CN107778338A - A kind of rebaudioside C isolation and purification methods - Google Patents
A kind of rebaudioside C isolation and purification methods Download PDFInfo
- Publication number
- CN107778338A CN107778338A CN201610190936.4A CN201610190936A CN107778338A CN 107778338 A CN107778338 A CN 107778338A CN 201610190936 A CN201610190936 A CN 201610190936A CN 107778338 A CN107778338 A CN 107778338A
- Authority
- CN
- China
- Prior art keywords
- rebaudioside
- isolation
- purification methods
- solution
- methods according
- 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
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
- C07H15/20—Carbocyclic rings
- C07H15/24—Condensed ring systems having three or more rings
- C07H15/256—Polyterpene radicals
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
- C07H1/06—Separation; Purification
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Saccharide Compounds (AREA)
Abstract
The invention discloses a kind of rebaudioside C isolation and purification methods, by using suitable process-scale chromatography post and solution, and optimize technique step, with high performance liquid chromatography detection, rebaudioside C is obtained from Stevioside raw material with reference to the method for column chromatography for separation and recrystallization, cost is low more economical, and purity is high, yield is high, and purity is more than 95%, and yield is more than 60%.
Description
Technical field
The present invention relates to a kind of rebaudioside C isolation and purification methods.
Background technology
Stevioside has been used more than 20 years in Asia as sweetener.2008, North America by ratify its as
Food additives.Therefore, stevioside and its effective purification process are that have very high business demand.It is main in STEVIA REBAUDIANA
The stevioside wanted includes Stevioside, rebaudioside C, content rebaudioside-A and rebaudioside D.Rebaudioside D and rebaudioside C
Sub-fraction therein is only account for, but they have purer sweet taste and unique application, thus there is higher business
Industry is worth.The isolation and purification method on content rebaudioside-A and stevioside has much in the prior art, to rebaudioside C's
Concern is seldom.
There is research to point out, although rebaudioside C bitter tastes are heavier, there is antitumor, lowering blood pressure and blood fat, fat-reducing etc.
Many effects, its potential using value have a great attraction.Therefore there is an urgent need to one kind can high-purity, high yield separation and it is pure
Change rebaudioside C method.
The content of the invention
The present invention is to solve the defects of prior art, there is provided a kind of rebaudioside C isolation and purification methods.For in solution
Technical problem is stated, the technical solution adopted by the present invention is：
A kind of rebaudioside C isolation and purification methods, comprise the following steps：
(1) raw material stevioside is dissolved in 3 times of amount deionized waters, then concentrated aqueous liquid to the 50% of about original volume,
Sample is made, above-mentioned sample is pumped into the process-scale chromatography column top for being filled with resinous polymer by (2), is mixed with water/acetone or acetonitrile
Solution gradient elutes, and the concentration gradient of the water and acetone/acetonitrile mixed solution is molten for 5%-35% water/acetone or acetonitrile
Liquid, the concentration gradient are percent by volume；
(3) the elution solution that Fractional Collections elutes from resinous polymer chromatographic column, then uses high-efficient liquid phase color respectively
Spectrum analysis detection, after rebaudioside C is all eluted in sample, stops elution, and chromatographic column continues to use pure water rinsing, balances
Pillar is in case reuse；
(4) the elution solution containing higher degree rebaudioside C in step (3) after analysis detects is passed through into NF membrane
Proper volume (solid contents is between 15-20%) is concentrated into, concentrate continues further to be concentrated with single-effect vacuum inspissator
To medicinal extract shape (ratio weighs about 1.13-1.15), medicinal extract is spray-dried to obtain rebaudioside C raw material dry powder.
5) above-mentioned dry powder is dissolved in aqueous acetone and is heated to all solids and is completely dissolved, then added equivalent to solution
2-3 times of acetone of volume, obtained mixed solution is stirred into more than 12h in 20-30 DEG C, finally obtains white crystal, mistake
Filter, dry rebaudioside C semi-finished product.
