CN106046150A - Apparatus for isolating collagen by ultrasonic wave - Google Patents
Apparatus for isolating collagen by ultrasonic wave Download PDFInfo
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- CN106046150A CN106046150A CN201610421284.0A CN201610421284A CN106046150A CN 106046150 A CN106046150 A CN 106046150A CN 201610421284 A CN201610421284 A CN 201610421284A CN 106046150 A CN106046150 A CN 106046150A
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- collagen protein
- cooling water
- ultrasound wave
- collagen
- cooling
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- 108010035532 Collagen Proteins 0.000 title claims abstract description 242
- 102000008186 Collagen Human genes 0.000 title claims abstract description 242
- 229920001436 collagen Polymers 0.000 title claims abstract description 34
- 241000251468 Actinopterygii Species 0.000 claims abstract description 84
- 238000001816 cooling Methods 0.000 claims abstract description 74
- 239000000498 cooling water Substances 0.000 claims description 95
- 238000000034 method Methods 0.000 claims description 89
- 238000002604 ultrasonography Methods 0.000 claims description 63
- 230000008569 process Effects 0.000 claims description 60
- 238000012360 testing method Methods 0.000 claims description 59
- 238000005204 segregation Methods 0.000 claims description 33
- 230000033228 biological regulation Effects 0.000 claims description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 18
- 230000002093 peripheral effect Effects 0.000 claims description 10
- 238000003860 storage Methods 0.000 claims description 3
- 239000002253 acid Substances 0.000 abstract description 31
- 238000000926 separation method Methods 0.000 abstract description 14
- 238000007599 discharging Methods 0.000 abstract 2
- 230000000903 blocking effect Effects 0.000 abstract 1
- 238000002955 isolation Methods 0.000 abstract 1
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 105
- 238000009210 therapy by ultrasound Methods 0.000 description 41
- 239000000203 mixture Substances 0.000 description 23
- 239000000284 extract Substances 0.000 description 18
- 230000000052 comparative effect Effects 0.000 description 14
- 239000002904 solvent Substances 0.000 description 13
- 108010010803 Gelatin Proteins 0.000 description 11
- 239000008273 gelatin Substances 0.000 description 11
- 229920000159 gelatin Polymers 0.000 description 11
- 235000019322 gelatine Nutrition 0.000 description 11
- 235000011852 gelatine desserts Nutrition 0.000 description 11
- 239000006228 supernatant Substances 0.000 description 11
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 10
- 238000005119 centrifugation Methods 0.000 description 8
- 238000004519 manufacturing process Methods 0.000 description 8
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 8
- 102000057297 Pepsin A Human genes 0.000 description 7
- 108090000284 Pepsin A Proteins 0.000 description 7
- 229940111202 pepsin Drugs 0.000 description 7
- 108090000765 processed proteins & peptides Proteins 0.000 description 7
- 238000012545 processing Methods 0.000 description 7
- 230000000694 effects Effects 0.000 description 6
- 239000003292 glue Substances 0.000 description 6
- 229920001184 polypeptide Polymers 0.000 description 6
- 102000004196 processed proteins & peptides Human genes 0.000 description 6
- 235000018102 proteins Nutrition 0.000 description 6
- 102000004169 proteins and genes Human genes 0.000 description 6
- 108090000623 proteins and genes Proteins 0.000 description 6
- 238000000605 extraction Methods 0.000 description 5
- 239000000499 gel Substances 0.000 description 5
- 239000000463 material Substances 0.000 description 5
- 239000002244 precipitate Substances 0.000 description 5
- 239000011780 sodium chloride Substances 0.000 description 5
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 4
- 238000001962 electrophoresis Methods 0.000 description 4
- 238000005194 fractionation Methods 0.000 description 4
- 210000002356 skeleton Anatomy 0.000 description 4
- 241001672694 Citrus reticulata Species 0.000 description 3
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 3
- 210000004204 blood vessel Anatomy 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 230000029087 digestion Effects 0.000 description 3
- 238000010438 heat treatment Methods 0.000 description 3
- 239000008213 purified water Substances 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 2
- 102000029816 Collagenase Human genes 0.000 description 2
- 108060005980 Collagenase Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 241000269795 Lateolabrax japonicus Species 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- 108010050808 Procollagen Proteins 0.000 description 2
