CN101029088A - Method for producing Matrimony vine polysaccharide - Google Patents
Method for producing Matrimony vine polysaccharide Download PDFInfo
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- CN101029088A CN101029088A CN 200710090210 CN200710090210A CN101029088A CN 101029088 A CN101029088 A CN 101029088A CN 200710090210 CN200710090210 CN 200710090210 CN 200710090210 A CN200710090210 A CN 200710090210A CN 101029088 A CN101029088 A CN 101029088A
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Abstract
Production of Barbary Wolfberry Fruit polysaccharide is carried out by taking Barbary Wolfberry Fruit as raw materials, soaking, crushing, extracting, enzyme hydrolysis reacting, column chromatography acting, depositing by alcohol, dealcoholizing and drying. It's cheap and simple, has short production period and better purity.
Description
Technical field
The present invention relates to biological products and chemical engineering technology, particularly a kind of method of from matrimony vine, extracting lycium barbarum polysaccharide.
Background technology
Matrimony vine (Lycium chinense Mill.) is a Chinese medicinal materials commonly used, is again protective foods, doctor's food homology.The another name of matrimony vine is a lot, is tribute fruit, blood matrimony vine, matrimony vine beans again, and the place of production is called " red fruit ", " son makes eye bright " etc.It is the mature fruit of defoliation bush plant of casket section matrimony vine, and machaka is because of cultivation forms the upright shrub with tree crown through artificial pruning, the thick diameter 10-20cm of single trunk stem is about plant height 1.5m, about crown diameter 1.6m, leaf alternate or cluster, lanceolar or long ellipticity lanceolar, corolla funnel-form, violet, fruit is a berry, redness, the likeness in form spindle more weighs down like carnelian.Be royal tribute in history, have " red treasured " laudatory title, be rich in vegetable polysaccharides, protein, vitamins and other nutritious components.The main effect of matrimony vine is nourishing liver and kidney, replenishing vital essence to improve eyesight; Be used for that consumptive disease smart is lost, soreness of waist and knee joint; Dizzy tinnitus, Nei Rexiao is thirsty, the deficiency of blood is sallow, and blurred vision is not clear.Successive dynasties expert in health preserving, physician value the tonic effect of matrimony vine very much.In Shennong's Herbal, just point out: obey hard muscles and bones for a long time; " Mingyi Bielu " meaning matrimony vine is good at " nourishing the essence and strengthening QI "; " Shiliao Bencao " also puts down in writing matrimony vine " the beneficial people of energy removes consumptive disease ".The effect of matrimony vine aspect nourishing liver and kidney, replenishing vital essence to improve eyesight not only praised highly by ancient Chinese medicine doctor and dietotherapy scholar.Modern pharmacological research proves, the contained major ingredient lycium barbarum polysaccharide of matrimony vine (Lycium chinense polysaccharide, be called for short LBP) be most important functional component in the matrimony vine, it is a kind of water-soluble polysaccharide, clearly this polysaccharide is a protein-polysaccharide, molecular weight is 22～25kD, and the protein main chain that polysaccharide side chain of being made up of six kinds of second carbon monose ARA, CLC, GAL, MAN, XYL and RHA and 18 seed amino acids are formed constitutes.Lycium barbarum polysaccharide is that wolfberry fruit is regulated immunity, the main active ingredient that delays senility, can improve people's immunologic function, promote body to produce the NK-killer cell, and improve its activity, can play antivirus action indirectly by the enhance immunity function, the neural internal secretion of equilibrium one immunological network there is vital role, can directly act on hypothalamus one hypophysis of human body, promote metabolic function, the adjusting of endocrine function and reproductive function, it is to malignant tumour, the prevention of acquired immune deficiency syndrome (AIDS) also can be played active effect with treatment, it can improve the elderly's fatiguability in addition, symptoms such as poor appetite and blurred vision, and have reducing blood-fat, anti-fatty liveranti-fatty liver, antitumor, the anti-ageing effect of waiting for a long time.
