CN107488224A - A kind of method that albumen is extracted with animal tissue - Google Patents
A kind of method that albumen is extracted with animal tissue Download PDFInfo
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- CN107488224A CN107488224A CN201710831202.4A CN201710831202A CN107488224A CN 107488224 A CN107488224 A CN 107488224A CN 201710831202 A CN201710831202 A CN 201710831202A CN 107488224 A CN107488224 A CN 107488224A
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- centrifugally operated
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
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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
- C07K1/145—Extraction; Separation; Purification by extraction or solubilisation
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- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biochemistry (AREA)
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- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Gastroenterology & Hepatology (AREA)
- Zoology (AREA)
- Toxicology (AREA)
- Analytical Chemistry (AREA)
- Peptides Or Proteins (AREA)
Abstract
The present invention relates to a kind of method that albumen is extracted with animal tissue, comprise the following steps：（1）By fresh animal tissue it is broken after with distillation water washing 8 10 times, remove blood constituent, then extrusion process is into pasty state；（2）Will（1）In tissue in add 0.5mol/L TRIS HCl pH7.6,1% NaTDC and 0.5g/mlPMSF and add micrococcus acid enzyme, stirred 1 hour after mixing；（3）Will（2）Obtained mixture centrifugally operated 30min, separate supernatant sediment；（4）Will（3）Middle addition 0.5mol/L TRIS HCl pH7.6,0.5g/mlPMSF washing is secondary, then centrifugally operated 30min, abandons supernatant, obtains tissue extracelluar matrix mixture；（5）Will（4）Obtained mixture stirs extraction after adding NaCl；It is an advantage of the invention that industrialized production, technological process be simple and production material wide material sources.
Description
Technical field
The invention belongs to extract protein techniques field, and in particular to a kind of method that albumen is extracted with animal tissue.
Background technology
There are hyaluronic acid, collagen etc. using more injection packing material in clinic at present, but they have
Have and be easy to be absorbed, the defects of bioactivity difference；Extracellular matrix is to be synthesized and be secreted into by zooblast extracellular, is distributed in
Cell surface or intercellular macromolecular, mainly some polysaccharide and albumen or proteoglycans；Extracellular matrix is not only cell
Provide a support structure and attachment site, and by the adhesion molecule with cell, growth regulator, associated proteins,
Proteolytic enzyme and enzyme inhibitor interaction, and the tune of adhesion, migration, propagation, differentiation and the gene expression to cell
Control plays important adjustment effect；Because extracellular matrix has above biological action, have and natural tissues organ phase
As three-D space structure, it is widely used in organizational project as timbering material；Successfully extracted at present extracellular
The histoorgan of matrix includes mucous membrane of small intestine, cardiac valves, blood vessel, skin, nerve, skeletal muscle, placenta, bladder and liver；Group
The source knitted is wide, abundant extracellular matrix and antibacterial peptide can be extracted from tissue, and then extract laminin, fibre
Even animal protein, the research such as albumen, collagen has shown that epimatrix can promote Various Tissues to regenerate as biomaterial.
The content of the invention
The invention aims to overcome the shortcomings of above-mentioned technical elements, and develop a kind of industrialized production, technique
Flow simply and production material wide material sources with animal tissue extraction albumen method.
The object of the present invention is achieved like this：A kind of method that albumen is extracted with animal tissue, comprises the following steps：
（1）By fresh animal tissue it is broken after with distillation water washing 8-10 time, removal blood constituent, then extrusion process is into paste
Shape；
（2）Will（1）In tissue in add 0.5mol/L TRIS-HCl pH7.6,1% NaTDC and 0.5g/mlPMSF
And micrococcus acid enzyme is added, stirred 1 hour after mixing；
（3）Will（2）Obtained mixture centrifugally operated 30min, separate supernatant sediment；
（4）Will（3）Middle addition 0.5mol/L TRIS-HCl pH7.6,0.5g/mlPMSF washing is secondary, then centrifugally operated
30min, supernatant is abandoned, obtain tissue extracelluar matrix mixture；
（5）Will（4）Obtained mixture stirs extraction after adding NaCl；
（6）Will（5）Mixture centrifugally operated 1 hour, obtain laminin after abandoning precipitation；
（7）Will（5）Mixture add 2mol/L aminocaproic acid and urea, centrifugally operated 1 hour, abandon precipitating, obtain it is fine even
Albumen；
（8）Will（5）Mixture add 0.5mol/L TRIS-HCl pH7.6, stood after stirring, after to be separated, abandon supernatant
Liquid, after taking out precipitated crystal, as collagen；
（9）Will（3）In after obtained supernatant dilution by ion exchange column, add 25mmol/L ammonium acetates wash away non-cationic into
Point, 10% acetic acid elution cation constituent is added, eluting peak is collected, is resuspended with 0.01% acetic acid, obtains animal derived antimicrobial peptide.
