CN108586308A - A kind of method that film combination refines both sexes alkaloid - Google Patents
A kind of method that film combination refines both sexes alkaloid Download PDFInfo
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- CN108586308A CN108586308A CN201810633249.4A CN201810633249A CN108586308A CN 108586308 A CN108586308 A CN 108586308A CN 201810633249 A CN201810633249 A CN 201810633249A CN 108586308 A CN108586308 A CN 108586308A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
- C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
- C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
- C07D207/10—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
- C07D207/16—Carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
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Abstract
A kind of method that film combination refines both sexes alkaloid, the invention provides a kind of novel both sexes Alkaloid separation methods, for the difference of the ingredients such as alkaloid, phenolic acid, saponin(e in both sexes alkaloid physico-chemical property and Chinese medicine, modifying ingredients existence, it introduces organic solvent and strengthens or weaken the active force between NF membrane, amplify both sexes alkaloid and other compositions nanofiltration active force otherness, to realize that purpose refines both sexes alkaloid.The invention fast implements the enrichment of both sexes alkaloid in complicated ingredient environment, and in separation process on heat-sensitive ingredients without influence, while the environmental pollution for avoiding the organic reagent generated by chromatographic isolation from bringing reduces energy consumption, saves pharmacy corporation production cost.
Description
Technical field
The present invention relates to a kind of both sexes alkaloid process for purification, and in particular to one kind being based on composition of alkaloids physical chemical characteristics
Both sexes alkaloid component is refined using membrane combination technology, belongs to field of medicaments.
Background technology
A kind of nitrogenous alkalinity that alkaloid is present in nature (predominantly plant, but some exists in animal) has
Machine compound has alkalinity more.Both sexes alkaloid has carboxyl or phenolic hydroxyl group compared to conventional alkaloid more in structure,
Also the physico-chemical property with acid compound while with alkalinity.
Traditional Chinese medicine ingredients complexity is various, there is flavones, saponin(e, alkaloid, phenolic acid, tannin, albumen, polysaccharide, inorganic ions etc.,
It is mostly used when being enriched with to composition of alkaloids and prepares chromatography, ion exchange resin, silica gel column chromatography etc., above-mentioned beneficiation technologies are deposited
In certain technological deficiency, the higher target component of relative purity can be obtained by preparing chromatography, but bioaccumulation efficiency and cost are inclined
Height, and follow-up enrichment positions solution recovery processing, can also cause ingredient transformation conversion；Ion exchange resin and chromatography color
Spectrum, separative efficiency is low, and the organic solvents such as chloroform, toluene for composition of alkaloids elution, not only to operation
Personnel bring injury, and pollution is also resulted in environment.How soda acid both sexes alkaloid to be carried out under room temperature or cryogenic conditions
Enrichment reduces organic solvent pollution, improves purification efficiency, is pharmaceuticals industry scientific and technical problem in the urgent need to address.
Membrane separation technique is to cut to realize separation by the molecule of membrane aperture, is had easy, efficient, excellent without fuel factor etc.
Gesture can targetedly remove macromolecular substances when to sample filtering, retain small molecule ingredient, but the choosing of membrane separation technique
Selecting property is poor.The present invention has carried out separation row of the composition of alkaloids under different solutions environment, dissociated state at the beginning of research
It for research, chances under low concentration organic solvent environment, the retention efficiency of solution amorph alkaloid, both sexes life can be strengthened
There are significant differences with other constituents of Chinese medicine with the correlation of solution for the dissociation behavior of alkaloids, to which purpose realizes soda acid two
Property alkaloid target the refined enrichment of property.
Based on above-mentioned background, provide it is a kind of to room temperature, low stain, low power consuming, the both sexes alkaloid in low period it is refined
Method retains its ultimate constituent structure, contributes to the development of the related work of component purification and new drug development.
Invention content
Goal of the invention：The object of the present invention is to provide a kind of films to combine the method for refining both sexes alkaloid, can be used for quickly
Detach the enrichment of both sexes alkaloid in complicated ingredient system.