Through patent retrieval, find patent " application number 200510046785.7 fermentation extraction of Chinese wolfberry fruit polysaccharide ", this disclosure of the Invention a kind of applied bioengineering technology extract the method for processing lycium barbarum polysaccharide, be to utilize the means that consume monose, disaccharide, nitrogenous substances in the Angel Yeast process of growth to improve the purity of lycium barbarum polysaccharide, whole technological process comprises pulverizing, soak, aerobic fermentation filters, decolouring, removal of impurities, vacuum distilling.Problems such as this method adopts zymotechnique, is prone to fermentation in the production process not exclusively, and impurity is many.Patent " 02115382.5 1 kinds of lycium barbarum polysaccharides of application number and its production and application ", disclose the preparation method of lycium barbarum polysaccharide: with the Chinese medicine matrimony vine is raw material, the hot water that adds 8～20 times 50～100 ℃, sheared 10～40 minutes squeezing, pressure expressed juice high speed centrifugation with high-speed shearing machine, the centrifugate molecular weight cut-off is 5～100,000 ultrafiltration membrance filter, ultrafiltrated is that 3000～10000 ultra-filtration membrane concentrates with molecular weight cut-off again, lyophilize, promptly.This method adopts membrane sepn, and yield is low in the production.
Summary of the invention
The object of the present invention is to provide a kind of technology of utilizing modern biological method to extract lycium barbarum polysaccharide, reach and improve lycium barbarum polysaccharide purity, the discharging of minimizing industrial waste, shortening production process, the purpose that reduces production costs.
The technical solution adopted for the present invention to solve the technical problems is: a kind of extraction working method of lycium barbarum polysaccharide is provided, specifically extracts procedure of processing and be:
1, the selection of raw material and pre-treatment: select new fresh fructus lycii or dried fruit of lycium barbarum for use, repeatedly to remove impurity, washing time was advisable with 30 seconds to 1 minute in the clear water washing;
2, soak: the volume ratio of matrimony vine and pure water is 1: the ratio soak at room temperature of 3-5, and the dried fruit of lycium barbarum soak time was advisable with 6～8 hours, and new fresh fructus lycii can suitably foreshorten to 2～3 hours;
3, fragmentation: matrimony vine after will soaking and soak solution inject pulverizer together and carry out fragmentation;
4, extract: the volume ratio in broken liquid of matrimony vine and pure water is 1: the ratio of 1-2 is extracted after adding pure water, and extracting temperature is 80-100 ℃, and pressure is 60-80KPa, and extraction time is 1-2 hour; Collect extracting solution, residue extracts 2 times with 10 multiple dose pure water again;
5, enzymolysis: united extraction liquid, the amount of pressing 0.05-0.5g/L adds polygalacturonase, is incubated enzymolysis 2-3 hour, the extremely boiling of heating extracting solution, enzymolysis reaction in 50-60 ℃;
6, concentrate: centrifugal behind the extracting liquid filtering, merge supernatant liquor, residue washs after-filtration with pure water, with filtrate and centrifugal after supernatant liquor merge, carry out concentrating under reduced pressure;
7, column chromatography: the solution after will concentrating is by the column chromatography resin post, post liquid volume 3-5 pure water doubly cleans resin with being equivalent to into, to remove the pigment in the solution, when effluent liquid shoals Huang, begin to collect effluent liquid, when the effluent liquid color is redly shoaled by pale yellow shoaling again, when the effluent liquid color when colourless, stop wash-out, resin column is used the ethanol clean of 75% above concentration again, the materials such as pigment of flush away absorption;
8, alcohol precipitation: concentrate eluant, add the ethanol of 95% above concentration in the concentrated solution, the dosage that adds alcohol is generally 5～7 times of concentrate eluant volume, adjusts the solution determining alcohol and reaches more than 80%, then, leaves standstill in 2-10 ℃ and fully to precipitate in 4～8 hours;
9, dealcoholysis: filter, collecting precipitation is with 95% concentration ethanol washing precipitation 2～3 times, with dissolved in purified water post precipitation concentrating under reduced pressure, to 25～30 mother-in-law U.S. degree medicinal extract;
10, drying: medicinal extract is carried out lyophilize, obtain the lycium barbarum polysaccharide powder after the pulverizing.
