CN108977488A - Skin is intensively repaired with elastin laminin and preparation method thereof - Google Patents
Skin is intensively repaired with elastin laminin and preparation method thereof Download PDFInfo
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- 102000016942 Elastin Human genes 0.000 title claims abstract description 77
- 108010014258 Elastin Proteins 0.000 title claims abstract description 77
- 229920002549 elastin Polymers 0.000 title claims abstract description 77
- 238000002360 preparation method Methods 0.000 title claims abstract description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 56
- 238000000034 method Methods 0.000 claims abstract description 23
- CDBYLPFSWZWCQE-UHFFFAOYSA-L sodium carbonate Substances [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 90
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 39
- 239000006228 supernatant Substances 0.000 claims description 34
- 238000003756 stirring Methods 0.000 claims description 32
- 239000000463 material Substances 0.000 claims description 28
- 238000006243 chemical reaction Methods 0.000 claims description 24
- 238000000502 dialysis Methods 0.000 claims description 22
- 102000004190 Enzymes Human genes 0.000 claims description 21
- 108090000790 Enzymes Proteins 0.000 claims description 21
- 229940088598 enzyme Drugs 0.000 claims description 21
- 239000006210 lotion Substances 0.000 claims description 19
- 230000008439 repair process Effects 0.000 claims description 19
- 238000005406 washing Methods 0.000 claims description 19
- 239000004365 Protease Substances 0.000 claims description 18
- 238000001914 filtration Methods 0.000 claims description 18
- 238000002156 mixing Methods 0.000 claims description 15
- 239000002270 dispersing agent Substances 0.000 claims description 14
- 239000012153 distilled water Substances 0.000 claims description 13
- 238000009777 vacuum freeze-drying Methods 0.000 claims description 13
- 239000002253 acid Substances 0.000 claims description 12
- 210000002808 connective tissue Anatomy 0.000 claims description 11
- 238000001816 cooling Methods 0.000 claims description 11
- 239000000203 mixture Substances 0.000 claims description 11
- 238000010792 warming Methods 0.000 claims description 11
- 235000005979 Citrus limon Nutrition 0.000 claims description 10
- 239000000796 flavoring agent Substances 0.000 claims description 10
- 235000019634 flavors Nutrition 0.000 claims description 10
- ODLHGICHYURWBS-LKONHMLTSA-N trappsol cyclo Chemical compound CC(O)COC[C@H]([C@H]([C@@H]([C@H]1O)O)O[C@H]2O[C@@H]([C@@H](O[C@H]3O[C@H](COCC(C)O)[C@H]([C@@H]([C@H]3O)O)O[C@H]3O[C@H](COCC(C)O)[C@H]([C@@H]([C@H]3O)O)O[C@H]3O[C@H](COCC(C)O)[C@H]([C@@H]([C@H]3O)O)O[C@H]3O[C@H](COCC(C)O)[C@H]([C@@H]([C@H]3O)O)O3)[C@H](O)[C@H]2O)COCC(O)C)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@@H]3O[C@@H]1COCC(C)O ODLHGICHYURWBS-LKONHMLTSA-N 0.000 claims description 10
- 239000004367 Lipase Substances 0.000 claims description 9
- 102000004882 Lipase Human genes 0.000 claims description 9
- 108090001060 Lipase Proteins 0.000 claims description 9
- 108090000526 Papain Proteins 0.000 claims description 9
- 108091005804 Peptidases Proteins 0.000 claims description 9
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 claims description 9
- 238000007710 freezing Methods 0.000 claims description 9
- 230000008014 freezing Effects 0.000 claims description 9
- 235000019421 lipase Nutrition 0.000 claims description 9
- 229940055729 papain Drugs 0.000 claims description 9
- 235000019834 papain Nutrition 0.000 claims description 9
- 235000019419 proteases Nutrition 0.000 claims description 9
- 238000000859 sublimation Methods 0.000 claims description 9
- 230000008022 sublimation Effects 0.000 claims description 9
- 229940097346 sulfobutylether-beta-cyclodextrin Drugs 0.000 claims description 9
- 238000005238 degreasing Methods 0.000 claims description 8
- 239000012065 filter cake Substances 0.000 claims description 8
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 6
- 235000011182 sodium carbonates Nutrition 0.000 claims description 6
- 239000003795 chemical substances by application Substances 0.000 claims description 5
- 238000004321 preservation Methods 0.000 claims description 5
- 238000005119 centrifugation Methods 0.000 claims description 4
- 235000014113 dietary fatty acids Nutrition 0.000 claims description 4
- 239000000194 fatty acid Substances 0.000 claims description 4
- 229930195729 fatty acid Natural products 0.000 claims description 4
- 150000004665 fatty acids Chemical class 0.000 claims description 4
- 230000007935 neutral effect Effects 0.000 claims description 3
- 230000003472 neutralizing effect Effects 0.000 claims description 3
- 244000248349 Citrus limon Species 0.000 claims 2
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 claims 1
- 239000005864 Sulphur Substances 0.000 claims 1
- 239000006185 dispersion Substances 0.000 claims 1
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 abstract description 7
- 229920002674 hyaluronan Polymers 0.000 abstract description 7
- 229960003160 hyaluronic acid Drugs 0.000 abstract description 7
- 230000000694 effects Effects 0.000 abstract description 6
- 230000032683 aging Effects 0.000 abstract description 4
- 239000003963 antioxidant agent Substances 0.000 abstract description 3
- 230000003078 antioxidant effect Effects 0.000 abstract description 3