Using high-speed refrigerated centrifuge in the centrifugally operated.
Beneficial effects of the present invention：It is an advantage of the invention that extract laminin, fibre using the tissue of wide material sources
Even albumen, collagen and antibacterial peptide, the method can be used for industrial mass production, and solving some methods can only be in laboratory
The problem of success, extensive technical support is provided for the extraction of albumen；Therefore, it is an advantage of the invention that industrialized production, work
Skill flow is simple and production material wide material sources.
Embodiment
Embodiment 1
A kind of method that albumen is extracted with animal tissue, comprises the following steps：（1）Fresh animal tissue is broken afterwards with steaming
Distilled water is washed 8-10 time, and removal blood constituent, then extrusion process is into pasty state；
（2）Will（1）In tissue in add 0.5mol/L TRIS-HCl pH7.6,1% NaTDC and 0.5g/mlPMSF
And micrococcus acid enzyme is added, stirred 1 hour after mixing；
（3）Will（2）Obtained mixture centrifugally operated 30min, separate supernatant sediment；
（4）Will（3）Middle addition 0.5mol/L TRIS-HCl pH7.6,0.5g/mlPMSF washing is secondary, then centrifugally operated
30min, supernatant is abandoned, obtain tissue extracelluar matrix mixture；
（5）Will（4）Obtained mixture stirs extraction after adding NaCl；
（6）Will（5）Mixture centrifugally operated 1 hour, obtain laminin after abandoning precipitation；
（7）Will（5）Mixture add 2mol/L aminocaproic acid and urea, centrifugally operated 1 hour, abandon precipitating, obtain it is fine even
Albumen；
（8）Will（5）Mixture add 0.5mol/L TRIS-HCl pH7.6, stood after stirring, after to be separated, abandon supernatant
Liquid, after taking out precipitated crystal, as collagen；
（9）Will（3）In after obtained supernatant dilution by ion exchange column, add 25mmol/L ammonium acetates wash away non-cationic into
Point, 10% acetic acid elution cation constituent is added, eluting peak is collected, is resuspended with 0.01% acetic acid, obtains animal derived antimicrobial peptide.
It is an advantage of the invention that the technique productions flow is simple, production material is simple, and the more original method of yield has very big
Improve；Therefore, it is an advantage of the invention that industrialized production, technological process be simple and production material wide material sources.
Embodiment 2
A kind of method that albumen is extracted with animal tissue, comprises the following steps：（1）Fresh animal tissue is broken afterwards with steaming
Distilled water is washed 8-10 time, and removal blood constituent, then extrusion process is into pasty state；
（2）Will（1）In tissue in add 0.5mol/L TRIS-HCl pH7.6,1% NaTDC and 0.5g/mlPMSF
And micrococcus acid enzyme is added, stirred 1 hour after mixing；
（3）Will（2）Obtained mixture centrifugally operated 30min, separate supernatant sediment；
（4）Will（3）Middle addition 0.5mol/L TRIS-HCl pH7.6,0.5g/mlPMSF washing is secondary, then centrifugally operated
30min, supernatant is abandoned, obtain tissue extracelluar matrix mixture；
（5）Will（4）Obtained mixture stirs extraction after adding NaCl；
（6）Will（5）Mixture centrifugally operated 1 hour, obtain laminin after abandoning precipitation；
（7）Will（5）Mixture add 2mol/L aminocaproic acid and urea, centrifugally operated 1 hour, abandon precipitating, obtain it is fine even
Albumen；
（8）Will（5）Mixture add 0.5mol/L TRIS-HCl pH7.6 again, stand after stirring, after to be separated, abandon
Supernatant, after taking out precipitated crystal, as collagen；
（9）Will（3）In after obtained supernatant dilution by ion exchange column, add 25mmol/L ammonium acetates wash away non-cationic into
Point, 10% acetic acid elution cation constituent is added, eluting peak is collected, is resuspended with 0.01% acetic acid, obtains animal derived antimicrobial peptide, it is described
Using high-speed refrigerated centrifuge in centrifugally operated.
Embodiment is unrestricted to further explanation of the invention, is existed for those of ordinary skills
Further conversion is done in the case of not departing from substantive content of the present invention to structure, and all these conversion should all belong to institute of the present invention
Attached scope of the claims.
