CN103936850B - Compound type degreasing liquid for extracting egg yolk immune globulin and application - Google Patents
Compound type degreasing liquid for extracting egg yolk immune globulin and application Download PDFInfo
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Abstract
The invention discloses compound type degreasing liquid for extracting egg yolk immune globulin and an application thereof. The compound type degreasing liquid for extracting the egg yolk immune globulin comprises the following components in percentage by weight: 0.05-0.2% of pectin, 0.01-0.15% of polysaccharide, 3-10% of ammonium sulfate and the balance of water, wherein polysaccharide is selected from one of carrageenan, carboxymethyl cellulose, methyl cellulose and dextran sulfate. The compound type degreasing liquid and ammonium sulfate are used for separating and purifying the IgY in the egg yolk, the extraction content of the obtained IgY is high, the purity of the IgY is relatively high, and the activity of the IgY is kept good. The IgY with the relatively high purity can be obtained by adopting the method only through twice centrifugation; the production cost and time are substantially reduced; favorable conditions are provided for large-scale industrial production of IgY extraction.
Description
Technical field
The present invention relates to protein separation technical field, extract for chicken yolk immunoglobulin in particular to a kind of
Compound go fat liquid and application.
Background technology
The bioactive substance value extracted from egg is high, has in aspects such as medicine, food, chemical industry, biologies
Widely purposes, is the focus in China's egg product scientific research exploitation over nearly 20 years.In recent years, with egg product research deeply and
The raising of isolation technics, people start protein components various in egg are studied, the γ-ovum especially in livitin
Yellow immunoglobulin (abbreviation igy), also known as chicken yolk antibody.Livitin is one of key protein in egg yolk, its tool
There is preparation process need not take a blood sample, the advantages of there is not cross reaction with the complement of rheumatoid factor and mammal, in disease
Prevention, food additive and medical domain are widely used.
Isolate and purify and in the technology of igy, generally adopt water dilution method, the organic solvent extracting sedimentation method, high molecular polymer to sink
The methods such as shallow lake method, ultrafiltration, chromatography are extracted and are separated igy.Although this several method can reach detached purpose to a certain extent,
But still there are shortcomings, although the igy purity obtaining as chromatography therein, ultrafiltration is higher, complex operation, time-consuming
Longer, operating condition is required also harsher, and the yield obtaining is not high；Method of organic solvent extraction, high molecular polymer
Although as polyethylene glycol precipitation yield is high but easily causing degeneration and the inactivation of protein molecule；Although water dilution method yield is relatively
High, safe but purity of protein that obtain is undesirable.Therefore, seek that simple to operate, with short production cycle, energy consumption is little, operation bar
Part is gentle, and the separation method being easy to amplify is always the important topic of research and development.
Content of the invention
It is an object of the invention to overcome exist in the existing technology isolating and purifying igy complex operation, time-consuming, low yield,
The defect of protein changeableness, provides a kind of simple to operate, low cost, yield high and protein active is intact for chicken
The compound of Yolk immunoglobulin extraction goes fat liquid and application.
For achieving the above object, designed by the present invention for chicken yolk immunoglobulin extract compound remove fat liquid,
Its each component and percentage by weight consist of: 0.05～0.2% pectin, 0.01～0.15% polysaccharide, 3～10% ammonium sulfate, balance of
Water；Described polysaccharide is selected from one of carrageenan, carboxymethyl cellulose, methylcellulose, dextran sulfate.
Above-mentioned compound go each component and percentage by weight in fat liquid preferably constitute for: 0.05～0.15% pectin, 0.05
～0.1% polysaccharide, 3～5% ammonium sulfate, balance of water.
Preferably, described polysaccharide is carrageenan, and weight percent content is 0.1%.
It is highly preferred that the weight percent content of described pectin be 0.1%, the weight percent content of ammonium sulfate be 3%.
Preferably, described compound go in fat liquid, also to include 5～20 μm of calcium chloride.Add cacl2Remove fat liquid
The effect either extracted amount or purity is all significantly improved of purification igy, possible cause is that calcium ion is mutual with polysaccharide
Effect, promotes the combination of polysaccharide and lipoprotein.
Compound go in fat liquid, also to include 10 μm of calcium chloride it is highly preferred that described.