- 108010077465 Tropocollagen Proteins 0.000 description 2
- 208000018756 Variant Creutzfeldt-Jakob disease Diseases 0.000 description 2
- 230000004913 activation Effects 0.000 description 2
- 235000001014 amino acid Nutrition 0.000 description 2
- 150000001413 amino acids Chemical class 0.000 description 2
- OHJMTUPIZMNBFR-UHFFFAOYSA-N biuret Chemical compound NC(=O)NC(N)=O OHJMTUPIZMNBFR-UHFFFAOYSA-N 0.000 description 2
- 208000005881 bovine spongiform encephalopathy Diseases 0.000 description 2
- 239000004202 carbamide Substances 0.000 description 2
- 235000013877 carbamide Nutrition 0.000 description 2
- 210000000845 cartilage Anatomy 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 229960002424 collagenase Drugs 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 102000038379 digestive enzymes Human genes 0.000 description 2
- 108091007734 digestive enzymes Proteins 0.000 description 2
- 230000007613 environmental effect Effects 0.000 description 2
- 229940088598 enzyme Drugs 0.000 description 2
- DNJIEGIFACGWOD-UHFFFAOYSA-N ethyl mercaptane Natural products CCS DNJIEGIFACGWOD-UHFFFAOYSA-N 0.000 description 2
- 210000003205 muscle Anatomy 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 210000003491 skin Anatomy 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 210000001835 viscera Anatomy 0.000 description 2
- DGVVWUTYPXICAM-UHFFFAOYSA-N β‐Mercaptoethanol Chemical compound OCCS DGVVWUTYPXICAM-UHFFFAOYSA-N 0.000 description 2
- CTRXDTYTAAKVSM-UHFFFAOYSA-N 3-{[ethyl({4-[(4-{ethyl[(3-sulfophenyl)methyl]amino}phenyl)(2-sulfophenyl)methylidene]cyclohexa-2,5-dien-1-ylidene})azaniumyl]methyl}benzene-1-sulfonate Chemical compound C=1C=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C(=CC=CC=2)S(O)(=O)=O)C=CC=1N(CC)CC1=CC=CC(S(O)(=O)=O)=C1 CTRXDTYTAAKVSM-UHFFFAOYSA-N 0.000 description 1
- 208000007212 Foot-and-Mouth Disease Diseases 0.000 description 1
- 241000710198 Foot-and-mouth disease virus Species 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- ZPIILPIOQFBBCP-UHFFFAOYSA-N SC(C)O.SC(C)O Chemical compound SC(C)O.SC(C)O ZPIILPIOQFBBCP-UHFFFAOYSA-N 0.000 description 1
- 101710172711 Structural protein Proteins 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 229940105402 brillant blue Drugs 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- NKLPQNGYXWVELD-UHFFFAOYSA-M coomassie brilliant blue Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=C1 NKLPQNGYXWVELD-UHFFFAOYSA-M 0.000 description 1
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- 238000004090 dissolution Methods 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 239000004744 fabric Substances 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- FBBDOOHMGLLEGJ-UHFFFAOYSA-N methane;hydrochloride Chemical compound C.Cl FBBDOOHMGLLEGJ-UHFFFAOYSA-N 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 230000003020 moisturizing effect Effects 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
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- 229920002401 polyacrylamide Polymers 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 235000021075 protein intake Nutrition 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 238000000527 sonication Methods 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
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- 208000011117 substance-related disease Diseases 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- -1 trihydroxy methyl amino Chemical group 0.000 description 1
- 238000002137 ultrasound extraction Methods 0.000 description 1
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Classifications
-
- 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, cold insoluble globulin [CIG]
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
- A23J1/00—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites
- A23J1/001—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites from waste materials, e.g. kitchen waste
- A23J1/002—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites from waste materials, e.g. kitchen waste from animal waste materials
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
- A23J1/00—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites
- A23J1/10—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites from hair, feathers, horn, skins, leather, bones, or the like
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L29/00—Foods or foodstuffs containing additives; Preparation or treatment thereof
- A23L29/20—Foods or foodstuffs containing additives; Preparation or treatment thereof containing gelling or thickening agents