The present invention, volume ratio the best of matrimony vine and pure water is 1: 4 in the step 1; In the step 2, it is that to soak Best Times be 2 hours 7 hours, new fresh fructus lycii that dried fruit of lycium barbarum soaks Best Times, and the time is too short can influence leaching yield, and long soaking time can influence production cycle or easy microbiological contamination; In the step 4, the optimum extraction temperature is that 90 ℃, pressure are 70KPa, and temperature is too high, pressure is crossed conference and destroyed active substance in the matrimony vine; In the step 5, the suitable consumption of polygalacturonase is 0.1-0.2g/L; In the step 7, column chromatography resin is selected from the macroporous adsorbent resin that model is D101 type, D201 type, DA201 type, AB-8 type, the synthetic polymeric adsorbent of small grain size that model is HP20SS or SP20SS; In the step 8, optimum temps is 4 ℃ when leaving standstill, the too high precipitation that is unfavorable for of temperature, and temperature is crossed the low energy consumption that then increases.
The lycium barbarum polysaccharide that preparation method of the present invention obtains is little sweet or tasteless, the powder that particle is tiny of golden yellow, flavor, and wherein the content of lycium barbarum polysaccharide is more than 40%.
The present invention extracts working method and has following advantage compared with prior art:
● reaction process is simple: the ethanol sedimentation extraction method step of present domestic application is miscellaneous, and only one of absolute ethanol washing just needs washing for several times, and column chromatographic technique of the present invention has reduced the step of ethanol precipitation;
● raw material consumption reduces, and ethanol sedimentation extraction method mass consumption dehydrated alcohol is a kind of invisible waste to extracting raw material;
● defectives such as production technique is simple, with short production cycle, technological process is beneficial to control, and the microbial fermentation of having avoided fermentation method is not exclusively, the yield of membrane sepn is low.
Embodiment
Embodiment 1
1, the selection of raw material and pre-treatment: select for use new fresh fructus lycii an amount of, repeatedly to remove impurity, washing time was advisable with 30 seconds to 1 minute in the clear water washing;
2, soak: in the volume ratio of matrimony vine and pure water is 1: 3 ratio soak at room temperature, and newly fresh fructus lycii can suitably foreshorten to 3 hours;
3, fragmentation: matrimony vine after will soaking and soak solution inject pulverizer together and carry out fragmentation;
4, extract: in the volume ratio of the broken liquid of matrimony vine and pure water is to extract after 1: 1 ratio is added pure water, and the extraction temperature is 80 ℃, and pressure is 60KPa, and extraction time is 1 hour; Collect extracting solution, residue extracts 2 times with 10 multiple dose pure water again;
5, enzymolysis: united extraction liquid, the amount of pressing 0.05g/L adds polygalacturonase, is incubated enzymolysis 2-3 hour, the extremely boiling of heating extracting solution, enzymolysis reaction in 50-60 ℃;
6, concentrate: centrifugal behind the extracting liquid filtering, merge supernatant liquor, residue washs after-filtration with pure water, with filtrate and centrifugal after supernatant liquor merge, carry out concentrating under reduced pressure;
7, column chromatography: the solution after will concentrating passes through column chromatography, model is the D201 resin column, post liquid volume 3-5 pure water doubly cleans resin with being equivalent to into, to remove the pigment in the solution, when effluent liquid shoals Huang, begin to collect effluent liquid, when the effluent liquid color is redly shoaled by pale yellow shoaling again, when the effluent liquid color when colourless, stop wash-out, resin column is used the ethanol clean of 75% above concentration again, the materials such as pigment of flush away absorption;
8, alcohol precipitation: concentrate eluant, add the ethanol of 95% above concentration in the concentrated solution, the dosage that adds alcohol is generally 5～7 times of concentrate eluant volume, adjusts the solution determining alcohol and reaches more than 80%, then, leaves standstill in 2 ℃ and fully to precipitate in 4 hours;
9, dealcoholysis: filter, collecting precipitation is with 95% concentration ethanol washing precipitation 2～3 times, with dissolved in purified water post precipitation concentrating under reduced pressure, to 25～30 mother-in-law U.S. degree medicinal extract;
10, drying: medicinal extract is carried out lyophilize, obtain the lycium barbarum polysaccharide powder after the pulverizing.