Technical solution：
A kind of method that film combination refines both sexes alkaloid, it is characterised in that：It comprises the steps of：A. to be separated two are detected
Property alkaloid isoelectric point, and calculate the pH and alkali of acidic-group in its both sexes alkaloid compound structure in solution amorph when
PH when property group journey solution amorph；B. extract to be separated is dissolved in purified water, it is super using molecular cut off 10KDa~100KDa
Filter membrane carries out pretreatment removal high molecular weight protein, polysaccharide, collects ultrafiltrate；C. by step b ultrafiltrates using water-soluble organic molten
Agent is gradually diluted to containing water-soluble organic solvent amount in solution and reaches 5%~40%, and then using 1%~10% mass concentration
Sodium hydrate aqueous solution adjusts the pH that pH is dissociated to step a neutral and alkali groups, using molecular cut off 100Da~800Da nanofiltrations
UF membrane, when trapped fluid volume can not continue to detach, purified water dilutes trapped fluid, after repeating 1~3 time, waits for trapped fluid volume
Collection terminal trapped fluid when can not continue to detach；D. the terminal trapped fluid in step c is gradually dilute using water-miscible organic solvent
It releases into solution containing water-soluble organic solvent amount and reaches 5%~40%, using the aqueous hydrochloric acid solution tune of 1%~10% mass concentration
The pH that acidic-group dissociates in section pH to step a waits for trapped fluid using molecular cut off 100Da~800Da nanofiltration UF membranes
When volume can not continue to detach, purified water dilutes trapped fluid, after repeating 1~3 time, is received when trapped fluid volume can not be detached persistently
Collect trapped fluid to get both sexes alkaloid position.
The ultrafiltration membrane material that a kind of film combination refines in the method for both sexes alkaloid is cellulose, polyether sulfone, vinegar
One kind in acid cellulose.
The water-miscible organic solvent that a kind of film combination refines in the method for both sexes alkaloid is methanol, ethyl alcohol, second
One kind in nitrile, tetrahydrofuran.
The nanofiltration membrane material that a kind of film combination refines in the method for both sexes alkaloid is poly- for organic solvent-resistant material
One kind in acid imide, polyamide, polyacrylonitrile.
Advantageous effect：(1) a kind of novel both sexes Alkaloid separation method is provided, is fast implemented in complicated ingredient environment
The enrichment of both sexes alkaloid, and in separation process on heat-sensitive ingredients without influence.(2) the organic examination generated by chromatographic isolation is avoided
The environmental pollution that agent is brought reduces energy consumption, saves pharmacy corporation production cost.
Description of the drawings
Fig. 1 is stachydrine reference substance chromatogram.
Fig. 2 is the chromatogram of stachydrine prepared by UF membrane.
Fig. 3 is stachydrine chromatogram prepared by resin separation.
Specific implementation mode
Form by the following examples is described in further detail the above of the present invention again, but should not be by this
The range for being interpreted as the above-mentioned theme of the present invention is only limitted to example below, and all technologies realized based on the above of the present invention are equal
Belong to the scope of the present invention.
Embodiment 1：
A kind of method that film combination refines both sexes alkaloid, wherein being as follows：
Step a is：The isoelectric point of both sexes alkaloid to be separated is detected, and is calculated sour in its both sexes alkaloid compound structure
Property group in solution amorph when pH and pH when basic group journey solution amorph；
Step b is：Extract to be separated is dissolved in purified water, using molecular cut off 10KDa~100KDa ultrafiltration membranes into
Row pretreatment removal high molecular weight protein, polysaccharide, collect ultrafiltrate；
Step c is：It is organic molten that step b ultrafiltrates using water-miscible organic solvent are gradually diluted to containing water-soluble in solution
Dosage reaches 5%~40%, and then adjusts alkali in pH to step a using the sodium hydrate aqueous solution of 1%~10% mass concentration
Property group dissociation pH wait for that trapped fluid volume can not continue to detach using molecular cut off 100Da~800Da nanofiltration UF membranes
When, purified water dilutes trapped fluid, after repeating 1~3 time, collection terminal trapped fluid when trapped fluid volume can not continue to detach；
Step d is：Terminal trapped fluid in step c is gradually diluted in solution using water-miscible organic solvent containing water-soluble
Property organic solvent amount reach 5%~40%, using 1%~10% mass concentration aqueous hydrochloric acid solution adjust pH to step a in acid
Property group dissociation pH wait for that trapped fluid volume can not continue to detach using molecular cut off 100Da~800Da nanofiltration UF membranes
When, purified water dilutes trapped fluid, and after repeating 1~3 time, trapped fluid is collected when trapped fluid volume can not continue to detach to get two
Property alkaloid position.
2 both sexes alkaloid stachydrine of embodiment refines
Motherwort medicinal material Aqueous extracts are taken, stachydrine is enriched with using embodiment 1 and cation exchange resin, it is pure with stachydrine
Degree, separation cycle are comparativeanalysis.