(6) above-mentioned semi-finished product are added in appropriate aqueous methanol, then heat to 55-60 DEG C it is completely molten to all solids
Solution temperature, is then down to and is stored at room temperature 5 hours, obtained white crystal is filtered out by solution.
(7) white crystal obtained in step (6) is placed at 70-80 DEG C and be dried in vacuo, finally use high performance liquid chromatography
Analyze dried crystalline content；
The resinous polymer is polystyrene/divinylbenzene polymer.Such as the CG-161m series plastics of ROHM AND HAAS
Or similar resin.
Preferably, the above-mentioned alleged equipment that eluent concentration is used is the nanofiltration film condensing device that molecular weight is 150..
Preferably, the weight of resinous polymer is 4-20 times of Stevioside weight in step (1) in step (2).
The beneficial effect that the present invention is reached：
The production method of the present invention can obtain rebaudioside C from stevioside raw material, and cost is low more economical, and purity
Height, yield is high, and purity is more than 95%, and yield is more than 60%.
Purity detecting, and production are carried out to the product of separation process all the time using HPLC analytical method in the present invention
Thing rebaudioside C and stevioside determination, high performance liquid chromatography detection equipment are by Shimadzu LC-10A systems and Shimadzu SPD-
10A multiwavelength detectors form, and pillar is 250mm*4.6mm i.d.5 μm C18 posts, what 32% acetonitrile and 68% water formed
Isocratic mobile phase, flow velocity 1.0ml/min, Detection wavelength 210nm, stevioside and rebaudioside C are quantified with reference substance.
Brief description of the drawings
Fig. 1 is the rebaudioside C high-efficient liquid phase chromatograms that the present invention isolates and purifies；
Fig. 2 production Technology route maps.
Embodiment
The invention will be further described below in conjunction with the accompanying drawings.Following examples are only used for clearly illustrating the present invention
Technical scheme, and can not be limited the scope of the invention with this.
Embodiment one
(1) C containing rebaudioside 15.2% raw material stevioside 50kg is added in 500L reactors, then added
150L deionized waters and stir be warming up to 70 DEG C, after solids is completely dissolved, this solution is released and is pumped into from reactor
The 50% of about original volume is concentrated into haplo-effect concentrator, sample is made.
(2) above-mentioned sample is pumped directly into and be filled with the top of the industrially prepared chromatographic column of resinous polymer, resinous polymer
Weight is 10 times of Stevioside weight in step (1).With water and the mixed solution gradient elution of acetone, the water mixes with acetone
The concentration gradient of solution is 10%-35% (volume ratio)
(3) the elution solution that Fractional Collections elutes from resinous polymer chromatographic column, then uses high-efficient liquid phase color respectively
Spectrum analysis detection, after rebaudioside C is all eluted in sample, stops elution.
(4) the elution solution containing higher degree rebaudioside C in step (3) after analysis detects is passed through into NF membrane
Device concentrate, it is to be concentrated to about original volume 20% when stop, concentrate is transferred to single-effect vacuum inspissator and is further concentrated into leaching
Paste (proportion 1.13-1.15), this medicinal extract must further contain rebaudioside C 35-45% Lai Baodi with spray dryer spray drying
Glucoside C raw materials for production.5) above-mentioned dry powder is dissolved in aqueous acetone and is heated to all solids and is completely dissolved, then add equivalent to
2-3 times of acetone of liquor capacity, obtained mixed solution is stirred into more than 12h in 20-30 DEG C, finally obtain white crystalline substance
Body, filtering, dry rebaudioside C semi-finished product.
(6) above-mentioned semi-finished product are added in appropriate aqueous methanol, then heat to 55-60 DEG C it is completely molten to all solids
Solution temperature, is then down to and is stored at room temperature 5 hours, obtained white crystal is filtered out by solution.
(7) white crystal obtained in step (6) is placed at 80 DEG C and be dried in vacuo, finally use efficient liquid phase chromatographic analysis
Dried crystalline content；The crystal is rebaudioside C, purity 95.3%, weight 4.65kg, yield 61%.
