhamnolipid and alkyl glycoside carboxyl betaine are respectively 0.01% and 0.04%.
The invention at least comprises the following beneficial effects:
the invention firstly mixes bupleurum powder with water containing rhamnolipid and alkyl glycoside carboxyl betaine, then freezes and breaks, and then adds cellulase, xylanase and pectase for enzymolysis. When freezing, the ice crystal grows and breaks, the ice crystal pierces the cell wall, and damages the structure of cellulose, which is beneficial to the enzymolysis of cellulose in the later stage. In the enzymolysis process, the compound surfactant composed of rhamnolipid and alkyl glycoside carboxyl betaine can avoid ineffective combination of cellulose, hemicellulose, pectic substance, cellulase, xylanase and pectic enzyme, increase the contact area of the cellulase, xylanase, pectic enzyme, cellulose, hemicellulose and pectic substance, improve the activity of the xylanase, and further improve the activity of the xylanase by the cooperation of rhamnolipid and alkyl glycoside carboxyl betaine, and simultaneously improve the activity of the cellulase and the pectic enzyme, thereby improving the integral enzymolysis efficiency, further promoting the rupture of the cell wall of bupleurum root and stem, fully releasing the saponin, improving the transfer rate of the saponin, carrying out repeated alcohol extraction, extracting the saponin in bupleurum, purifying the saponin by macroporous resin in the later stage, and improving the purity of the bupleurum saponin. In the alcohol extraction process, the existence of the surfactants rhamnolipid and alkyl glycoside carboxyl betaine can reduce the interfacial tension of solid and liquid phases, greatly enhance the wetting and dissolution exudation capacity of the surfactant on bupleurum substrates, and greatly improve the transfer rate of saponin in bupleurum as a whole.
Additional advantages, objects, and features of the invention will be set forth in part in the description which follows and in part will become apparent to those having ordinary skill in the art upon examination of the following or may be learned from practice of the invention.
Detailed Description
The present invention is described in further detail below with reference to examples to enable those skilled in the art to practice the same by referring to the description.
The experimental methods described in the following embodiments are conventional methods unless otherwise indicated, and the reagents and materials are commercially available.
Example 1
A method for extracting saikosaponin from bupleurum, comprising:
step one, crushing bupleurum into bupleurum powder, soaking the bupleurum powder in water containing rhamnolipid and alkyl glycoside carboxyl betaine, freezing the bupleurum powder, crushing the bupleurum powder into ice residues by using a crusher, adding cellulase, xylanase and pectase for enzymolysis after the ice residues are melted at room temperature, inactivating the enzyme, and freeze-drying the obtained product; alkyl glycoside carboxybetaine an alkyl glycoside carboxybetaine zwitterionic surfactant with an authorized publication number of CN111013484B and the alkyl glycoside carboxybetaine prepared in example 2 of the preparation method thereof are adopted.
Step two, adding ethanol solution with the volume fraction of 60% and the weight of which is 6 times that of the bupleurum processed in the step one, carrying out reflux extraction for 2 times at 70 ℃ for 1h each time, and combining the extracting solutions of each time;
step three, the combined extracting solution is put on a macroporous adsorption resin column, sequentially washed by alkaline water and pure water, discarded by washing liquid, eluted by ethanol solution with the volume fraction of 60%, and the eluent is collected;
and step four, concentrating the eluent under reduced pressure, and freeze-drying to obtain the saikosaponin powder.
According to the invention, bupleurum powder is firstly mixed with water containing rhamnolipid and alkyl glycoside carboxyl betaine, then frozen and crushed, and then cellulase, xylanase and pectase are added for enzymolysis, when frozen, ice crystal grows, and when crushed, the ice crystal pierces cell walls, so that the structure of cellulose is damaged, and the enzymolysis of cellulose in later stage is facilitated. In the enzymolysis process, the compound surfactant composed of rhamnolipid and alkyl glycoside carboxyl betaine can avoid ineffective combination of cellulose, hemicellulose, pectic substance, cellulase, xylanase and pectic enzyme, increase the contact area of the cellulase, xylanase, pectic enzyme, cellulose, hemicellulose and pectic substance, improve the activity of the xylanase, and further improve the activity of the xylanase by the cooperation of rhamnolipid and alkyl glycoside carboxyl betaine, and simultaneously improve the activity of the cellulase and the pectic enzyme, thereby improving the integral enzymolysis efficiency, further promoting the rupture of the cell wall of bupleurum root and stem, fully releasing the saponin, improving the transfer rate of the saponin, carrying out repeated alcohol extraction, extracting the saponin in bupleurum, purifying the saponin by macroporous resin in the later stage, and improving the purity of the bupleurum saponin. In the alcohol extraction process, the existence of the surfactants rhamnolipid and alkyl glycoside carboxyl betaine can reduce the interfacial tension of solid and liquid phases, greatly enhance the wetting and dissolution exudation capacity of the surfactant on bupleurum substrates, and greatly improve the transfer rate of saponin in bupleurum as a whole.