Present invention also offers a kind of above-mentioned compound method going fat liquid to extract chicken yolk immunoglobulin, its step is such as
Under:
(1) by egg pretreatment, separate and obtain egg yolk liquid；
(2) egg yolk liquid is removed fat liquid by the ratio of volume 1:1～10 with being used for the compound of chicken yolk immunoglobulin extraction
Example mix homogeneously, adjusts ph to 4.5～6.0 simultaneously, and time of repose is 2～12h, obtains mixed liquor；
(3) by mixed liquor under the rotating speed of 8000～12000 × g frozen centrifugation 20min, take supernatant；
(4) add ammonium sulfate in supernatant while stirring, reach 20 to final concentration in supernatant for the ammonium sulfate～
35wt%, after being sufficiently stirred for, frozen centrifugation 25min under the rotating speed of 10000～14000 × g, collects precipitation；
(5) precipitation is dissolved in distilled water, after stirring, 4～10 DEG C of dialysis desalinations, lyophilization is dry to obtain final product chicken yolk immune ball
Albumen.
Preferably, in described step (1), the method for egg pretreatment is: chooses Fresh Egg, is separated using the yellow and white
Device, separates and obtains egg yolk, rinsed well egg yolk with distilled water, after being placed on filter paper rolling around and blotting, punctures egg yolk
Film, collects egg yolk liquid.
Preferably, described step (2) is that with being used for the compound of chicken yolk immunoglobulin extraction, egg yolk liquid is gone fat liquid
In the ratio mix homogeneously of volume 1:6～8, adjust ph to 4.8～5.2, time of repose is 6～8h, obtains mixed liquor simultaneously.
Main component in yolk is protein and fat, and its ratio is 1: 2.Most of protein is lipoprotein, exists
In yolk protein, water insoluble, only α, β, γ-livitin is water miscible, and igy is γ-livitin.Cause
This, the lipid how removing high concentration igy is isolated and purified it is critical that.This method first adopt anion polysaccharide with
The aqueous two-phase of inorganic salts configuration goes fat liquid to separate igy, then uses the technology path that ammonium sulfate is purified, obtain a kind of efficiently, safely,
The method of few, efficient, the safe igy separation and Extraction of low cost, step.The method of this separating-purifying has following several advantage:
First, polysaccharide, can be used as a kind of food additive used as a kind of material being widely present in nature, and its mild condition has
Beneficial to the native conformation keeping protein and biological activity；Second, anion polysaccharide can pass through ionic bond, electrostatic force, hydrogen
Key to form complex to remove most lipids and lipoprotein in egg yolk liquid with lipoprotein.The electric charge of polysaccharide is close simultaneously
Degree, conformational structure, more and sulfate group all can lead to the difference of igy separating effect to the decomposition constant of carbonyl, and multiple polysaccharide are joined
Close using mutually promoting, go fat effect be substantially better than traditional water dilution method and individually a kind of preparation of polysaccharide remove fat liquid；The
Three, as the one kind of middle neutral salt of saltouing, its temperature coefficient is little and dissolubility is cheap and easy to get greatly, and subsection efect is than other for ammonium sulfate
Salt good it is not easy to cause protein denaturation.Meanwhile, add a small amount of ammonium sulfate to going scalable ionic strength in fat liquid, promote many
Sugar and the precipitation of lipoprotein.
Beneficial effects of the present invention: for chicken yolk immunoglobulin extract compound to go fat liquid to separate with ammonium sulfate pure
Change igy in egg yolk, the igy extracted amount that obtains is high, purity is higher, activity keeps good, the method only needs the two times centrifugal can
Obtain the igy of higher degree, significantly reduce production cost and time, be that the large-scale industrial production that igy extracts provides
Advantage.Meanwhile, polysaccharide can form the polymer safety non-toxic of aqueous two-phase compared with other, or even can be used for food additive,
Be conducive to keeping native conformation and the biological activity of igy, purification igy out is particularly suited for food or the igy of pharmaceutical grade is big
Large-scale production.