Embodiment 2
1, the selection of raw material and pre-treatment: select for use dried fruit of lycium barbarum an amount of, repeatedly to remove impurity, washing time was advisable with 30 seconds to 1 minute in the clear water washing;
2, soak: in the volume ratio of matrimony vine and pure water is 1: 4 ratio soak at room temperature, and newly fresh fructus lycii can suitably foreshorten to 7 hours;
3, fragmentation: matrimony vine after will soaking and soak solution inject pulverizer together and carry out fragmentation;
4, extract: in the volume ratio of the broken liquid of matrimony vine and pure water is to extract after 1: 2 ratio is added pure water, and the extraction temperature is 90 ℃, and pressure is 70KPa, and extraction time is 1 hour; Collect extracting solution, residue extracts 2 times with 10 multiple dose pure water again;
5, enzymolysis: united extraction liquid, the amount of pressing 0.2g/L adds polygalacturonase, is incubated enzymolysis 2-3 hour, the extremely boiling of heating extracting solution, enzymolysis reaction in 50-60 ℃;
6, concentrate: centrifugal behind the extracting liquid filtering, merge supernatant liquor, residue washs after-filtration with pure water, with filtrate and centrifugal after supernatant liquor merge, carry out concentrating under reduced pressure;
7, column chromatography: the solution after will concentrating passes through column chromatography, model is the D101 resin column, post liquid volume 3-5 pure water doubly cleans resin with being equivalent to into, to remove the pigment in the solution, when effluent liquid shoals Huang, begin to collect effluent liquid, when the effluent liquid color is redly shoaled by pale yellow shoaling again, when the effluent liquid color when colourless, stop wash-out, resin column is used the ethanol clean of 75% above concentration again, the materials such as pigment of flush away absorption;
8, alcohol precipitation: concentrate eluant, add the ethanol of 95% above concentration in the concentrated solution, the dosage that adds alcohol is generally 5～7 times of concentrate eluant volume, adjusts the solution determining alcohol and reaches more than 80%, then, leaves standstill in 4 ℃ and fully to precipitate in 6 hours;
9, dealcoholysis: filter, collecting precipitation is with 95% concentration ethanol washing precipitation 2～3 times, with dissolved in purified water post precipitation concentrating under reduced pressure, to 25～30 mother-in-law U.S. degree medicinal extract;
10, drying: medicinal extract is carried out lyophilize, obtain the lycium barbarum polysaccharide powder after the pulverizing.
Embodiment 3
1, the selection of raw material and pre-treatment: select for use dried fruit of lycium barbarum an amount of, repeatedly to remove impurity, washing time was advisable with 30 seconds to 1 minute in the clear water washing;
2, soak: in the volume ratio of matrimony vine and pure water is 1: 5 ratio soak at room temperature, and newly fresh fructus lycii can suitably foreshorten to 8 hours;
3, fragmentation: matrimony vine after will soaking and soak solution inject pulverizer together and carry out fragmentation;
4, extract: in the volume ratio of the broken liquid of matrimony vine and pure water is to extract after 1: 2 ratio is added pure water, and the extraction temperature is 100 ℃, and pressure is 80KPa, and extraction time is 1 hour; Collect extracting solution, residue extracts 2 times with 10 multiple dose pure water again;
5, enzymolysis: united extraction liquid, the amount of pressing 0.5g/L adds polygalacturonase, is incubated enzymolysis 2-3 hour, the extremely boiling of heating extracting solution, enzymolysis reaction in 50-60 ℃;
6, concentrate: centrifugal behind the extracting liquid filtering, merge supernatant liquor, residue washs after-filtration with pure water, with filtrate and centrifugal after supernatant liquor merge, carry out concentrating under reduced pressure;
7, column chromatography: the solution after will concentrating passes through column chromatography, model is the synthetic adsorption column of the small grain size of HP20SS, post liquid volume 3-5 pure water doubly cleans resin with being equivalent to into, to remove the pigment in the solution, when effluent liquid shoals Huang, begin to collect effluent liquid, when the effluent liquid color is redly shoaled by pale yellow shoaling again, when the effluent liquid color when colourless, stop wash-out, resin column is used the ethanol clean of 75% above concentration again, the materials such as pigment of flush away absorption;
8, alcohol precipitation: concentrate eluant, add the ethanol of 95% above concentration in the concentrated solution, the dosage that adds alcohol is generally 5～7 times of concentrate eluant volume, adjusts the solution determining alcohol and reaches more than 80%, then, leaves standstill in 10 ℃ and fully to precipitate in 8 hours;
9, dealcoholysis: filter, collecting precipitation is with 95% concentration ethanol washing precipitation 2～3 times, with dissolved in purified water post precipitation concentrating under reduced pressure, to 25～30 mother-in-law U.S. degree medicinal extract;
10, drying: medicinal extract is carried out lyophilize, obtain the lycium barbarum polysaccharide powder after the pulverizing.