- A23L29/275—Foods or foodstuffs containing additives; Preparation or treatment thereof containing gelling or thickening agents of animal origin, e.g. chitin
- A23L29/281—Proteins, e.g. gelatin or collagen
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
Abstract
An apparatus for isolating collagen by ultrasonic wave is provided to reduce acid use and to improve the yield of collagen. An apparatus for isolating collagen by ultrasonic wave comprises: a sample tank containing fish skin; a separation unit which separates collagen and supplies the collagen to the sample tank; an ultrasonic wave generation unit which is connected to the separation unit to generate ultrasonic wave; a cooling unit for blocking heat generated during collagen isolation; a sample feeding tube for supplying the fish skin from the sample tank to the separation unit; a sample discharging tube for supplying the isolated collagen to the sample tank; a circulation pump for circulating the fish skin and collagen along the sample feeding tube and sample discharging tube; and a circulation amount controller.
Description
The application is filing date December in 2011 30, and Application No. 201180063095.6, invention and created name is
The divisional application of the Chinese invention patent application " utilizing the method and device of ultrasonic extraction collagen protein ".
Technical field
The invention relates to a kind of collagen protein segregation apparatus utilizing ultrasound wave.
Background technology
Collagen protein is that one is included in nearly all tissues such as skin, blood vessel, skeleton, internal organs, and accounts for constituting body
The main protein of the 30% of the protein of body.About 40% constituted in the collagen protein of human body is present in skin, and 20% comprises
In skeleton and cartilage, except this outside, it is widely distributed in blood vessel, internal organs etc..This collagen protein is the most always with bright
The forms such as glue eat, and this gelatin is decomposed by the enzyme in digestive tract recently and absorbs with polypeptide, amino acid whose form, and these are many
Peptide, aminoacid improve immunologic function, promote that the regeneration of cell makes joint strong, and maintain the metabolic activation of skin
And moisturizing power, thus beautifying skin is played remarkable effect.Collagen protein mainly extracts from the skin of domestic animal, skeleton, joint etc.,
But immediate cause BSE (Bovine Spongiform Encephalopathy, bovine spongiform encephalopathy), foot and mouth disease etc. generation and to animality
The impression of collagen protein is poor, and the value of the marine collagen albumen therefore extracted from the skin of Fish, squama etc. is gradually increased.
On the other hand, used the chemical substance of acid, alkali, salt etc. such as to extract collagen protein in the past, but exist and therefore produce
The problem points such as environmental pollution and waste water processing cost.
Therefore, reality is for requiring to research and develop following method: the most do not use chemical substance, and can environmental protection
Ground extracts collagen protein.
Summary of the invention
It is an object of the invention to provide a kind of method that utilization extracts collagen protein by ultrasonic Treatment, by chemicals
The usage amount of matter minimizes and makes output capacity maximally extract method and the device thereof of collagen protein from fish skin.
In order to reach described purpose, in a specific example of the present invention, it is provided that a kind of method extracting collagen protein, its bag
Containing following steps: the 1st step, it is in the acetic acid solution of 0.01～0.5M, and fish skin carries out the ultrasound wave of 0.1 to 10 hour
Process and obtain collagen protein extract；And second step, it is from described collagen protein extract, separates collagen protein and warp
The fish skin of ultrasonic Treatment；And the described fish skin through ultrasonic Treatment is repeated the 1st step and second step.It addition, it is described
Ultrasonic Treatment can realize under the frequency of 20kHz, and the amplitude of described ultrasound wave can be 75～85%, and described ultrasonic Treatment can
Perform at 0 to 10 DEG C, described collagen protein extract centrifugation is obtained supernatant, add sodium chloride to this supernatant
And make precipitation can extract or separate collagen protein (NaCl), in the case of carrying out 8 described 1st steps and second step, super
The amplitude of sound wave can perform 24 hours with 20～40%.
