CN102312021A - Preparation method of Curdlan oligomers - Google Patents
Preparation method of Curdlan oligomers Download PDFInfo
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- CN102312021A CN102312021A CN2010102136794A CN201010213679A CN102312021A CN 102312021 A CN102312021 A CN 102312021A CN 2010102136794 A CN2010102136794 A CN 2010102136794A CN 201010213679 A CN201010213679 A CN 201010213679A CN 102312021 A CN102312021 A CN 102312021A
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- acid
- derain
- oligosaccharides
- curdlan
- oligomers
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Abstract
The invention discloses a preparation method of Curdlan oligomers. The method comprises the following steps of: fully dissolving Curdlan polysaccharide into a solvent to obtain a homogeneous solution; adding acid at a temperature in a range of 80-140 DEG C, wherein a final concentration of the acid in the solution is 0.1-10 M; reacting for 30-500 min; adding organic solvent sediments which are 3-5 times as much as the volume of reaction liquid and meanwhile, stirring and filtering to obtain a milky solid; washing the solid by the organic solvent 1-4 times to obtain the Curdlan oligomers with an average molecular weight of 340-4000 Da, which is measured by an efficient liquid phase detection after drying. The degradation efficiency of the preparation method reaches more than 85%. The Curdlan oligomers are oligomers formed by connecting glucose molecules through beta-(1->3) glucosidic bonds; the degree of polymerization (Degree of Polymerization, DP) of the Curdlan oligomers is 2-24; a molecular formula of the Curdlan oligomers is (C6H12O6)n, wherein n is equal to 2-24. The average molecular weight of the Curdlan oligomers is 340-4000 Da. The preparation method provided by the invention has a simple and controllable preparation process, short reaction time and good degradation efficiency, and is environmentally friendly and suitable for industrial production.
Description
Technical field
But the present invention relates to the Derain oligosaccharides, but but the method for specifically a kind of acid system degraded Derain polysaccharide preparation Derain oligosaccharides.
Background technology
But the Derain polysaccharide is by the exocellular polysaccharide that Alcaligenes faecalis var myxogenes 10c3 fermentation produces, and is that substruction is that glucose passes through β-(1 → 3) glycosidic link and is formed by connecting, and is β-1, a member of 3-VISOSE family.Up to now, this polysaccharide is the unique a kind of ramose β-1 that do not contain of occurring in nature, the 3-VISOSE.Yet, but because the Derain polysaccharide is water insoluble, seriously hindered its application in food, medicine, fodder additives and field of materials.
But the Derain oligosaccharides also is called as β-1, and 3-Portugal oligosaccharides is the polysaccharide degradation product, water-soluble enhancing greatly.Along with oligosaccharides research receives people's attention day by day, but the biological function of Derain oligosaccharides and verivate thereof and physiologically active are disclosed gradually.After Shimizu (2001) passes through to compare the 26S Proteasome Structure and Function of Nutriflora P and wooden poly oligosaccharide, but think that the Derain oligosaccharides possibly also have the adjusting intestinal microflora, as the function of beneficial source of students.Simultaneously, also having the potential market that is developed as medical research process midbody is worth.
Although but the Derain oligosaccharides has the important physical function; But yet since the Derain oligosaccharides have special tertiary structure and form characteristics such as gel is water insoluble; But there is not report to use the Derain oligosaccharides to be applied to plant biological resistance inductor and growth-promoting activity research at present as yet; Main bottleneck problem is exactly but that Derain oligosaccharides degraded difficulty is big, and the preparation process is loaded down with trivial details, and efficient is lower.Report such as Grandpierre (2008) uses sulfuric acid or trifluoroacetic acid hydrolysis 90h, and the degraded productive rate has only 25%.And but chemosynthesis Derain oligosaccharides technical difficulty is big, and particularly the oligosaccharide monomer more than DP＞7 is synthetic, because of mode of connection and the steric configuration that relates to sugar unit, and a series of problems such as the protection of reactive group, difficulty and cost all make us hanging back.
But if there is not effectively to prepare the method for Derain oligosaccharides, will be unfavorable for studying oligosaccharide structure and bioactive functions, inquire into the relation between Derain oligosaccharides and verivate and the disease generation profoundly but hinder from the mechanism aspect.
Summary of the invention
The present invention adopts cheap raw material, but clear structure Derain polysaccharide, but use the reasonable preparation technology of acid system degraded and the technical object that the separation and purification means realize preparation Derain oligosaccharides; Its advantage can accurately be measured and used, and uses homogeneous reaction system, improves the degradation efficiency of oligosaccharides; Degradation efficiency is up to more than 85%; The present invention has overcome oligosaccharides effectively and has prepared the difficult problem in the process, has improved the yield of oligosaccharides, but lays a good foundation for preparing a large amount of Derain oligosaccharides raw materials.As glycan molecule, can degrade at occurring in nature fully, free from environmental pollution, have positive effect to promoting environmental friendliness and green agriculture.
