CN102936292A - Preparation method of lycium barbarum polysaccharide having high antioxidant activity - Google Patents
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Abstract
The invention relates to a preparation method of a lycium barbarum polysaccharide having high antioxidant activity, which comprises the following steps: in the preparation process, performing ultrafine grinding pretreatment on wolfberry fruit, performing ethanol pretreatment, extracting with water, performing vacuum concentration, performing ethanol precipitation, removing proteins, and performing ethanol fractional precipitation treatment to obtain the lycium barbarum polysaccharide having high antioxidant activity. According to the preparation method, the wolfberry fruit is used as the raw material, the ultrafine grinding technology is used to reduce the molecular weight of the polysaccharide; the preparation process of the lycium barbarum polysaccharide having high antioxidant activity is simple and convenient to operate; and the ethanol fractional precipitation method adopted by the preparation method has the advantages of high yield and low cost, thereby being hopeful to realize industrial application.
Description
Technical field
The invention belongs to matrimony vine deep process technology field, especially a kind of auxiliary method for preparing the active lycium barbarum polysaccharide of high anti-oxidation of superfine technique that adopts.
Background technology
Matrimony vine (Lycium chinense Mill) belongs to Solanaceae, is the rare traditional Chinese medicine of the traditional dietotherapeutic of China, and its flavor is sweet, property flat, the function that has invigorating the liver and kidney, benefiting essence-blood and make eye bright.The pharmacology of matrimony vine and health-care effect and biologically active substance polysaccharide wherein have much relations.Modern scientific research shows, lycium barbarum polysaccharide has the immunity of adjusting, removes free radical, suppresses tumour, delays senility, reducing blood-fat, hypoglycemic and fatigue-resisting function, has wide development prospect.
The structure of polysaccharide is the important factor that affects its biologic activity, and its physico-chemical property such as viscosity, solubleness etc. are also determining active polysaccharide to a certain extent in addition.Polysaccharide is water-soluble to be most important condition of its performance biologic activity, there are some researches show, by reducing molecular mass, introducing side chain or side chain is suitably modified, all can improve polysaccharide solubleness, thereby strengthen its activity.
The polysaccharide molecule quality is larger, molecular volume is larger, be unfavorable for that polysaccharide leap cell multiplex film obstacle enters performance biologic activity in the organism, but be not that the polysaccharide molecule quality is more low better yet, because molecular mass is excessively low, can't form and produce active paradigmatic structure, the people such as Alban have studied the relation that molecular mass and sulfation are condensed the polysaccharide anticoagulant active, think that anticoagulant active and relative molecular mass are the dumbbell shape curved line relation; The scope of best relative molecular mass that different polysaccharide produces biologic activity is different, the people such as Gao report, and the sulfation polysaccharide that condenses is (7～11) * 10 at relative molecular mass
3In, along with relative molecular mass raises, its anticoagulant active has the trend of enhancing, the really poly-polysaccharide Levans of certain that from bacterium, separates, and relative molecular mass reaches 2.1 * 10
5The time to suppress the activity of tumor growth the strongest.
Studies show that the molecular weight of polysaccharide anti-oxidant activity and polysaccharide, monose composition, glucuronic acid content, glycosidic link type, chain conformation equimolecular structure have substantial connection.And molecular size range is an important structure characteristic parameter that affects the anti-oxidant activity power, and within the specific limits, molecular weight is lower, and then anti-oxidant activity is stronger.For example, the people such as mountain, Zhou village adopt ultrasonic technology to extract bar shaped spot laver amylose, and its molecular weight reduces, the corresponding raising of polysaccharide oxidation-resistance; The Auricularia polysaccharide that Ceng Weicai etc. adopt microwave assisting method to extract, molecular weight is 2.77 * 10
4Da, experiment shows that its resistance of oxidation is more remarkable.
The development of modern processing and application become more meticulous the pre-treatment of polysaccharide raw material more, such as, the application of superfine grinding and nano-pulverization technology can destroy cellularstructure greatly, even breaks cell walls, the polysaccharide that is present in cell walls inside is come out, improve the extraction yield of polysaccharide.Application by processing technology has changed polysaccharide when extracting yield, also just change the constructional features such as the molecular composition in the polyoses extract, average weight, glycosidic link ratio, the degree of branching, and then changed processing characteristics and the biological activity thereof such as water-soluble, rheological properties, thermal characteristic, colloid property of polyoses extract.