In a concrete example, it is provided that utilize described method and the collagen protein that extracts.
In a concrete example, it is provided that a kind of method extracting collagen protein, it comprises the steps of: the 1st step, its be
In acid solution, fish skin is carried out ultrasonic Treatment and obtains collagen protein extract；And second step, it is from described collagen
Protein extract, separates collagen protein and the fish skin through ultrasonic Treatment；And repeat to hold to the described fish skin through ultrasonic Treatment
Row the 1st step and second step.
In a concrete example, it is provided that a kind of collagen protein segregation apparatus utilizing ultrasound wave, its spy is just to comprise: test portion
Case, it houses fish skin；Separative element, its reception is supplied from the described fish skin of described test portion case and is separated into collagen protein, by institute
The described collagen protein separated supplies again to described test portion case；Ultrasonic wave generator unit, it is connected to described separative element, with
Just the ultrasound wave in order to described fish skin to be separated into described collagen protein is produced.
The collagen protein segregation apparatus utilizing described ultrasound wave further includes to produce when intercepting the described collagen protein of separation
Heat, to described separative element side supply cooling water cooling end, described separative element can comprise: process portion, and it receives supply
From the fish skin of described test portion case, this fish skin is separated into collagen protein；And the 1st cooling water inflow and outflow portion, it is from described process
The outer peripheral face in portion separates certain intervals and closes the outside in described process portion, thus provide can make to be supplied from described cold
But the space of the described cooling current into and out in portion.
Described test portion case can comprise: reservoir, and it stores described fish skin；And the 2nd cooling water inflow and outflow portion, it is from described
The outer peripheral face of reservoir separates certain intervals and closes the outside of described reservoir, thus provides and can make to be supplied from described cooling end
The space of described cooling current into and out.
Described ultrasonic wave generator unit can comprise: at least one ticker, its be combined in described process portion outer peripheral face and
Produce ultrasound wave；And vibration regulation controller, it regulates the output degree of described ticker.
Described ticker can be following multiple ticker: be isolated from each other in the range of 5～6cm configuration, in order to mutually it
Between will not be affected by ultrasound wave.
The frequency of the ultrasound wave produced by described ticker can be 20kHz.
The output of described vibration regulation controller can be 0.1～1000W.
Described cooling end can comprise: cooling body；1st cooling water flows into pipe, and it is from the lower lateral institute of described cooling body
The downside stating the 1st cooling water inflow and outflow portion extends, in order to cooling water can flow into described 1st cooling water from described cooling body
Outflow portion side inflow；1st cooling water flow out of pipe, and it is from the upper lateral described cooling body in described 1st cooling water inflow and outflow portion
Upside extend, in order to cooling water can from described 1st cooling water inflow and outflow portion flow out to described cooling body side；2nd cooling
Water flows into pipe, and it extends from the downside in the lower the most described 2nd cooling water inflow and outflow portion of described cooling body, in order to cooling water
Can be from described cooling body to described 2nd cooling water inflow and outflow portion side inflow；And the 2nd cooling water flow out of pipe, it is from the described 2nd
The upside of the upper lateral described cooling body in cooling water inflow and outflow portion extends, in order to cooling water can be from described 2nd cooling current
Enter outflow portion to flow out to described cooling body side.
The collagen protein segregation apparatus utilizing described ultrasound wave can further include: test portion supply pipe, it is by described reservoir
Downside is connected with each other with the downside in described process portion, in order to can be from described reservoir to side, described process portion supply fish skin；And examination
Material discharge pipe, the upside in described process portion is connected with each other by it with the upside of described reservoir, in order to can be to described examination bin side
Discharge the collagen protein utilizing described separative element and separate.