Brief description
Fig. 1 is the electrophoretogram of the compound igy going fat liquid separation and Extraction of variety classes；
Fig. 2 is the western blots design sketch of the compound igy going fat liquid separation and Extraction of variety classes；
Fig. 3 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of a1 group；
Fig. 4 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of a2 group；
Fig. 5 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of a3 group；
Fig. 6 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of b1 group；
Fig. 7 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of b2 group；
Fig. 8 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of b3 group；
Fig. 9 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of c1 group；
Figure 10 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of c2 group；
Figure 11 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of c3 group；
Figure 12 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of d1 group；
Figure 13 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of d2 group；
Figure 14 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of d3 group；
Figure 15 is the compound effectiveness comparison figure going fat liquid that igy is isolated and purified of e group；
Figure 16 is the effectiveness comparison figure of different types of compound igy activity influence going fat liquid to isolate and purify.
Specific embodiment
Below in conjunction with the drawings and specific embodiments, the present invention is described in further detail.
Embodiment 1
It is formulated for the compound of chicken yolk immunoglobulin extraction and removes fat liquid, wherein polysaccharide is selected carrageenan, prepared one
The compound of the carrageenan containing different final concentrations for the group goes fat liquid, numbering a1 group, and one group of preparation contains answering of the pectin of different final concentrations
Mould assembly removes fat liquid, numbering a2 group, and one group of preparation contains the compound of the ammonium sulfate of different final concentrations and removes fat liquid, and numbering a3 group is above-mentioned
Compound go fat liquid be aqueous solution, its each component comprising and weight percent content as shown in the table:
| A1 group | Pectin | Carrageenan | Ammonium sulfate |
| a11 | 0.1% | 0.01% | 3% |
| a12 | 0.1% | 0.05% | 3% |
| a13 | 0.1% | 0.1% | 3% |
| a14 | 0.1% | 0.15% | 3% |
| a15 | 0.1% | 0.2% | 3% |
| A2 group | Pectin | Carrageenan | Ammonium sulfate |
| a21 | 0.01% | 0.1% | 3% |
| a22 | 0.05% | 0.1% | 3% |
| a23 | 0.1% | 0.1% | 3% |
| a24 | 0.15% | 0.1% | 3% |
| a25 | 0.2% | 0.1% | 3% |
| A3 group | Pectin | Carrageenan | Ammonium sulfate |
| a31 | 0.1% | 0.1% | 0 |
| a32 | 0.1% | 0.1% | 3% |
| a33 | 0.1% | 0.1% | 5% |
| a34 | 0.1% | 0.1% | 7% |
| a35 | 0.1% | 0.1% | 10% |
During preparation, above-mentioned soluble in water being sufficiently mixed of each component is uniformly obtained final product.
Embodiment 2
It is formulated for the compound of chicken yolk immunoglobulin extraction and removes fat liquid, wherein polysaccharide selects carboxymethyl cellulose,
Prepare the compound of one group of carboxymethyl cellulose containing different final concentrations and go fat liquid, numbering b1 group, one group of preparation contains different dense eventually
The compound of the pectin of degree goes fat liquid, numbering b2 group, and one group of preparation contains the compound of the ammonium sulfate of different final concentrations and removes fat liquid, compiles
Number b3 group, above-mentioned compound go fat liquid to be aqueous solution, its each component comprising and weight percent content are as shown in the table:
| B1 group | Pectin | Carboxymethyl cellulose | Ammonium sulfate |
| b11 | 0.1% | 0.01% | 3% |
| b12 | 0.1% | 0.05% | 3% |
| b13 | 0.1% | 0.1% | 3% |
| b14 | 0.1% | 0.15% | 3% |
| b15 | 0.1% | 0.2% | 3% |
| B2 group | Pectin | Carboxymethyl cellulose | Ammonium sulfate |
| b21 | 0.01% | 0.1% | 3% |
| b22 | 0.05% | 0.1% | 3% |
| b23 | 0.1% | 0.1% | 3% |
| b24 | 0.15% | 0.1% | 3% |
| b25 | 0.2% | 0.1% | 3% |
| B3 group | Pectin | Carboxymethyl cellulose | Ammonium sulfate |
| b31 | 0.1% | 0.1% | 0 |
| b32 | 0.1% | 0.1% | 3% |
| b33 | 0.1% | 0.1% | 5% |
| b34 | 0.1% | 0.1% | 7% |
| b35 | 0.1% | 0.1% | 10% |
During preparation, above-mentioned soluble in water being sufficiently mixed of each component is uniformly obtained final product.