In the method for extracting the saikosaponin from the bupleurum, in water containing rhamnolipid and alkyl glycoside carboxyl betaine, the mass fractions of the rhamnolipid and the alkyl glycoside carboxyl betaine are respectively 0.005% and 0.03%.
In the method for extracting the saikosaponin from the bupleurum, cellulase is added according to 15FPU/g bupleurum powder, xylanase is added according to 200U/g bupleurum powder, and pectase is added according to 300U/g bupleurum powder.
In the method for extracting saikosaponin from bupleurum, the freezing temperature in the first step is-30 ℃ and the freezing time is 20 hours.
In the method for extracting saikosaponin from bupleurum, naOH is adopted to adjust the pH value to 9 in the second step.
In the method for extracting saikosaponin from bupleurum, the macroporous adsorption resin column is D101.
In the method for extracting saikosaponin from bupleurum, the alkaline water is 2% NaOH aqueous solution by mass percent.
In the method for extracting saikosaponin from bupleurum, the bupleurum is crushed into 40-mesh bupleurum powder in the first step.
In the method for extracting saikosaponin from bupleurum, the enzymolysis temperature is 30 ℃ and the enzymolysis time is 2 hours.
Example 2
A method for extracting saikosaponin from bupleurum, comprising:
step one, crushing bupleurum into bupleurum powder, soaking the bupleurum powder in water containing rhamnolipid and alkyl glycoside carboxyl betaine, freezing the bupleurum powder, crushing the bupleurum powder into ice residues by using a crusher, adding cellulase, xylanase and pectase for enzymolysis after the ice residues are melted at room temperature, inactivating the enzyme, and freeze-drying the obtained product; alkyl glycoside carboxybetaine an alkyl glycoside carboxybetaine zwitterionic surfactant with an authorized publication number of CN111013484B and the alkyl glycoside carboxybetaine prepared in example 2 of the preparation method thereof are adopted.
Step two, adding ethanol solution with the volume fraction of 70% which is 7 times of the weight of the bupleurum processed in the step one, adjusting the pH value to be alkaline, carrying out reflux extraction for 3 times at 75 ℃ for 1.5h each time, and combining the extracting solutions of the times;
step three, the combined extracting solution is put on a macroporous adsorption resin column, sequentially washed by alkaline water and pure water, discarded by washing liquid, eluted by ethanol solution with the volume fraction of 65%, and the eluent is collected;
and step four, concentrating the eluent under reduced pressure, and freeze-drying to obtain the saikosaponin powder.
According to the invention, bupleurum powder is firstly mixed with water containing rhamnolipid and alkyl glycoside carboxyl betaine, then frozen and crushed, and then cellulase, xylanase and pectase are added for enzymolysis, when frozen, ice crystal grows, and when crushed, the ice crystal pierces cell walls, so that the structure of cellulose is damaged, and the enzymolysis of cellulose in later stage is facilitated. In the enzymolysis process, the compound surfactant composed of rhamnolipid and alkyl glycoside carboxyl betaine can avoid ineffective combination of cellulose, hemicellulose, pectic substance, cellulase, xylanase and pectic enzyme, increase the contact area of the cellulase, xylanase, pectic enzyme, cellulose, hemicellulose and pectic substance, improve the activity of the xylanase, and further improve the activity of the xylanase by the cooperation of rhamnolipid and alkyl glycoside carboxyl betaine, and simultaneously improve the activity of the cellulase and the pectic enzyme, thereby improving the integral enzymolysis efficiency, further promoting the rupture of the cell wall of bupleurum root and stem, fully releasing the saponin, improving the transfer rate of the saponin, carrying out repeated alcohol extraction, extracting the saponin in bupleurum, purifying the saponin by macroporous resin in the later stage, and improving the purity of the bupleurum saponin. In the alcohol extraction process, the existence of the surfactants rhamnolipid and alkyl glycoside carboxyl betaine can reduce the interfacial tension of solid and liquid phases, greatly enhance the wetting and dissolution exudation capacity of the surfactant on bupleurum substrates, and greatly improve the transfer rate of saponin in bupleurum as a whole.
