CN110343190A - A kind of synchronous method for extracting hericium erinaceum polysaccharide and protein - Google Patents
A kind of synchronous method for extracting hericium erinaceum polysaccharide and protein Download PDFInfo
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- CN110343190A CN110343190A CN201910772489.7A CN201910772489A CN110343190A CN 110343190 A CN110343190 A CN 110343190A CN 201910772489 A CN201910772489 A CN 201910772489A CN 110343190 A CN110343190 A CN 110343190A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/30—Extraction; Separation; Purification by precipitation
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0003—General processes for their isolation or fractionation, e.g. purification or extraction from biomass
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
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Abstract
The invention discloses a kind of synchronous methods for extracting hericium erinaceum polysaccharide and protein, using homogenate extraction method, polysaccharide and protein in combined extracting Hericium erinaceus；Using protein near isoelectric point the characteristics of coagulation, by the protein mixed liquor of extraction through isoelectric precipitation, polysaccharide and Separation of Proteins are realized, technological parameter optimizes, and polysaccharide and extraction rate of protein and purity is high are conducive to promote and apply.
Description
Technical field
The invention belongs to effective component extractive technique fields, and in particular to a kind of synchronous to extract hericium erinaceum polysaccharide and protein
Method.
Background technique
Hericium erinaceus (Hericium erinaceus) belong to fungus circle Basidiomycota monkey mushroom mesh, have significantly to disease of digestive system
Curative effect can effectively treat gastric ulcer, chronic gastritis and duodenal ulcer, there is preferable inhibition to make stomach, intestinal canal tumour
With there is the title of " mountain delicacy hedgehog hydnum, seafood delights shark's fin ".Hericium erinaceus is rich in polysaccharide, protein, cellulose, amino acid, including 8 kinds of human bodies
Essential amino acid.Hericium erinaceum polysaccharide has protective effect to stomach lining, can promote digestion, also has and improves immunity, anti-aging, drop
Blood glucose and other effects.The common extracting method of hericium erinaceum polysaccharide has hot water to extract, soda acid extracts, enzyme extracts and microwave radiation exaraction
Deng.Traditional extraction process temperature is high, speed is slow, destroys to heat-sensitive ingredients big.In recent years, homogenate extraction technology is widely used in
In terms of natural product extraction, it utilizes the mechanical shear stress and mixing power of high speed, quickly and effectively destroys plant cell tissue, leads to
Crossing local decompression's osmosis diffuses to effective component in solvent rapidly, has the advantages that extraction rate is fast, temperature is low etc., can have
Effect ground retains heat-sensitive ingredients and activity.
Summary of the invention
The purpose of the present invention is to provide a kind of synchronous methods for extracting hericium erinaceum polysaccharide and protein, using homogenate extraction
Method, polysaccharide and protein in combined extracting Hericium erinaceus；Using protein near isoelectric point the characteristics of coagulation, by the albumen of extraction
Matter mixed liquor realizes polysaccharide and Separation of Proteins through isoelectric precipitation, and technological parameter optimizes, polysaccharide and extraction rate of protein and
Purity is high is conducive to promote and apply.
To achieve the above object, the present invention adopts the following technical scheme:
A kind of synchronous method for extracting hericium erinaceum polysaccharide and protein, extracts polysaccharide and protein using flash method, using etc. electricity
Point separating polyose and protein.
The following steps are included:
(1) 40 DEG C of hericium erinaceus fruiting body 16 h of baking are crushed, and -20 DEG C store for future use；
(2) 10 g Hericium erinaceus powder are taken, distilled water is added, are extracted using flash method, are filtered, centrifugation obtains hericium erinaceum polysaccharide and egg
White matter extracts mixed liquor；
(3) adjustment mixed liquor pH value is centrifugally separating to obtain the supernatant and protein precipitation of Thick many candies to isoelectric point.
Hericium erinaceus powder size 40-120 mesh extracts voltage 100-200V, extraction time 20-120 s, liquid-to-solid ratio 14-22:
1 mL/g, isoelectric pH=3.65.
The beneficial effects of the present invention are: use homogenate extraction method, polysaccharide and protein in combined extracting Hericium erinaceus；It utilizes
Protein the characteristics of coagulation, by the protein mixed liquor of extraction through isoelectric precipitation, realizes polysaccharide and albumen near isoelectric point
Matter separation, technological parameter optimization, polysaccharide and extraction rate of protein and purity is high, are conducive to promote and apply.
Detailed description of the invention
Fig. 1 is influence curve figure of the extraction time to hericium erinaceum polysaccharide and protein yield；
Fig. 2 is influence curve figure of the liquid-to-solid ratio to hericium erinaceum polysaccharide and protein yield；
Fig. 3 is influence curve figure of the powder granularity to hericium erinaceum polysaccharide and protein yield；
Fig. 4 is to extract voltage to the influence curve figure of hericium erinaceum polysaccharide and protein yield；
Fig. 5 is the SDS-PAGE electrophoretic analysis result of Hericium erinaceus protein；
Fig. 6 is the infrared spectrogram of hericium erinaceum polysaccharide (HEP).
