CN106008739B - The preparation method of oyster polysaccharide - Google Patents

Abstract

The present invention disclose it is a kind of without ethanol precipitation, reliable, at low cost, the high oyster polysaccharide of purity the preparation method of safe operation, carry out in accordance with the following steps：Oyster meat is subjected to homogenized, extracts 10min with 10 ~ 15 times of boiling water of volume/mass, intermittent stirring, filtering obtain filtrate；It is 4 ~ 5 to adjust filtrate pH, stands 12h, and 6000rpm centrifuges 20 min, collects supernatant；It is 7 ~ 8 to adjust supernatant pH, and 6 h are digested under the conditions of 20 ~ 30 DEG C with the complex enzyme of supernatant mass percent 0.5% ~ 1%, and it by unit of activity ratio is 3 that the complex enzyme, which is marine alkaline proteinase, papain, flavor protease and trypsase,：3：2：2 ratios mix；Enzymolysis liquid 6000rpm is centrifuged into 20min, collects enzymolysis supernatant；The ultrafiltration membrance filter that supernatant passes through molecular cut off 50kDa will be digested, trapped fluid after dried film obtains oyster polysaccharide.

Description

The preparation method of oyster polysaccharide

Technical field

The method that the present invention relates to a kind of to prepare oyster polysaccharide from oyster, it is especially a kind of without ethanol precipitation, operation Securely and reliably, at low cost, the high oyster polysaccharide of purity preparation method.

Background technology

Oyster is one of the Important Economic cultivated shellfish in China, has kind of oyster kind about more than 20 in China's Coastal Areas, Such as Pacific oyster（Crassostrea gigas）, Crassostrea rivularis (Crassostrea rivularis), pleat oyster (Crassostrea plicatula), ostrea talienwhanensis Crosse (Crassostrea talienwhanensis) and close squama oyster (Ostrea denselamellosa) etc. kinds.Oyster contains abundant nutriment, in dry product protein account for about 45%~ 57%, fat accounts for about 7%~11%, and polysaccharide accounts for about 19%~38%.Promote cell metabolism research shows that oyster polysaccharide has, change Mercy is dirty, liver and sanguimotor function, is active constituent important in oyster.Currently, the preparation method of oyster polysaccharide has It is a variety of, such as the application for a patent for invention of Chinese Patent Application No. 201510322757.7, disclose a kind of " ocean oyster biological polyoses Extract preparation method ", it comprises the concrete steps that：" it is rubbed after cleaning up oyster fresh meat, adds the water homogenate of 20-30 times of quality volume, PH is adjusted, is added 0.5%（Mass fraction）Protease, in 50-60 DEG C of insulation reaction 3-5h；Again by centrifuge, it is heavy to collect It forms sediment to get oyster crude extract；Oyster crude extract is sufficiently stirred dissolving in 15-25 times of water, is stood, not by centrifuge removal Molten object；Clear liquid is filtered by cardboard filter device, 2-4 times of 95% ethyl alcohol of volume is then slowly added in filtrate, at 3-5 DEG C Under the conditions of it is 12-16 hours static, then centrifuge, precipitation taken out, vacuum dried, obtain oyster polysaccharide ".Although not disclosing tool The protease of body needs consuming energy but it can be seen that its enzymolysis process need to carry out under 50-60 DEG C of hot conditions（Thermal energy）, Increase production cost.Nevertheless, still polysaccharide cannot be thoroughly digested with the pilotaxitic texture that protein combines makes it detach, only High concentration can be used（95%）Ethyl alcohol makes the polysaccharide precipitation in liquid, and ethyl alcohol has safety during large-scale industrial production Hidden danger.

Invention content

The present invention is provided a kind of without ethanol precipitation, behaviour to solve the above-mentioned technical problem present in the prior art Make the preparation method of safe and reliable, the at low cost high oyster polysaccharide of purity.