Superfine technique is a new and high technology that develops rapidly in the world in recent years.Superfine grinding refers to utilize machinery or hydrokinetic method material particles to be crushed to the process of 1nm-100 μ m.Material owing to have surface effects, small-size effect, quantum effect and macro quanta tunnel effect, possesses some special physico-chemical properties that general particle does not have, such as good solubility, dispersiveness, adsorptivity etc. after superfine grinding.The people such as Li utilize high frequency vibrating dynamic formula micronizing to make the Rhizoma amorphophalli glucomannan of miniaturization, change has all occured in Rhizoma amorphophalli glucomannan crystalline structure and the swelling character of finding miniaturization, and the sample of ball milling has reduced the body weight of obesity mice significantly, and the content of tri-glyceride, glucose and high-density lipoprotein (HDL) also all has remarkable reduction in the blood, illustrates that the miniaturization Rhizoma amorphophalli glucomannan can be used for the exploitation of diet products.The bright grade of plum utilizes superfine communication technique that Pachymose and Poria powder are carried out the miniaturization processing, the variation of its physico-chemical property before and after the comparison process, identify through the infrared scan collection of illustrative plates, Pachymose infrared absorption pattern after the micronizing changes, the polysaccharide solubility rate increases, and this is conducive to improve the processing characteristics of Poria cocos in food.
Because the polysaccharide multiple good physicochemical property and the biologic activity that have, thereby can be used as thickening material, stablizer, functional factor, fat subsitutes, membrane-forming agent etc. and be widely used in food-processing.For molecular structure and the biological activity of polysaccharide remarkably influenced is arranged and produce various new and high technologies that polysaccharide adopts, and then have influence on the application characteristic of polysaccharide in food-processing.Therefore, be necessary to carry out in a deep going way processing technology for lycium barbarum polysaccharide structure and bioactive impact, develop the production technology of high antioxidant lycium barbarum polysaccharide, thereby instruct production and the Applications in food of lycium barbarum polysaccharide.
By retrieval, find two pieces of patent documentations related to the present invention.
1, a kind of lycium barbarum polysaccharide extracting method and lycium barbarum polysaccharide weight reduction product (CN1670036) mainly may further comprise the steps: wolfberry fruit ethanolic soln pre-treatment; Through pretreated wolfberry fruit distilled water extraction; Extracting solution is processed with polymeric amide, DEAE-cellulose (OH-) and SephadexG-25 gel chromatographic columns.The lycium barbarum polysaccharide weight reduction product is take lycium barbarum polysaccharide as functional component, is equipped with corresponding auxiliary material, makes according to routine techniques.The method not only can improve the purity of lycium barbarum polysaccharide, and can utilize this polysaccharide to make weight reduction product.In addition, because lycium barbarum polysaccharide weight reduction product provided by the invention is to be made by pure natural substance, thereby the patient has no side effect after taking, and fat-reducing effect is obvious.
2, a kind of enzymolysis process (CN101069562) for preparing the high-immunity regulation activity lycium barbarum polysaccharide take the matrimony vine schlempe as raw material, its technology characteristics is: with the first polysaccharide in the water extraction and alcohol precipitation method extraction matrimony vine schlempe; Adding concentration is 0.004～0.01% saccharifying enzyme liquid, in 30～50 ℃ of hydrolysis 15～40min; Hydrolyzed solution is boiled 10min, add again 95% ethanol precipitation (alcohol precipitation concentration is 80%), left standstill 10～18 hours; The centrifugal 15min of 3000r/min, taking precipitate, vacuum-drying namely gets brown high-immunity regulation activity enzymolysis lycium barbarum polysaccharide (weight-average molecular weight 1～30,000).The present invention is take the matrimony vine schlempe as raw material, improved resource utilization, compared with prior art, having utilized physiologically active and its molecular weight of lycium barbarum polysaccharide in close relations, changed its molecular weight by zymolysis technique, is the lycium barbarum polysaccharide of high-immunity regulation activity with the first polysaccharide enzymolysis of matrimony vine, this technique is simple, easy to operate, the lycium barbarum polysaccharide yield is high, and immunoregulatory activity is strong.