The collagen protein segregation apparatus utilizing described ultrasound wave can further include: circulating pump, and it possesses and supplies at described test portion
On the prolongation path of pipe, in order to can forcibly make fish skin and described collagen protein along described test portion supply pipe and described test portion row
Go out pipe circulation；And circulating load regulation controller, it regulates the driver of described circulating pump, in order to fish skin described in scalable and institute
State the circulating load of collagen protein.
" fish skin " of the present invention refers to, comprises from availability halobiontic marine collagen albumen, has removed Fish
Squama.The characteristic being somebody's turn to do " fish skin " is when only carrying out ultrasonic Treatment without acid solvent, it is difficult to cause collagen fabric
Structure change, thus in order to realize the separation of collagen protein, it is necessary to need acid solvent.In the present invention, at acid solvent
In the presence of parallel ultrasonic Treatment, therefore in the acid solvent of Cmin, it is also possible to higher output capacity dissolution glue
Former albumen.Therefore, can significantly reduce the acid amount used when separating collagen protein.
" collagen protein " of the present invention is as the fibre constituting the conjunctive tissue of animal, skeleton, muscle, skin, cartilage, blood vessel etc.
Dimension shape structural protein, basic structural unit is tropocollagen (tropocollagen), and refers to have and be about by molecular weight
The triple helices structure that 3/100000ths polypeptide are constituted, 3 polypeptide chains are stabilisation each other by hydrogen bond knot, if but heating,
The gelatin cutting off these minority bonds and become on random coil, thus physical property can be changed.
" ultrasonic Treatment " of the present invention is that ultrasonic energy makes the particle vibration of destination object destroy object or make non-
Activation, in biochemical, mainly use for the purpose of destroying cell membrane and discharge cellular content, and is not limited thereto, make
Sound wave with the vibration number higher than audio frequency region (about below 20kHz).
[effect of invention]
The collagen protein extracting method of the present invention can pass through to repeat to carry out ultrasonic Treatment, and more existing collagen protein carries
Access method reduces the concentration of acid solution, extracts collagen protein with higher output capacity.It addition, according to the collagen protein of the present invention
Extracting method, can directly with original basic structure extract high molecular collagen protein and and the hydrolyzate shape of noncollagen protein
State.
Accompanying drawing explanation
Fig. 1 to Fig. 4 is the method that represents and utilize embodiment 1 to 4 to separate the collagen protein in the case of collagen protein
Separate output capacity.
Fig. 5 is the separation output capacity representing the collagen protein in the case of utilizing the method for comparative example 1 to 4 to separate.
Fig. 6 is the maximum output capacity representing the collagen protein in the case of utilizing the method for embodiment 1 to 4 to separate.
Fig. 7 is the maximum output capacity representing the collagen protein in the case of utilizing the method for comparative example 1 to 4 to separate.
Fig. 8 is the method that represents and utilize embodiment 1 and 3 to separate the collagen protein separated in the case of collagen protein
SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis, polyacrylamide
Amine gel electrophoresis) pattern.
Fig. 9 is the method that represents and utilize comparative example 2 to 4 to separate the collagen protein separated in the case of collagen protein
SDS-PAGE pattern.
Figure 10 is to represent to carry out ultrasonic Treatment in the presence of acid solvent, until representing and carrying out with the acetic acid with 0.5M
Time required in the case of till processing identical output capacity for 24 hours.
Figure 11 is in the case of making separated collagen protein and collagen protein gelatinization, assesses eupepsy and proves glue
The result of former albumen.
Figure 12 is the chart of the extraction output capacity of the collagen protein representing the number of repetition according to ultrasonic Treatment.
Figure 13 is the SDS-PAGE pattern representing the collagen protein separated according to the number of repetition of ultrasonic Treatment.
Figure 14 is the outline ideograph of the collagen protein segregation apparatus utilizing ultrasound wave of one embodiment of the invention.
Figure 15 is the outline ideograph of the separative element of the collagen protein segregation apparatus utilizing ultrasound wave of Figure 14.
Figure 16 is the collagen protein segregation apparatus utilizing ultrasound wave representing and using Figure 14, separates the method for collagen protein
Precedence diagram.