Resinous polymer alleged by above is ROHM AND HAAS Amberlite 1600
Embodiment two
Target material：60kg steviosides, wherein purchase gained, Stevioside 24%, content rebaudioside-A 27%, rebaudioside
C15.5%, surplus are impurity.
Chromatographic column：High pressure industrial prepares layer post, column dimensions 800*3000mm, filling 1000L Rohm
HaasAmberlite1600。
Experimental procedure：
(1) C containing rebaudioside 15.5% raw material stevioside 60kg is added in 500L reactors, then added
180L deionized waters and stir be warming up to 75 DEG C, after solids is completely dissolved, this solution is released and is pumped into from reactor
The 50% of about original volume is concentrated into haplo-effect concentrator, sample is made.
(2) above-mentioned sample is pumped directly into and be filled with the top of the industrially prepared chromatographic column of resinous polymer, resinous polymer
Weight is 8 times of Stevioside weight in step (1).With water and the mixed solution gradient elution of acetone, the water mixes with acetone
The concentration gradient of solution is 10%-35% (volume ratio)
(3) the elution solution that Fractional Collections elutes from resinous polymer chromatographic column, then uses high-efficient liquid phase color respectively
Spectrum analysis detection, after rebaudioside C is all eluted in sample, stops elution.
(4) the elution solution containing higher degree rebaudioside C in step (3) after analysis detects is passed through into NF membrane
Device concentrate, it is to be concentrated to about original volume 20% when stop, concentrate is transferred to single-effect vacuum inspissator and is further concentrated into leaching
Paste (proportion 1.13-1.15), this medicinal extract must further contain rebaudioside C 35-45% Lai Baodi with spray dryer spray drying
Glucoside C raw materials for production.
(5) above-mentioned dry powder is dissolved in aqueous acetone and is heated to all solids and is completely dissolved, then added equivalent to molten
2-3 times of acetone of liquid product, stirs more than 12h by obtained mixed solution in 20-30 DEG C, finally obtains white crystal,
Filtering, dry rebaudioside C semi-finished product.
(6) above-mentioned semi-finished product are added in appropriate aqueous methanol, then heat to 55-60 DEG C it is completely molten to all solids
Solution temperature, is then down to and is stored at room temperature 5 hours, obtained white crystal is filtered out by solution.
(7) white crystal obtained in step (6) is placed at 80 DEG C and be dried in vacuo, finally use efficient liquid phase chromatographic analysis
Dried crystalline content；The crystal is rebaudioside C, purity 95.5%, weight 5.76kg, yield 62%.
The high performance liquid chromatography spectrogram of stevioside and rebaudioside C after isolating and purifying above is shown in Fig. 1.
Described above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, without departing from the technical principles of the invention, some improvement and deformation can also be made, these are improved and deformation
Also it should be regarded as protection scope of the present invention.
Claims (8)
1. a kind of rebaudioside C isolation and purification methods, it is characterised in that comprise the following steps：
(1) raw material Stevioside is added in the deionized water of triplication, is then heated to 70 DEG C and is completely dissolved to all solids, so
Obtained solution is concentrated into small size afterwards sample is made, above-mentioned sample is pumped into the process-scale chromatography for being filled with resinous polymer by (2)
Column top, with water and acetone/acetonitrile mixed solution gradient elution, the water and the concentration gradient of acetone/acetonitrile mixed solution are
5%-35% water and acetone/acetonitrile solution, the concentration gradient is volume % ratios；
(3) the elution solution that Fractional Collections elutes from resinous polymer chromatographic column, then respectively with high performance liquid chromatography point
Analysis detection, after rebaudioside C is all eluted in sample, stop elution；
(4) it is the elution solution containing high content rebaudioside C fractions in step (3) after analysis detects is dense by NF membrane
Proper volume (solid contents is between 15-20%) is reduced to, concentrate is further concentrated into medicinal extract with single-effect vacuum inspissator
Shape (ratio weighs about 1.13-1.15), medicinal extract are spray-dried to obtain rebaudioside C raw material dry powder；
(5) above-mentioned dry powder is dissolved in aqueous acetone and is heated to all solids and is completely dissolved, then added equivalent to solution body
2-3 times long-pending of acetone, obtained mixed solution is stirred into more than 12h in 20-30 DEG C, finally obtains white crystal, filtered,
Dry rebaudioside C semi-finished product；
(6) above-mentioned semi-finished product are added in appropriate aqueous methanol, then heat to 55-60 DEG C and be completely dissolved to all solids, with
Solution temperature is down to afterwards and is stored at room temperature 5 hours, obtained white crystal is filtered out；
(7) white crystal obtained in step (6) is placed at 70-80 DEG C and be dried in vacuo, finally use efficient liquid phase chromatographic analysis
Dried crystalline content.