By contrast, there are the different of essence in patent application of the present invention from above-mentioned two pieces of patent publication us.
Summary of the invention
The object of the invention is to overcome the deficiencies in the prior art part, provide a kind of technique simple, easy to operate, prepared lycium barbarum polysaccharide yield is high, anti-oxidant activity is strong, adopts the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation of superfine grinding pre-treatment.
The objective of the invention is to be achieved through the following technical solutions:
The preparation method of the active lycium barbarum polysaccharide of a kind of high anti-oxidation, wolfberry fruit is through the superfine grinding pre-treatment in preparation process.
And described wolfberry fruit is after the superfine grinding pre-treatment, and the wolfberry fruit particle of 90-99% is less than 700nm.
And described superfine grinding pre-treatment concrete steps are: wolfberry fruit is through coarse reduction, and at 45 ℃ of lower oven drying at low temperature 3-5h, 5-7h is pulverized in the impact grinding of high energy nanometer.
And concrete steps are as follows:
⑴ superfine grinding pre-treatment: wolfberry fruit is through coarse reduction, oven drying at low temperature, and the impact grinding of high energy nanometer is pulverized, and makes wolfberry fruit particle more than 90% less than 700nm after the pulverizing;
⑵ ethanol pre-treatment: adding volume fraction to the matrimony vine of processing through superfine grinding is 80% aqueous ethanolic solution, and feed liquid ratio g:mL is 1:2-4, refluxing extraction, and abandoning supernatant, lower floor's insolubles repeats aforesaid operations, obtains the wolfberry fruit after the pre-treatment;
⑶ water extraction: water with through the wolfberry fruit of pre-treatment proportionally mL:g be after 4 ~ 6:1 mixes, after the water-bath lixiviate, filter, collect filtrate, insolubles repetition aforesaid operations 2 times, merging filtrate;
⑷ vacuum concentration: vacuum tightness＜0.1MPa is concentrated to soluble solid content greater than 10% with filtrate among the step ⑶ under 45 ~ 55 ℃, gets concentrated solution;
⑸ ethanol precipitation: slowly add ethanol in concentrated solution under agitation condition, make the final volume concentration of ethanol greater than 80%, leave standstill more than 6 hours, after the separation, collect solid matter, volatilization is removed ethanol and is got the matrimony vine Crude polysaccharides;
⑹ removal of protein: adopt polymeric amide chromatogram column technique or Sevag method deproteination matter, being concentrated to massfraction is more than 5%, and vacuum lyophilization gets lycium barbarum polysaccharide;
⑺ ethanol precipitation: it is 2.5% lycium barbarum polysaccharide solution that lycium barbarum polysaccharide is made into mass concentration, adds while stirring volumetric concentration and be the ethanol more than 95%, makes the ethanol volumetric concentration reach 60%, leave standstill more than 6 hours, separate, collect supernatant liquor, continuing to add concentration is 95%(v/v) above ethanol, reach 80%(v/v to alcohol concn), leave standstill more than 6 hours, separate collecting precipitation, ethanol is removed in volatilization, gets lycium barbarum polysaccharide after the lyophilize.
And wolfberry fruit is through coarse reduction among the ⑴ in the described step, and at 45 ℃ of lower oven drying at low temperature 4h, 6h is pulverized in the impact grinding of high energy nanometer.
And, in the described step among the ⑵ feed liquid ratio g:mL be 1:3, in 80 ℃ of water-bath refluxing extraction 0.5h, abandoning supernatant, the lower floor insolubles repeats last time and operates 4 times.
And the condition of ⑶ water-bath lixiviate is 95 ℃ of water-bath lixiviate 95min in the described step.
And it is centrifugal more than 10 minutes to be separated into 3400r/min in the described step among ⑸ or the ⑺, or filtration treatment.
And, in the described step among the ⑹ polymeric amide chromatogram column technique except the concrete steps of albumen be: it is 1%-2% that the matrimony vine Crude polysaccharides is dissolved to mass concentration, and it is added the polymeric amide chromatographic column, applied sample amount is the 15%-20% of column capacity, with water elution, collect elutriant, being concentrated to massfraction is more than 5%.
