CN104327184A - Preparation method of yolk globulin powder - Google Patents
Preparation method of yolk globulin powder Download PDFInfo
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- CN104327184A CN104327184A CN201410508752.9A CN201410508752A CN104327184A CN 104327184 A CN104327184 A CN 104327184A CN 201410508752 A CN201410508752 A CN 201410508752A CN 104327184 A CN104327184 A CN 104327184A
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Abstract
The invention discloses a preparation method of a yolk globulin powder. The preparation method includes following steps: (1) dissolving yolk in deionized water, stirring the yolk and performing precipitation to extract a raw liquid; (2) performing salting-out precipitation to the raw liquid for removing residual fat, precipitating protein and adding water for reducing the protein, and performing plate-frame filtration to remove impurities for obtaining a clear liquid; (3) purifying the clear liquid through membrane separation for desalination and dehydration; and (4) performing micro-filtration for sterilization, and performing low-temperature spray drying to obtain the yolk globulin powder. The method can be carried out continuously. A production process is easy to control. A finally product is stable in quality and performance, is low in cost, can achieve industrial production and is easy to popularize.
Description
Technical field
The present invention relates to a kind of preparation method of livetin powder, be specifically related to the techniques such as one adopts water dilution method combination to saltout and extract active substance Yolk immune globulin in yolk, and prepared the method for livetin powder by ultrafiltration purification.
Background technology
Containing abundant biologically active substance and the material, particularly yolk mesolecithal solid content about 50% with nutritive value in birds, beasts and eggs, protein about 16%, lipid about 32%, inorganics and carbohydrate are respectively 2% and 1%.Vitellin(Vt) majority is phosphorprotein and lipoprotein, is probably divided into low-density lipoprotein, ovoglobulin and phosvitin and high-density lipoprotein (HDL).The exploitation of current yolk mainly concentrate on the composition such as Oil of egg yolk, Ovum Gallus domesticus Flavus lecithin, exploitation for Yolk immune globulin also has certain report, in yolk, immunoglobulin (Ig) can improve people's immunizing power, the gastrointestinal tract disease such as prevention and therapy gastrointestinal tract infection, diarrhoea, be particularly suitable for the immunocompromised persons such as baby, immune deficient patients and the elderly, have wide range of applications.
At present for the lab scale that the extraction of Yolk immune globulin is on basis, laboratory with exploitation, the research of synthesization exploitation is less, few at the research report of commercial production scale pilot scale and lab scale.The technology commonly used in its preparation process comprise saltout, chromatography, freezing, centrifugal, ultrafiltration, supercritical extraction, micro-filtration, dialysis, the technology such as freeze-drying.But easily cause protein inactivation and sex change in aforesaid method ubiquity production process and cause the defect of character instability, and production process step is many, batch is little, reagent consumption is large, the rate of recovery is low and cause the problem that is unfavorable for that technology is produced.
Summary of the invention
The object of the invention is to overcome above-mentioned defect in prior art, the techniques such as one adopts water dilution method combination to saltout are provided to extract active substance Yolk immune globulin in yolk, and the method for livetin powder is prepared by ultrafiltration purification, the method good separating effect, product purity are high, and operating procedure is simple, step is simple, can continuity operation, be easy to amplificationizations production thus be suitable for suitability for industrialized production.
The scheme solved the problems of the technologies described above of the present invention is: a kind of preparation method of livetin powder, and its concrete grammar comprises:
(1) get yolk, by yolk: the weight kg of deionized water: volume L adds deionized water than 1 ~ 5:1, after stirring 3 ~ 10 minutes, input settling tank, leave standstill 4 ~ 6 hours, obtain supernatant liquor;
(2) supernatant liquor is extracted into stirred pot, makes supernatant liquor be in whipped state, stir speed (S.S.) is 50 ~ 70r/min, in supernatant liquor, 4:1 ratio adds salts solution by volume simultaneously, close after salts solution adds completely and stir, supernatant liquor leaves standstill 3 ~ 4 hours, makes albumen precipitation;
(3) after albumen precipitates completely, upper liquid degrease is extracted, in upper liquid: deionized water volume ratio 1:5 ratio adds deionized water reduction albumen, obtains reducing protein liquid, prepares Plate Filtration;
(4) be mounted on plate-and-frame filter press by filter cloth, after diatomite precoating, the protein liquid that will reduce is by Plate Filtration, and remove impurity, permeate is clear liquor;
(5) be separated through ultra-filtration membrane desalination by clear liquor, molecular weight cut-off is 50000 dalton;
(6) concentrated solution is carried out low temperature micro-filtration degerming, microfiltration membrane adopts 0.22 micron pore size.
(7) MF permeate liquid is carried out drying under low-temperature spray drying tower, obtain major product livetin powder.