- 235000006708 antioxidants Nutrition 0.000 abstract description 3
- 230000003020 moisturizing effect Effects 0.000 abstract description 3
- 238000007781 pre-processing Methods 0.000 abstract 1
- 210000003491 skin Anatomy 0.000 description 52
- 239000004744 fabric Substances 0.000 description 13
- 238000012360 testing method Methods 0.000 description 13
- 239000000159 acid neutralizing agent Substances 0.000 description 12
- 244000131522 Citrus pyriformis Species 0.000 description 8
- 238000001035 drying Methods 0.000 description 8
- 238000004108 freeze drying Methods 0.000 description 7
- 210000001519 tissue Anatomy 0.000 description 7
- JEIPFZHSYJVQDO-UHFFFAOYSA-N iron(III) oxide Inorganic materials O=[Fe]O[Fe]=O JEIPFZHSYJVQDO-UHFFFAOYSA-N 0.000 description 6
- 239000007788 liquid Substances 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 241000699666 Mus <mouse, genus> Species 0.000 description 5
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 4
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 241000699670 Mus sp. Species 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- -1 hydroxypropyl Chemical group 0.000 description 3
- 210000000056 organ Anatomy 0.000 description 3
- 239000002994 raw material Substances 0.000 description 3
- 230000002500 effect on skin Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 238000011049 filling Methods 0.000 description 2
- 238000005286 illumination Methods 0.000 description 2
- 210000004072 lung Anatomy 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 108090000765 processed proteins & peptides Proteins 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- 244000025254 Cannabis sativa Species 0.000 description 1
- 229920000858 Cyclodextrin Polymers 0.000 description 1
- 239000004375 Dextrin Substances 0.000 description 1
- 229920001353 Dextrin Polymers 0.000 description 1
- 238000008157 ELISA kit Methods 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 239000001116 FEMA 4028 Substances 0.000 description 1
- 102000004157 Hydrolases Human genes 0.000 description 1
- 108090000604 Hydrolases Proteins 0.000 description 1
- 238000012449 Kunming mouse Methods 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 210000000577 adipose tissue Anatomy 0.000 description 1
- 125000003275 alpha amino acid group Chemical group 0.000 description 1
- 230000003064 anti-oxidating effect Effects 0.000 description 1
- 210000000709 aorta Anatomy 0.000 description 1
- 210000003433 aortic smooth muscle cell Anatomy 0.000 description 1
- 210000001367 artery Anatomy 0.000 description 1
- 229960004853 betadex Drugs 0.000 description 1
- 238000003763 carbonization Methods 0.000 description 1
- 210000003321 cartilage cell Anatomy 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000006835 compression Effects 0.000 description 1
- 238000007906 compression Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 235000019425 dextrin Nutrition 0.000 description 1
- 229920001971 elastomer Polymers 0.000 description 1
- 239000000806 elastomer Substances 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 230000007071 enzymatic hydrolysis Effects 0.000 description 1
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 description 1
- 210000000630 fibrocyte Anatomy 0.000 description 1
- 230000007760 free radical scavenging Effects 0.000 description 1
- 238000000265 homogenisation Methods 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 210000003041 ligament Anatomy 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 230000035882 stress Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 238000009281 ultraviolet germicidal irradiation Methods 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
-
- 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
-
- 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
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
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- Chemical & Material Sciences (AREA)
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- Life Sciences & Earth Sciences (AREA)
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- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Biochemistry (AREA)
- Zoology (AREA)
- Biophysics (AREA)
- Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Wood Science & Technology (AREA)
- Toxicology (AREA)
- Gastroenterology & Hepatology (AREA)
- Biotechnology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Microbiology (AREA)
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- General Engineering & Computer Science (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Cosmetics (AREA)
Abstract
The invention discloses a kind of intensive reparation elastin laminins and preparation method thereof of skin, and the preparation method comprises the following steps: (1) pre-processing；(2) it digests；(3) it dialyses；(4) dry.Skin of the present invention is intensively repaired with elastin laminin and preparation method thereof, method is simply easy to implement, it is environment friendly and pollution-free, by carrying out ungrease treatment to ox paxwax surface, the enzymolysis efficiency and purity of elastin laminin are improved, obtained elastin laminin has effects that anti-oxidant, moisturizing, the hyaluronic acid and water content that can be obviously improved in skin, aging skin can intensively be repaired, can be used for skin care item and health care product, have a good application prospect.
