CN104031165A - Grifola frondosa polysaccharide and extraction and purification method thereof, and application of Grifola frondosa polysaccharide as tobacco humectant - Google Patents
Grifola frondosa polysaccharide and extraction and purification method thereof, and application of Grifola frondosa polysaccharide as tobacco humectant Download PDFInfo
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- CN104031165A CN104031165A CN201410292982.6A CN201410292982A CN104031165A CN 104031165 A CN104031165 A CN 104031165A CN 201410292982 A CN201410292982 A CN 201410292982A CN 104031165 A CN104031165 A CN 104031165A
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- grifolan
- grifola frondosa
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Abstract
The invention relates to a Grifola frondosa polysaccharide and an extraction and purification method thereof. The method comprises the following steps: carrying out Soxhlet extraction on pulverized dry Grifola frondosa with aether, volatilizing the residue, extracting with boiling water, concentrating the boiling water extracting solution, centrifugating, taking the supernate, carrying out alcohol precipitation, standing, centrifugating, collecting the precipitate, washing, dissolving in distilled water, adding 0.1-0.2% papain in a 50+/-5 DEG C water bath, standing for 2-4 hours, removing the protein by a Sevage process, decolorizing with activated carbon, concentrating under reduced pressure, cooling, carrying out running water dialysis, carrying out alcohol precipitation, standing, and drying the precipitate at 60-70 DEG C to obtain the crude polysaccharide. The test proves that the Grifola frondosa polysaccharide has favorable physical and sensory humidifying actions on cigarettes, can lower the irritation and impure gas of the cigarettes, can enhance the smoothness and comfort of the cigarette smoke, and can be used as an excellent-performance tobacco humectant in cigarettes.
Description
Technical field
The invention belongs to humectant technical field, be specifically related to a kind of grifolan, extracting and purifying method and the application as tobacco humectant thereof.
Background technology
Current, in China's cigarette industry, the main polyhydroxy substances such as glycerine, propylene glycol, sorbyl alcohol that adopt are as humectant, use the main purpose of this type of humectant to be to maintain the water ratio of pipe tobacco in the course of processing, improve the resistance to processibility of pipe tobacco, but above-mentioned humectant to finished cigarettes water ratio maintain and suck comfort level to improve effect unsatisfactory.
Polysaccharide is to be polymerized by a large amount of monose, and the hydroxyl that they are a large amount of and water molecules form hydrogen bond, can make it have stronger water-holding power.At present, at home and abroad there is no the relevant report that grifolan is applied in cigarette as tobacco humectant.
Summary of the invention
The object of the invention is to provide a kind of grifolan, extracting and purifying method and the application as tobacco humectant thereof.
For achieving the above object, the present invention adopts following technical scheme:
A kind of extracting and purifying method of grifolan, comprise the steps: the dry Grifola frondosa ether Soxhlet of pulverizing to extract after (generally adding ether by solid-liquid ratio 1g:5-8ml), residue volatilizes and extracts with boiling water, boiling water extraction liquid is concentrated, centrifugal, getting supernatant liquor alcohol precipitation leaves standstill, recentrifuge collecting precipitation thing, after throw out washing, dissolve with distilled water, add again the papoid of weight of precipitate 1-2 ‰ to place 2-4h in 50 ± 5 DEG C of water-baths, adopt Sevage method deproteination, concentrating under reduced pressure after activated carbon decolorizing, after cooling, carry out flowing water dialysis in order to remove small molecular weight impurity, after dialysis finishes, alcohol precipitation leaves standstill again, be deposited in 60-70 DEG C and be dried to obtain brown Crude polysaccharides.
Further preferably, by Crude polysaccharides chromatographic separation purifying twice, obtain refining polysaccharide; Twice separation and purification elutriant used is respectively 0.5 mol/L NaCl solution and 0.05 mol/L NH
4hCO
3solution.
Utilize said extracted purification process to prepare the grifolan of gained.
Utilize said extracted purification process to prepare the application of gained grifolan as tobacco humectant.
