CN103435714B - The preparation of the thick polysaccharide of a kind of fig and purification process and purposes - Google Patents
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Abstract
The present invention is the preparation method of the thick polysaccharide of a kind of fig, and its step is as follows: get that pulled figs is some to be shredded, with 95% alcohol reflux degrease and pigment, after suction filtration, residue removes small molecular weight impurity with 70% alcohol reflux; Then get residue, as the extraction raw material of fig polysaccharide; Get residue, after water extracts under microwave condition as solvent, suction filtration, gets filtrate and is concentrated into 100Ml with rotary evaporation, adds and leaves standstill 24h after 95% ethanol, gets precipitation after centrifugal, and precipitation is by Sevage method except after deproteinized, and freeze drying, obtains the thick polysaccharide of fig; When extraction, the solid-liquid ratio 1:20-23 of residue and water; Microwave power is 550W, and extraction time is 13min, and the 3.50-3.60 that alcohol precipitation is extracting liquid volume by amount of alcohol doubly. The invention also discloses purification process and the purposes of the thick polysaccharide of fig.
Description
Technical field
The present invention relates to a kind of preparation method of polysaccharide, particularly the preparation method of the thick polysaccharide of a kind of fig; The present inventionAlso relate to the purification process of the thick polysaccharide of fig that preceding method makes; The invention still further relates to the thick polysaccharide of fig or figPolysaccharide sterling is in the purposes of preparing in alpha-glucosidase restrainer.
Background technology
Fig Ficuscarica.L is subordinate to Moraceae Ficus, and edible part is its dry holder, and not only taste is sweetU.S., also has nutritive value and medical value widely. It is reported, fig can be used for invigorating stomach and promoting bower movements, subdhing swelling and detoxicating, and can press downTumour processed, raising immunity of organisms. Fig is rich in polysaccharide, and polysaccharide has multiple physiologically active, as antitumor, immunological regulation,Antibacterial, anti-oxidant, Inhibiting α-glucosidase etc. All can from document, find experimental basis about the biologically active such as antitumor,But it suppresses effect nobody research of glucuroide, and alpha-glucosidase energy catalysis α-Isosorbide-5-Nitrae-hydrolysis of glycoside bond, is littleThe enzyme of the oligosaccharides hydrolysis such as the interior maltose of intestines, sucrose, the activity of Inhibiting α-glucosidase, can make the generation of glucose and absorption subtractSlow, reduce Postprandial peak glucose. Therefore the alpha-glucosidase restrainer novel type-II diabetes medication that is a class, can be used for after the mealBlood glucose-control, keeps patient's postprandial blood sugar steady.
Summary of the invention
Technical problem to be solved by this invention is for the deficiencies in the prior art, and a kind of method advantages of simple, behaviour are providedMake the preparation method of the thick polysaccharide of fig convenient, yield is high.
Another technical problem to be solved by this invention has been to provide the purifying of the thick polysaccharide of fig prepared by said methodMethod.
The fig that the thick polysaccharide of fig that the method for the invention makes or described purification process purifying obtain is manySugar sterling can be for the preparation of alpha-glucosidase restrainer.
Technical problem to be solved by this invention is to realize by following technical scheme. The present invention is a kind of without flowerThe preparation method of the thick polysaccharide of fruit, is characterized in, its step is as follows: get that pulled figs is some to be shredded, with 95% alcohol reflux degrease andPigment, after suction filtration, residue removes small molecular weight impurity with 70% alcohol reflux; Then get residue, as the extraction raw material of fig polysaccharide;Get residue, after water extracts under microwave condition as solvent, suction filtration, gets filtrate and is concentrated into 100Ml with rotary evaporation, adds 95%After ethanol, leave standstill 24h, get precipitation after centrifugal, precipitation is by Sevage method except after deproteinized, and freeze drying, obtains fig slightly manySugar; When extraction, the solid-liquid ratio 1:20-23 of residue and water; Microwave power is 550W, and extraction time is 13min, alcohol precipitation amount of alcoholFor 3.50-3.60 times of extracting liquid volume.