In the method for extracting the saikosaponin from the bupleurum, in water containing rhamnolipid and alkyl glycoside carboxyl betaine, the mass fractions of the rhamnolipid and the alkyl glycoside carboxyl betaine are respectively 0.01 percent and 0.04 percent.
In the method for extracting the saikosaponin from the bupleurum, the cellulase is added according to 17FPU/g bupleurum powder, the xylanase is added according to 220U/g bupleurum powder, and the pectinase is added according to 450U/g bupleurum powder.
In the method for extracting saikosaponin from bupleurum, the freezing temperature in the first step is-25 ℃ and the freezing time is 25 hours.
In the method for extracting saikosaponin from bupleurum, in the second step, naOH is adopted to adjust the pH value to 9-11.
In the method for extracting saikosaponin from bupleurum, the macroporous adsorption resin column is D101.
In the method for extracting saikosaponin from bupleurum, the alkaline water is 2% NaOH aqueous solution by mass percent.
In the method for extracting saikosaponin from bupleurum, the bupleurum is crushed into 50-mesh bupleurum powder in the first step.
In the method for extracting the saikosaponin from the bupleurum, the enzymolysis temperature is 40 ℃ and the enzymolysis time is 2.5h.
Example 3
A method for extracting saikosaponin from bupleurum, comprising:
step one, crushing bupleurum into bupleurum powder, soaking in water containing rhamnolipid and alkyl glycoside carboxyl betaine, which is 5 times the weight of the bupleurum powder, freezing, crushing into ice residues by using a crusher, adding cellulase, xylanase and pectase for enzymolysis after the ice residues are melted at room temperature, and freeze-drying after enzyme deactivation; alkyl glycoside carboxybetaine an alkyl glycoside carboxybetaine zwitterionic surfactant with an authorized publication number of CN111013484B and the alkyl glycoside carboxybetaine prepared in example 2 of the preparation method thereof are adopted.
Step two, adding ethanol solution with the volume fraction of 80% and the weight of which is 8 times that of the bupleurum processed in the step one, adjusting the pH value to be alkaline, carrying out reflux extraction for 3 times at 80 ℃ for 2 hours each time, and combining the extracting solutions of the times;
step three, the combined extracting solution is put on a macroporous adsorption resin column, sequentially washed by alkaline water and pure water, discarded by washing liquid, eluted by ethanol solution with the volume fraction of 70%, and the eluent is collected;
and step four, concentrating the eluent under reduced pressure, and freeze-drying to obtain the saikosaponin powder.
According to the invention, bupleurum powder is firstly mixed with water containing rhamnolipid and alkyl glycoside carboxyl betaine, then frozen and crushed, and then cellulase, xylanase and pectase are added for enzymolysis, when frozen, ice crystal grows, and when crushed, the ice crystal pierces cell walls, so that the structure of cellulose is damaged, and the enzymolysis of cellulose in later stage is facilitated. In the enzymolysis process, the compound surfactant composed of rhamnolipid and alkyl glycoside carboxyl betaine can avoid ineffective combination of cellulose, hemicellulose, pectic substance, cellulase, xylanase and pectic enzyme, increase the contact area of the cellulase, xylanase, pectic enzyme, cellulose, hemicellulose and pectic substance, improve the activity of the xylanase, and further improve the activity of the xylanase by the cooperation of rhamnolipid and alkyl glycoside carboxyl betaine, and simultaneously improve the activity of the cellulase and the pectic enzyme, thereby improving the integral enzymolysis efficiency, further promoting the rupture of the cell wall of bupleurum root and stem, fully releasing the saponin, improving the transfer rate of the saponin, carrying out repeated alcohol extraction, extracting the saponin in bupleurum, purifying the saponin by macroporous resin in the later stage, and improving the purity of the bupleurum saponin. In the alcohol extraction process, the existence of the surfactants rhamnolipid and alkyl glycoside carboxyl betaine can reduce the interfacial tension of solid and liquid phases, greatly enhance the wetting and dissolution exudation capacity of the surfactant on bupleurum substrates, and greatly improve the transfer rate of saponin in bupleurum as a whole.