Preferably, in step (1), deionized water temperature is 0 ~ 4 DEG C, specific conductivity≤0.3;
Preferably, remain egg yolk liquid when supernatant liquor being extracted in step (2) stirred pot and can prepare yolk powder; Salts solution adopts saturated sodium-chloride salts solution/ammoniumsulphate soln, and saturation concentration is 33%, churning time 5-10min;
Preferably, when step (3) carries out Plate Filtration, filter pressure is 0.35-0.6MPa;
Preferably, in step (4) by Penetration ration in the smooth installation of 350---400 order filter cloth on a filter press, first with air in excavator in water cycle to pressure filter, 150 order fineness diatomite, at 0.35-0.5MPa, all should be coated on filter cloth by top hole pressure in advance;
Preferably, in step (5), water cycle desalination can also can be added in ultra-filtration membrane desalination sepn process, or on-line monitoring;
Preferably, when in step (6), low temperature micro-filtration is degerming, control temperature is below 10 DEG C, and pressure difference is 0.02-0.08MPa; Or below control temperature 8-16 DEG C, pressure difference is 0.04-0.06Mpa;
Preferably, when carrying out drying under low-temperature spray drying tower in step (7), inlet temperature 100-140 DEG C, temperature out 60-80 DEG C, temperature of charge 20-40 DEG C, mass flow 6-8%.
In technical solution of the present invention, certain applications technology such as micro-filtration can adopt the methods such as Plate Filtration to substitute, simultaneously freeze drying technology high value-added product as drug development utilize in benefit better, and the technology such as spraying dry can be adopted to complete for gourmet food batching.
Membrane separation purification desalting and dewatering, film is the material with selective separation function, utilizes that the selective separation of film realizes the separation of the different components of feed liquid, purifying, concentrated process are called membrane sepn.It is from the different of traditional filtering, and film can be separated in molecular range, and this process is a kind of physical process, and do not need change and interpolation auxiliary agent that phase occurs, separation efficiency is high, energy consumption is low.Micro-filtration (MF) is also known as millipore filtration, its ultimate principle is sieve aperture sepn process, according to the separation characteristic of millipore filtration, the range of application of millipore filtration mainly retains particulate, bacterium and other pollutents from gas phase and liquid phase, to reach purification, separation, concentrated object, the material of microfiltration membrane is divided into organic and inorganic two large classes, and the former is as cellulose acetate, polypropylene, polycarbonate, polysulfones, polymeric amide etc., and the latter is as pottery and metal etc.
Spraying dry is by contacting with drying medium after spraying gun atomization the liquid containing solid substance, complete evaporation drying in short period of time and obtain the operation of dry product, low temperature spray drying can be continuous to the processing of livetin powder, throughput is large, time is short, and can the advantages such as original composition does not destroy be kept, thus demonstrate very large development potentiality.Spray-drying process can be divided into and first feed liquid is atomized into droplet, and droplet contacts mixing and flowing and most end form, becomes the stages such as particulate product with drying medium.Air is by strainer and well heater, and enter the air distributor at moisture eliminator top, warm air evenly enters moisture eliminator in the shape of a spiral.Feed liquid by material fluid bath through filter by the centrifugal atomizer being pumped to moisture eliminator top, make feed liquid be sprayed into minimum misty liquid droplets, feed liquid is with warm air and flow and contact, moisture evaporates rapidly, at very short time inner drying finished product, finished product is discharged by bottom and cyclonic separator, and waste gas is discharged by blower fan.
The inventive method can be carried out in serialization, and reaction conditions is easy to control, end product quality stable performance, and equipment used is simple, and little by enzyme amount, cost is low, is easy to promote.
Embodiment
The present invention is further described below in conjunction with embodiment.
Embodiment 1
Get 4 kilograms, yolk, add the deionized water of 4 times of volumes, stir and input settling tank after 5 minutes, leave standstill 4 hours, obtain supernatant liquor, supernatant liquor is extracted into stirred pot, residue egg yolk liquid can prepare yolk powder; Supernatant liquor is in whipped state, and stir speed (S.S.) is 50 ~ 70r/min, and by volume/weight ratio 4:1 ratio adds salts solution, and close after salts solution adds completely and stir, supernatant liquor leaves standstill 3 ~ 4 hours, makes albumen precipitation.After albumen precipitates completely, extract upper liquid degrease.In upper liquid: deionized water volume ratio 1:5 ratio adds deionized water reduction albumen, prepares Plate Filtration.Be mounted to by filter cloth on plate-and-frame filter press, after diatomite precoating, the protein liquid that will reduce is by Plate Filtration, and filter pressure 0.35-0.6Mpa, removing impurity permeate is clear liquor.Be separated through ultra-filtration membrane desalination by clear liquor, molecular weight cut-off is 50000 dalton.Adopt 0.22 μm of filter membrane to carry out low temperature micro-filtration to concentrated solution degerming, temperature controls below 10 DEG C, and pressure difference is 0.02-0.08Mpa.MF permeate liquid is carried out drying under low-temperature spray drying tower, inlet temperature 100-140 DEG C, temperature out 60-80 DEG C, temperature of charge 20-40 DEG C, mass flow 6-8%.Obtain major product livetin powder.