For realizing above-mentioned purpose, the technical scheme that the present invention adopts is:
But a kind of acid system degraded prepares the method for Derain oligosaccharides.But homogeneous phase solution is processed in the abundant dissolving in solvent of Derain polysaccharide, can effectively be avoided the problem of the insufficient degraded that causes of dissolving insufficient (yield has only 27%) in water solution system; Under 80-140 ℃ of condition, the acid solution reaction 30-500min that adds concentration and be 0.1M-5M can make yield reach more than 85.12%, and the adjusting PH with base of 0.1M-5M will further improve yield to 6-8; Reach more than 90.2%, be concentrated into the 1/5-5/5 of former reaction volume, add organic solvent deposit; Stir simultaneously, filter, obtain Off-white solid; Solid is with organic solvent washing 1-4 time; But dry back is used performance liquid to detect and is obtained the Derain oligosaccharides of molecular-weight average at 340-4000Da, for glucose molecule through the linear molecule that β-(1 → 3) glycosidic link is formed by connecting, do not contain side chain branch; What be different from other sources contains side chain ramose β-1,3-VISOSE; But the polymerization degree of Derain oligosaccharides is 2-24, and molecular formula is (C
6H
12O
6) n, n=2-24, but the molecular-weight average of described Derain oligosaccharides is shown in the following figure at its structural formula of 340-4000Da:
Described solvent is generally one or more mixed solutions that solvent is alkali lye, DMSO 99.8MIN., formic acid, cadmium oxalic acid, liquor kalii iodide, and the ratio of itself and polysaccharide is 5: 1-30: 1.Described acid is one or more mixing acid of hydrochloric acid, sulfuric acid, nitric acid, acetic acid, formic acid, trifluoroacetic acid and oxalic acid.Described alkali is one or more mixed bases of sodium hydroxide, Pottasium Hydroxide, yellow soda ash, sodium hydrogencarbonate, salt of wormwood, saleratus.Described organic solvent is one or more the mixture in ether, acetone, methyl alcohol, ethanol, sherwood oil, the normal hexane.
But the present invention is an abundant dissolving Derain polysaccharide in specific solvent, but obtains the Derain oligosaccharides that the polymerization degree is 2-24 through the acid system degraded, and is efficient and processing condition are simple, improved the preparation efficiency of oligosaccharides greatly.Be used for agriculture prodn, can improve the output of crop, the inducing plant disease resistance strengthens the natural immunity of plant, is eco-friendly green bio agricultural chemicals.
Simultaneously, but proved also that through grape cell disease resistance experimental result the effect of linearity Derain oligosaccharides inducing plant resistance is better than the VISOSE in other sources of containing branched structure.
The present invention has following advantage:
But the preparation method of Derain oligosaccharides of the present invention and the product of preparation have very big advantage with relevant patent with publishing an article, the main performance as follows:
1. the acid system of the present invention's employing is simple to operate, and condition is controlled, and by product is few, and experimental repeatability is good;
2. degradation effect can reach more than 85% in homogeneous system, be higher than far away Grandpierre etc. (Enzymatic and chemical degradation of curdlan targeting the production of beta-(1 → 3) oligoglucans [J].
Carbohydrate Polymers71 (2): the 277-286.) result of the acidolysis efficient 25% of report; And for they reacted 90h, this experimental period cycle was short, and the equipment and instrument requirement is low, technology is simple, yield is high, product purity is high; Whole process is green, has the good prospect of suitability for industrialized production;
3. but the Derain oligosaccharides of the present invention's preparation can be degraded at occurring in nature as glycan molecule fully, and is free from environmental pollution.
Description of drawings
But Fig. 1 detects the component concentration figure of the Derain oligosaccharides of acid system preparation for the present invention uses performance liquid
But Fig. 2 uses ground substance assistant laser desorption ionization flight time mass spectrum (MALDI-TOF MS) for the present invention the collection of illustrative plates of detection Derain oligosaccharides
Embodiment
Below in conjunction with embodiment the present invention is described further:
Embodiment 1: but the preparation method of Derain oligosaccharides (A)
But take by weighing 5g Derain polysaccharide and in 50ml potassiumiodide, cadmium ethylenediamine solution (ratio is 1: 1), be stirred well to dissolving fully, under 90 ℃ (can operate under the 80-140 ℃ of condition), slowly be added dropwise to the oxalic acid mixing solutions 3ml of 0.5M, behind the reaction 60min; Mixing solutions (V/V=1) deposition with 50mL ethanol and acetone; Stir simultaneously, obtain off-white powder, suction filtration; Obtain 4.25g in 50 ℃ of following vacuum-dryings, productive rate reaches 85.12%.