In the preparation method of the thick polysaccharide of fig of the present invention, most preferred technical characterictic is: when extraction, residue andThe solid-liquid ratio 1:22.45 of water; Alcohol precipitation is extracting liquid volume by amount of alcohol 3.55 times.
The invention also discloses a kind of purification process of the thick polysaccharide of fig that method makes as described in above technical scheme,Be characterized in, its step is as follows:
(1) ion exchange cellulose DEAE-52 is after distilled water is fully swelling, with the NaOH of 0.5M and the HCl of 0.5MAlternate treatment is used distilled water cyclic washing for several times again, until its pH to 6 fills post, adopts 2.0cm × 30cm normal pressure sandCore glass post, it is stand-by that the ion column installing carries out balance 24 hours with distilled water again;
(2) get the thick polysaccharide of fig and be mixed with the solution of mass concentration as 1% taking distilled water; Then fig polysaccharide is moltenThe DEAE-52 ion exchange column that on liquid, balance is good, loading volume is 5mL, uses 0-1MNaCl200mL with 50mL/hFlow velocity cross post, collect with the amount of every pipe 5mL with automatic fraction collector, with the every pipe of phenolsulfuric acid colorimetric method for determiningTotal sugar concentration is made elution curve, collects identical component according to elution curve, obtains without flower in 40 DEG C of reduced pressure concentrations freeze dryingFruit polysaccharide;
(3) SephacrylS-300HR softly washs for several times, reduces pressure after degassed through distilled water and fills post, adopt 1.6cm ×80cm post, with distilled water balance; The fig polysaccharide that step (2) is obtained is dissolved in deionized water, and compound concentration is 15mg/mLPolysaccharide solution, on SephacrylS-300HR molecular sieve column that balance is good, applied sample amount is 2mL, by deionized water with 6The flow velocity of mL/h carries out wash-out; Collect with fraction collector, survey the total sugar concentration of every pipe by phenolsulfuric acid colorimetric method, thenMerge identical component according to elution curve, desalt after dialysis, 40 DEG C of reduced pressure concentration postlyophilizations; As above operation is carried out pure repeatedlyChange several times freeze drying and obtain fig polysaccharide sterling.
Below the inventive method is made a more detailed description.
1.1 materials and reagent
Fig dry fruit is purchased from Hai Lianlu market, Lianyungang; Alpha-glucosidase (lot number: EC3.2.1.20); 4-Nitrophenols-α-D-glucopyranoside (PNPG, lot number: 026K1516); Acarbose (lot number: lot16869); Below all purchaseFrom Sigma company; 4-nitrophenol (PNP, Shanghai brilliant pure reagent Co., Ltd); Sulfuric acid, glucose, phenol, ethanol etc. areDomestic analysis is pure.
2 methods
Preparation and the assay of the thick polysaccharide of 2.1 fig (CFCP)
2.1.1 calibration curve
Adopt with glucose as a standard product of phenolsulfuric acid method, in the absorbance of 490nm colorimetric estimation polysaccharide extraction liquid,Carry out linear regression with absorbance (A) and solution polysaccharide mass concentration (c), obtain regression equation and be: A=14.568c – 0.0082, r=0.9972. Result shows, glucose is good at 7.5～52.5 μ g/mL and absorbance linear relationship.
2.1.2CFCP extract
Due to defects such as classical water extracting method are consuming time, and yield is low, slightly carry in process at fig polysaccharide, adopt microwaveMethod is extracted, for obtaining maximum polysaccharide yield, and the optimization that has adopted response surface method to extract each factor. Adopt figFour factors that polysaccharide yield impact is larger, add the volume multiple of alcohol when microwave power, microwave time, solid-liquid ratio and alcohol precipitation.
Pretreatment before extracting: get that pulled figs is some to be shredded, with 95% alcohol reflux degrease and pigment, residue use after suction filtration70% alcohol reflux removes small molecular weight impurity, then gets residue, as the extraction raw material of fig polysaccharide. Get residue water as moltenAfter agent is extracted under microwave condition, suction filtration, gets filtrate and is concentrated into 100mL left and right with rotary evaporation, adds several times volume 95% ethanolThe rear 24h that leaves standstill, gets precipitation after centrifugal, and precipitation is by Sevage method except after deproteinized, and freeze drying, obtains the thick polysaccharide of figCFCP. And be mixed with certain density CFCP solution, stand-by.