In the method for extracting the saikosaponin from the bupleurum, in water containing rhamnolipid and alkyl glycoside carboxyl betaine, the mass fractions of the rhamnolipid and the alkyl glycoside carboxyl betaine are respectively 0.015% and 0.05%.
In the method for extracting the saikosaponin from the bupleurum, cellulase is added according to 20FPU/g bupleurum powder, xylanase is added according to 250U/g bupleurum powder, and pectase is added according to 600U/g bupleurum powder.
In the method for extracting saikosaponin from bupleurum, the freezing temperature in the first step is-20 ℃ and the freezing time is 30 hours.
In the method for extracting saikosaponin from bupleurum, naOH is adopted to adjust the pH value to 14 in the second step.
In the method for extracting saikosaponin from bupleurum, the macroporous adsorption resin column is D101.
In the method for extracting saikosaponin from bupleurum, the alkaline water is 2% NaOH aqueous solution by mass percent.
In the method for extracting saikosaponin from bupleurum, the bupleurum is crushed into 60-mesh bupleurum powder in the first step.
In the method for extracting saikosaponin from bupleurum, the enzymolysis temperature is 50 ℃ and the enzymolysis time is 3 hours.
Comparative example 1
The difference from example 2 is that in the first step, bupleurum is crushed into bupleurum powder, soaked in water with the weight of 4 times of bupleurum powder, frozen, crushed into ice residue by a crusher, and after the ice residue is melted at room temperature, cellulase, xylanase and pectase are added for enzymolysis, and freeze drying is carried out after enzyme deactivation; otherwise, the same as in example 2 was used.
Comparative example 2
The difference from example 2 is that in step one, bupleurum is crushed into bupleurum powder, soaked in water containing rhamnolipid, the weight of which is 4 times of that of bupleurum powder, frozen, crushed into ice residue by a crusher, and after the ice residue is melted at room temperature, cellulase, xylanase and pectase are added for enzymolysis, and after enzyme deactivation, freeze drying is carried out; in the water containing rhamnolipid, the mass fractions of the rhamnolipid are respectively 0.04%. Otherwise, the same as in example 2 was used.
Comparative example 3
The difference from example 2 is that in the first step, bupleurum is crushed into bupleurum powder, soaked in water containing alkyl glycoside carboxyl betaine, frozen, crushed into ice residue by a crusher, and after the ice residue is melted at room temperature, cellulase, xylanase and pectase are added for enzymolysis, enzyme deactivation and freeze drying are carried out; in the water containing the alkyl glycoside carboxybetaines, the mass fractions of the alkyl glycoside carboxybetaines are respectively 0.04 percent. Otherwise, the same as in example 2 was used.
Comparative example 4
The difference from example 2 is that the mass fraction of the alkyl glycoside carboxybetaine is 0.035%. Otherwise, the same as in example 2 was used.
Comparative example 5
The difference from example 2 is that the mass fraction of the alkyl glycoside carboxybetaine is 0.045%. Otherwise, the same as in example 2 was used.
Test 1
The transfer rate of saikosaponin, the content of saikosaponin in saikosaponin powder of example 2 and comparative examples 1 to 5 were measured, and the results are shown in Table 1.
TABLE 1 transfer rate of saikosaponin and content of saikosaponin in saikosaponin powder
| Transfer rate of saikosaponin/% | Saikosaponin content/% | |
| Example 2 | 89 | 92 |
| Comparative example 1 | 56 | 75 |
| Comparative example 2 | 67 | 80 |
| Comparative example 3 | 75 | 82 |
| Comparative example 4 | 83 | 86 |
| Comparative example 5 | 89 | 92 |
As is clear from Table 1, the transfer rate of saikoside is higher than that of comparative example 1 in example 2, because the rhamnolipid and alkyl glycoside carboxybetaine form a compound surfactant in example 2, ineffective combination of cellulose, hemicellulose, pectic substance, cellulase, xylanase and pectic enzyme can be avoided, the contact area of the cellulose, xylanase, pectic enzyme, cellulose, hemicellulose and pectic substance can be increased, the rhamnolipid can improve the activity of xylanase, the activity of the rhamnoside can be further improved by cooperation with the alkyl glycoside carboxybetaine, the activity of the cellulase and the pectic enzyme can be simultaneously improved, the overall enzymolysis efficiency is further improved, the cell wall rupture of saikoside can be further promoted, the transfer rate of the saponin can be fully released, the existence of the rhamnolipid and the alkyl glycoside carboxybetaine serving as surfactants can be reduced, the interfacial tension of solid-liquid phase can be greatly enhanced, the wetting and dissolution capacities of the substrate of the bupleurum can be greatly improved, and the transfer rate of the saikoside can be greatly improved as a whole.