2. a kind of rebaudioside C isolation and purification methods according to claim 1, it is characterised in that use nano filter membrance device
Eluent is reclaimed into reuse repeatedly.
3. a kind of rebaudioside C isolation and purification methods according to claim 1, it is characterised in that used in concentrated liquid
Equipment is single-effect vacuum ager.
A kind of 4. rebaudioside C isolation and purification methods according to claim 1, it is characterised in that the resinous polymer
For polystyrene/divinylbenzene polymer.
5. a kind of rebaudioside C isolation and purification methods according to claim 1, it is characterised in that during the chromatographic column is
High pressure industrial preparative separation chromatographic column.
A kind of 6. rebaudioside C isolation and purification methods according to claim 5, it is characterised in that the mesohigh industry
The operating pressure of preparative separation chromatographic column is 1.5-3.0Mpa.
A kind of 7. rebaudioside C isolation and purification methods according to claim 1, it is characterised in that the column Mobile
The elution flow rate of phase is 2-5 bed volume (2-5BV)/h per hour.
8. a kind of rebaudioside C isolation and purification methods according to claim 1, it is characterised in that resin gathers in step (2)
The weight of compound is 4-20 times of Stevioside weight in step (1).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610190936.4A CN107778338A (en) | 2016-08-29 | 2016-08-29 | A kind of rebaudioside C isolation and purification methods |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610190936.4A CN107778338A (en) | 2016-08-29 | 2016-08-29 | A kind of rebaudioside C isolation and purification methods |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107778338A true CN107778338A (en) | 2018-03-09 |
Family
ID=61387707
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610190936.4A Pending CN107778338A (en) | 2016-08-29 | 2016-08-29 | A kind of rebaudioside C isolation and purification methods |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107778338A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108530503A (en) * | 2018-05-17 | 2018-09-14 | 苏州赛分科技有限公司 | A kind of isolation and purification method of dulcoside B |
| CN111440220A (en) * | 2020-05-09 | 2020-07-24 | 浙江天草生物科技股份有限公司 | Method for preparing high-purity stevioside RC from mother liquor sugar |
| CN112480127A (en) * | 2020-12-11 | 2021-03-12 | 无锡福祈制药有限公司 | Novel method for producing mitomycin |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101628924A (en) * | 2009-08-21 | 2010-01-20 | 天津美伦医药集团有限公司 | Process for extracting rebaudioside C in stevioside |
| CN102030788A (en) * | 2011-01-05 | 2011-04-27 | 沈阳天峰生物制药有限公司 | Method for preparing rebaudioside C |
| CN102127130A (en) * | 2011-01-14 | 2011-07-20 | 青岛润浩甜菊糖高科有限公司 | Purification method of stevioside RC (rebaudioside C) |
| CN103122015A (en) * | 2011-11-18 | 2013-05-29 | 国际香料和香精公司 | Method for purifying rebaudioside c |
| CN103965271A (en) * | 2013-01-25 | 2014-08-06 | 沈阳天峰生物制药有限公司 | Method for preparing rebaudioside C from stevia sugar |
-
2016
- 2016-08-29 CN CN201610190936.4A patent/CN107778338A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101628924A (en) * | 2009-08-21 | 2010-01-20 | 天津美伦医药集团有限公司 | Process for extracting rebaudioside C in stevioside |
| CN102030788A (en) * | 2011-01-05 | 2011-04-27 | 沈阳天峰生物制药有限公司 | Method for preparing rebaudioside C |
| CN102127130A (en) * | 2011-01-14 | 2011-07-20 | 青岛润浩甜菊糖高科有限公司 | Purification method of stevioside RC (rebaudioside C) |
| CN103122015A (en) * | 2011-11-18 | 2013-05-29 | 国际香料和香精公司 | Method for purifying rebaudioside c |