Table 1 fig polysaccharide extracts response surface analysis factor and level
Table 2 Box-Behnken design and response
The regression equation being gone out by Box-Behnken experimental fit is: Y=10.0332+0.1688X1+0.3285X2+0.4118X3-0.2475X4-0.2950X1 2-1.0071X2 2-0.7270X3 2-0.7709X4 2+0.2498X1X2-0.0878X1X3-0.1013X1X4+0.6615X2X3-0.3713X2X4-0.3510X3X4
In formula, Y is polysaccharide yield (%), X1For liquid ratio (g/mL), X2For microwave power (W), X3For ethanol volume integralNumber (%), X4For extraction time (min).
The every variance analysis of regression equation is in table 3.
Table 3 analysis of variance table
Note: * * Pr > F value is less than 0.0001 for highly significant; * Pr > F value is less than 0.05 for significantly
Can find out, while describing being related between each factor and response with above-mentioned regression equation, dependent variable and all fromLinearity between variable
Relation is (r=17.75/18.05=0.9834) significantly, and the level of signifiance of model is far smaller than 0.05, nowQuadratic regression variance model is highly significant. The results of analysis of variance every from regression equation it can also be seen that, equationMistake intend littlely, show that this equation is good to experimental result matching, experimental error is little, therefore, available this regression equation replacesThe true point of experiment carries out analysis and prediction to experimental result.
To regression equation go single order partially inverse equal 0, arrange to obtain response when maximum, X1=0.49，X2=0.50，X3=0.59, X4=-0.45. The optimum process condition that corresponding fig polysaccharide extracts is: solid-liquid ratio 1:22.45, microwave power550W, extraction time 12.9min, alcohol precipitation is extract by amount of alcohol 3.55 times, with this understanding, the yield of fig polysaccharideCan reach 10.33% in theory. For the ease of operation, optimum extraction condition is modified to: solid-liquid ratio 1:20, microwave power is 550W,Extraction time is 13 minutes, and alcohol precipitation ethanol volume multiple is 3.5 times. Adopt above-mentioned optimal conditions, carried out extraction fig manyThe confirmatory experiment of sugar, parallel 3 times, the average recovery rate obtaining is 10.24%, substantially conforms to predicted value, and response surface method pair is describedThe optimization of fig polysaccharide extraction conditions is feasible.
Compare the fig polysaccharide yield of Microwave Extraction method and traditional water extraction. Result water extraction 2h consuming time, extracts 2Inferior, yield is 8.87%, illustrates that microwave method extracts polysaccharide, with respect to traditional water extraction have consuming time less, that energy-conservation, yield is high etc. is excellentPoint. Polyoses content in thick polysaccharide is high, not only can make full use of raw material, can also make follow-up purifying work more easy to operate.
2.1.3CFCP middle determination of polysaccharide
Get the CFCP52.0124mg after freeze drying, be dissolved in 100mL volumetric flask, and be adjusted to scale with pure water. Shake upAfter, the accurate 5mL that draws, in 10mL volumetric flask, and mends to scale with pure water. Shake up, get 1mL and carry out quantitative assay, average threeInferior, obtain A=0.23, bringing calibration curve into, to obtain concentration in colorimetric cylinder be 0.01635ug/mL. So polyoses content is in FCP0.01635 × 10 × 1 × 10 × 20=32.7018mg. So polyoses content is 32.7018/52.0124 × 100%=in CFCP62.76%。
The purifying of 2.2CFCP
For obtaining the fig polysaccharide that purity is higher, further adopt column chromatography to carry out purifying.