The transfer rate of saikosaponin in example 2 is higher than that of comparative example 2 because only a surfactant of rhamnolipid, which can improve the enzymolysis efficiency to some extent but has limited improvement degree, and much less combination of rhamnolipid and alkyl glycoside carboxybetaine is adopted in comparative example 2, so that the transfer rate of saikosaponin is higher than that of comparative example 1, and example 2 is lower.
The transfer rate of saikoside is higher in example 2 than in comparative example 3 because only the surfactant of alkyl glycoside carboxybetaine, is used in comparative example 3, and although the enzymatic hydrolysis efficiency is improved to some extent, the degree of improvement is not more than when rhamnolipid and alkyl glycoside carboxybetaine are combined, and thus the transfer rate of saikoside is lower than in example 2. As is clear from comparison of comparative example 3 and comparative example 2, the transfer rate of saikoside is higher by using only the surfactant of alkyl glycoside carboxybetaine than by using only the surfactant of rhamnolipid.
The higher transfer rate of saikosaponin in example 2 than comparative example 4 is because the amount of alkyl glycoside carboxybetaine added in comparative example 4 is lower than that in example 2, indicating that the decrease in the amount of alkyl glycoside carboxybetaine decreases the transfer rate of saikosaponin.
The transfer rate of saikosaponin in example 2 was the same as that of comparative example 5, indicating that the increase in the amount of alkyl glycoside carboxybetaine did not increase the transfer rate of saikosaponin, and thus the optimum amount of alkyl glycoside carboxybetaine was 0.04%.
Although embodiments of the present invention have been disclosed above, it is not limited to the details and embodiments shown, it is well suited to various fields of use for which the invention is suited, and further modifications may be readily made by one skilled in the art, and the invention is therefore not to be limited to the particular details and examples shown and described herein, without departing from the general concepts defined by the claims and the equivalents thereof.
Claims (10)
1. The method for extracting saikosaponin from bupleurum root is characterized by comprising the following steps:
step one, crushing bupleurum into bupleurum powder, soaking the bupleurum powder in water containing rhamnolipid and alkyl glycoside carboxyl betaine, freezing the bupleurum powder, crushing the bupleurum powder into ice residues by using a crusher, adding cellulase, xylanase and pectase for enzymolysis after the ice residues are melted at room temperature, inactivating the enzyme, and freeze-drying;
step two, adding ethanol solution with the volume fraction of 60-80% which is 6-8 times of the weight of the bupleurum processed in the step one, adjusting the pH value to be alkaline, carrying out reflux extraction for 2-3 times at 70-80 ℃ for 1-2 h each time, and combining the extracting solutions of each time;
step three, the combined extracting solution is put on a macroporous adsorption resin column, then is washed by alkaline water and pure water in sequence, the washing liquid is discarded, then is eluted by ethanol solution with the volume fraction of 60-70%, and the eluent is collected;
and step four, concentrating the eluent under reduced pressure, and freeze-drying to obtain the saikosaponin powder.
2. The method for extracting saikosaponin from bupleurum root according to claim 1, wherein in the water containing rhamnolipid and alkyl glycoside carboxybetaine, the mass fractions of rhamnolipid and alkyl glycoside carboxybetaine are 0.005-0.015% and 0.03-0.05%, respectively.
3. The method for extracting saikosaponin from bupleurum root of claim 2, wherein cellulase is added according to 15-20 FPU/g bupleurum powder, xylanase is added according to 200-250U/g bupleurum powder, and pectase is added according to 300-600U/g bupleurum powder.
4. The method for extracting saikosaponin from bupleurum root according to claim 1, wherein the freezing temperature in the first step is-30 to-20 ℃ and the freezing time is 20-30 h.
5. Such asThe method for extracting saikosaponin from bupleurum root as defined in claim 1, wherein in the second step, naOH and Na are adopted 2 CO 3 Or KOH to adjust the pH to 9-14.
6. The method of extracting saikosaponin from bupleurum of claim 1, wherein the macroporous adsorbent resin column is D101.
7. The method for extracting saikosaponin from bupleurum according to claim 1, wherein the alkaline water is 2% NaOH aqueous solution by mass fraction.
8. The method of extracting saikosaponin from bupleurum according to claim 1, wherein in the first step, bupleurum is crushed into bupleurum powder of 40-60 meshes.