And, in the described step among the ⑹ Sevag method except the concrete steps of albumen be: get the matrimony vine Crude polysaccharides, add water and dissolve fully, add successively chloroform and propyl carbinol, polysaccharide soln: chloroform: the volume ratio of propyl carbinol is 25:5:1, violent jolting 20min standing demix, inclining supernatant liquor, discards lower floor's organic solution and middle denatured protein layer, the supernatant liquor repetitive operation for several times, until without till the denatured protein appearance, collect supernatant liquor, being concentrated in vacuo to massfraction is more than 5%.
Advantage of the present invention and positively effect are:
1, preparation method of the present invention adopts superfine technique to reduce polysaccharide molecular weight take matrimony vine as raw material, and the technique of preparation high antioxidant lycium barbarum polysaccharide is simple, easy to operate.
2, the ethanol precipitation method that adopts of this preparation method has the advantage of the high and low cost of yield, is expected to realize industrial applications.
3, the preparation-obtained lycium barbarum polysaccharide yield of preparation method of the present invention is high, and anti-oxidant activity is strong, and with the lycium barbarum polysaccharide comparison without the superfine grinding pre-treatment, the lycium barbarum polysaccharide that present method obtains is to the half-inhibition concentration (IC of DPPH free radical
50) reduce more than 60%, to the IC of ABTS free radical
50Value reduces more than 70%.
Embodiment
Below in conjunction with embodiment, the present invention is further described, and following embodiment is illustrative, is not determinate, can not limit protection scope of the present invention with following embodiment.
Employed reagent is the commercially available prod if no special instructions in the embodiment of the invention; Employed test method is ordinary method if no special instructions; Employed instrument is conventional instrument if no special instructions.
Embodiment 1:
The preparation method of the active lycium barbarum polysaccharide of a kind of high anti-oxidation, concrete steps are as follows:
⑴ superfine grinding pre-treatment: wolfberry fruit is through coarse reduction, and at 45 ℃ of lower oven drying at low temperature 3h, 5h is pulverized in the impact grinding of high energy nanometer, makes wolfberry fruit particle more than 90% less than 700nm after the pulverizing.
⑵ ethanol pre-treatment: take by weighing 200g through the matrimony vine of above-mentioned processing treatment, adding 80%(V/V) aqueous ethanolic solution, feed liquid ratio g:mL is 1:2(g matrimony vine ultrafine powder: the v ethanolic soln), in 80 ℃ of water-bath refluxing extraction 0.5h, abandoning supernatant, lower floor's insolubles repeats last time and operates 4 times, obtains the wolfberry fruit of pre-treatment;
⑶ water extraction: through the wolfberry fruit of Ethanol Treatment, ethanol is removed in volatilization, adds distilled water, material/liquor ratio is (4 ~ 6): 1(mL distilled water: the g wolfberry fruit), in 95 ℃ of water-bath lixiviate 95min, filter, collect filtrate, insolubles repeats aforesaid operations 2 times, merges to collect filtrate;
⑷ vacuum concentration: at vacuum tightness＜0.1MPa, 45 ~ 55 ℃ lower concentrated, and the soluble solid of extracting solution is reached more than 10%, gets concentrated solution;
⑸ ethanol precipitation: slowly add ethanol under agitation condition in concentrated solution, make alcohol concn leave standstill 12h greater than 80%, 4 ℃, filter, collect solid matter, ethanol is removed in volatilization, namely gets the matrimony vine Crude polysaccharides;
⑹ removal of protein: with the matrimony vine Crude polysaccharides that obtains, adding distil water dissolves fully, get matrimony vine Crude polysaccharides solution, (matrimony vine Crude polysaccharides solution: chloroform: the volume ratio of propyl carbinol is 25:5:1 to add Sevag reagent,), violent jolting 20min leaves standstill, centrifugal 15min under rotating speed 3400r/min, inclining supernatant liquor, discards lower floor's organic solution and middle protein denaturation layer, the supernatant liquor repetitive operation for several times, until without till the denatured protein appearance, collect supernatant liquor, being concentrated to massfraction is more than 5%, through lyophilize, get lycium barbarum polysaccharide;
⑺ ethanol precipitation: lycium barbarum polysaccharide is mixed with 2.5% the aqueous solution, stir and to add 95%(V/V down) ethanol to alcohol concn reaches 60%(v/v), 4 ℃ leave standstill 6h, filtration, collect supernatant liquor, continue to add ethanol to its concentration in the supernatant liquor and reach 80%(v/v), 4 ℃ leave standstill 6h, filter, collecting precipitation, ethanol is removed in volatilization, and lyophilize gets the active lycium barbarum polysaccharide of high anti-oxidation.