Figure 17 is the collagen protein segregation apparatus utilizing ultrasound wave representing and using Figure 14, in the case of separating collagen protein
The chart of collagen protein output capacity.
Figure 18 is to represent by SDS-electrophoresis method, analyze use Figure 14 the collagen protein segregation apparatus utilizing ultrasound wave and
The chart of the result of the collagen protein separated.
[explanation of symbol]
100 collagen protein segregation apparatuss
110 test portion casees
111 reservoir
113 the 2nd cooling water inflow and outflow portions
130 separative elements
131 process portions
133 the 1st cooling water inflow and outflow portions
150 ultrasonic wave generator unit
151 tickers
153 vibration regulation controllers
170 cooling ends
171 cooling bodies
180a test portion supply pipe
180b test portion discharge pipe
191 circulating pumps
192 circulating load regulation controllers
Detailed description of the invention
Hereinafter, according to following embodiment, the present invention is explained.But, following embodiment only illustrates the present invention, this
Bright content is not limited to following embodiment.
Embodiment
The pre-treatment of embodiment 1. test portion
Test portion is to receive the freezing fish skin of the Lateolabrax japonicus (Cuvier et Va-lenciennes) (Lateolabracis) being supplied from O Sung fish farm (stock) and use.Fish skin is residual in removal
After staying Lateolabrax japonicus (Cuvier et Va-lenciennes) (Lateolabracis) muscle inside a part and squama, frozen water is utilized to clean and go the removal of impurity, thus big with 1.0cm × 1.0cm
Little essence is cut.In order to fully remove the salt soluble protein being attached to fish skin, at the 0.5M adding 20 times relative to fish skin weight
NaCl solution after, be stirred equably and 6, under 000rpm, carry out the centrifugation of 10 minutes, thus remove supernatant.
Repeat 3 operations as above, and all operations is all carried out below 4 DEG C.Again clear to take care of the Purified Water below 4 DEG C
After washing the precipitate obtained by centrifugation, add the cold ethanol of about 20 times of weight relative to precipitate and one side at 4 DEG C
Stir the defat of 24 hours one sides, thus it is set to refine test portion.Refined test portion keeping, below-20 DEG C, is taken out when needed
Use.
The extraction of embodiment 2. collagen protein
2-1. utilizes the extraction of the collagen protein that ultrasonic Treatment carries out
After the acetic acid of 0.01 to the 0.5M of the fish skin about 200 times of weight of interpolation prepared in described embodiment 1,4
With the condition of table 1 below at DEG C, carry out ultrasonic Treatment.The on/off of pulse is to carry out with 20sec/20sec.With 6,
After 000rpm carries out the centrifugation of 10 minutes to the viscosity solution obtained after carrying out ultrasonic Treatment, separate supernatant, will
NaCl adds to supernatant in the way of becoming 5wt%, thus obtains white precipitate.It is centrifuged described white precipitate separating
And after dialysing into Purified Water, carry out freeze-dried and extract collagen protein.
Table 1
The extraction of 2-2. acid-soluble collagen albumen
With the condition of table 2 below, the acetic acid relative to the 0～0.5M of test portion weight interpolation 200 times, thus one side is at 4 DEG C
24 hours one sides of lower stirring are extracted.In the centrifugation carrying out 10 minutes with the 6,000rpm viscosity solution to being obtained
After, separate supernatant, and NaCl is added to supernatant in the way of becoming 5wt%, thus obtain white precipitate.To described
After white precipitate is centrifuged separating and dialyses into Purified Water, carry out freeze-dried and extract collagen protein.