2.2.1 ion exchange cellulose DEAE-52 is after distilled water is fully swelling, with the NaOH of 0.5M and 0.5MHCl alternate treatment is used distilled water cyclic washing for several times again, until its pH fills post in 6 left and right, (2.0cm × 30cm is normalPress core glass column), it is stand-by that the ion column installing carries out balance 24 hours with distilled water again. Fig through dialysis freeze-drying is slightly manySugar CFCP1g is mixed with the solution of mass concentration as 1% taking distilled water. Then by balance on fig polysaccharide CFCP solutionGood DEAE-52 ion exchange column, loading volume is 5mL, crosses post with 0-1MNaCl200mL with the flow velocity of 50mL/h, usesAutomatically fraction collector is collected with the amount of every pipe 5mL, washes with the total sugar concentration of the every pipe of phenolsulfuric acid colorimetric method for determiningDe-curve, collects identical component according to elution curve, collects 13 pipes to 32 pipes, obtains nothing in 40 DEG C of reduced pressure concentrations freeze dryingFlowers and fruits polysaccharide FCP. Elution curve is shown in Fig. 1.
2.2.2SephacrylS-300HR molecular sieve column chromatography
SephacrylS-300HR softly washs for several times, reduces pressure after degassed through distilled water and fills post (1.6cm × 80Cm), with distilled water balance. The fig polysaccharide FCP obtaining by ion exchange column is dissolved in to deionized water, and compound concentration isThe polysaccharide solution of 15mg/mL, on SephacrylS-300HR molecular sieve column that balance is good, applied sample amount is 2mL, spend fromSub-water carries out wash-out with the flow velocity of 6mL/h. Collect with fraction collector, survey the total of every pipe by phenolsulfuric acid colorimetric methodSugar concentration, then merge identical component according to elution curve, collect 27 pipes to 43 pipes, desalt after dialysis, cold after 40 DEG C of reduced pressure concentrationsFreeze-drying is dry. As above operation is repeatedly carried out purifying freeze drying several times and is obtained fig polysaccharide sterling FCP1. Elution curve is shown in Fig. 2.
2.2.3 fig polysaccharide FCP1 purity checking
Preparation concentration is 10mg/mL fig polysaccharide sterling FCP solution, gets the upper processed good SephacrylS-of 2mL300HR post, elution curve is shown in Fig. 3. Visible eluting peak is single symmetrical peak, illustrates that FCP1 is the polysaccharide of character homogeneous.
2.2.4 product characterizes
2.2.4.1 get sterling FCP1 and be mixed with on a small quantity the solution of 0.1mg/mL, scanning spectra under ultraviolet, 260nm, 280nmDo not absorb, show in polysaccharide not containing impurity such as albumen.
2.2.4.2 get the fig polysaccharide sample F CP12mg of purifying, use KBr pressed disc method in 4000-400cm-1 intervalScan to obtain infrared spectrogram. See Fig. 4.
Visible in Fig. 4, at 3448cm-1The strong absworption peak at place is that the stretching vibration of the alcoholic extract hydroxyl group-OH in polysaccharide molecule is inhaledReceive peak, 2940cm-1The absworption peak of vicinity is the stretching vibration peak of C-H, at 1588cm-1The absworption peak at place is-bending of OH shakesMoving absworption peak. At 1378cm-1Place is C-H flexural vibrations absworption peaks. At 1106cm-1The strong absworption peak of vicinity proves alcohol hydroxylThe angle vibration absorption peak of base-OH. At 846cm-1There is the characteristic absorption peak of α-type glycosidic bond in place, illustrates that this fig polysaccharide isα-polysaccharide.
2.3 alpha-glucosidases suppress determination of activity
2.3.1 alpha-glucosidase suppresses activity test method
Taking PNPG as substrate, reaction system is: phosphate buffer (the pH value 6.8) 0.6mL of 0.1mol/L, and α-Glucuroide 0.1mL, after 37 DEG C of insulation 15min, adds 0.05mL testing sample solution and 0.2mL substrate PNPG, 37DEG C reaction 15min, add 1mol/LNa2CO3Solution 0.5mL cessation reaction, measures A value at 405nm place. By following formulaCalculate inhibiting rate, wherein sample contrasts only with buffer solution and sample solution, and control group is not containing sample, and blank is only with buffer solution. PositiveControl group replaces sample with positive control drug acarbose.