By detecting:
The yield of the active lycium barbarum polysaccharide of preparation-obtained high anti-oxidation is the 1.5-1.8g/100g raw material; Its number average molecular-weight average is 540,000, and the alditol content in the molecule is 41.2%, and neutral sugar content is 28.8%, and protein content is 1.6%, is comprised of Fucose, pectinose, wood sugar, glucose, seminose and semi-lactosi.The lycium barbarum polysaccharide that obtains with aforesaid method is to the half-inhibition concentration (IC of DPPH free radical
50) reduce and (to be reduced to below the 3.5mg/mL by 9.0mg/mL) more than 60%, to the IC of ABTS free radical
50The value reduction (is reduced to below the 0.9mg/mL by 3.2mg/mL) more than 70%.
Embodiment 2:
The preparation method of the active lycium barbarum polysaccharide of a kind of high anti-oxidation, step is:
⑴ superfine grinding pre-treatment: wolfberry fruit is through coarse reduction, and at 45 ℃ of lower oven drying at low temperature 4h, 6h is pulverized in the impact grinding of high energy nanometer.Make the wolfberry fruit particle of 90-99% less than between the 700nm after the pulverizing.
⑵ aqueous ethanolic solution ethanol pre-treatment: through the matrimony vine adding 80%(V/V of superfine grinding processing), the feed liquid ratio is 1:3g/v, in 80 ℃ of water-bath refluxing extraction 0.5h, abandoning supernatant, lower floor's insolubles repeats last time and operates 4 times, obtains the wolfberry fruit of pre-treatment;
⑶ water extraction: water is (4 ~ 6) with the ratios of the many sons of matrimony vine of process pre-treatment: 1(mL:g), in 95 ℃ of water-bath lixiviate 95min, filter, collect filtrate, insolubles repetition aforesaid operations 2 times merges and collects filtrate;
⑷ vacuum concentration: vacuum tightness＜0.1MPa is concentrated to extracting solution more than 10% under 45 ~ 55 ℃;
⑸ ethanol precipitation: slowly add ethanol under agitation condition in concentrated solution, make alcohol concn leave standstill 12h greater than 80%, 4 ℃, 3400r/min is centrifugal, collects solid matter, and ethanol is removed in volatilization, gets the matrimony vine Crude polysaccharides.
⑹ removal of protein: get a certain amount of Crude polysaccharides, adding distil water dissolves fully, adding Sevag reagent (solution: chloroform: propyl carbinol=25:5:1), violent jolting 20min leaves standstill, centrifugal 15min under rotating speed 3400r/min, collect supernatant liquor, supernatant liquor repetitive operation several is until without till the denatured protein appearance.Collect supernatant liquor, the vacuum concentration lyophilize gets lycium barbarum polysaccharide;
⑺ ethanol precipitation: accurately take by weighing the refining lycium barbarum polysaccharide of 1g, add the 40mL dissolved in distilled water, add 95%(V/V under stirring) to transfer pure volume fraction be 60% to ethanol, 4 ℃ leave standstill 6h, and the centrifugal 15min of 3400r/min collects supernatant liquor, continue to add ethanol in the supernatant liquor, make its concentration reach 80% (v/v), 4 ℃ leave standstill 6h, centrifugal, collecting precipitation, ethanol is removed in volatilization, and lyophilize gets the active lycium barbarum polysaccharide of high anti-oxidation.
By detecting:
The yield of the active lycium barbarum polysaccharide of preparation-obtained high anti-oxidation is the 1.5-1.8g/100g raw material; Its number average molecular-weight average is 540,000, and the alditol content in the molecule is 41.2%, and neutral sugar content is 28.8%, and protein content is 1.6%, is comprised of Fucose, pectinose, wood sugar, glucose, seminose and semi-lactosi.The lycium barbarum polysaccharide that obtains with aforesaid method is to the half-inhibition concentration (IC of DPPH free radical
50) reduce and (to be reduced to below the 3.5mg/mL by 9.0mg/mL) more than 60%, to the IC of ABTS free radical
50The value reduction (is reduced to below the 0.9mg/mL by 3.2mg/mL) more than 70%.