Technical solution of the invention is：A kind of preparation method of oyster polysaccharide, it is characterised in that successively according to as follows Step carries out：

A. fresh decladding oyster is gone after internal organ to carry out homogenized, is extracted with 10 ~ 15 times of boiling water of volume/mass 10min, intermittent stirring, filtering, obtains filtrate；

B. it is 4 ~ 5 to adjust filtrate pH, stands 12h, and 6000rpm centrifuges 20 min, collects supernatant；

C. it is 7 ~ 8 to adjust supernatant pH, with the complex enzyme of supernatant mass percent 0.5% ~ 1% at 20 ~ 30 DEG C with supernatant 0.5% ~ 1% complex enzyme of liquid mass percent digests 6 h under the conditions of 20 ~ 30 DEG C, under the conditions of digest 6 h, the complex enzyme is The flavor protease and 6000u/ of the marine alkaline proteinase of 6000u/mg, the papain of 6000u/mg, 15000 u/mg Mg trypsase is 3 by unit of activity ratio：3：2：2 ratios mix；

D. enzymolysis liquid 6000rpm is centrifuged into 20min, collects enzymolysis supernatant；

E. the ultrafiltration membrance filter that supernatant passes through molecular cut off 50kDa will be digested, trapped fluid after dried film obtains Oyster polysaccharide.

The present invention is to be configured to compound protease in proportion using four kinds of protease to digest oyster homogenate supernatant, Various enzyme interactings can make in oyster polysaccharide and protein pilotaxitic texture at normal temperatures（20~30℃）Enzymolysis separation, makes collection Supernatant in contain the higher oyster polysaccharide of concentration, be not necessarily to ethanol precipitation, only need to pass through molecular cut off 50kDa ultrafiltration membrane Enzyme and enzymolysis product in filtering removal enzymolysis liquid, you can the oyster polysaccharide of purity >=96% is obtained, reliable with safe operation, At low cost, the advantages that purity is high.

The pilotaxitic texture that polysaccharide in oyster is combined with protein can be separated well, be not necessarily to ethanol precipitation, only need through Cross the ultrafiltration membrance filter of molecular cut off 50kDa, you can with the smaller enzyme and enzymolysis product of removal molecular weight, obtain purity >= 96% oyster polysaccharide has many advantages, such as that safe operation is reliable, at low cost, purity is high.

Specific implementation mode

A. fresh decladding oyster goes after internal organ to carry out homogenized, with 15 times（Volume/mass）Boiling water extracts 10min, Intermittent stirring, filtering obtain filtrate, and filter residue is reprocessed 2 times with same method, and 3 filtrates are merged；

B. it is 5 by the filtrate second acid for adjusting pH of collection, stands 12h, 6000rpm centrifuges 20 min, collects supernatant；

C. it is 7 to adjust supernatant pH with sodium hydroxide, and supernatant is added in the complex enzyme of supernatant mass percent 1% In, room temperature（20~30℃）6 h are digested, the complex enzyme is by marine alkaline proteinase (6000u/mg), papain （6000u/mg）, flavor protease（15000 u/mg）And trypsase（6000u/mg）It is 3 by unit of activity ratio：3：2：2 ratios Example mixing；

D. by obtained enzymolysis liquid, 6000rpm centrifuges 20min again, obtains enzymolysis supernatant；

E. the enzymolysis supernatant of acquisition is passed through into ultrafiltration membrane（Molecular cut off 50kDa）, the trapped fluid after film excessively is carried out Spray drying, obtains oyster polysaccharide.Oyster polysaccharide purity >=96% is measured through Phenol sulfuric acid procedure.

Claims (1)

1. a kind of preparation method of oyster polysaccharide, it is characterised in that carry out in accordance with the following steps successively：

A. fresh decladding oyster is gone after internal organ to carry out homogenized, is extracted with 10 ~ 15 times of boiling water of volume/mass 10min, intermittent stirring, filtering, obtains filtrate；

B. it is 4 ~ 5 to adjust filtrate pH, stands 12h, and 6000rpm centrifuges 20 min, collects supernatant；

C. it is 7 ~ 8 to adjust supernatant pH, is digested under the conditions of 20 ~ 30 DEG C with the complex enzyme of supernatant mass percent 0.5% ~ 1% 6h, the complex enzyme are the flavor of the marine alkaline proteinase of 6000u/mg, the papain of 6000u/mg, 15000 u/mg It is 3 that protease and 6000u/mg trypsase, which press unit of activity ratio,：3：2：2 ratios mix；

D. enzymolysis liquid 6000rpm is centrifuged into 20min, collects enzymolysis supernatant；

E. the ultrafiltration membrance filter that supernatant passes through molecular cut off 50kDa will be digested, trapped fluid after dried film obtains oyster Polysaccharide.