Embodiment 2
Get 20 kilograms, yolk, add the deionized water of 4 times of volumes, stir and input settling tank after 10 minutes, leave standstill 6 hours, obtain supernatant liquor, after supernatant liquor is removed, residue egg yolk liquid is for the preparation of yolk powder; Adding ammonium sulfate (sodium-chlor) to saturation concentration is 33%, is that 50 ~ 70r/min stirs 5-10min with speed.Close after salts solution adds completely and stir, supernatant liquor leaves standstill 3 ~ 4 hours, makes albumen precipitation.After albumen precipitates completely, extract upper liquid degrease.In upper liquid: deionized water volume ratio 1:5 ratio adds deionized water reduction albumen, prepares Plate Filtration.Be mounted to by filter cloth on plate-and-frame filter press, after diatomite precoating, the protein liquid that will reduce is by Plate Filtration, and filter pressure 0.35-0.6Mpa, poromerics is 0.8 micron, and removing impurity permeate is clear liquor.Be separated through ultra-filtration membrane desalination by clear liquor, molecular weight cut-off is 50000 dalton.Adopt 0.22 μm of filter membrane to carry out low temperature micro-filtration to concentrated solution degerming, temperature controls below 8-16 DEG C, and pressure difference is 0.04-0.06Mpa.MF permeate liquid is carried out drying under low-temperature spray drying tower, inlet temperature 100-140 DEG C, temperature out 60-80 DEG C, temperature of charge 20-40 DEG C, mass flow 6-8%.Obtain major product livetin powder.
Product property and detection:
The purity of immune globulin IgY sample adopts Lab-Image software to calculate according to SDS-PAGE electrophoretic image, and the protein content in sample adopts Xylene Brilliant Cyanine G RNA isolation kit to measure, by purity and protein content according to the following formulae discovery rate of recovery.
Immune globulin IgY activity adopts euzymelinked immunosorbent assay (ELISA) to detect, adopt the light absorption value drawing standard curve of the standard substance of a series of known activity and concentration, according to keeping active immune globulin IgY content in the light absorption value calculation sample of testing sample, immune globulin IgY total content in middle total protein content and immune globulin IgY purity calculation sample per sample, active conservation rate is according to following formulae discovery.
Measure and calculation the results are shown in Table 1, adopts the immunoglobulin (Ig) rate of recovery of the inventive method separation and purification gained, purity, active conservation rate all higher.
Table 1: the IgY rate of recovery after extraction and isolation, purity, active conservation rate calculation result
| Embodiment | The rate of recovery (%) | Purity (%) | Active conservation rate (%) |
| Embodiment 1 | 94.32±0.03％ | 77.55±0.01％ | 96.32±0.06％ |
| Embodiment 2 | 92.32±0.02％ | 75.35±0.04％ | 93.32±0.05％ |
The protein powder that the present embodiment 1 obtains is colourless extremely light yellow lens, and main component is protein 95%, moisture 2.8%, grey matter 2.2%, and wherein the content of Yolk immune globulin IgY is 75.3%.After depositing 2 years under room temperature after vacuum packaging, activity decrease is less than 4%, low-temp storage 6-7, and activity decrease is less than 2%.
The protein powder that the present embodiment 2 obtains is colourless extremely light yellow lens, and main component is protein 94%, moisture 3.4%, grey matter 2.6%, and wherein the content of Yolk immune globulin IgY is 78.3%.After depositing 2 years under room temperature after vacuum packaging, activity decrease is less than 4%, low-temp storage 6-7, and activity decrease is less than 2%.
Claims (9)
1. a preparation method for livetin powder, comprising:
(1) get yolk, by yolk: the weight kg of deionized water: volume L adds deionized water than 1 ~ 5:1, after stirring 3 ~ 10 minutes, input settling tank, leave standstill 4 ~ 6 hours, obtain supernatant liquor;
(2) supernatant liquor is extracted into stirred pot, makes supernatant liquor be in whipped state, stir speed (S.S.) is 50 ~ 70r/min, in supernatant liquor, 4:1 ratio adds salts solution by volume simultaneously, close after salts solution adds completely and stir, supernatant liquor leaves standstill 3 ~ 4 hours, makes albumen precipitation;
(3) after albumen precipitates completely, upper liquid degrease is extracted, in upper liquid: deionized water volume ratio 1:5 ratio adds deionized water reduction albumen, obtains reducing protein liquid, prepares Plate Filtration;
(4) be mounted on plate-and-frame filter press by filter cloth, after diatomite precoating, the protein liquid that will reduce is by Plate Filtration, and remove impurity, permeate is clear liquor;
(5) be separated through ultra-filtration membrane desalination by clear liquor, molecular weight cut-off is 50000 dalton;
(6) concentrated solution is carried out low temperature micro-filtration degerming, microfiltration membrane adopts 0.22 micron pore size,
(7) MF permeate liquid is carried out drying under low-temperature spray drying tower, obtain major product livetin powder.