The present invention：Isoelectric point by detecting stachydrine is 7.60 or so, and then calculates stachydrine using the method for titration
The pH that acid and alkaline group dissociates in structure is 10.50, and pH when basic group solution amorph is 5.50；Motherwort Aqueous extracts are taken, are used
The ultrafiltration membrane of molecular cut off 50KDa cellulose materials carries out pretreatment removal high molecular weight protein, polysaccharide, collects ultrafiltrate；It is super
Filtrate uses ethyl alcohol to be diluted to containing concentration of alcohol as 25%, the sodium hydrate aqueous solution adjusting pH to 10.50 of 5% mass concentration,
Using molecular cut off 500Da polyacrylonitrile material nanofiltration UF membranes, when trapped fluid volume can not continue to detach, purified water is dilute
Trapped fluid is released, after being repeated 2 times, collection terminal trapped fluid when trapped fluid volume can not continue to detach；Terminal trapped fluid is used
Ethyl alcohol is gradually diluted in solution reaches 40% containing ethyl alcohol, adjusts pH to 5.50 using the aqueous hydrochloric acid solution of 5% mass concentration, adopts
With retention molecule 300Da nanofiltration UF membranes, when trapped fluid volume can not continue to detach, purified water dilutes trapped fluid, is repeated 2 times
Afterwards, trapped fluid is collected when trapped fluid volume can not continue to detach to get stachydrine position.Waters2695 high performance liquid chromatography
Instrument, 6000 evaporative light scattering detector of Alltech detect the stachydrine of stachydrine reference substance and refining spearation, water respectively
Soviet Union's alkali reference substance chromatogram is shown in Fig. 1, and the chromatogram of the both sexes alkaloid stachydrine of enrichment is shown in Fig. 2, and then calculates stachydrine purity
It the results are shown in Table 1；
Cation exchange resin processes take motherwort extracting solution 1000mL to use, using 1000mL732 type cation exchange trees
Fat adsorption refining refines the detection of position liquid phase, and chromatogram is shown in Fig. 3, and then calculates stachydrine purity and the results are shown in Table 1.
Table 1
| Process for purification | Stachydrine purity (%) | Separation cycle |
| Embodiment 1 | 77.6 | 3 hours |
| Resin cation adsorbs | 42.5 | 20 hours |
By changing stachydrine existence, adjustment and active force before NF membrane, amplification stachydrine and its in motherwort
Difference between the effects between allogene alkali, sterol, liposoluble ingredient and NF membrane, realization stachydrine and its in motherwort of Objective
The quick separating of his constituents, purity 77.6%.Because being operated using room temperatureization in subtractive process, stachydrine is hardly deposited
It is lost in conversion.It is adsorbed using resin cation, wherein loading absorption, elution hitless operation, separation cycle is more than 20 small
When, and separating liquid needs thermal concentration, because to both sexes alkaloid can not characteristic, stachydrine in prepared TA position
Purity is 42.5%, has apparent technical advance using the method in embodiment 1.
3 both sexes alkaloid trigonelline of embodiment refines
Faenum graecum medicinal material Aqueous extracts are taken, trigonelline are enriched with using embodiment 1 and cation exchange resin, with trigonelline
Purity, production cycle are comparativeanalysis.
The present invention：Isoelectric point by detecting trigonelline is 6.10 or so, and then calculates wood using the method for titration
The pH that acid and alkaline group dissociates in alkali structure is 8.50, and pH when basic group solution amorph is 4.00；Faenum graecum Aqueous extracts are taken, are used
The ultrafiltration membrane of molecular cut off 100KDa cellulose materials carries out pretreatment removal high molecular weight protein, polysaccharide, collects ultrafiltrate；It is super
Filtrate uses methanol dilution to being 30% containing methanol concentration, and the sodium hydrate aqueous solution of 5% mass concentration adjusting pH to 8.50 is adopted
With molecular cut off 800Da polyamide material nanofiltration UF membranes, when trapped fluid volume can not continue to detach, purified water dilution is cut
Liquid is stayed, after being repeated 2 times, collection terminal trapped fluid when trapped fluid volume can not continue to detach；Terminal trapped fluid is used into methanol
Gradually be diluted in solution and reach 35% containing methanol, using 5% mass concentration aqueous hydrochloric acid solution adjust pH to 4.00, using cut
Molecule 300Da nanofiltration UF membranes are stayed, when trapped fluid volume can not continue to detach, purified water dilutes trapped fluid, after being repeated 3 times,
Trapped fluid is collected when trapped fluid volume can not continue to detach to get trigonelline position.Waters2695 high performance liquid chromatography
Instrument, PAD UV detector detect and calculate trigonelline purity and the results are shown in Table 2；
Cation exchange resin processes take faenum graecum extracting solution 1000mL to use, using 1000mL732 type cation exchange trees
Fat adsorption refining, refined position calculate trigonelline purity and the results are shown in Table 2.