But high-efficient liquid phase technique detection and ground substance assistant laser desorption ionization flight time mass spectrum proof gained material are the Derain oligosaccharides after testing.
Embodiment 2: but the preparation method of Derain oligosaccharides (B)
But take by weighing 5g Derain polysaccharide in the 50ml DMSO 99.8MIN., be stirred well to dissolving fully in the liquor kalii iodide (ratio is 1: 1), under 90 ℃ (can operate under the 80-140 ℃ of condition), slowly be added dropwise to the trifluoroacetic acid mixing solutions 3ml of 0.5M; Behind the reaction 60min,, neutralize with mixed ammonium/alkali solutions with the mixed ammonium/alkali solutions neutralization; Be concentrated into 10ml at 45 ℃ of (can under 30-150 ℃, operate) rotary evaporations, be light yellow transparent liquid, with mixing solutions (V/V=1) deposition of 50mL ethanol and acetone; Stir simultaneously, obtain off-white powder, suction filtration; But get Derain oligosaccharides 4.51g in 50 ℃ of following vacuum-dryings, productive rate reaches 90.2%.
But raw material Derain polysaccharide of the present invention is so far, unique a kind of β-1 that does not contain branched structure that nature is found, 3-VISOSE; Be different from the VISOSE that other contain branched structure; For example yeast glucan (β-1, the 3-main chain has β-1,6-glucose side chain; Isolation of soluble yeast beta-glucans that inhibit human monocyte phagocytosi s mediated by beta-glucanreceptors [J] .The Journal of Immunology; 1986,137 (10), 3270); Compare and relate to the method for from yeast, extracting VISOSE with patent 200410046141.3; The method operating process that the present invention adopts is simpler; Do not contain mannosans in the raw material that extracts; Can not introduce the interference of mannosans, need not loaded down with trivial details step process such as ultrafiltration, annealing, process is simple to operation; Degradation efficiency is up to more than 85%, more than the purity to 98%.And the primary product that the present invention obtains is the oligosaccharides of molecular weight (340-4000Da), is far smaller than the molecular weight (2 * 10 of yeast glucan
5-2 * 10
6), but therefore the Derain oligosaccharides has better water-solubility.And aspect activity research, but cell more is prone to the lower Derain oligosaccharides of identification molecular weight, so the biological activity of its embodiment is better than the high-molecular weight yeast glucan; More help studying the relation between carbohydrate molecule structure and the biological activity.
The present invention adopts cheap raw material, but clear structure Derain polysaccharide, but use the reasonable preparation technology of acid system degraded and the technical object that the separation and purification means realize preparation Derain oligosaccharides; Its advantage can accurately be measured and used, and uses homogeneous reaction system, improves the degradation efficiency of oligosaccharides, and degradation efficiency, can be degraded at occurring in nature as glycan molecule up to more than 85% fully, and is free from environmental pollution, has positive effect to promoting environmental friendliness and green agriculture.
Claims (5)
1. but acid system degraded Derain polysaccharide prepares the method for oligosaccharides, it is characterized in that: process homogeneous phase solution but will the Derain polysaccharide in solvent, fully dissolve, under 80-140 ℃ of condition; Add acid, the final concentration of acid in solution is 0.1-10M, reaction 30-500min; The organic solvent deposit that adds 3-5 times of volumetric reaction liquid stirs simultaneously, filters; Obtain Off-white solid, solid is with organic solvent washing 1-4 time, but obtains molecular-weight average after the drying at 340-4000Da Derain oligosaccharides; But described Derain oligosaccharides be glucose molecule through β-(1 → 3) glycosidic link linear molecule that is formed by connecting, do not contain side chain branch, what be different from other sources contains side chain ramose β-1,3-VISOSE; (but Degree of Polymerization DP) is 2-24 to the polymerization degree of Derain oligosaccharides, and molecular formula is (C
6H
12O
6) n, n=2-24, its structural formula is shown in the following figure:
2. according to the method for claim 1, it is characterized in that: but the solvent of said dissolving Derain polysaccharide is one or more mixing solutionss in cadmium ethylenediamine, potassiumiodide, alkali lye, DMSO 99.8MIN., the formic acid.
3. according to the method for claim 1, it is characterized in that: described acid is one or more mixing acid in hydrochloric acid, sulfuric acid, nitric acid, acetic acid, formic acid, trifluoroacetic acid and the oxalic acid.
4. according to the method for claim 1, it is characterized in that: said organic solvent is one or more the mixture in ether, acetone, methyl alcohol, ethanol, sherwood oil, the normal hexane.