Embodiment 3:
The preparation method of the active lycium barbarum polysaccharide of a kind of high anti-oxidation, concrete steps are as follows:
⑴ superfine grinding pre-treatment: wolfberry fruit is through coarse reduction, and at 45 ℃ of lower oven drying at low temperature 7h, 6h is pulverized in the impact grinding of high energy nanometer, makes wolfberry fruit particle more than 90% less than 700nm after the pulverizing.
⑵ ethanol pre-treatment: take by weighing 200g through the matrimony vine of above-mentioned processing treatment, adding 80%(V/V) aqueous ethanolic solution, feed liquid ratio g:mL is 1:4(g matrimony vine ultrafine powder: the v ethanolic soln), in 80 ℃ of water-bath refluxing extraction 0.5h, abandoning supernatant, lower floor's insolubles repeats last time and operates 4 times, obtains the wolfberry fruit of pre-treatment;
⑶ water extraction: through the wolfberry fruit of Ethanol Treatment, ethanol is removed in volatilization, adds distilled water, material/liquor ratio is (4 ~ 6): 1(mL distilled water: the g wolfberry fruit), in 95 ℃ of water-bath lixiviate 95min, filter, collect filtrate, insolubles repeats aforesaid operations 2 times, merges to collect filtrate;
⑷ vacuum concentration: at vacuum tightness＜0.1MPa, 45 ~ 55 ℃ lower concentrated, and the soluble solid of extracting solution is reached more than 10%, gets concentrated solution;
⑸ ethanol precipitation: slowly add ethanol under agitation condition in concentrated solution, make alcohol concn leave standstill 12h greater than 80%, 4 ℃, filter, collect solid matter, ethanol is removed in volatilization, namely gets the matrimony vine Crude polysaccharides;
⑹ removal of protein: with the matrimony vine Crude polysaccharides that obtains, adding distil water dissolves fully, to mass concentration be 1%-2%, get matrimony vine Crude polysaccharides solution, it is added the polymeric amide chromatographic column, applied sample amount is the 15%-20% of column capacity, with water elution, elution flow rate is column volume of 4h-5h, detects with the phenol sulfuric acid method, begins to collect elutriant when having sugar to flow out with moving phase, to no longer include sugar during by wash-out till, it is more than 5% that collected elutriant is concentrated in vacuo to massfraction, through lyophilize, gets lycium barbarum polysaccharide;
⑺ ethanol precipitation: lycium barbarum polysaccharide is mixed with 2.5% the aqueous solution, stir and to add 95%(V/V down) ethanol to alcohol concn reaches 60%(v/v), 4 ℃ leave standstill 6h, filtration, collect supernatant liquor, continue to add ethanol to its concentration in the supernatant liquor and reach 80%(v/v), 4 ℃ leave standstill 6h, filter, collecting precipitation, ethanol is removed in volatilization, and lyophilize gets the active lycium barbarum polysaccharide of high anti-oxidation.
By detecting:
The yield of the active lycium barbarum polysaccharide of preparation-obtained high anti-oxidation is the 1.5-1.8g/100g raw material; Its number average molecular-weight average is 540,000, and the alditol content in the molecule is 41.2%, and neutral sugar content is 28.8%, and protein content is 1.6%, is comprised of Fucose, pectinose, wood sugar, glucose, seminose and semi-lactosi.The lycium barbarum polysaccharide that obtains with aforesaid method is to the half-inhibition concentration (IC of DPPH free radical
50) reduce and (to be reduced to below the 3.5mg/mL by 9.0mg/mL) more than 60%, to the IC of ABTS free radical
50The value reduction (is reduced to below the 0.9mg/mL by 3.2mg/mL) more than 70%.
Claims (10)
1. the preparation method of the active lycium barbarum polysaccharide of a high anti-oxidation, it is characterized in that: wolfberry fruit is through the superfine grinding pre-treatment in preparation process.
2. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 1, it is characterized in that: described wolfberry fruit is after the superfine grinding pre-treatment, and the wolfberry fruit particle of 90-99% is less than 700nm.
3. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 1, it is characterized in that: described superfine grinding pre-treatment concrete steps are: wolfberry fruit is through coarse reduction, and at 45 ℃ of lower oven drying at low temperature 3-5h, 5-7h is pulverized in the impact grinding of high energy nanometer.