Claims (2)
1. a kind of method that albumen is extracted with animal tissue, comprises the following steps：（1）By the broken use afterwards of fresh animal tissue
Distill water washing 8-10 time, removal blood constituent, then extrusion process is into pasty state；
（2）Will（1）In tissue in add 0.5mol/L TRIS-HCl pH7.6,1% NaTDC and 0.5g/mlPMSF
And micrococcus acid enzyme is added, stirred 1 hour after mixing；
（3）Will（2）Obtained mixture centrifugally operated 30min, separate supernatant sediment；
（4）Will（3）Middle addition 0.5mol/L TRIS-HCl pH7.6,0.5g/mlPMSF washing is secondary, then centrifugally operated
30min, supernatant is abandoned, obtain tissue extracelluar matrix mixture；
（5）Will（4）Obtained mixture stirs extraction after adding NaCl；
（6）Will（5）Mixture centrifugally operated 1 hour, obtain laminin after abandoning precipitation；
（7）Will（5）Mixture add 2mol/L aminocaproic acid and urea, centrifugally operated 1 hour, abandon precipitating, obtain it is fine even
Albumen；
（8）Will（5）Mixture add 0.5mol/L TRIS-HCl pH7.6, stood after stirring, after to be separated, abandon supernatant
Liquid, after taking out precipitated crystal, as collagen；
（9）Will（3）In after obtained supernatant dilution by ion exchange column, add 25mmol/L ammonium acetates wash away non-cationic into
Point, 10% acetic acid elution cation constituent is added, eluting peak is collected, is resuspended with 0.01% acetic acid, obtains animal derived antimicrobial peptide.
A kind of 2. method that albumen is extracted with animal tissue according to claim 1, it is characterised in that：The centrifugally operated
In using high-speed refrigerated centrifuge.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710831202.4A CN107488224A (en) | 2017-09-15 | 2017-09-15 | A kind of method that albumen is extracted with animal tissue |
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| CN201710831202.4A CN107488224A (en) | 2017-09-15 | 2017-09-15 | A kind of method that albumen is extracted with animal tissue |
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| CN107488224A true CN107488224A (en) | 2017-12-19 |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108812639A (en) * | 2018-07-05 | 2018-11-16 | 广州昕生医学材料有限公司 | A kind of extracellular matrix freeze-dried powder |
| WO2023212256A1 (en) * | 2022-04-28 | 2023-11-02 | Aufbau Medical Innovations Limited | Affinity capture of extracellular matrix bodies |
| WO2023212255A1 (en) * | 2022-04-28 | 2023-11-02 | Aufbau Medical Innovations Limited | Isolating extracellular matrix bodies |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4511653A (en) * | 1981-11-26 | 1985-04-16 | Foundation Merieux | Process for the industrial preparation of collagenous materials from human placental tissues, human collagenous materials obtained and their application as biomaterials |
| CN105154508A (en) * | 2015-09-27 | 2015-12-16 | 重庆都好生物科技有限公司 | Antimicrobial peptide preparation method |
-
2017
- 2017-09-15 CN CN201710831202.4A patent/CN107488224A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4511653A (en) * | 1981-11-26 | 1985-04-16 | Foundation Merieux | Process for the industrial preparation of collagenous materials from human placental tissues, human collagenous materials obtained and their application as biomaterials |
| CN105154508A (en) * | 2015-09-27 | 2015-12-16 | 重庆都好生物科技有限公司 | Antimicrobial peptide preparation method |
Non-Patent Citations (2)
| Title |
|---|
| 无: "层粘连蛋白（Laminin，LN）的分离", 《HTTPS://WWW.BIOMART.CN/EXPERIMENT/430/488/489/37650.HTM?WD=%E5%B1%82%E7%B2%98%E8%BF%9E%E8%9B%8B%E7%99%BD%E7%9A%84%E6%8F%90%E5%8F%96&POSITIONID=1》 * |
| 陈留存等: "家蝇抗菌肽的分离纯化及性质研究", 《山 东 大 学 学 报( 自然科学版)》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108812639A (en) * | 2018-07-05 | 2018-11-16 | 广州昕生医学材料有限公司 | A kind of extracellular matrix freeze-dried powder |
| WO2023212256A1 (en) * | 2022-04-28 | 2023-11-02 | Aufbau Medical Innovations Limited | Affinity capture of extracellular matrix bodies |
| WO2023212255A1 (en) * | 2022-04-28 | 2023-11-02 | Aufbau Medical Innovations Limited | Isolating extracellular matrix bodies |
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| PB01 | Publication | ||
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| TA01 | Transfer of patent application right |
Effective date of registration: 20181220 Address after: 450000 East U Valley of Hanghai East Road North, 25th Street West and East U Valley of Zhengzhou Economic and Technological Development Zone, Zhengzhou City, Henan Province, 36 East District of Zhengzhou Jingkai International Enterprise Port Project (Phase I) Applicant after: Henan Eln Biotechnology Co., Ltd. Address before: Room 302, 20 Hongqi Street, Huancui District, Weihai City, Shandong Province Applicant before: Wu Guang |
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| RJ01 | Rejection of invention patent application after publication | ||
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Application publication date: 20171219 |