Grifola frondosa is food, medicine dual-purpose gill fungus bacterium, and between the autumn in summer, Chang Yesheng is around chestnut.Sporophore endoplasm, the short coralliform branch that is of handle, is overlapped into clump, and its outward appearance is graceful, stacked like chrysanthemum; Its smell, delicate fragrance overflow, refreshing; Its meat is tender and crisp tasty and refreshing, and hundred eat and do not mind; Its nutrition has good health-care effect and very high pharmaceutical use.As a kind of high-class healthy food, be in fashion the markets such as Japan, Singapore in recent years.The present invention has obtained highly purified refining grifolan by extracting purifying, and carried out physics humid keeping performance and tested and inherent sensory quality assessment, find that it has good physics and sense organ humectation effect to cigarette, and can reduce pungency and the assorted gas of cigarette, improve mellow and full sense and the comfortableness of cigarette smoke, the tobacco humectant that can be used as a kind of excellent property is applied in cigarette, and has determined its Optimum in cigarette.
Compared to the prior art, beneficial effect of the present invention: 1) in the leaching process of grifolan, utilize alcohol precipitation, protease hydrolysis, Sevage except albumen and dialyse and remove the method for small molecular phase combination, effectively reduce the impurity in grifolan, for further separation and purification lays the foundation.2) adopt gel chromatography column chromatography for separation purifying twice, obtain highly purified refining grifolan.Whole extraction purge process is easy and simple to handle, and raw material and the reagent of use are cheap and easy to get.3), through evidence, the humectation successful of grifolan of the present invention to tobacco product, therefore has significant industrial application value and wide popularizing application prospect.
Embodiment
The present invention is described further by the following examples, but protection scope of the present invention is not limited to this.
test main agents and instrument used:
Ether, ethanol, acetone (Chinese Suzhou chemical reagent company limited, analytical pure), DEAE-Celluose and Sephadex G200 gel (sigma company of the U.S.), experimental water is distilled water.
BS200SWE1 type electronic balance (sensibility reciprocal: 0.0001g, Beijing match Doris balance company limited); R-215 type Rotary Evaporators (BUCHI company of Switzerland); V-700 vacuum pump (BUCHI company of Switzerland); DLSB-20/60 DEG C of subcooling recycle pump (Shanghai Mei Qiang plant and instrument company limited); KDM type regulating temp. electrothermal cover (Shandong Hua Lu electric heating Instrument Ltd.); 85-2 type constant temperature blender with magnetic force (Shanghai Si Le instrument plant), ZK-82B type electric vacunm drying case (Shanghai Shen Guang instrument company limited), CHRIST Freeze Drying Equipment (BUCHI company of Switzerland), TSKGEL G2000 PW efficient gel chromatographic column (Sigma company of the U.S.), Agilent LC1100 type high performance liquid phase instrument and differential refraction detector (Agilent company of the U.S.).
embodiment 1
An extracting and purifying method for grifolan, comprises the steps: the dry Grifola frondosa of the 300g pulverizing 1800ml ether Soxhlet to extract 2h in order to remove degrease, and residue volatilizes.Residue after volatilizing is used 750ml boiling water extraction 3 times again, merges the boiling water extraction liquid of 3 gained, concentrated, centrifugal, gets supernatant liquor, carries out alcohol precipitation, hold over night (12h), recentrifuge collecting precipitation thing with the dehydrated alcohol of 3 times of volumes of supernatant liquor.After throw out respectively washs 3 times with 300ml ethanol, 300ml acetone successively, dissolve with distilled water, then add the papoid of weight of precipitate 1 ‰ to place 2h in 50 DEG C of water-baths, adopt Sevage method deproteination.Add 8g gac to be placed in 70 DEG C of water-baths and heat 3h, remove by filter gac, concentrating under reduced pressure, after naturally cooling, pack flowing water in dialysis tubing into and dialyse 48h in order to remove small molecular weight impurity, after dialysis finishes, again add the dehydrated alcohol of the rear 3 times of amounts of polysaccharide soln volume of dialysis to carry out alcohol precipitation, hold over night (12h), be deposited in 70 DEG C and be dried to obtain 14.4g Crude polysaccharides, yield is 4.8%.