Table 2
| Distinguish | Acetic acid concentration (M) |
| Comparative example 1 | 0 |
| Comparative example 2 | 0.01 |
| Comparative example 3 | 0.1 |
| Comparative example 4 | 0.5 |
2-3. utilize the extraction repeating the collagen protein that ultrasonic Treatment is carried out
After the acetic acid of the 0.01M adding about 200 times of weight at the fish skin that prepared in described embodiment 1, at 4 DEG C with
The amplitude of 40% to 80% carries out the ultrasonic Treatment of 3 hours.The on/off of pulse is to carry out with 20sec/20sec.With 6,
After the 000rpm viscosity solution to being obtained after carrying out ultrasonic Treatment carries out the centrifugation of 10 minutes, separate supernatant (glue
Former albumen), and after the acetic acid of the 0.01M again adding 200 times of weight to precipitation (residue), with 40% to 80% at 4 DEG C
Amplitude carry out the ultrasonic Treatment of 3 hours.Separate collagen protein by centrifugation, and again add to residual residue
The acetic acid of 0.01M and repetition operation is set to altogether 4 times to 8 times, thus separate collagen protein.Altogether through 4 to 8 as inciting somebody to action as mentioned above
The supernatant that subjob is collected is freeze-dried and extracts collagen protein (with reference to Figure 12).
The mensuration of embodiment 3. collagen protein output capacity
Utilize Biuret (biuret) method (Gornall, A, G. etc., 1949), supernatant is measured protein content, on this
Clear liquid be under 15,000 × g, the test portion obtained in described manufacture example and comparative example is carried out 20 minutes centrifugation and
Obtain.Collagen protein output capacity is after total protein content is carried out ultrasonic Treatment, carrys out basis with the ratio of protein content
Following formula and calculate.
Collagen protein output capacity (%)=(protein concentration/total protein concentration in supernatant) × 100
Fig. 1 to Fig. 5 will be shown according to the collagen protein output capacity of the time of process.Fig. 1 to 4 represents according to manufacturing example 1 to 4
The output capacity of the collagen protein of acetic acid concentration, Fig. 5 represents the output capacity of the collagen protein of the acid concentration according to comparative example 1 to 5.Root
According to Fig. 1 to 4, it is expressed as owing to carrying out ultrasonic Treatment, and the output capacity of collagen protein less carries out Fig. 5 of ultrasonic Treatment more
Increase soon.Especially, show as accelerating along with amplitude becomes speed that is big and that increase.According to Fig. 5, not carrying out ultrasonic Treatment and
In the case of only processing with the acetic acid of the low concentration of 0.01M, collagen protein does not almost separate, but according to Fig. 1, when
When processing with the ultrasound wave that amplitude is 20% under 20kHz, even if in the low concentration of acetic acid of 0.01M, dividing of collagen protein
Also begin to increase from amount.It addition, according to Fig. 2 to 4, show as concentration the highest, then the amplification of collagen protein is the biggest, and shows as
Under conditions of identical, amplitude is the biggest, then it gathers way the fastest.It is therefore contemplated that the separating power of collagen protein depends on acetic acid
Concentration and amplitude of ultrasonic and increase.
It addition, the output capacity of the collagen protein obtained repeating to extract by ultrasound wave is shown in Figure 12.Carried out with 3 hours
8 amplitudes be the collagen protein of the ultrasonic Treatment of 40% output capacity relatively by conventional method i.e. in the acetic acid of 0.01M,
The output capacity of the collagen protein extracting 24 hours increases by more than 3 times.And, carrying out 4 amplitudes with 3 hours is the ultrasound wave of 80%
In the method i.e. acetic acid of 0.01M that the output capacity of collagen protein that processes is more conventional, extract the output of the collagen protein of 12 hours
Rate about increases about 2 times.Therefore, it is known that when carrying out ultrasound wave and repeating to extract, the output capacity of collagen protein carries with ultrasound wave
Take number of times to be directly proportional.
The maximum output capacity of ＜ collagen protein measures ＞
Utilize the method identical with described manufacture example 1, measure the output capacity of collagen protein, when output capacity increases, pass through
Following formula calculates its speed (ki).
Ki=(nt-no)
Nt: carry out the dissolubility after the ultrasonic Treatment of t hour
No: the dissolubility before ultrasonic Treatment
T: ultrasonic treatment time
The results are shown in Fig. 6 and 7.Fig. 6 represents and carries out, with ultrasound wave, the feelings that process in the acid solvent manufacturing example 1 to 4
Condition and the collagen protein maximum output capacity in the case of only processing with the acid solvent of comparative example 1 to 4.In figure 6, respectively
Mark represents 0% (●), 20% (zero), 40% (), 60% () and the amplitude of 80% (■).Fig. 7 represents that collagen protein produces
Go out gathering way of rate.