Inhibiting rate P (%)=1-
2.3.2 the mensuration of alpha-glucosaccharase enzyme activity
According to adopted reaction system, the phosphate buffer of 0.1mol/L (pH6.8) 0.6mL, alpha-glucosidase0.1mL, after 37 DEG C of insulation 15min, adds 200 μ LPNPG, 37 DEG C of reaction 15min, then add 1mol/LNa2CO3MoltenLiquid 0.5mL cessation reaction, measures absorption value at 405nm place.
Wherein enzyme activity unit definition: at 37 DEG C, under pH value 6.8 conditions, in 1min, hydrolysis PNPG discharges 1 μ moLPNPRequired enzyme amount (U).
2.3.3 the making of calibration curve
With phosphate buffer (pH6.8) preparation 1000 μ moL/L-1PNP solution, accurate absorption 2,1.75,1.5 respectively,1.25,1,0.75,0.5,0mL to colorimetric cylinder, and add respectively 1mol/LNa2CO3Solution 0.5mL, is settled to 5mL, mixedEven, the final concentration that makes PNP is 400,350,300,250,200,150,100,50 and 0 μ molL-1. Under 405nm, measure A value,Surveying 3 groups averages. Taking A value as ordinate, PNP concentration is abscissa, makes calibration curve.
2.3.4FCP alpha-glucosidase is suppressed to determining of type
Under certain inhibitor concentration condition, adding substrate, its concentration is 0,0.5,1.0,1.5,2.5,4mmol/L, measuresEnzyme activity. Change inhibitor concentration, be respectively 0,4.3482mg/mL, can obtain the enzyme under a series of different concentration of substrate conditionsVigor. Press Lineweave-Burk mapping and determine inhibition type.
3 results and discussion
3.1 calibration curve
Taking PNP concentration (X) as abscissa, A value is ordinate (Y), and drawing standard curve, obtains regression equation Y=0.0022X-0.0088 (r=0.9987), linear relationship is good, and linear concentration range is 0 ~ 400 μ molL-1。
The inhibitory action of 3.2 variable concentrations FCP to alpha-glucosidase
Experimental result shows, along with the rising of FCP concentration, suppresses activity and improves thereupon. And taking FCP concentration (mg/mL) asAbscissa, inhibiting rate (%) is ordinate, draws Trendline, and obtains regression equation Y=9.8746X+2.8243(r=0.9985),Calculating 503nhibiting concentration is Ic50=4.7775mg/mL。
3.6 suppress determining of type
3.6.1km the mensuration of value
The amount of the PNP that reaction rate (V) can produce with hydrolysis substrate in the unit interval represents, reaction rate in experimentLight absorption value by the PNP that generates in the unit interval in assaying reaction system is tried to achieve.
Regression equation is: Y=39.697X+29.921 (r=0.9580) can be obtained by formula, and km is 1.2599 × 10-3moL/L，Maximum reaction velocity is 33.42moL/Lmin.
3.6.2 suppressing type determines
Known according to figure, reaction rate Vmax diminishes with the increase of inhibitor concentration, and Michaelis constant Km keeps notBecome, therefore infer, FCP belongs to noncompetitive to the inhibitory action of alpha-glucosidase restrainer and suppresses.
Brief description of the drawings
Fig. 1 is the DEAE-52 column chromatography elution curve of the thick polysaccharide CFCP of fig;
Fig. 2 is the SephacrylS-300HR column chromatography elution curve of fig polysaccharide FCP;
Fig. 3 is the SephacrylS-300HR column chromatography elution curve of fig polysaccharide FCP1;
Fig. 4 is the infrared spectrogram of fig polysaccharide FCP1.
Detailed description of the invention
Below further describe concrete technical scheme of the present invention, so that those skilled in the art understands furtherThe present invention, and do not form the restriction to its right.