Claims (7)
1, a kind of preparation method of lycium barbarum polysaccharide is characterized in that: extract lycium barbarum polysaccharide from wolfberry fruit, concrete steps are:
1), the selection and the pre-treatment of raw material: select new fresh fructus lycii or dried fruit of lycium barbarum for use, repeatedly to remove impurity, washing time was advisable with 30 seconds to 1 minute in the clear water washing;
2), soak: the volume ratio in matrimony vine and pure water is 1: the ratio soak at room temperature of 3-5, the dried fruit of lycium barbarum soak time was advisable with 6～8 hours, newly fresh fructus lycii can suitably foreshorten to 2～3 hours;
3), fragmentation: matrimony vine after will soaking and soak solution inject pulverizer together and carry out fragmentation;
4), extract: the volume ratio in broken liquid of matrimony vine and pure water is 1: the ratio of 1-2 is extracted after adding pure water, and extracting temperature is 80-100 ℃, and pressure is 60-80KPa, and extraction time is 1-2 hour; Collect extracting solution, residue extracts 2 times with 10 multiple dose pure water again;
5), enzymolysis: united extraction liquid, the amount of pressing 0.05-0.5g/L adds polygalacturonase, in 50-60 ℃ of insulation enzymolysis 2-3 hour, the heating extracting solution was to boiling, enzymolysis reaction;
6), concentrate: centrifugal behind the extracting liquid filtering, merge supernatant liquor, residue washs after-filtration with pure water, with filtrate and centrifugal after supernatant liquor merge, carry out concentrating under reduced pressure;
7), column chromatography: the solution after will concentrating is by the column chromatography resin post, post liquid volume 3-5 pure water doubly cleans resin with being equivalent to into, to remove the pigment in the solution, when effluent liquid shoals Huang, begin to collect effluent liquid, when the effluent liquid color is redly shoaled by pale yellow shoaling again, when the effluent liquid color when colourless, stop wash-out, resin column is used the ethanol clean of 75% above concentration again, the materials such as pigment of flush away absorption;
8), alcohol precipitation: concentrate eluant, add the ethanol of 95% above concentration in the concentrated solution, the dosage that adds alcohol is generally 5～7 times of concentrate eluant volume, adjusts the solution determining alcohol and reaches more than 80%, then, leaves standstill in 2-10 ℃ and fully to precipitate in 4～8 hours;
9), dealcoholysis: filter, collecting precipitation is with 95% concentration ethanol washing precipitation 2～3 times, with dissolved in purified water post precipitation concentrating under reduced pressure, to 25～30 mother-in-law U.S. degree medicinal extract;
10), drying: medicinal extract is carried out lyophilize, obtain the lycium barbarum polysaccharide powder after the pulverizing.
2, as the preparation method of right 1 described a kind of lycium barbarum polysaccharide, it is characterized in that: volume ratio the best of matrimony vine and pure water is 1: 4 in the step 1).
3, as the preparation method of right 1 described a kind of lycium barbarum polysaccharide, it is characterized in that: step 2) in, it is that to soak Best Times be 2 hours for 7 hours, new fresh fructus lycii that structure Qi dry fruit soaks Best Times.
4, as the preparation method of right 1 described a kind of lycium barbarum polysaccharide, it is characterized in that: in the step 4), the optimum extraction temperature is that 90 ℃, pressure are 70KPa.
5, as the preparation method of right 1 described a kind of lycium barbarum polysaccharide, it is characterized in that: in the step 5), the suitable consumption of polygalacturonase is 0.1-0.2g/L.
6, as the preparation method of right 1 described a kind of lycium barbarum polysaccharide, it is characterized in that: in the step 7), column chromatography resin is selected from the macroporous adsorbent resin that model is D101 type, D201 type, DA201 type, AB-8 type, the synthetic polymeric adsorbent of small grain size that model is HP20SS or SP20SS.
7, as the preparation method of right 1 described a kind of lycium barbarum polysaccharide, it is characterized in that: in the step 8), optimum temps is 4 ℃ when leaving standstill.
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