Description
Technical field
The present invention relates to technical field of biochemistry, and in particular to a kind of skin is intensively repaired with elastin laminin and its preparation side
Method.
Background technique
Elastin laminin is a kind of important extracellular matrix protein, and chief component is elastomer.It largely exists
In the tissue and organ that the frequent stress such as lung, main artery and skin of mammal deforms, major function is for place group
The ability for providing with organ and resisting Repeated Compression and deformation is provided.The content of elastin laminin in the tissue is because of elastic work needed for tissue
Energy degree is different and different, and content is minimum in skin, about 2-3%；Content is placed in the middle in lung and aorta, respectively 10-25%
And 30-50%；Content highest in ligament, about 80%.Elastin laminin is mainly by fibrocyte, aortic smooth muscle cells simultaneously
With produced by cartilage cell, the cell of generation due to locating organ is different and difference.
Elastin laminin contains distinctive amino acid structure, is three spiral knots being cross-linked to form by two α chains and a β chain
Structure has good elasticity and ductility due to its fine and close structure and hydrophobicity, and can resist the enzyme of most of hydrolase
Solution.However due to the structure of elastin laminin raw material, enzymatic hydrolysis process can be hindered, it is possible to create miscellaneous peptide, and influence the flavor of product.
It is often rich in lipid material in elastin laminin tissue, influences the using effect of elastin laminin.Application No. is
In 2013102946564 patent of invention, it is used as raw material to prepare elastin laminin using ox neck is tough, impregnates in acetone preliminary de-
Rouge, then removing fat is boiled with oxalic acid solution, but acetone is low toxicity material, it is volatile, it is unfavorable for environmental protection in actual production, grass
Though acid, which boils, can remove partial fat, the loss of partial elastic albumen is also resulted in.
Summary of the invention
Aiming at the above shortcomings existing in the prior art, technical problem to be solved by the invention is to provide a kind of skin is close
Collect reparation elastin laminin and preparation method thereof, method is simply easy to implement, environment friendly and pollution-free, by ox paxwax surface into
Row ungrease treatment improves the enzymolysis efficiency and purity of elastin laminin, and obtained elastin laminin has effects that anti-oxidant, moisturizing,
The hyaluronic acid and water content that can be obviously improved in skin.
Object of the present invention is to what is be achieved through the following technical solutions:
A kind of skin intensively repairs the preparation method with elastin laminin, comprising the following steps:
(1) it pre-processes；
(2) it digests；
(3) it dialyses；
(4) vacuum freeze drying.
A kind of skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into (1-4) mm × (1-4) mm × (1-4)
The block of mm mixes 10-20 parts of blocks and 40-80 parts of 3-6wt% aqueous sodium carbonates, at 40-60 DEG C with 100-300
Revs/min stirring 20-40 minute, filtering, be washed with water to washing lotion pH be it is neutral, drain surface moisture, obtain pretreated feedstock；
(2) digest: pretreated feedstock and 40-80 part water mixed, with 100-300 revs/min stirring 5-10 minutes, addition
It is 7.0-8.5 that aqueous sodium carbonate that 0.01-0.1 parts of trypsase are 5wt% with mass fraction, which adjusts pH, 45-55 DEG C with
100-300 revs/min stirring 2-4 hours, add 0.02-0.2 parts of mixed enzymes, with mass fraction be 10% lemon acid for adjusting pH
For 6.0-7.0,50-60 DEG C with 100-300 revs/min stirring 4-6 hours, then be warming up to 90-100 DEG C of heat preservation 2-5 minutes, drop naturally
Temperature to 15-30 DEG C, with 4000-6000 revs/min centrifugation 20-30 minutes, collect supernatant；
(3) it dialyses: supernatant is dialysed 36-72 hours in bag filter, dialysis medium is distilled water, the body for medium of dialysing
Product is 8-12 times of supernatant volume, and every primary dialysis medium of 10-15 hour replacement obtains dialyzate；
(4) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin.