Get after the dissolving of 1.5g Crude polysaccharides distilled water, by DEAE-Celluose gel chromatography column chromatography for separation (5.0 × 50cm), taking 0.5mol/L NaCl solution as moving phase (being elutriant), flow velocity is 2.5ml/min, adopt phenol sulfuric acid method to develop the color to chromatographic solution, collect main chromatographic peak, freeze-drying.Then by freeze-drying gained solid 0.05mol/L NH
4hCO
3solution dissolves, by Sephadex G200 gel chromatographic columns chromatographic separation (5.0 × 50cm) again, with 0.05mol/L NH
4hCO
3solution is moving phase (being elutriant), and flow velocity is 2.0ml/min, adopts phenol sulfuric acid method to develop the color to chromatographic solution, collects main chromatographic peak, and freeze-drying obtains the refining grifolan solid of 365mg, yield 24.3%, and LT1 sugar content is 96% after testing.
embodiment 2
An extracting and purifying method for grifolan, comprises the steps: the dry Grifola frondosa of the 300g pulverizing 2000ml ether Soxhlet to extract 4h in order to remove degrease, and residue volatilizes.Residue after volatilizing is used 750ml boiling water extraction 3 times again, merges the boiling water extraction liquid of 3 gained, concentrated, centrifugal, gets supernatant liquor, carries out alcohol precipitation with the dehydrated alcohol of 3 times of volumes of supernatant liquor, leaves standstill 12h, recentrifuge collecting precipitation thing.After throw out respectively washs 3 times with 300ml ethanol, 300ml acetone successively, dissolve with distilled water, then add the papoid of weight of precipitate 2 ‰ to place 4h in 50 DEG C of water-baths, adopt Sevage method deproteination.Add 8g gac to be placed in 70 DEG C of water-baths and heat 4h, remove by filter gac, concentrating under reduced pressure, after naturally cooling, pack flowing water in dialysis tubing into and dialyse 48h in order to remove small molecular weight impurity, after dialysis finishes, again add the dehydrated alcohol of the rear 3 times of amounts of polysaccharide soln volume of dialysis to carry out alcohol precipitation, hold over night, be deposited in 70 DEG C and be dried to obtain 15.6g Crude polysaccharides, yield is 5.2%.
Get after the dissolving of 1.5g Crude polysaccharides distilled water, by DEAE-Celluose gel chromatography column chromatography for separation (5.0 × 50cm), taking 0.5mol/L NaCl solution as moving phase (being elutriant), flow velocity is 2.5ml/min, adopt phenol sulfuric acid method to develop the color to chromatographic solution, collect main chromatographic peak, freeze-drying.Then by freeze-drying gained solid 0.05mol/L NH
4hCO
3solution dissolves, by Sephadex G200 gel chromatographic columns chromatographic separation (5.0 × 50cm) again, with 0.05mol/L NH
4hCO
3solution is moving phase (being elutriant), and flow velocity is 2.0ml/min, adopts phenol sulfuric acid method to develop the color to chromatographic solution, collects main chromatographic peak, and freeze-drying obtains the refining grifolan solid of 349mg, yield 23.3%, and LT1 sugar content is 96.2% after testing.
application test: embodiment 1 prepares the applied research of the refining grifolan of gained as tobacco humectant
1, physics humid keeping performance test
For investigating the physics humid keeping performance of grifolan to cigarette, taking the blank cigarette shreds of Bright Yellow (great Jin circle) as carrier, same grifolan glycerine, propylene glycol are carried out respectively together with sorbyl alcohol to physics humid keeping performance contrast test research, prepare respectively massfraction and be 5% the above-claimed cpd aqueous solution.The blank cigarette shreds first 1000g being mixed is placed in 22 ± 1 DEG C of temperature, relative humidity 60 ± 2% environment balance 48h.Then equivalent takes five parts of pipe tobaccos, and every part of 100g evenly applies the 4g above-claimed cpd aqueous solution respectively in pipe tobacco, and contrasting blank sample is that pipe tobacco evenly applies 4g water.Then five parts of pipe tobaccos are placed in respectively and under 22 ± 1 DEG C, the environment of relative humidity 40 ± 2%, investigate it and separate wet process, and the variation of instant water ratio is analyzed, investigate its physics humectation effect (adding the moisture content of drying base in pipe tobacco 72h after different samples changes), the results are shown in Table 1.