2. the preparation method of livetin powder according to claim 1, is characterized in that, in described step (1), deionized water temperature is 0 ~ 4 DEG C, specific conductivity≤0.3.
3. the preparation method of livetin powder according to claim 1, is characterized in that, in described step (2), salts solution adopts saturated sodium-chloride salts solution/ammoniumsulphate soln, and saturation concentration is 33%, churning time 5-10min.
4. the preparation method of livetin powder according to claim 1, is characterized in that, remaining egg yolk liquid when supernatant liquor being extracted in step (2) stirred pot can prepare yolk powder.
5. the preparation method of livetin powder according to claim 1, is characterized in that, when step (3) carries out Plate Filtration, filter pressure is 0.35-0.6MPa.
6. the preparation method of livetin powder according to claim 1, it is characterized in that, in described step (4) by Penetration ration in the smooth installation of 350---400 order filter cloth on a filter press, first with air in excavator in water cycle to pressure filter, 150 order fineness diatomite, at 0.35-0.5MPa, are evenly coated on filter cloth by top hole pressure in advance.
7. the preparation method of livetin powder according to claim 1, is characterized in that, can also can add water cycle desalination in step (5) in ultra-filtration membrane desalination sepn process, or on-line monitoring.
8. the preparation method of livetin powder according to claim 1, is characterized in that, when in step (6), low temperature micro-filtration is degerming, control temperature is below 10 DEG C, and pressure difference is 0.02-0.08MPa; Or below control temperature 8-16 DEG C, pressure difference is 0.04-0.06Mpa.
9. the preparation method of livetin powder according to claim 1, is characterized in that, when carrying out drying under low-temperature spray drying tower in step (7), and inlet temperature 100-140 DEG C, temperature out 60-80 DEG C, temperature of charge 20-40 DEG C, mass flow 6-8%.
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Cited By (7)
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| CN107083269A (en) * | 2017-04-01 | 2017-08-22 | 新疆喀什昆仑翠翎鸽业有限责任公司 | One kind is without BaP pigeon eggs butter and preparation method thereof |
| CN108586597A (en) * | 2018-04-27 | 2018-09-28 | 姜权 | Fish promote ingest mature peptide and its application |
| CN110003319A (en) * | 2019-04-16 | 2019-07-12 | 成都大学 | A kind of method and product based on a variety of egg yolk proteins of aqueous phase separation combined extracting |
| CN111150066A (en) * | 2020-02-19 | 2020-05-15 | 苟春虎 | Epidemic prevention immune peptide |
| CN112569200A (en) * | 2020-12-30 | 2021-03-30 | 江苏协合转化医学研究院有限公司 | Effervuline effervescent tablet and preparation method thereof |
| CN113514294A (en) * | 2021-04-22 | 2021-10-19 | 济南海关技术中心 | Egg Sudan red standard sample and preparation method thereof |
| CN115428956A (en) * | 2022-08-08 | 2022-12-06 | 赛法特（长沙）生物技术有限公司 | Compound protein powder and preparation method thereof |
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107083269A (en) * | 2017-04-01 | 2017-08-22 | 新疆喀什昆仑翠翎鸽业有限责任公司 | One kind is without BaP pigeon eggs butter and preparation method thereof |
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| CN110003319A (en) * | 2019-04-16 | 2019-07-12 | 成都大学 | A kind of method and product based on a variety of egg yolk proteins of aqueous phase separation combined extracting |
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| CN111150066A (en) * | 2020-02-19 | 2020-05-15 | 苟春虎 | Epidemic prevention immune peptide |
| CN112569200A (en) * | 2020-12-30 | 2021-03-30 | 江苏协合转化医学研究院有限公司 | Effervuline effervescent tablet and preparation method thereof |
| CN113514294A (en) * | 2021-04-22 | 2021-10-19 | 济南海关技术中心 | Egg Sudan red standard sample and preparation method thereof |
| CN113514294B (en) * | 2021-04-22 | 2024-06-07 | 济南海关技术中心 | Sudan red standard sample in eggs and preparation method thereof |
| CN115428956A (en) * | 2022-08-08 | 2022-12-06 | 赛法特（长沙）生物技术有限公司 | Compound protein powder and preparation method thereof |
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