CN103467611A - Preparation method of carragheenan sulfated polysaccharides - Google Patents
Preparation method of carragheenan sulfated polysaccharides Download PDFInfo
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Abstract
The invention discloses a preparation method of carragheenan sulfated polysaccharides, and belongs to the medicine processing field. The carragheenan sulfated polysaccharides can be used for exploiting novel anti-influenza virus medicines, and prepared through the steps of carrying out microwave radiation acidolysis and sulfation reactions to obtain sulfated polysaccharides having molecular weights respectively being 1000-3000Da, 5000-8000Da and 10000-12000Da and having specific sulfate radical substitution values, and purifying and drying through adopting nanofiltration and vacuum lyophilization. A product obtained through the method has a yield of above 80% and a concentrated molecular weight, and the method is simple to operate and has a good repeatability. The product and the production method can be widely used in the active polysaccharide function research field and the pharmaceutical healthcare product field.
Description
Technical field
The present invention relates to a kind of polyose medicament processing technology, is to utilize carrageenin to prepare the method for the controlling sulfate polyose of a kind of specified molecular weight and specific sulfuric acid substitution value concretely.
Background technology
Marine alga is quantity and the maximum class of kind in marine plant, increasingly wide to their bioactive research.The seaweed bio active substance is broadly divided into two kinds, and a kind of is intercellular viscous polysaccharide, mainly comprises alginic acid in brown alga, fucoidin, agar-agar, carrageenin etc. in red algae; Another kind is molecular weight, after absorption, can directly or indirectly affect the internal metabolism material, mainly comprises halogen compound, terpenoid, bromine phenolic compound, Resorcinol, marine alga tannin and laminine etc.The Sargassum polysaccharides that up to the present, can carry out suitability for industrialized production has algin, agar-agar, carrageenin, fork Furcellaria gum etc.The carrageenin extracted from the red algae carrageen is a kind of galactan sulfate, and it has no adverse effects the experiment in vitro proof to normal cell, but the energy blocking virus is to absorption, the intrusion of cell.The carrageenin of the specified molecular weight obtained by acid degradation and sulfation has anti-influenza activity preferably, and the space conformation of the molecular size range of the activity of carrageenin polysaccharide and polysaccharide molecule, sulfate radical content and polysaccharide has larger contacting.
The molecular weight of polysaccharide is to affect one of its bioactive important factor, because the conformation that can the molecular weight of polysaccharide have reflected to a certain extent solubleness, the viscosity of polysaccharide and be formed at the acceptor coupling, for example (molecular-weight average is 4 kDa ~ 5 kDa to low-molecular-weight heparin, distribution range is 1 kDa ~ 10 kDa) anti thrombotic action be better than unfractionated heparin (30 kDa), there are the characteristics such as bioavailability is high, Half-life in vivo is long, bleeding tendency is little, oral easy absorption.In fact, when polysaccharide and receptors bind, only have several oligosaccharides and receptors bind in its molecule, therefore with enzyme, want to die, there is the active centre of one or several oligose fragment in its molecule, at present, find and find that new small molecules oligosaccharides has become the development trend of research polysaccharide according to active centre.Given this, select suitable degradation method to obtain the Low-molecular-weight polysaccharide widow bglii fragment of narrow distribution to screening the Low-molecular-weight polysaccharide medicine and finding that active centre is significant.
The method of polysaccharide degraded has a variety of, as acid degradation, oxidative degradation, ultrasonic wave (assisting) degraded and microwave (assisting) degraded etc., wherein the microwave assistant degradation method is because heating rapidly the even fast degradation that is conducive to polysaccharide with homogeneous heating by microwave, thereby can obtain the Low-molecular-weight polysaccharide of narrow molecular weight distributions or oligosaccharides and noticeable, have easy to operate, pollute low, yield high.At present, the degraded of carrageenin also is only limited to enzyme liberating, oxidative degradation and acid degradation, and relevant microwave assistant degradation method is also more rare for the preparation of small molecules amount carrageenin polysaccharide.
Summary of the invention
The invention discloses a kind of preparation method of carrageenan sulfated esterificated polysaccharide, purpose is to provide prioritization scheme for small molecules amount carrageenin Sulfate of polysaccharide preparation technology, make the target product molecular weight more concentrated, efficiency of pcr product is high, simple to operate, reproducible, thus starting material provided for the bioactivity research of carrageenan sulfated polysaccharide.
A kind of carrageenan sulfated esterificated polysaccharide of the present invention, be to be made by carrageenin purifying drying after microwave radiation acidolysis and sulfuric acid esterification, has specific molecular weight distribution and sulfate radical substitution value, and concrete preparation method is as follows:
(1) microwave radiation acidolysis
A. the carrageenin raw material is made into to 4 ~ 8% Colloidal fluids, preparation amount 600 mL, Microwave Pretreatment 8 ~ 15 min, obtain the colloidal solution of homogeneous transparent; In the situation that pH 2.0, with 750 W microwave treatment 10 ~ 30 min;
When microwave treatment time is 25 ~ 35 min, polysaccharide molecular weight is 1000～3000 Da; When microwave treatment time is 15 ~ 25 min, polysaccharide molecular weight is 5000～8000 Da; When microwave treatment time is 10 ~ 15 min, polysaccharide molecular weight is 10000～12000 Da;
B. after acidolysis reaction finishes, add the 1mol/L sodium hydroxide solution, the pH value is pulled back to 4.5, when temperature drops to normal temperature, regulate pH to 6.0 with alkali again;
C. acid hydrolysis solution adopts rotary evaporation to be concentrated, the polysaccharide of molecular weight 10000 ~ 20000 Da is concentrated into 8 ~ 12%, and the polysaccharide that molecular weight is less than 10000 Da is concentrated into 12 ~ 18%, after concentrating, carries out spraying drying, sprinkler pressure is 1.5 ~ 3 kg, operating pressure 4 ~ 6 kg;
(2) sulfuric acid esterification
Adopting gradient increased temperature to carry out sulfuric acid esterification, is 3 ~ 5:1 in the mol ratio of esterifying agent chlorsulfonic acid and polysaccharide, reacts 4 h, and the polysaccharide of 3 kinds of molecular weight reacts under the gradient alternating temperature of 45 ℃ ~ 80 ℃, and the sulfate radical substitution value is 1.13 ~ 1.95;
(3) purifying drying
Purifying process is the nanofiltration membrane that adopts 250Da, is mainly used in removing SO
4 2-and mineral ion, drying process is vacuum lyophilization.
The controlling sulfate polyose prepared by aforesaid method has good antiviral activity, can be used for development of new resisiting influenza virus class medicine.
Beneficial effect of the present invention:
The preparation method of the carrageenan sulfated esterificated polysaccharide of the present invention is clear and definite specific fractionated molecule amount and sulfate radical substitution value, optimized the preparation technology of different molecular weight carrageenin polysaccharide and sulphating derivative thereof, and main advantage is embodied in following three aspects:
(1) effectively improve preparation efficiency and the molecular weight intensity of specified range small molecules amount carrageenin polysaccharide, obtained gratifying effect;
(2) determined the microwave acidolysis condition of specific small molecules weight range, that the additive methods such as the relative enzymolysis process of preparation method have is easy to operate, pollute low, yield high;
(3) optimizing and controlling method of the sulfuric acid esterification gradient increased temperature of taking, make the carrageenin Sulfate of polysaccharideization more even.Research for the carrageenan sulfated polysaccharide antiviral activity of small molecules amount is significant.
embodiment:
1,the microwave radiation acidolysis
(1) the carrageenin raw material is made into to 5% Colloidal fluid, preparation amount 600mL, Microwave Pretreatment 10 min, obtain the colloidal solution of homogeneous transparent; In the situation that pH 2.0, with 750 W microwave treatment 10 ~ 30 min.
A. when microwave treatment time is 30 min, polysaccharide molecular weight is 1000～3000 Da;
B. when microwave treatment time is 20 min, polysaccharide molecular weight is 5000～8000 Da;
C. when microwave treatment time is 12 min, polysaccharide molecular weight is 10000～12000 Da.
(2) after acidolysis reaction finishes, add 1 mol/L sodium hydroxide solution, the pH value is pulled back to 4.5, when temperature drops to normal temperature, regulate pH to 6.0 with alkali again.
(3) acid hydrolysis solution adopts rotary evaporation to be concentrated, and the polysaccharide of molecular weight 10000 ~ 20000 Da is concentrated into 10%, and the polysaccharide that molecular weight is less than 10000 Da is concentrated into 15%, after concentrating, carries out spraying drying, and sprinkler pressure is 2 kg, operating pressure 5 kg.
,sulfuric acid esterification
(1) when the mol ratio of esterifying agent chlorsulfonic acid and polysaccharide is 4:1, while reacting 4 h, the polysaccharide of different molecular weight and temperature of reaction are controlled, and the sulfate radical substitution value is as follows:
(2) different esterifying agent chlorsulfonic acids and polysaccharide mol ratio, the sulfuric acid substitution value result of the polysaccharide of different molecular weight, temperature of reaction and time is as follows:
3,the purifying drying
The purifying process of controlling sulfate polyose is the nanofiltration membrane that adopts 250Da, is mainly used in removing SO
4 2-and mineral ion; Drying process is vacuum lyophilization.
The controlling sulfate polyose yield that utilizes this explained hereafter is 83.2%, and molecular weight is concentrated, and can be widely used in the exploitation of anti influenza class medicine.
Claims (3)
1. a carrageenan sulfated esterificated polysaccharide, is characterized in that this product is to be made by carrageenin purifying drying after microwave radiation acidolysis and sulfuric acid esterification, has specific molecular weight distribution and sulfate radical substitution value, and concrete preparation method is as follows:
(1) microwave radiation acidolysis
A. the carrageenin raw material is made into to 4 ~ 8% Colloidal fluids, preparation amount 600 mL, Microwave Pretreatment 8 ~ 15 min, obtain the colloidal solution of homogeneous transparent; In the situation that pH 2.0, with 750 W microwave treatment 10 ~ 30 min;
B. after acidolysis reaction finishes, add the 1mol/L sodium hydroxide solution, the pH value is pulled back to 4.5, when temperature drops to normal temperature, regulate pH to 6.0 with alkali again;
C. acid hydrolysis solution adopts rotary evaporation to be concentrated, the polysaccharide of molecular weight 10000 ~ 20000 Da is concentrated into 8 ~ 12%, and the polysaccharide that molecular weight is less than 10000 Da is concentrated into 12 ~ 18%, after concentrating, carries out spraying drying, sprinkler pressure is 1.5 ~ 3 kg, operating pressure 4 ~ 6 kg;
(2) sulfuric acid esterification
Adopting gradient increased temperature to carry out sulfuric acid esterification, is 3 ~ 5:1 in the mol ratio of esterifying agent chlorsulfonic acid and polysaccharide, reacts 4 h, and the polysaccharide of 3 kinds of molecular weight reacts under the gradient alternating temperature of 45 ℃ ~ 80 ℃, and the sulfate radical substitution value is 1.13 ~ 1.95;
(3) purifying drying
Purifying process is the nanofiltration membrane that adopts 250Da, is mainly used in removing SO
4 2-and mineral ion, drying process is vacuum lyophilization.
2. use 750 W microwave treatment 10 ~ 40 min as described as claim 1 step (1), is characterized in that when microwave treatment time is 25 ~ 35 min, and polysaccharide molecular weight is 1000～3000 Da; When microwave treatment time is 15 ~ 25 min, polysaccharide molecular weight is 5000～8000 Da; When microwave treatment time is 10 ~ 15 min, polysaccharide molecular weight is 10000～12000 Da.
3. controlling sulfate polyose as claimed in claim 1, is characterized in that, described controlling sulfate polyose has good antiviral activity, can be used for development of new resisiting influenza virus class medicine.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107011454A (en) * | 2017-04-10 | 2017-08-04 | 浙江海洋大学 | A kind of sea cucumber fucoidan preparation method of the high sulphation of low molecule amount |
| CN111825777A (en) * | 2020-07-13 | 2020-10-27 | 山东众山生物科技有限公司 | Method for preparing heparinoids from chondroitin |
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2013
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107011454A (en) * | 2017-04-10 | 2017-08-04 | 浙江海洋大学 | A kind of sea cucumber fucoidan preparation method of the high sulphation of low molecule amount |
| CN107011454B (en) * | 2017-04-10 | 2019-06-18 | 浙江海洋大学 | A kind of sea cucumber fucoidan preparation method of the high sulphation of low molecular weight |
| CN111825777A (en) * | 2020-07-13 | 2020-10-27 | 山东众山生物科技有限公司 | Method for preparing heparinoids from chondroitin |
| CN111825777B (en) * | 2020-07-13 | 2022-05-27 | 山东众山生物科技有限公司 | Method for preparing heparinoids from chondroitin |
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