| CN103965271A (en) * | 2013-01-25 | 2014-08-06 | 沈阳天峰生物制药有限公司 | Method for preparing rebaudioside C from stevia sugar |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108530503A (en) * | 2018-05-17 | 2018-09-14 | 苏州赛分科技有限公司 | A kind of isolation and purification method of dulcoside B |
| CN108530503B (en) * | 2018-05-17 | 2021-05-07 | 苏州赛分科技有限公司 | Method for separating and purifying rebaudioside C |
| CN111440220A (en) * | 2020-05-09 | 2020-07-24 | 浙江天草生物科技股份有限公司 | Method for preparing high-purity stevioside RC from mother liquor sugar |
| CN111440220B (en) * | 2020-05-09 | 2023-05-05 | 浙江天草生物科技股份有限公司 | Method for preparing high-purity stevioside RC from mother liquor sugar |
| CN112480127A (en) * | 2020-12-11 | 2021-03-12 | 无锡福祈制药有限公司 | Novel method for producing mitomycin |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103408602B (en) | A kind of method being separated preparation four kinds of glycoside chemical reference substances from ZANGYINCHEN | |
| CN107778338A (en) | A kind of rebaudioside C isolation and purification methods | |
| CN105566414B (en) | The method that four kinds of flavone glycosides are isolated and purified from waxberry flesh | |
| WO2016082122A1 (en) | Preparation method for medicinal chlorogenic acid | |
| CN104892717B (en) | A kind of technical grade preparative liquid chromatography separation method of momordica glycoside V | |
| CN107098942A (en) | A kind of method of kaempferia galamga glycosides in Subcritical Water Extraction radish leaves | |
| CN107674116A (en) | A kind of purification process of Pneumocandin B0 | |
| CN107936033A (en) | The method that industrially prepared liquid chromatogram isolates and purifies koumine | |
| CN108003211A (en) | A kind of method of quick separating ganoderic acid in accessory substance from ganoderma lucidum | |
| CN100434148C (en) | Process for separating and purifying organic compound by macroporous absorption resin-reverse osmosis membrane | |
| CN104628801A (en) | Process for extracting and separating linarin from wild chrysanthemum | |
| CN105017367B (en) | A kind of method separating lanosterol and lanostenol | |
| CN103265521A (en) | Preparation method of demethylated beilidifolin | |
| CN104961716B (en) | Method for separating high-purity lactone type lovastatin from fermentum rubrum powder | |
| CN104844550B (en) | A kind of method that osthole and imperatorin are isolated and purified from Fructus Cnidii | |
| CN104844547B (en) | A kind of high efficiency extraction of barbaloin and grading purification method | |
| CN108373474B (en) | A bilobalide compound extracted from folium Ginkgo and its preparation method | |
| CN105820213B (en) | The method for efficiently separating purifying knob not Kangding | |
| CN107382943B (en) | Method for subcritical water extraction of dihydroquercetin in sorghum bran | |
| CN102276570B (en) | Method for purifying epigallo catechin gallate (EGCG) | |
| CN102503998B (en) | Method for rapidly separating quercetin from flos albiziae | |
| CN105037452B (en) | A kind of process for purification of quick preparation high-purity Fondaparinux sodium | |
| CN106008441B (en) | A kind of purification process of high-purity EGC | |
| CN101353294A (en) | Separation and purification method of high-content resveratrol | |
| CN103408616A (en) | Preparation method of Brucea javanica glucoside A |
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 | ||
| DD01 | Delivery of document by public notice | ||
| DD01 | Delivery of document by public notice |
Addressee: Patents of Stilwell Biotechnology (Suzhou) Co.,Ltd. The person in charge Document name: Deemed notice of withdrawal |
|
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20180309 |