A kind of skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into (1-4) mm × (1-4) mm × (1-4)
The block of mm mixes 10-20 parts of blocks and 40-80 parts of 3-6wt% aqueous sodium carbonates, at 40-60 DEG C with 100-300
Revs/min stirring 20-40 minute, filtering, be washed with water to washing lotion pH be it is neutral, drain surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 40-80 parts of water are mixed, and 0.01-0.05 parts of lipase, 0.1-0.3 parts points are added
Powder, 0.1-0.3 parts of fatty acid neutralizing agents, it is 8-10 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, in 40-
60 DEG C with 100-300 revs/min stirring 90-180 minutes, reaction process maintain reaction pH be 8-10, end of reaction, filtering, filter cake
Washed 1-3 times with 25-35 parts of 3-6%wt aqueous sodium carbonates, be washed with water wash to washing lotion pH for neutrality, drain surface moisture,
Obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 40-80 parts of water mixing are stirred with 100-300 revs/min
It mixes 5-10 minutes, 0.01-0.1 parts of trypsase is added, it is 7.0- that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH,
8.5,45-55 DEG C with 100-300 revs/min stirring 2-4 hours, add 0.02-0.2 parts of mixed enzymes, be with mass fraction
10% lemon acid for adjusting pH is 6.0-7.0,50-60 DEG C with 100-300 revs/min stirring 4-6 hours, then be warming up to 90-100 DEG C
Heat preservation 2-5 minute, be naturally cooling to 15-30 DEG C, with 4000-6000 revs/min centrifugation 20-30 minutes, collection supernatant；
(4) it dialyses: supernatant is dialysed 36-72 hours in bag filter, dialysis medium is distilled water, the body for medium of dialysing
Product is 8-12 times of supernatant volume, and every primary dialysis medium of 10-15 hour replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin.
The condition of the vacuum freeze drying are as follows: material thickness 3-6mm sets pre-freezing temperature as -20~-25 DEG C, works as sample
Product temperature degree is kept for 1-2 hours after dropping to set temperature, sets sublimation temperature as 5-15 DEG C, resolution temperature is 30-35 DEG C, vacuum degree
10-30Pa, drying time 10-20 hour.
The mixed enzyme is papain and flavor protease (1-3) in mass ratio: the mixture of (1-3).
The dispersing agent is hydroxypropyl-β-cyclodextrin and/or Sulfobutyl ether β _ cyclodextrin.
Preferably, the dispersing agent is the mixture of hydroxypropyl-β-cyclodextrin and Sulfobutyl ether β _ cyclodextrin, the hydroxypropyl
The mass ratio of group-beta-cyclodextrin and Sulfobutyl ether β _ cyclodextrin is (1-5): 1.
The fat acid neutralizing agent is sodium carbonate and/or sodium hydroxide.
A kind of skin is intensively repaired with elastin laminin, is prepared using the above method.
Skin of the present invention, which is intensively repaired, uses elastin laminin and preparation method thereof, and method is simply easy to implement, environment friendly and pollution-free,
By carrying out ungrease treatment to ox paxwax surface, the enzymolysis efficiency and purity of elastin laminin are improved, obtained elastin laminin tool
There is the effect of anti-oxidant, moisturizing, the hyaluronic acid and water content that can be obviously improved in skin can intensively repair aging skin
It is multiple, it can be used for skin care item and health care product, have a good application prospect.
Specific embodiment
In the present invention, if not refering in particular to, all devices and raw material is commercially available or the industry is common are following
Method in embodiment is unless otherwise instructed conventional method in that art.
Bag filter, American Association carbonization bag filter, molecular cut off 1000D.
Lipase, Pangbo Bioengineering Co Ltd, Nanning, 50,000 U/g, model: PB26.
Trypsase, Xi'an Jim Press bioengineering Co., Ltd, 50,000 U/g, model: Jim Press -1121.
Papain, Pangbo Bioengineering Co Ltd, Nanning, 50,000 U/g of enzyme activity, model: PB01.
Flavor protease, Pangbo Bioengineering Co Ltd, Nanning, 50,000 U/g of enzyme activity, model: PB03.
Hydroxypropyl-β-cyclodextrin, No. CAS: 128446-35-5, the Wuhan source Dong Kang Science and Technology Ltd. provides.
Sulfobutyl ether β _ cyclodextrin, No. CAS: 25167-62-8, the Wuhan source Dong Kang Science and Technology Ltd. provides.
Scavenging ability of DPPH free radical measurement: by the intensive reparation elastin laminin of skin obtained in embodiment, according to merchant
" excusing from death pretreatment rice protein prepares anti-oxidation peptide " of pretty strong, Ma Haile etc. is measured removing DPPH free radical, measures
DPPH free radical scavenging activity of skin when intensively reparation with elastin laminin concentration is 1.0g/L.
Skin intensively influence experiment of the reparation elastin laminin to mice aging model: KM mouse, SPF grades, male, weight
18-22g, it is normal to feed quarantine observation 7 days, it is randomly divided into 9 groups, every group 10, respectively normal group, model group, experimental group 1-
7.Elastin laminin obtained in embodiment 1-7 is diluted to 5.0mg/mL with distilled water respectively, obtains the test sample of experimental group 1-7
Product.