Embodiment 1, the preparation method of the thick polysaccharide of a kind of fig, its step is as follows: get that pulled figs is some to be shredded, use95% alcohol reflux degrease and pigment, after suction filtration, residue removes small molecular weight impurity with 70% alcohol reflux; Then get residue, as nothing flowerThe extraction raw material of fruit polysaccharide; Get residue, after water extracts under microwave condition as solvent, suction filtration, gets filtrate rotary evaporationBe concentrated into 100Ml, add and leave standstill 24h after 95% ethanol, get precipitation after centrifugal, precipitation by Sevage method except after deproteinized, freezing dryDry, obtain the thick polysaccharide of fig; When extraction, the solid-liquid ratio 1:20 of residue and water; Microwave power is 550W, and extraction time is13min, alcohol precipitation is extracting liquid volume by amount of alcohol 3.50 times.
Embodiment 2, the preparation method of the thick polysaccharide of a kind of fig, its step is as follows: get that pulled figs is some to be shredded, use95% alcohol reflux degrease and pigment, after suction filtration, residue removes small molecular weight impurity with 70% alcohol reflux; Then get residue, as nothing flowerThe extraction raw material of fruit polysaccharide; Get residue, after water extracts under microwave condition as solvent, suction filtration, gets filtrate rotary evaporationBe concentrated into 100Ml, add and leave standstill 24h after 95% ethanol, get precipitation after centrifugal, precipitation by Sevage method except after deproteinized, freezing dryDry, obtain the thick polysaccharide of fig; When extraction, the solid-liquid ratio 1:23 of residue and water; Microwave power is 550W, and extraction time is13min, alcohol precipitation is extracting liquid volume by amount of alcohol 3.60 times.
Embodiment 3, the preparation method of the thick polysaccharide of a kind of fig, its step is as follows: get that pulled figs is some to be shredded, use95% alcohol reflux degrease and pigment, after suction filtration, residue removes small molecular weight impurity with 70% alcohol reflux; Then get residue, as nothing flowerThe extraction raw material of fruit polysaccharide; Get residue, after water extracts under microwave condition as solvent, suction filtration, gets filtrate rotary evaporationBe concentrated into 100Ml, add and leave standstill 24h after 95% ethanol, get precipitation after centrifugal, precipitation by Sevage method except after deproteinized, freezing dryDry, obtain the thick polysaccharide of fig; When extraction, the solid-liquid ratio 1:22.45 of residue and water; Alcohol precipitation is extracting liquid volume by amount of alcohol3.55 doubly.
Embodiment 4, a kind of purification process of the thick polysaccharide of fig that method makes as described in embodiment 1 or 2, its step asUnder:
(1) ion exchange cellulose DEAE-52 is after distilled water is fully swelling, with the NaOH of 0.5M and the HCl of 0.5MAlternate treatment is used distilled water cyclic washing for several times again, until its pH to 6 fills post, adopts 2.0cm × 30cm normal pressure sandCore glass post, it is stand-by that the ion column installing carries out balance 24 hours with distilled water again;
(2) get the thick polysaccharide of fig and be mixed with the solution of mass concentration as 1% taking distilled water; Then fig polysaccharide is moltenThe DEAE-52 ion exchange column that on liquid, balance is good, loading volume is 5mL, uses 0-1MNaCl200mL with 50mL/hFlow velocity cross post, collect with the amount of every pipe 5mL with automatic fraction collector, with the every pipe of phenolsulfuric acid colorimetric method for determiningTotal sugar concentration is made elution curve, collects identical component (collecting 13 pipes to 32 pipes), in 40 DEG C of reduced pressure concentrations also according to elution curveFreeze drying obtains fig polysaccharide;
(3) SephacrylS-300HR softly washs for several times, reduces pressure after degassed through distilled water and fills post, adopt 1.6cm ×80cm post, with distilled water balance; The fig polysaccharide that step (2) is obtained is dissolved in deionized water, and compound concentration is 15mg/mLPolysaccharide solution, on SephacrylS-300HR molecular sieve column that balance is good, applied sample amount is 2mL, by deionized water with 6The flow velocity of mL/h carries out wash-out; Collect with fraction collector, survey the total sugar concentration of every pipe by phenolsulfuric acid colorimetric method, thenMerge identical component (collecting 27 pipes to 43 pipes) according to elution curve, desalt after dialysis, 40 DEG C of reduced pressure concentration postlyophilizations; AsUpper operation is repeatedly carried out the freeze drying of purifying several times and is obtained fig polysaccharide sterling.