Specific embodiment
In order to make content of the present invention easily facilitate understanding, With reference to embodiment to of the present invention
Technical solution is described further, but the present invention is not limited only to this.
Synchronous extraction process
Sample treatment: hericium erinaceus fruiting body toasts 16 h in heated-air drying baking oven under the conditions of 40 DEG C, and pulverizer is crushed,
It is stored for future use under the conditions of -20 DEG C.
It extracts process: taking 10 g Hericium erinaceus sample powders that distilled water is added by liquid-to-solid ratio, mentioned under setup parameter using flash method
It takes, is repeatedly centrifuged through filter-cloth filtering, adjust hericium erinaceum polysaccharide and Protein Extraction mixed liquor pH value to isoelectric point (pH 3.65),
It is centrifugally separating to obtain the supernatant and protein precipitation of Thick many candies.
Protein identification
Hericium erinaceum polysaccharide and protein mixed liquor are centrifuged obtained protein precipitation and are redissolved to obtain turbid solution using distilled water, then pass through from
The isolated supernatant of the heart is used for sodium dodecyl sulfate-polypropylene acrylamide gel (SDS-PAGE) electrophoresis.
Polysaccharide identification
It is Hericium erinaceus Thick many candies liquid that hericium erinaceum polysaccharide and protein mixed liquor, which are centrifuged obtained supernatant, and 1/3 liquor capacity is added
The decoloration of D315 macroreticular resin, 4 h of magnetic agitation takes filtrate after filtering.Filtrate moves in 2500 Da bag filters and dialyses, and 4 DEG C
Dialyse 48 h in distilled water, and it is primary that every 4-5 h changes water.It is put into 12 h of precooling in -4 DEG C of refrigerators after rotary evaporation concentration, then is passed through
Freeze-drying obtains HEP powder.It takes 3 mg HEP powder and 0.3 g KBr ground and mixed uniform, suppresses thin slice, carry out Fourier
Transform infrared spectroscopy analysis, scanning wavelength are 4000-400 cm-1。
Single factor experiment
1. influence of the extraction time to polysaccharide and protein yield
Influence of the extraction time to hericium erinaceum polysaccharide and protein yield is as shown in Figure 1, as seen from the figure as extraction time is from 20
S increases to 60 s, and hericium erinaceum polysaccharide and protein yield all dramatically increase.Extraction time is more than hericium erinaceum polysaccharide after 60 s
Yield decline, it may be possible to since the time is longer, the energy that homogenate extraction cutter head is saved bit by bit is more and generates heat, and causes the decomposition of polysaccharide,
Yield is caused to reduce；Protein yield is dramatically increased from 20-60 s, and protein variation of yield is little after 60 s；Time is more than
Homogenate extraction reaches balance after 60 s, and it is little on the influence of the yield of protein to extend the time；Therefore extraction time is advisable with 60 s.
2. influence of the liquid-to-solid ratio to polysaccharide and protein yield
As shown in Figure 2, hericium erinaceum polysaccharide yield is reduced afterwards as the increase of liquid-to-solid ratio first increases, and protein yield gradually increases.
Polysaccharide yield reduces after liquid-to-solid ratio 18:1 mL/g, it may be possible to since liquid-to-solid ratio is excessively high, cause dilution to solution, reduce
The touch opportunity of Hericium erinaceus and the shearing of flash cutter head, while excessively high liquid-to-solid ratio will increase subsequent concentration difficulty；Therefore liquid material
Than taking 18:1 mL/g preferable.
3. influence of the powder granularity to polysaccharide and protein yield
From the figure 3, it may be seen that polysaccharide yield first increases and reduces afterwards with the reduction of powder granularity, albumen yield slowly declines.Due to monkey
Head mushroom dry powder crosses that sieve pore is excessively tiny, so that the partial size of effective component becomes smaller in hericium erinaceum polysaccharide, causes surface that can increase and is inhaled
It is attached to the surface of solids or subtle intergranular stick to each other, is allowed to not readily dissolve, polysaccharide yield is caused to reduce；Powder granularity is smaller,
Protein yield is lower, it may be possible to which, since the emulsification enhancing of protein and fat causes, change effect reaches flat after 100 mesh
Weighing apparatus, yield no longer significantly reduce；Therefore powder granularity takes 100 mesh of sieving to be advisable.
4. extracting influence of the voltage to polysaccharide and protein yield
As shown in Figure 4, with the increase of homogenate extraction voltage, hericium erinaceum polysaccharide yield is gradually increased, and protein must take the lead in reducing
After rise.Homogenate extraction voltage increase will lead to that flash cutter head revolving speed is too fast, and mechanical energy is converted into thermal energy, lead to solution temperature liter
Height, solution colour are deepened, and cause the denaturation of polysaccharide and protein, being allowed to lose activity contradicts with test objective.Comprehensively consider
Polysaccharide and protein yield, extract voltage is advisable with 100 V.