Embodiment 3
It is formulated for the compound of chicken yolk immunoglobulin extraction and removes fat liquid, wherein polysaccharide selects methylcellulose, system
The compound of the standby one group methylcellulose containing different final concentrations goes fat liquid, numbering c1 group, and one group of preparation contains different final concentrations
The compound of pectin goes fat liquid, numbering c2 group, and the compound of ammonium sulfate that one group of preparation contains different final concentrations goes fat liquid, numbering c3
Group, above-mentioned compound go fat liquid be aqueous solution, its each component comprising and weight percent content as shown in the table:
| C1 group | Pectin | Methylcellulose | Ammonium sulfate |
| c11 | 0.1% | 0.01% | 3% |
| c12 | 0.1% | 0.05% | 3% |
| c13 | 0.1% | 0.1% | 3% |
| c14 | 0.1% | 0.15% | 3% |
| c15 | 0.1% | 0.2% | 3% |
| C2 group | Pectin | Methylcellulose | Ammonium sulfate |
| c21 | 0.01% | 0.1% | 3% |
| c22 | 0.05% | 0.1% | 3% |
| c23 | 0.1% | 0.1% | 3% |
| c24 | 0.15% | 0.1% | 3% |
| c25 | 0.2% | 0.1% | 3% |
| C3 group | Pectin | Methylcellulose | Ammonium sulfate |
| c31 | 0.1% | 0.1% | 0 |
| c32 | 0.1% | 0.1% | 3% |
| c33 | 0.1% | 0.1% | 5% |
| c34 | 0.1% | 0.1% | 7% |
| c35 | 0.1% | 0.1% | 10% |
During preparation, above-mentioned soluble in water being sufficiently mixed of each component is uniformly obtained final product.
Embodiment 4
It is formulated for the compound of chicken yolk immunoglobulin extraction and removes fat liquid, wherein polysaccharide selects dextran sulfate, system
The compound of the standby one group dextran sulfate containing different final concentrations goes fat liquid, numbering d1 group, and one group of preparation contains different final concentrations
The compound of pectin goes fat liquid, numbering d2 group, and the compound of ammonium sulfate that one group of preparation contains different final concentrations goes fat liquid, numbering d3
Group, above-mentioned compound go fat liquid be aqueous solution, its each component comprising and weight percent content as shown in the table:
| D1 group | Pectin | Dextran sulfate | Ammonium sulfate |
| d11 | 0.1% | 0.01% | 3% |
| d12 | 0.1% | 0.05% | 3% |
| d13 | 0.1% | 0.1% | 3% |
| d14 | 0.1% | 0.15% | 3% |
| d15 | 0.1% | 0.2% | 3% |
| D2 group | Pectin | Dextran sulfate | Ammonium sulfate |
| d21 | 0.01% | 0.1% | 3% |
| d22 | 0.05% | 0.1% | 3% |
| d23 | 0.1% | 0.1% | 3% |
| d24 | 0.15% | 0.1% | 3% |
| d25 | 0.2% | 0.1% | 3% |
| D3 group | Pectin | Dextran sulfate | Ammonium sulfate |
| d31 | 0.1% | 0.1% | 0 |
| d32 | 0.1% | 0.1% | 3% |
| d33 | 0.1% | 0.1% | 5% |
| d34 | 0.1% | 0.1% | 7% |
| d35 | 0.1% | 0.1% | 10% |
During preparation, above-mentioned soluble in water being sufficiently mixed of each component is uniformly obtained final product.