5%D- galactolipin 1.0g/kg is subcutaneously injected in model group and the daily nape of experimental group, normally organizes daily nape and subcutaneously infuses
Penetrate isometric physiological saline.Experimental group and model group mouse carry out UV irradiation simultaneously, and UV illumination wavelength is 350-400nm, shine
Penetrate the time 40min/ days, light source is 40 cm vertical eminences away from mouse distance, continues modeling 40 days, naive mice normal illumination
Raising, modeling the 11st day starts, and given the test agent is given in daily stomach-filling to experimental group respectively, and pharmaceutical quantities are 50mL/kg, normal group and
The model group then isometric distilled water of daily stomach-filling, continuous feeding 30 days.
Moisture content of skin: mouse is put to death, and takes 2cm × 2cm back skin tissues, scrapes off light yellow subcutaneus adipose tissue,
Weighing, obtains m1, 12 hours dry at 80 DEG C, weighing obtains m2, then skin moisture content are as follows: (m1-m2)/m1× 100%.
After measuring skin moisture content, the physiological saline for being equivalent to the pre-cooling of 9 times of the tissue block weight is taken with graduated cylinder, pours into dress
It in the beaker of organized block, then shreds tissue block as early as possible with eye scissors and pours into homogenate tube, grind 5min using refiner, make
Organize homogenization.10% homogenate prepared is centrifuged 30min with 5000r/min, takes 4 DEG C of preservations of supernatant to be measured.
Measure hyaluronic acid contents in skin: using hyaluronic acid ELISA kit, (homogeneous biotechnology (Shanghai) is limited
Company provides), illustrate to measure hyaluronic acid contents in mouse skin in strict accordance with kit.
Embodiment 1
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) it digests: pretreated feedstock and 60 parts of water being mixed, is stirred 6 minutes with 200 revs/min, 0.03 part of tryptose is added
It is 8.0 that the aqueous sodium carbonate that enzyme is 5wt% with mass fraction, which adjusts pH, is stirred 3 hours at 50 DEG C with 200 revs/min, then plus
Entering 0.06 part of mixed enzyme, the lemon acid for adjusting pH for being 10% with mass fraction is 6.5, it is stirred 5 hours at 55 DEG C with 200 revs/min,
It is warming up to 98 DEG C again and keeps the temperature 3 minutes, is naturally cooling to 25 DEG C, is centrifuged 30 minutes with 5000 revs/min, collects supernatant；
(3) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(4) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The mixed enzyme is papain and the flavor protease mixture of 2:1 in mass ratio.
Embodiment 2
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 60 parts of water are mixed, and 0.03 part of lipase, 0.2 part of fatty acid neutralizing agent is added,
It is 9 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, is stirred 120 minutes at 50 DEG C with 200 revs/min, reaction process
Maintaining reaction pH is 9, and end of reaction, using 100 mesh filter-cloth filterings, filter cake is washed 2 times with 30 parts of 5%wt aqueous sodium carbonates,
Be washed with water wash to washing lotion pH for neutrality, drain surface moisture, obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 60 parts of water mixing stir 6 points with 200 revs/min
0.03 part of trypsase is added in clock, and it is 8.0 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, at 50 DEG C with 200
Rev/min stirring 3 hours, add 0.06 part of mixed enzyme, the lemon acid for adjusting pH for being 10% with mass fraction is 6.5, at 55 DEG C
It is stirred 5 hours with 200 revs/min, then is warming up to 98 DEG C and keeps the temperature 3 minutes, be naturally cooling to 25 DEG C, be centrifuged 30 points with 5000 revs/min
Clock collects supernatant；
(4) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The mixed enzyme is papain and the flavor protease mixture of 2:1 in mass ratio.
The fat acid neutralizing agent is sodium carbonate.
Embodiment 3
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 60 parts of water are mixed, 0.03 part of lipase, 0.2 part of dispersing agent, 0.2 part of rouge is added
It is 9 that the aqueous sodium carbonate that fat acid neutralizing agent is 5wt% with mass fraction, which adjusts pH, stirs 120 points at 50 DEG C with 200 revs/min
Clock, it is 9 that reaction process, which maintains reaction pH, and end of reaction, using 100 mesh filter-cloth filterings, filter cake is water-soluble with 30 parts of 5%wt sodium carbonate
Liquid wash 2 times, be washed with water wash to washing lotion pH for neutrality, drain surface moisture, obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 60 parts of water mixing stir 6 points with 200 revs/min
0.03 part of trypsase is added in clock, and it is 8.0 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, at 50 DEG C with 200
Rev/min stirring 3 hours, add 0.06 part of mixed enzyme, the lemon acid for adjusting pH for being 10% with mass fraction is 6.5, at 55 DEG C
It is stirred 5 hours with 200 revs/min, then is warming up to 98 DEG C and keeps the temperature 3 minutes, be naturally cooling to 25 DEG C, be centrifuged 30 points with 5000 revs/min
Clock collects supernatant；
(4) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The mixed enzyme is papain and the flavor protease mixture of 2:1 in mass ratio.
The dispersing agent is hydroxypropyl-β-cyclodextrin.