Table 2
| Process for purification | Trigonelline purity (%) | Separation cycle |
| Embodiment 1 | 70.4 | 3 hours |
| Resin cation adsorbs | 24.3 | 25 hours |
By being compared with ion exchange resin, in embodiment 1 using organic solvent adjust both sexes alkaloid and NF membrane it
The isolation technics of intermolecular forces is with the obvious advantage, the trigonelline purity in prepared alkaloid position be 70.4%, higher than sun from
Trigonelline prepared by sub- resin adsorption, at the same ultrafiltration-nanofiltration combination separation short implementation cycle, organic reagent dosage well below
Resin detaches, which can carry out purpose separation to both sexes alkaloid, and the subtractive process of room temperature does not generate ingredient damage
It loses.
Claims (4)
1. a kind of method that film combination refines both sexes alkaloid, it is characterised in that：It comprises the steps of：
A. the isoelectric point of both sexes alkaloid to be separated is detected, and it is in solution to calculate acidic-group in its both sexes alkaloid compound structure
PH when pH and basic group journey solution amorph when amorph；
B. extract to be separated is dissolved in purified water, pretreatment is carried out using molecular cut off 10KDa~100KDa ultrafiltration membranes
Except high molecular weight protein, polysaccharide, ultrafiltrate is collected；
C. step b ultrafiltrates containing water-soluble organic solvent amount in solution is gradually diluted to using water-miscible organic solvent to reach
5%~40%, and then pH is adjusted to step a neutral and alkali group solutions using the sodium hydrate aqueous solution of 1%~10% mass concentration
From pH, using molecular cut off 100Da~800Da nanofiltration UF membranes, when trapped fluid volume can not continue to detach, purified water
Trapped fluid is diluted, after repeating 1~3 time, collection terminal trapped fluid when trapped fluid volume can not continue to detach；
D. the terminal trapped fluid in step c is gradually diluted to containing water-soluble organic solvent in solution using water-miscible organic solvent
Amount reaches 5%~40%, and adjusting acidic-group in pH to step a using the aqueous hydrochloric acid solution of 1%~10% mass concentration dissociates
PH, using molecular cut off 100Da~800Da nanofiltration UF membranes, when trapped fluid volume can not continue to detach, purified water is dilute
Trapped fluid is released, after repeating 1~3 time, trapped fluid is collected when trapped fluid volume can not continue to detach to get both sexes alkaloid portion
Position.
2. a kind of method that film combination refines both sexes alkaloid according to claim 1, it is characterised in that：In step b
Ultrafiltration membrane material is one kind in cellulose, polyether sulfone, cellulose acetate.
3. a kind of method that film combination refines both sexes alkaloid according to claim 1, it is characterised in that：Step c and step
Water-miscible organic solvent in rapid d is one kind in methanol, ethyl alcohol, acetonitrile, tetrahydrofuran.
4. a kind of method that film combination refines both sexes alkaloid according to claim 1, it is characterised in that：Step c and step
Nanofiltration membrane material in rapid d is one kind in organic solvent-resistant material polyimides, polyamide, polyacrylonitrile.
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| CN201810633249.4A CN108586308B (en) | 2018-06-15 | 2018-06-15 | Method for refining amphoteric alkaloid by membrane combination |
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| CN201810633249.4A CN108586308B (en) | 2018-06-15 | 2018-06-15 | Method for refining amphoteric alkaloid by membrane combination |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103830298A (en) * | 2014-03-26 | 2014-06-04 | 南京中医药大学 | Method for preparing panax notoginseng saponins by adopting membrane coupling technology |
| CN106045890A (en) * | 2016-05-11 | 2016-10-26 | 南京泽朗医药科技有限公司 | A stachydrine preparing method |
| CN106631767A (en) * | 2017-01-04 | 2017-05-10 | 南京中医药大学 | Method for high-efficiency enrichment of phenolic acid components in traditional Chinese medicine through nanofiltration |
-
2018
- 2018-06-15 CN CN201810633249.4A patent/CN108586308B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103830298A (en) * | 2014-03-26 | 2014-06-04 | 南京中医药大学 | Method for preparing panax notoginseng saponins by adopting membrane coupling technology |
| CN106045890A (en) * | 2016-05-11 | 2016-10-26 | 南京泽朗医药科技有限公司 | A stachydrine preparing method |
| CN106631767A (en) * | 2017-01-04 | 2017-05-10 | 南京中医药大学 | Method for high-efficiency enrichment of phenolic acid components in traditional Chinese medicine through nanofiltration |
Non-Patent Citations (1)
| Title |
|---|
| 李存玉等: "超滤-纳滤联用优化益母草生物碱的浓缩工艺", 《中国中药杂志》 * |
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