4. according to claim 1 to the preparation method of the active lycium barbarum polysaccharide of 3 each described high anti-oxidations, it is characterized in that: concrete steps are as follows:
⑴ superfine grinding pre-treatment: wolfberry fruit is through coarse reduction, oven drying at low temperature, and the impact grinding of high energy nanometer is pulverized, and makes the wolfberry fruit particle of 90-99% less than 700nm after the pulverizing;
⑵ ethanol pre-treatment: adding volume fraction to the matrimony vine of processing through superfine grinding is the aqueous ethanolic solution of 70-90%, and feed liquid ratio g:mL is 1:2-4, refluxing extraction, and abandoning supernatant, lower floor's insolubles repeats aforesaid operations, obtains the wolfberry fruit after the pre-treatment;
⑶ water extraction: water with through the wolfberry fruit of pre-treatment proportionally ml:g be after 4 ~ 6:1 mixes, after the water-bath lixiviate, filter, collect filtrate, insolubles repetition aforesaid operations 2 times, merging filtrate;
⑷ vacuum concentration: vacuum tightness＜0.1MPa is concentrated to soluble solid content greater than 10% with filtrate among the step ⑶ under 45 ~ 55 ℃, gets concentrated solution;
⑸ ethanol precipitation: slowly add ethanol in concentrated solution under agitation condition, make the final volume concentration of ethanol greater than 80%, leave standstill more than 6 hours, after the separation, collect solid matter, volatilization is removed ethanol and is got the matrimony vine Crude polysaccharides;
⑹ removal of protein: adopt polymeric amide chromatogram column technique or Sevag method deproteination matter, being concentrated to massfraction is more than 5%, and vacuum lyophilization gets lycium barbarum polysaccharide;
⑺ ethanol precipitation: it is 2.5% lycium barbarum polysaccharide solution that lycium barbarum polysaccharide is made into mass concentration, adds while stirring volumetric concentration and be the ethanol more than 95%, makes the ethanol volumetric concentration reach 60%, leave standstill more than 6 hours, separate, collect supernatant liquor, continue to add the ethanol of volumetric concentration more than 95%, reach volumetric concentration 80% to alcohol concn, leave standstill more than 6 hours, separate collecting precipitation, ethanol is removed in volatilization, gets lycium barbarum polysaccharide after the lyophilize.
5. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 4, it is characterized in that: wolfberry fruit is through coarse reduction among the ⑴ in the described step, and at 45 ℃ of lower oven drying at low temperature 4h, 6h is pulverized in the impact grinding of high energy nanometer.
6. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 4 is characterized in that: in the described step among the ⑵ feed liquid ratio g:mL be 1:3, in 80 ℃ of water-bath refluxing extraction 0.5h, abandoning supernatant, lower floor's insolubles repeats last time and operates 4 times.
7. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 4, it is characterized in that: the condition of ⑶ water-bath lixiviate is 90-100 ℃ of water-bath lixiviate 90-100min in the described step.
8. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 4 is characterized in that: be separated into the centrifugal 10-50min of 3400r/min or filtration treatment in the described step among ⑸ or the ⑺.
9. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 4, it is characterized in that: in the described step among the ⑹ polymeric amide chromatogram column technique except the concrete steps of albumen be: it is 1%-2% that the matrimony vine Crude polysaccharides is dissolved to mass concentration, it is added the polymeric amide chromatographic column, applied sample amount is the 15%-20% of column capacity, with water elution, collect elutriant, being concentrated to massfraction is 5-20%.
10. the preparation method of the active lycium barbarum polysaccharide of high anti-oxidation according to claim 4, it is characterized in that: in the described step among the ⑹ Sevag method except the concrete steps of albumen be: get the matrimony vine Crude polysaccharides, adding water dissolves fully, add successively chloroform and propyl carbinol, polysaccharide soln: chloroform: the volume ratio of propyl carbinol is 25:5:1, violent jolting 20min standing demix, incline and supernatant liquor, discard lower floor's organic solution and middle denatured protein layer, the supernatant liquor repetitive operation for several times, until without till the denatured protein appearance, collect supernatant liquor, being concentrated in vacuo to massfraction is 5-20%.