The fat acid neutralizing agent is sodium carbonate.
Embodiment 4
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 60 parts of water are mixed, 0.03 part of lipase, 0.2 part of dispersing agent, 0.2 part of rouge is added
It is 9 that the aqueous sodium carbonate that fat acid neutralizing agent is 5wt% with mass fraction, which adjusts pH, stirs 120 points at 50 DEG C with 200 revs/min
Clock, it is 9 that reaction process, which maintains reaction pH, and end of reaction, using 100 mesh filter-cloth filterings, filter cake is water-soluble with 30 parts of 5%wt sodium carbonate
Liquid wash 2 times, be washed with water wash to washing lotion pH for neutrality, drain surface moisture, obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 60 parts of water mixing stir 6 points with 200 revs/min
0.09 part of trypsase is added in clock, and it is 8.0 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, at 40 DEG C with 200
Rev/min stirring 8 hours, then be warming up to 98 DEG C keep the temperature 3 minutes, be naturally cooling to 25 DEG C, with 5000 revs/min be centrifuged 30 minutes, receive
Collect supernatant；
(4) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The dispersing agent is hydroxypropyl-β-cyclodextrin.
The fat acid neutralizing agent is sodium carbonate.
Embodiment 5
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 60 parts of water are mixed, 0.03 part of lipase, 0.2 part of dispersing agent, 0.2 part of rouge is added
It is 9 that the aqueous sodium carbonate that fat acid neutralizing agent is 5wt% with mass fraction, which adjusts pH, stirs 120 points at 50 DEG C with 200 revs/min
Clock, it is 9 that reaction process, which maintains reaction pH, and end of reaction, using 100 mesh filter-cloth filterings, filter cake is water-soluble with 30 parts of 5%wt sodium carbonate
Liquid wash 2 times, be washed with water wash to washing lotion pH for neutrality, drain surface moisture, obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 60 parts of water mixing stir 6 points with 200 revs/min
Clock adds 0.09 part of mixed enzyme, and the lemon acid for adjusting pH for being 10% with mass fraction is 6.5, is stirred at 55 DEG C with 200 revs/min
It mixes 8 hours, then is warming up to 98 DEG C and keeps the temperature 3 minutes, be naturally cooling to 25 DEG C, be centrifuged 30 minutes with 5000 revs/min, collect supernatant
Liquid；
(4) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The mixed enzyme is papain and the flavor protease mixture of 2:1 in mass ratio.
The dispersing agent is hydroxypropyl-β-cyclodextrin.
The fat acid neutralizing agent is sodium carbonate.
Embodiment 6
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 60 parts of water are mixed, 0.03 part of lipase, 0.2 part of dispersing agent, 0.2 part of rouge is added
It is 9 that the aqueous sodium carbonate that fat acid neutralizing agent is 5wt% with mass fraction, which adjusts pH, stirs 120 points at 50 DEG C with 200 revs/min
Clock, it is 9 that reaction process, which maintains reaction pH, and end of reaction, using 100 mesh filter-cloth filterings, filter cake is water-soluble with 30 parts of 5%wt sodium carbonate
Liquid wash 2 times, be washed with water wash to washing lotion pH for neutrality, drain surface moisture, obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 60 parts of water mixing stir 6 points with 200 revs/min
0.03 part of trypsase is added in clock, and it is 8.0 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, at 50 DEG C with 200
Rev/min stirring 3 hours, add 0.06 part of mixed enzyme, the lemon acid for adjusting pH for being 10% with mass fraction is 6.5, at 55 DEG C
It is stirred 5 hours with 200 revs/min, then is warming up to 98 DEG C and keeps the temperature 3 minutes, be naturally cooling to 25 DEG C, be centrifuged 30 points with 5000 revs/min
Clock collects supernatant；
(4) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The mixed enzyme is papain and the flavor protease mixture of 2:1 in mass ratio.
The dispersing agent is Sulfobutyl ether β _ cyclodextrin.
The fat acid neutralizing agent is sodium carbonate.
Embodiment 7
Skin intensively repairs the preparation method with elastin laminin, includes the following steps, described part is parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into 2mm × 2mm × 2mm block, will
15 parts of blocks and 60 parts of 5wt% aqueous sodium carbonate mixing, are stirred 30 minutes with 200 revs/min at 50 DEG C, are filtered using 100 mesh
Cloth filtering is washed with water to washing lotion pH as neutrality, drains surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 60 parts of water are mixed, 0.03 part of lipase, 0.2 part of dispersing agent, 0.2 part of rouge is added
It is 9 that the aqueous sodium carbonate that fat acid neutralizing agent is 5wt% with mass fraction, which adjusts pH, stirs 120 points at 50 DEG C with 200 revs/min
Clock, it is 9 that reaction process, which maintains reaction pH, and end of reaction, using 100 mesh filter-cloth filterings, filter cake is water-soluble with 30 parts of 5%wt sodium carbonate
Liquid wash 2 times, be washed with water wash to washing lotion pH for neutrality, drain surface moisture, obtain pure elastin laminin material；
(3) digest: the pure elastin laminin material that step (2) is obtained and 60 parts of water mixing stir 6 points with 200 revs/min
0.03 part of trypsase is added in clock, and it is 8.0 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, at 50 DEG C with 200
Rev/min stirring 3 hours, add 0.06 part of mixed enzyme, the lemon acid for adjusting pH for being 10% with mass fraction is 6.5, at 55 DEG C
It is stirred 5 hours with 200 revs/min, then is warming up to 98 DEG C and keeps the temperature 3 minutes, be naturally cooling to 25 DEG C, be centrifuged 30 points with 5000 revs/min
Clock collects supernatant；
(4) it dialyses: supernatant is dialysed 48 hours in bag filter, dialysis medium is distilled water, the volume for medium of dialysing
It is 10 times of supernatant volume, every primary dialysis medium of 12 hours replacement obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin, the vacuum
The condition of freeze-drying are as follows: material thickness 5mm sets pre-freezing temperature as -25 DEG C, keeps after sample temperature drops to set temperature
1.5 hours, sublimation temperature is set as 10 DEG C, resolution temperature is 35 DEG C, vacuum degree 20Pa, drying time 15 hours.
The mixed enzyme is papain and the flavor protease mixture of 2:1 in mass ratio.
The dispersing agent is the mixture of hydroxypropyl-β-cyclodextrin and Sulfobutyl ether β _ cyclodextrin, the hydroxy propyl-Beta-ring
The mass ratio of dextrin and Sulfobutyl ether β _ cyclodextrin is 3:1.
The fat acid neutralizing agent is sodium carbonate.
Test case 1
The removing DPPH ability of the intensive reparation elastin laminin of the skin that embodiment is prepared is tested, specifically
Test result is shown in Table 1.
1 skin of table intensively reparation with elastin laminin remove DPPH aptitude tests result table
| Remove DPPH ability, % | |
| Embodiment 1 | 47.6 |
| Embodiment 2 | 58.3 |
| Embodiment 3 | 67.1 |
| Embodiment 4 | 54.4 |
| Embodiment 5 | 59.2 |
| Embodiment 6 | 65.3 |
| Embodiment 7 | 73.8 |
Test case 2
Skin is mouse skin moisture content, transparent intensively in influence experiment of the reparation elastin laminin to mice aging model
Matter acid content test result table is shown in Table 2-3.Data difference compares using the t method of inspection between each group, and P < 0.05 is that difference has system
Meter learns meaning.Compared with model group, experimental group data P < 0.05, compared to the blank group, model group data P < 0.05.
2 skin water cut test result table of table
3 hyaluronic acid contents test result table of table
Test case 3
The improvement skin effects of the intensive reparation elastin laminin of the skin that embodiment is prepared are tested, specific to survey
Test result is shown in Table 4.
Test method: by 210 30-40 years old healthy womens, skin is stemness, is randomly divided into 7 groups, every group of 30 people,
The skin of embodiment 1-7 intensively reparation elastin laminin is taken respectively, is taken once a day, each 3g, the warm water with 80 DEG C is molten
Solution is taken later, and Time of Administration 90 days.
Testing standard:
Effective: skin is obviously improved, and microgroove significantly reduces, and skin keeps wet.
Effective: skin makes moderate progress, and microgroove is reduced, and skin keeps relatively wet.
Invalid: skin does not have any improvement.
Table 4 improves skin effects test result table
Claims (9)
1. a kind of skin intensively repairs the preparation method with elastin laminin, which comprises the following steps:
(1) it pre-processes；
(2) it digests；
(3) it dialyses；
(4) dry.
2. a kind of skin intensively repairs the preparation method with elastin laminin, which is characterized in that include the following steps, described part is
Parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into (1-4) mm × (1-4) mm × (1-4) mm
Block mixes 10-20 parts of blocks and 40-80 parts of 3-6wt% aqueous sodium carbonates, at 40-60 DEG C of stirring 20-40 points
Clock, filtering are washed with water to washing lotion pH as neutrality, drain surface moisture, obtain pretreated feedstock；
(2) it digests: pretreated feedstock and 40-80 parts of water being mixed, stirred 5-10 minutes, 0.01-0.1 parts of trypsase are added,
The aqueous sodium carbonate for being 5wt% with mass fraction adjust pH be 7.0-8.5,45-55 DEG C stirring 2-4 hours, add
The lemon acid for adjusting pH that 0.02-0.2 parts of mixed enzymes are 10% with mass fraction is 6.0-7.0,50-60 DEG C stirring 4-6 hours,
Be warming up to 90-100 DEG C of heat preservation 2-5 minutes again, be naturally cooling to 15-30 DEG C, with 4000-6000 revs/min centrifugation 20-30 minutes,
Collect supernatant；
(3) it dialyses: supernatant is dialysed 36-72 hours in bag filter, dialysis medium is distilled water, and the volume for medium of dialysing is
8-12 times of supernatant volume, every primary dialysis medium of 10-15 hour replacement, obtains dialyzate；
(4) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin.