Embodiment 5
It is formulated for the compound of chicken yolk immunoglobulin extraction and removes fat liquid, wherein polysaccharide is selected carrageenan, prepared one
The cacl containing different final concentrations for the group2Compound remove fat liquid, numbering e group, above-mentioned compound go fat liquid to be aqueous solution, it comprises
Each component and content are as shown in the table:
| E group | Pectin | Carrageenan | Ammonium sulfate | cacl2 |
| e1 | 0.1wt% | 0.1wt% | 3wt% | 0 |
| e2 | 0.1wt% | 0.1wt% | 3wt% | 1μm |
| e3 | 0.1wt% | 0.1wt% | 3wt% | 5μm |
| e4 | 0.1wt% | 0.1wt% | 3wt% | 10μm |
| e5 | 0.1wt% | 0.1wt% | 3wt% | 15μm |
| e6 | 0.1wt% | 0.1wt% | 3wt% | 20μm |
During preparation, above-mentioned soluble in water being sufficiently mixed of each component is uniformly obtained final product.
Test example 1
By the various embodiments described above be obtained compound go fat liquid that egg yolk is extracted, respectively obtain corresponding igy, carry
Take step as follows:
First, sample pre-treatments: choose Fresh Egg, using egg-white egg-yellow separator, separate and obtain egg yolk, will with distilled water
Egg yolk is rinsed well, is placed in rolling around on filter paper and blots, and punctures membrane of yolk afterwards, collects egg yolk liquid.
2nd, the isolating and purifying of protein:
(1) acquisition of water-soluble component
Egg yolk liquid is separately added into the above-mentioned compound of 5 times of volumes and removes fat liquid, stir, adjust ph with 1m hcl and arrive
5.0,4 DEG C stand overnight, and after 10000 × g frozen centrifugation 20min, obtain the clarified supernatant i.e. water-soluble component containing igy (wsf).
(2) ammonium sulfate precipitation method purification igy
Add ammonium sulfate while stirring in the supernatant of above-mentioned acquisition, reach 35% to its final concentration, after being sufficiently stirred for,
12000 × g frozen centrifugation 25min, collects precipitation.Precipitation is redissolved the distilled water of certain volume, load bag filter (retention molecule
Amount 8000-14000kd) in, stirring, 4 DEG C of dialysis desalinations, lyophilization is dry to obtain final product igy.
Inspection example
Qualitative, purity, the extracted amount of target protein, the mensure of activity
(1) sodium dodecyl sulfate-polyacrylamide gel electrophoresiss (sds-page) are adopted to measure analysis target protein
Molecular weight, analysis condition used is: 5% concentration glue, 10% separation gel.By above-mentioned corresponding embodiment a13 to be determined,
The lyophilized powder of the igy of b13, c13, d13 is dissolved in distilled water, takes 80 microlitres of solution after fully dissolving, and adds 20 microlitres of egg
White sample-loading buffer (5 times of concentration), after mixing, boiling water bath heats about 5 minutes, takes 10 microlitres to be injected in loading wells after cooling, dense
Contracting glue constant voltage 80v, separation gel constant electrophoresis 120v carries out electrophoresis.After the completion of electrophoresis, first by fixative (50% ethanol,
10% acetic acid) gel is fixed 30 minutes, (r-250 content 0.1%, contains simultaneously then to use Coomassie brilliant blue r-250 solution
25% ethanol, 8% acetic acid) dye 30 minutes at 45 DEG C, it is transferred in destaining solution (25% ethanol and 8% acetic acid) de- after the completion of dyeing
Color, to background color takes off, band is clear.The electrophoretogram of gained is as shown in Figure 1.
(2) adopt western blots, with the anti-chicken igg of two anti-horseradish peroxidase mark rabbits to embodiment a13, b13,
The corresponding igy of c13, d13 is hybridized, qualitative objective albumen.Western blots result such as Fig. 2 shows, these four are compound
Go fat liquid extract separate the target protein that obtains can the successfully anti-chicken igg hybridization with two anti-horseradish peroxidase mark rabbits, enter
The target protein that one step demonstrates this experiment purification is igy.
(3) igy corresponding to each embodiment above-mentioned, adopts using gel-pro analyzer software according to sds-page electricity
The purity of igy in sample is extrapolated in swimming map analysis.In sample, the content of igy passes through light absorption value under determination sample 280nm, 1.33 works
Measure for molar extinction coefficient.Calculated by formula a=ε bc.
The compound comparative result removing the corresponding igy of fat liquid a13, b13, c13, d13 of homopolysaccharide is not shown in Table 1.