9. The method for extracting saikosaponin from bupleurum root of claim 1, wherein the enzymolysis temperature in the first step is 30-50 ℃ and the enzymolysis time is 2-3 h.
10. The method for extracting saikosaponin from bupleurum root according to claim 2, wherein the mass fractions of rhamnolipid and alkyl glycoside carboxybetaine in water containing rhamnolipid and alkyl glycoside carboxybetaine are 0.01% and 0.04%, respectively.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310191883.8A CN116019848A (en) | 2023-03-02 | 2023-03-02 | Method for extracting saikosaponin from bupleurum |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310191883.8A CN116019848A (en) | 2023-03-02 | 2023-03-02 | Method for extracting saikosaponin from bupleurum |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116019848A true CN116019848A (en) | 2023-04-28 |
Family
ID=86076141
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310191883.8A Pending CN116019848A (en) | 2023-03-02 | 2023-03-02 | Method for extracting saikosaponin from bupleurum |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116019848A (en) |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110141591A (en) * | 2019-05-23 | 2019-08-20 | 佛山市欧若拉生物科技有限公司 | A kind of saikoside preparation method |
| CN111013484A (en) * | 2019-11-25 | 2020-04-17 | 南京科技职业学院 | Alkyl glycoside carboxyl betaine type zwitterionic surfactant and preparation method thereof |
-
2023
- 2023-03-02 CN CN202310191883.8A patent/CN116019848A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110141591A (en) * | 2019-05-23 | 2019-08-20 | 佛山市欧若拉生物科技有限公司 | A kind of saikoside preparation method |
| CN111013484A (en) * | 2019-11-25 | 2020-04-17 | 南京科技职业学院 | Alkyl glycoside carboxyl betaine type zwitterionic surfactant and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| 李慧婷等: "浸提辅助剂在中药提取中的应用及研究进展", 中国实验方剂学杂志, vol. 23, no. 23, pages 212 - 219 * |
| 赵余庆主编: "《中药及天然产物提取制备关键技术》", vol. 1, 中国医药科技出版社, pages: 132 - 133 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102924423B (en) | Method for extracting cyanidin from banana peel | |
| CN101607997B (en) | Method for extracting polysaccharide compound of traditional Chinese medicine cassia seed | |
| CN110295209B (en) | Technological method for extracting alginate oligosaccharides by enzymatic hydrolysis of gulfweed | |
| CN102876732B (en) | Method for preparing high-added-value sugar alcohols by efficiently using wood fiber raw materials | |
| CN102382205A (en) | Method for extracting pectin from banana peels | |
| CN101955548B (en) | Method for enzyme-assisted extraction of fructus tribuli polysaccharide | |
| CN102040417A (en) | Plant anti-freezing fluid | |
| CN106349405A (en) | Method for extracting pectin from shaddock peel through enzymolysis and ultrasonic waves | |
| CN108048495B (en) | Biological extraction method of resveratrol | |
| CN111909791A (en) | Preparation method of rose essential oil and rose essential oil | |
| CN105385741A (en) | Preparation method for grateloupia filicina polypeptide solution | |
| CN111802408A (en) | Plant defense protective agent for preventing fruit tree freezing injury and preparation method thereof | |
| CN103462140B (en) | Low frequency ultrasonic-assisted desugaring and deacidification method for concentrated waxberry juice | |
| CN116019848A (en) | Method for extracting saikosaponin from bupleurum | |
| CN108300747B (en) | Method for improving saccharified efficiency of silvergrass enzymolysis by utilizing sodium chlorite pretreatment | |
| CN111304266A (en) | Biological enzymolysis fermentation process for aloe | |
| CN108998491A (en) | A method of extracting protein in the peony seeds dregs of rice | |
| CN111961525A (en) | Method for extracting acer truncatum fruit edible oil | |
| CN105132119A (en) | Method for preparing boxthorn seed oil through enzymolysis | |
| CN111057117B (en) | Comprehensive utilization method of immature bitter orange | |
| CN102181076B (en) | Method for cleanly preparing starch containing abundant flavones and comprehensively utilizing radix puerariae resource | |
| CN104397842B (en) | A kind of pepper freezes peeling method | |
| CN106811382A (en) | A kind of preparation method of olive wine | |
| CN108047288B (en) | Preparation method of geniposide | |
| CN110066841B (en) | Method for improving extraction yield of active ingredient albiflorin of red peony root |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |