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Radial flow chromatographic separation and purification method for biological polysaccharide

A chromatographic separation and purification method technology, applied in the field of separation and purification of biological polysaccharides using macroporous resin radial flow chromatography technology, can solve the problems of complex separation and purification process, adverse effects of drug quality, operator and environmental hazards, and achieve simple process , shorten the separation and purification time, and facilitate the operation

Inactive Publication Date: 2010-12-15
HARBIN INST OF TECH AT WEIHAI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, most of the separation and purification strategies of biological polysaccharides are polysaccharide extraction supernatants are heated and concentrated—ethanol precipitationcentrifugation—to obtain crude polysaccharides—dialysis to remove small molecular impurities—hydrogen peroxide decolorization or activated carbon decolorization—Sevage method or trichloroacetic acid method to remove Miscellaneous protein—axial flow ion exchange chromatography or gel chromatography purification—eluent concentration—ethanol precipitationcentrifugation—precipitation and freeze drying—finally purified polysaccharides. It can be seen that the separation and purification process of traditional methods is complicated and the amount of chemical solvents large, poor environmental protection effect, low production efficiency, high production cost, and difficult to industrialized production. At the same time, due to the use of a large amount of organic solvents in the preparation process of polysaccharides, it not only causes harm to the operator and the environment, but also the residue of harmful substances affects the quality of the drug. Also cause a certain degree of adverse effects, therefore, the development of simple, efficient, clean, green separation and refining technology is a top priority
[0003] Radial flow chromatography technology is a new type of high-efficiency chromatography technology. Due to its radial flow principle, it can increase flow rate, reduce pressure, large capacity and load, easy to operate, and the chromatographic parameters obtained by laboratory or pilot test are stable. It can be directly applied to industrial production. At present, radial flow chromatography is mainly used to separate protein drugs. Unfortunately, there is no method for separating and purifying biological polysaccharides using radial flow chromatography.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] A macroporous resin radial flow chromatography for directly separating and purifying exopolysaccharides from the fermentation supernatant of medicinal fungi, first centrifuging the liquid fermentation product of medicinal fungi at 4000 rpm for 10 minutes, collecting the supernatant, and then , the fermentation supernatant containing fungal polysaccharides is directly loaded on a 500 ml macroporous resin radial chromatographic column at a flow rate of 60 ml / min. Flow rate is 50 milliliters / minute, collects 530 milliliters of eluents, is the polysaccharide solution that contains purification, through measuring with phenol-sulfuric acid method, the recovery rate of polysaccharide is 85.2%, through spectrophotometry, the pigment content in the reclaimed polysaccharide The reduction can reach 82.3%. As determined by the Bradford method, the protein content in the recovered polysaccharide can be reduced by up to 85.6%, so that both the recovery rate and the purity of the polys...

Embodiment 2

[0018] A macroporous resin radial flow chromatography for directly separating and purifying intracellular polysaccharides from the supernatant of mycelium enzymolysis. First, the mycelium enzymolysis product is centrifuged at 6000 rpm for 10 minutes to collect the supernatant, and then, The enzymolysis supernatant containing fungal polysaccharides is directly loaded on a 500 ml macroporous resin radial chromatographic column at a flow rate of 120 ml / min. 200 milliliters / min, collect 800 milliliters of eluents, namely contain the polysaccharide solution of purification, through measuring with phenol-sulfuric acid method, the recovery rate of polysaccharide is 98.1%, measure through spectrophotometry, the pigment content reduction in reclaiming polysaccharide can be As determined by the Bradford method, the protein content in the recovered polysaccharide can be reduced by up to 75.6%, so that the recovery rate and purity of the polysaccharide are significantly increased.

Embodiment 3

[0020] A macroporous resin radial flow chromatography for directly separating and purifying polysaccharides from fungal fermentation supernatants. First, the fungal liquid fermentation product is centrifuged at 5000 rpm for 10 minutes to collect the supernatant. The fermentation supernatant is directly loaded on a 500 ml macroporous resin radial chromatographic column at a flow rate of 90 ml / min, with a sample volume of 500 ml, and then eluted with distilled water at a flow rate of 100 ml / min , collect 650 milliliters of eluents, namely contain the polysaccharide solution of purification, through measuring with phenol-sulfuric acid method, the recovery rate of polysaccharide is 94.9%, measure through spectrophotometry, the pigment content in reclaiming polysaccharide reduces and can reach 79.2%, As determined by the Bradford method, the protein content in the recovered polysaccharide can be reduced by up to 82.6%, so that both the recovery rate and the purity of the polysacchar...

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Abstract

The invention relates to a radial flow chromatographic separation and purification method for biological polysaccharide, which comprises the following steps of: centrifuging a medical fungi liquid fermentation product or a mycelium (fruiting body) enzymolysis product and collecting supernatant or collecting supernatant which comprises polysaccharide and is extracted from plants, animals, bacteriaand yeast; directly loading the supernatant on a macroporous resin radial chromatographic column of 500ml at the flow rate of 60 to 120ml per minute; eluting the macroporous resin radial chromatographic column by using distilled water or deionized water at the flow rate of 50 to 200ml per minute; and collecting 500 to 800ml of eluent, namely solution containing purified polysaccharide. The polysaccharide is directly separated and purified from the enzymolysis supernatant and the fermentation supernatant of the medical fungi mycelium and the supernatant extracted from the plants, animals, bacteria and yeast by a one-step method; and the biological polysaccharide separation and purification method has the advantages of simple process, convenient operation, no use of an organic solvent, highproduction efficiency and obvious purification effect.

Description

Technical field: [0001] The invention relates to technical fields such as separation science and engineering, biopharmaceuticals, etc., and specifically relates to a method for separating and purifying biological polysaccharides by utilizing macroporous resin radial flow chromatography technology. Background technique: [0002] Biological polysaccharides (including fungi, bacteria, plants, animals, etc.) have attracted extensive attention at home and abroad because of their remarkable anti-tumor, immune regulation, anti-oxidation, anti-aging and other biological activities. At present, most of the separation and purification strategies of biological polysaccharides are polysaccharide extraction supernatants are heated and concentrated—ethanol precipitationcentrifugation—to obtain crude polysaccharides—dialysis to remove small molecular impurities—hydrogen peroxide decolorization or activated carbon decolorization—Sevage method or trichloroacetic acid method to remove Miscel...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08B37/00B01D15/08
Inventor 闫培生张炳照曹立新肖伟
Owner HARBIN INST OF TECH AT WEIHAI
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