The compound result of calculation going fat liquid to isolate and purify igy extracted amount, purity of table 1 variety classes
Upper table shows, in the case of other conditions identical, using the compound igy purity going fat liquid to extract of carrageenan
Higher.
(4) different the compound of group go fat liquid to igy extraction effect as shown in Fig. 3～15, and in figure shows, a13 is compound
Go fat liquid (0.1% pectin, 0.1% carrageenan and 3% ammonium sulfate) effect preferably, on this basis, add calcium chloride optimization of C/C composites,
When addition is 10, best results.
(5) igy activity is using enzyme-linked immunoassay method (elisa) detection.A series of standard substance with known activity and concentration
Light absorption value draw standard curve, guarantor in sample is calculated according to the light absorption value of a1 group, b1 group, c1 group, d1 group respectively corresponding igy
Hold the igy content of activity.As shown in figure 16, in this several anion polysaccharide, compound from carrageenan goes fat liquid the warmest
With.
Claims (9)
1. a kind of remove fat liquid for the compound of chicken yolk immunoglobulin extraction, its each component and percentage by weight consist of:
0.05～0.2% pectin, 0.01～0.15% polysaccharide, 3～10% ammonium sulfate, balance of water；Described polysaccharide is selected from carrageenan, carboxylic
One of methylcellulose, methylcellulose, dextran sulfate.
2. according to claim 1 be used for chicken yolk immunoglobulin extract compound remove fat liquid it is characterised in that: each group
Divide and percentage by weight consists of: 0.05～0.15% pectin, 0.05～0.1% polysaccharide, 3～5% ammonium sulfate, balance of water.
3. according to claim 1 be used for chicken yolk immunoglobulin extract compound remove fat liquid it is characterised in that: described
Polysaccharide is carrageenan, and weight percent content is 0.1%.
4. according to claim 3 be used for chicken yolk immunoglobulin extract compound remove fat liquid it is characterised in that: described
The weight percent content of pectin is 0.1%, the weight percent content of ammonium sulfate is 3%.
5. a kind of remove fat liquid for the compound of chicken yolk immunoglobulin extraction, its each group is divided into: 0.1wt% pectin,
0.1wt% carrageenan, 3wt% ammonium sulfate, 5～20 μm of calcium chloride, balance of water.
6. according to claim 5 be used for chicken yolk immunoglobulin extract compound remove fat liquid it is characterised in that: described
Calcium chloride content is 10 μm.
7. a kind of using the method going fat liquid to extract chicken yolk immunoglobulin compound described in claim 1, its step is as follows:
(1) by egg pretreatment, separate and obtain egg yolk liquid；
(2) egg yolk liquid is gone fat liquid to mix in the ratio of volume 1:1～10 with being used for the compound of chicken yolk immunoglobulin extraction
Close uniformly, adjust ph to 4.5～6.0 simultaneously, time of repose is 2～12h, obtains mixed liquor；
(3) by mixed liquor under the rotating speed of 8000～12000 × g frozen centrifugation 20min, take supernatant；
(4) add ammonium sulfate in supernatant while stirring, reach 20～35wt% to final concentration in supernatant for the ammonium sulfate,
After being sufficiently stirred for, frozen centrifugation 25min under the rotating speed of 10000～14000 × g, collects precipitation；
(5) precipitation is dissolved in distilled water, after stirring, 4～10 DEG C of dialysis desalinations, lyophilization obtains final product chicken yolk immunoglobulin.
8. according to claim 7 compound go fat liquid extract chicken yolk immunoglobulin method it is characterised in that: described
In step (1), the method for egg pretreatment is: chooses Fresh Egg, using egg-white egg-yellow separator, separates and obtain egg yolk, with steaming
Egg yolk is rinsed well by distilled water, after being placed on filter paper rolling around and blotting, punctures membrane of yolk, collects egg yolk liquid.
9. according to claim 7 compound go fat liquid extract chicken yolk immunoglobulin method it is characterised in that: described
Step (2) is to go fat liquid to mix in the ratio of volume 1:6～8 with being used for the compound of chicken yolk immunoglobulin extraction egg yolk liquid
Close uniformly, adjust ph to 4.8～5.2 simultaneously, time of repose is 6～8h, obtains mixed liquor.
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| CN103936850A (en) | 2014-07-23 |
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