Response surface optimization
The optimum condition of model prediction homogenate extraction hericium erinaceum polysaccharide is obtained by software are as follows: 80 s of extraction time, liquid-to-solid ratio
16.64:1 mL/g, 80 mesh of powder granularity, hericium erinaceum polysaccharide yield is 5.577% under this condition.In view of practical operation is to above-mentioned
Model prediction optimal conditions is modified, final polysaccharide optimal conditions are as follows: 80 s of extraction time, liquid-to-solid ratio 17:1 mL/g, powder
80 mesh of granularity carries out 3 parallel tests verifying with this condition, and polysaccharide yield is 5.842 ± 0.159% in Hericium erinaceus, with theory
Predicted value 5.577% is very close, illustrates to carry out process optimization with certain reality to the extraction of hericium erinaceum polysaccharide with the model
Border operability.
Product identification
1. protein
The Hericium erinaceus protein of homogenate extraction is subjected to SDS-PAGE electrophoretic analysis, as a result as shown in Figure 5.As seen from the figure, exist
66.4 kDa and 44.3 kDa nearby respectively have an obvious band, illustrate that protein content is higher.It was found from Unipot database
(https: //www.uniprot.org), Hericium erinaceus protein molecular weight are mainly pigment near 66.4 kDa, and slightly
It is mainly tubulin and ribalgilase lower than 44.3 kDa；The relative molecular mass of most protein is lower than 14.3 kDa,
As a result with Humans etc.[18]What is measured is similar.Related Hericium erinaceus albumen Quality Research is less both at home and abroad, and Lee etc. is thick to Hericium erinaceus
Albumen carries out SDS-PAGE electrophoretic analysis, it is found that the relative molecular mass of a small amount of fungal immunomodulatory protein is 10-14 kDa.
Humans etc. is the study found that acidic protein of the relative molecular mass lower than 14.3 kDa is primarily involved in the Hericium erinaceus albumen extracted
Translation, Ribosome Structure and biology generation, transporting pathway.
2. polysaccharide
The infrared spectroscopy of the hericium erinaceum polysaccharide (HEP) of homogenate extraction is as shown in Figure 6.HEP is in 3400 cm-1Nearby have than wider
Strong absworption peak, 2930 cm-1Nearby there are weak absorbing peak, 1400-1200 cm-1Weak absorbing peak belong to the characteristic absorption of polysaccharide
Peak.3400 cm-1Wider peak is the stretching vibration of hydroxyl O-H, 2925 cm-1Locating weaker absorption peak is the flexible of methyl C-H
Vibration[21]；1401 cm-1The peak at place is the common stretching vibration of C-H and C-O；1151 cm-1The peak at place is that the side chain of C-OH is flexible
Vibration；1078 cm-1With 1038 cm-1Peak be glycosidic bond C-O-C stretching vibration, 918 cm-1The absorption peak at place is due to D-
The asymmetric stretching vibration of pyranoid ring generates.Li Shanshan etc. forms hericium erinaceum polysaccharide yield, the monosaccharide of different extraction processes
Studies have shown that Hericium erinaceus is made of glucose, galactolipin, fucose and mannose.This research institute obtains HEP through hydrolysis and derivatization
Laggard promoting the circulation of qi analysis of hplc is handled, other than eight kinds of common monosaccharide, there are also the unknown peak with potential researching value, cards
The active constituent that flash method extracts energy effective protection natural products is illustrated.
The foregoing is merely presently preferred embodiments of the present invention, all equivalent changes done according to scope of the present invention patent with
Modification, is all covered by the present invention.
Claims (4)
1. a kind of synchronous method for extracting hericium erinaceum polysaccharide and protein, it is characterised in that: extract polysaccharide and egg using flash method
White matter, using isoelectric point separating polyose and protein.
2. according to the method described in claim 1, it is characterized by comprising following steps:
(1) 40 DEG C of hericium erinaceus fruiting body 16 h of baking are crushed, and -20 DEG C store for future use；
(2) 10 g Hericium erinaceus powder are taken, distilled water is added, are extracted using flash method, are filtered, centrifugation obtains hericium erinaceum polysaccharide and egg
White matter extracts mixed liquor；
(3) adjustment mixed liquor pH value is centrifugally separating to obtain the supernatant and protein precipitation of Thick many candies to isoelectric point.
3. according to the method described in claim 2, it is characterized by: Hericium erinaceus powder size 40-120 mesh, extracts voltage 100-
200V, extraction time 20-120 s, liquid-to-solid ratio 14-22:1 mL/g, isoelectric pH=3.65.
4. according to the method described in claim 3, it is characterized by: 80 mesh of Hericium erinaceus powder size, extracts voltage 100V, extraction
80 s of time, liquid-to-solid ratio 17:1 mL/g, isoelectric pH=3.65.
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| CN113951367A (en) * | 2021-11-16 | 2022-01-21 | 安徽中志科技有限公司 | Edible fungus protein powder with high organic selenium content, preparation method and application |
| CN115197339A (en) * | 2022-07-13 | 2022-10-18 | 佛山中国发明成果转化研究院 | Method for combined extraction of protein and polysaccharide |
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