The physics humectation effect comparison of table 1, grifolan, glycerine, propylene glycol and sorbyl alcohol
The test-results of table 1 shows: add the pipe tobacco of grifolan, glycerine, four kinds of compounds of propylene glycol and sorbyl alcohol compared with blank pipe tobacco, moisture loss speed is slower, this is because the contained hydrophilic radical hydroxyl of polyol can form hydrogen bond with water and water be fettered, delay scattering and disappearing of moisture, improved the physics humid keeping performance of pipe tobacco; Wherein the physics humid keeping performance of grifolan is best, is secondly propylene glycol and glycerine, is finally sorbyl alcohol.
2, sense organ humid keeping performance test
Make solvent with 20v% aqueous ethanolic solution, it is 1% solution that grifolan is made into massfraction.Get respectively the above-mentioned solution containing grifolan of 0.100g, 0.500g, 1.000g, 5.000g, 10.000g and 20.000g, then evenly spray is added in five parts of blank cigarette shreds of not spiced 100g Bright Yellow respectively, roll, respectively pick out 100 same weight cigarette, the climatic chamber balance 48h that is placed in 22 DEG C ± 1 DEG C of temperature, humidity 60% ± 2%, smokes panel test.Control sample is the blank cigarette of Bright Yellow, and control sample is balance 48h under uniform temp, humidity condition equally.Smoking result is in table 2.
The grifolan smoking result of table 2 Different adding amount
Smoking result by table 2 can find out, grifolan shows as and makes flue gas obviously become mellow and full, soft the Main Function of cigarette, and comfortable taste promotes, and assorted gas and pungency reduce.When grifolan consumption hour, it is not clearly to the effect of cigarette; In the time that its consumption exceedes 1 ‰, there is the phenomenon that spine sense increase, concentration reduce; In the time that its consumption is 1 ‰, sucking quality is best.Therefore, best results add 0.1g grifolan in 100g cigarette time, optimum consumption is 1 ‰.
Claims (4)
1. the extracting and purifying method of a grifolan, it is characterized in that, after comprising the steps: the dry Grifola frondosa ether Soxhlet of pulverizing to extract, residue volatilizes and extracts with boiling water, boiling water extraction liquid is concentrated, centrifugal, getting supernatant liquor alcohol precipitation leaves standstill, recentrifuge collecting precipitation thing, after throw out washing, dissolve with distilled water, add again the papoid of 1-2 ‰ to place 2-4h in 50 ± 5 DEG C of water-baths, adopt Sevage method deproteination, concentrating under reduced pressure after activated carbon decolorizing, after cooling, carry out flowing water dialysis, after dialysis finishes, alcohol precipitation leaves standstill again, be deposited in 60-70 DEG C and be dried to obtain Crude polysaccharides.
2. the extracting and purifying method of grifolan as claimed in claim 1, is characterized in that, by Crude polysaccharides chromatographic separation purifying twice, obtains refining polysaccharide; Twice separation and purification elutriant used is respectively 0.5 mol/L NaCl solution and 0.05 mol/L NH
4hCO
3solution.
3. utilize extracting and purifying method described in claim 1 or 2 to prepare the grifolan of gained.
Described in claim 3 grifolan as the application of tobacco humectant.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106551422A (en) * | 2016-11-30 | 2017-04-05 | 湖北中烟工业有限责任公司 | Preparation method and application of the cigarette with Semen Ziziphi Spinosae extractum |
Citations (1)
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| CN101736053A (en) * | 2010-02-01 | 2010-06-16 | 南京泽朗医药科技有限公司 | Technique for extracting Grifola frondosa water-soluble polysaccharide |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101736053A (en) * | 2010-02-01 | 2010-06-16 | 南京泽朗医药科技有限公司 | Technique for extracting Grifola frondosa water-soluble polysaccharide |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106551422A (en) * | 2016-11-30 | 2017-04-05 | 湖北中烟工业有限责任公司 | Preparation method and application of the cigarette with Semen Ziziphi Spinosae extractum |
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Address after: 450000 Yulin South Road, Henan, Zheng Dong, No. 16 South Road, Zhengzhou Patentee after: China Tobacco Henan Industrial Co., Ltd. Address before: 450000 Zhengzhou agricultural road, Henan, No. 29 Patentee before: China Tobacco Henan Industrial Co., Ltd. |