5. according to the method for claim 1, it is characterized in that: before adopting organic solvent deposit, with reaction solution with in the alkaline solution with accent system pH to 5-7, with the yield that further improves oligosaccharides;
Alkaline solution is one or more mixed bases of 0.1-10M sodium hydroxide, Pottasium Hydroxide, yellow soda ash, sodium hydrogencarbonate, salt of wormwood, saleratus.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103524760A (en) * | 2013-10-10 | 2014-01-22 | 江苏大学 | Deoxycholic group-oxidized curdlan polysaccharide nanoparticle and preparation method thereof |
| CN104087531A (en) * | 2014-07-03 | 2014-10-08 | 江苏一鸣生物科技有限公司 | Alcaligenes faecalis mutant strain and method for preparing curdlan by using alcaligenes faecalis mutant strain |
| CN105566510A (en) * | 2015-12-18 | 2016-05-11 | 南京理工大学 | Glucooligosaccharide and preparation method and application thereof |
| CN110004140A (en) * | 2019-02-25 | 2019-07-12 | 华东师范大学 | A method of improving curdlan Screening of strain with high productivity efficiency |
| CN111333748A (en) * | 2017-01-04 | 2020-06-26 | 苏州大学 | β -1, 3-glucuronic acid oligosaccharide, and preparation method and application thereof |
| CN112251480A (en) * | 2020-10-16 | 2021-01-22 | 福建环海生物科技股份有限公司 | Production process of tremella oligosaccharide |
Citations (3)
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|---|---|---|---|---|
| JPH02243697A (en) * | 1989-03-17 | 1990-09-27 | Meiji Seika Kaisha Ltd | Production of oligosaccharide |
| CN1491966A (en) * | 2002-10-21 | 2004-04-28 | 宁波天安生物材料有限公司 | Curdlan treating method for synthetic reaction |
| JP2005110675A (en) * | 2003-09-16 | 2005-04-28 | Meiji Univ | Method for producing low-molecular polysaccharide and/or oligosaccharide, and functional low-molecular polysaccharide and/or oligosaccharide |
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2010
- 2010-06-30 CN CN2010102136794A patent/CN102312021A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02243697A (en) * | 1989-03-17 | 1990-09-27 | Meiji Seika Kaisha Ltd | Production of oligosaccharide |
| CN1491966A (en) * | 2002-10-21 | 2004-04-28 | 宁波天安生物材料有限公司 | Curdlan treating method for synthetic reaction |
| JP2005110675A (en) * | 2003-09-16 | 2005-04-28 | Meiji Univ | Method for producing low-molecular polysaccharide and/or oligosaccharide, and functional low-molecular polysaccharide and/or oligosaccharide |
Non-Patent Citations (1)
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103524760A (en) * | 2013-10-10 | 2014-01-22 | 江苏大学 | Deoxycholic group-oxidized curdlan polysaccharide nanoparticle and preparation method thereof |
| CN103524760B (en) * | 2013-10-10 | 2016-01-20 | 江苏大学 | A kind of Septochol base-oxidation Curdlan polysaccharide nanoparticle and preparation method |
| CN104087531A (en) * | 2014-07-03 | 2014-10-08 | 江苏一鸣生物科技有限公司 | Alcaligenes faecalis mutant strain and method for preparing curdlan by using alcaligenes faecalis mutant strain |
| CN104087531B (en) * | 2014-07-03 | 2016-07-06 | 江苏一鸣生物科技有限公司 | One strain Bacillus foecalis alkaligenes mutant and the method preparing curdlan with it |
| CN105566510A (en) * | 2015-12-18 | 2016-05-11 | 南京理工大学 | Glucooligosaccharide and preparation method and application thereof |
| CN111333748A (en) * | 2017-01-04 | 2020-06-26 | 苏州大学 | β -1, 3-glucuronic acid oligosaccharide, and preparation method and application thereof |
| CN111333748B (en) * | 2017-01-04 | 2022-06-21 | 苏州大学 | Beta-1, 3-glucuronic acid oligosaccharide, and preparation method and application thereof |
| CN110004140A (en) * | 2019-02-25 | 2019-07-12 | 华东师范大学 | A method of improving curdlan Screening of strain with high productivity efficiency |
| CN110004140B (en) * | 2019-02-25 | 2022-03-11 | 华东师范大学 | Method for improving screening efficiency of curdlan high-yield strains |
| CN112251480A (en) * | 2020-10-16 | 2021-01-22 | 福建环海生物科技股份有限公司 | Production process of tremella oligosaccharide |
| CN112251480B (en) * | 2020-10-16 | 2022-03-08 | 福建环海生物科技股份有限公司 | Production process of tremella oligosaccharide |
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Application publication date: 20120111 |