Purification process purifying described in the thick polysaccharide of fig or embodiment 4 that described in embodiment 1 or 2 or 3, method makesThe fig polysaccharide sterling obtaining can be for the preparation of alpha-glucosidase restrainer.
Claims (1)
1. a preparation method for the thick polysaccharide of fig, is characterized in that, its step is as follows: get that pulled figs is some to be shredded, use95% alcohol reflux degrease and pigment, after suction filtration, residue removes small molecular weight impurity with 70% alcohol reflux; Then get residue, as nothingThe extraction raw material of flowers and fruits polysaccharide; Get residue, after water extracts under microwave condition as solvent, suction filtration, gets filtrate rotary evaporationBe concentrated into 100mL, add and leave standstill 24h after 95% ethanol, get precipitation after centrifugal, precipitation by Sevage method except after deproteinized, freezing dryDry, obtain the thick polysaccharide of fig; When extraction, the solid-liquid ratio 1:22.45 of residue and water; Microwave power is 550W, and extraction time is13min, alcohol precipitation is extracting liquid volume by amount of alcohol 3.55 times;
Its purification process step is as follows:
(1) ion exchange cellulose DEAE-52 is after distilled water is fully swelling, with the NaOH of 0.5M and alternately place of the HCl of 0.5MReason is used distilled water cyclic washing for several times again, until its pH to 6 fills post, adopts 2.0cm × 30cm normal pressure core glass column,It is stand-by that the ion column installing carries out balance 24 hours with distilled water again;
(2) get the thick polysaccharide of fig and be mixed with the solution of mass concentration as 1% taking distilled water; Then by fig polysaccharide solutionThe DEAE-52 ion exchange column that balance is good, loading volume is 5mL, the flow velocity mistake with 0-1MNaCl200mL with 50mL/hPost, collects with the amount of every pipe 5mL with automatic fraction collector, with the total sugar concentration of the every pipe of phenolsulfuric acid colorimetric method for determiningMake elution curve, collect identical component according to elution curve, obtain fig polysaccharide in 40 DEG C of reduced pressure concentrations freeze drying;
(3) SephacrylS-300HR softly washs for several times, reduces pressure after degassed through distilled water and fills post, adopts 1.6cm × 80cmPost, with distilled water balance; The fig polysaccharide that step (2) is obtained is dissolved in deionized water, the polysaccharide that compound concentration is 15mg/mLSolution, on SephacrylS-300HR molecular sieve column that balance is good, applied sample amount is 2mL, by deionized water with 6mL/h'sFlow velocity carries out wash-out; Collect with fraction collector, survey the total sugar concentration of every pipe by phenolsulfuric acid colorimetric method, then according to washingDe-curve merges identical component, desalts after dialysis, and 40 DEG C of reduced pressure concentration postlyophilizations; As above it is some that purifying is carried out in operation repeatedlyInferior freeze drying obtains fig polysaccharide sterling.
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| CN104031164A (en) * | 2014-06-26 | 2014-09-10 | 河南中烟工业有限责任公司 | Ficus carica polysaccharide, extraction and purification method and application of ficus carica polysaccharide as tobacco humectant |
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| CN102504041A (en) * | 2011-11-17 | 2012-06-20 | 云南金九地生物科技有限公司 | Separation and purification method for dendrobium polysaccharide |
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| CN101591398A (en) * | 2009-06-30 | 2009-12-02 | 华南理工大学 | A kind of method for quickly purifying of momordica polysaccharide crude product |
| CN102504041A (en) * | 2011-11-17 | 2012-06-20 | 云南金九地生物科技有限公司 | Separation and purification method for dendrobium polysaccharide |
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| 响应面分析法优选黄花菜多糖提取工艺;李盈蕾 等;《中国酿造》;20101231(第7期);第72-75页，第1.2.2节 * |
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