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Cited By (11)
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| CN103980377A (en) * | 2014-05-20 | 2014-08-13 | 河南中烟工业有限责任公司 | Lycium barbarum polysaccharide, extracting and purifying method and application thereof as tobacco humectant |
| CN104126867A (en) * | 2014-07-18 | 2014-11-05 | 云南中烟工业有限责任公司 | Method for extracting bolete polysaccharide and application of bolete polysaccharide in tobacco products |
| CN104193846A (en) * | 2014-09-17 | 2014-12-10 | 上海交通大学 | Method for extracting polysaccharide from lycium barbarum and application of polysaccharide |
| CN104530252A (en) * | 2014-12-25 | 2015-04-22 | 上海交通大学 | Purifying process of wolfberry polysaccharide |
| CN104623310A (en) * | 2015-01-09 | 2015-05-20 | 郑州师范学院 | Method for processing fresh dendrobium devonianum stem powder |
| CN105218694A (en) * | 2015-10-28 | 2016-01-06 | 青海尤尼克生物技术有限公司 | A kind of Lycium barbarum polysaccharide extract and preparation method thereof |
| CN105831637A (en) * | 2016-03-28 | 2016-08-10 | 重庆理工大学 | Anti-radiation mulberry, wolfberry and jujube composite, preparation method and application thereof |
| CN106387902A (en) * | 2016-09-20 | 2017-02-15 | 王晓临 | Health care food capable of improving digestive tract disease symptoms |
| CN106496348A (en) * | 2016-11-29 | 2017-03-15 | 南京中医药大学 | There are the preparation method and applications of the Fructus Lycii refined polysaccharide of antioxidant activity |
| CN107163162A (en) * | 2017-07-06 | 2017-09-15 | 西北大学 | A kind of refined method of LBP-X |
| CN110106221A (en) * | 2019-05-28 | 2019-08-09 | 山东圣洲海洋生物科技股份有限公司 | Sea cucumber oligopeptides and the preparation method and application thereof |
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| CN103980377A (en) * | 2014-05-20 | 2014-08-13 | 河南中烟工业有限责任公司 | Lycium barbarum polysaccharide, extracting and purifying method and application thereof as tobacco humectant |
| CN104126867A (en) * | 2014-07-18 | 2014-11-05 | 云南中烟工业有限责任公司 | Method for extracting bolete polysaccharide and application of bolete polysaccharide in tobacco products |
| CN104126867B (en) * | 2014-07-18 | 2015-10-21 | 云南中烟工业有限责任公司 | The extracting method of speciosus polysaccharide and the purposes in tobacco product thereof |
| CN104193846A (en) * | 2014-09-17 | 2014-12-10 | 上海交通大学 | Method for extracting polysaccharide from lycium barbarum and application of polysaccharide |
| CN104530252A (en) * | 2014-12-25 | 2015-04-22 | 上海交通大学 | Purifying process of wolfberry polysaccharide |
| CN104623310A (en) * | 2015-01-09 | 2015-05-20 | 郑州师范学院 | Method for processing fresh dendrobium devonianum stem powder |
| CN105218694A (en) * | 2015-10-28 | 2016-01-06 | 青海尤尼克生物技术有限公司 | A kind of Lycium barbarum polysaccharide extract and preparation method thereof |
| CN105831637A (en) * | 2016-03-28 | 2016-08-10 | 重庆理工大学 | Anti-radiation mulberry, wolfberry and jujube composite, preparation method and application thereof |
| CN106387902A (en) * | 2016-09-20 | 2017-02-15 | 王晓临 | Health care food capable of improving digestive tract disease symptoms |
| CN106496348A (en) * | 2016-11-29 | 2017-03-15 | 南京中医药大学 | There are the preparation method and applications of the Fructus Lycii refined polysaccharide of antioxidant activity |
| CN107163162A (en) * | 2017-07-06 | 2017-09-15 | 西北大学 | A kind of refined method of LBP-X |
| CN110106221A (en) * | 2019-05-28 | 2019-08-09 | 山东圣洲海洋生物科技股份有限公司 | Sea cucumber oligopeptides and the preparation method and application thereof |
| CN110106221B (en) * | 2019-05-28 | 2022-10-14 | 山东圣洲海洋生物科技股份有限公司 | Sea cucumber oligopeptide and preparation method and application thereof |
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