3. a kind of skin intensively repairs the preparation method with elastin laminin, which is characterized in that include the following steps, described part is
Parts by weight:
(1) pre-process: fresh ox paxwax removes fat and connective tissue, is cut into (1-4) mm × (1-4) mm × (1-4) mm
Block mixes 10-20 parts of blocks and 40-80 parts of 3-6wt% aqueous sodium carbonates, at 40-60 DEG C of stirring 20-40 points
Clock, filtering are washed with water to washing lotion pH as neutrality, drain surface moisture, obtain pretreated feedstock；
(2) degreasing: pretreated feedstock and 40-80 parts of water are mixed, and 0.01-0.05 parts of lipase, 0.1-0.3 parts of dispersions are added
Agent, 0.1-0.3 parts of fatty acid neutralizing agents, it is 8-10 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH, in 40-60
DEG C stirring 90-180 minute, reaction process maintain reaction pH be 8-10, end of reaction, filter, 25-35 parts of 3-6%wt of filter cake
Aqueous sodium carbonate washs 1-3 time, be washed with water wash to washing lotion pH be it is neutral, drain surface moisture, obtain pure elastin laminin material
Material；
(3) it digests: the pure elastin laminin material that step (2) is obtained and 40-80 parts of water mixing, with 100-300 revs/min of stirring 5-
10 minutes, 0.01-0.1 parts of trypsase are added, it is 7.0-8.5 that the aqueous sodium carbonate for being 5wt% with mass fraction, which adjusts pH,
45-55 DEG C stirring 2-4 hours, add 0.02-0.2 parts of mixed enzymes, the lemon acid for adjusting pH for being 10% with mass fraction is
6.0-7.0,50-60 DEG C stirring 4-6 hours, then be warming up to 90-100 DEG C of heat preservation 2-5 minutes, be naturally cooling to 15-30 DEG C, with
4000-6000 revs/min centrifugation 20-30 minutes, collect supernatant；
(4) it dialyses: supernatant is dialysed 36-72 hours in bag filter, dialysis medium is distilled water, and the volume for medium of dialysing is
8-12 times of supernatant volume, every primary dialysis medium of 10-15 hour replacement, obtains dialyzate；
(5) dry: dialyzate being subjected to vacuum freeze drying, obtains skin intensively reparation elastin laminin.
4. skin as claimed in claim 2 or claim 3 intensively repairs the preparation method with elastin laminin, which is characterized in that described true
The condition of vacuum freecing-dry are as follows: material thickness 3-6mm sets pre-freezing temperature as -20~-25 DEG C, when sample temperature drops to setting
It is kept for 1-2 hours after temperature, sets sublimation temperature as 5-15 DEG C, resolution temperature is 30-35 DEG C, vacuum degree 10-30Pa, when dry
Between 10-20 hours.
5. skin as claimed in claim 2 or claim 3 intensively repairs the preparation method with elastin laminin, which is characterized in that described mixed
Synthase is papain and flavor protease (1-3) in mass ratio: the mixture of (1-3).
6. skin as claimed in claim 3 intensively repairs the preparation method with elastin laminin, which is characterized in that the fatty acid
Neutralizer is sodium carbonate and/or sodium hydroxide.
7. skin as claimed in claim 3 intensively repairs the preparation method with elastin laminin, which is characterized in that the dispersing agent
For hydroxypropyl-β-cyclodextrin and/or Sulfobutyl ether β _ cyclodextrin.
8. skin as claimed in claim 7 intensively repairs the preparation method with elastin laminin, which is characterized in that the dispersing agent
For the mixture of hydroxypropyl-β-cyclodextrin and Sulfobutyl ether β _ cyclodextrin, the hydroxypropyl-β-cyclodextrin and sulphur butyl-β-ring paste
The mass ratio of essence is (1-5): 1.
9. a kind of skin is intensively repaired with elastin laminin, which is characterized in that be prepared into using any one of claim 1-8 method
It arrives.
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| CN109680031A (en) * | 2019-03-08 | 2019-04-26 | 福建康是美生物科技有限公司 | A kind of extracting method of Elastin peptide |
| CN111481458A (en) * | 2020-04-10 | 2020-08-04 | 上海清冬闻晓电子商务有限公司 | Nutrition restoration composition for enhancing postoperative effect of medical and beauty treatment |
| CN111955592A (en) * | 2020-08-31 | 2020-11-20 | 中恩（天津）医药科技有限公司 | Preparation method of elastin peptide |
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