According to Fig. 6, the concentration of acetic acid more increases the amplitude of i.e. ultrasound wave and more increases, then the maximum output capacity table of collagen protein
Now the highest, maximum output capacity when acetic acid concentration is 0.1M is similar to for maximum output capacity during 0.5M, therefore can speculate such as
Lower situation: even if acetic acid concentration increases to more than 0.5M, the increase of maximum output capacity is the most insignificant.It addition, without
Acetic acid and in the case of carrying out ultrasonic Treatment, cause the separation of collagen protein the most hardly.Entering merely with acid solvent
In the case of the comparative example that row processes, along with acetic acid concentration increase, maximum output capacity also increases, but gathers way and be slower than parallel
The situation of ultrasonic Treatment.It addition, in the case of acetic acid concentration is 0M, the separation of collagen protein will not be caused, thus understands
Acid is needed in the separation of collagen protein.
According to Fig. 7, observe in detail the amplitude of the ultrasound wave involving impact that gathers way on collagen protein output capacity with
The relation of acetic acid concentration, result, no matter under the conditions of which kind of, all shows line relationship, and the relational expression of each line relationship is such as
Under.That is, the acetic acid under 0.01M is y=0.0198x+0.2296 with the relational expression of amplitude, under 0.1M, and y=0.0418x+
0.6832, under 0.5M, y=0.044x+1.2633.According to this formula, gradient is different according to acetic acid concentration, compares
The acetic acid of 0.01M, gradient is added above at 0.1M, thus in the presence of the acetic acid of more than 0.1M, causes collagen rapidly
The separation of albumen.It addition, compare the acetic acid of 0.1M with 0.5M, gradient is almost identical, thus in the second of more than 0.5M
Under acid concentration, the output capacity of collagen protein will not be gathered way and cause large effect by acetic acid concentration.
The SDS-PAGE pattern of the collagen protein that embodiment 4. is separated
Utilize SDS electrophoresis (SDS-PAGE), the son of the collagen protein that research obtains according to described manufacture example and comparative example
Unit (subunit) forms.SDS-PAGE is by Lammli method (Lammli, V.K. etc., 1970), utilizes the slab of 7.5%
Gel (slab gel) and perform.The carbamide of 8M is added to the collagen protein test portion separated in described manufacture example and comparative example
(urea), 2% mercaptoethanol (mercaptoethanol), Tris-HCl (the trihydroxy methyl amino of SDS and 20mM of 2%
Methane hydrochloride salt) (pH value is 8.0) dissolve, thus heat 2 minutes at 100 DEG C.Fixing (necessarily) and staining (dye
Color) it is the method by Neuhoff (Neuhoff V. etc., 1988), utilize Coomassie brilliant blue (Coomassie brillant
Blue) implement.The result of the manufacture example of the ultrasonic Treatment utilizing amplitude to be 40% is shown in Fig. 8, make acetic acid concentration different and
The result of the comparative example 2 to 4 processed is shown in Fig. 9.
According to Fig. 8, in the presence of the acetic acid of 0.01M, when the ultrasound wave utilizing amplitude to be 40% processes, little 4
Be initially observed the composition being equivalent to collagen protein time after, along with ultrasonic treatment time elongated and observe effectively α 1, α 2,
β and γ chain (chain).This tendency is the polymeric composition also observed in the top of gel and be speculated as collagen protein.
In the case of processing with the amplitude of 60%, overall tendency is identical with the situation carrying out processing with the amplitude of 20%.But,
Along with ultrasonic treatment time is elongated and it is initially observed the composition of the catabolite being speculated as collagen protein.Particularly, exist
When carrying out the ultrasonic Treatment of 24 hours under the acetic acid of 0.5M, find following situation: the i.e. α of main subelement 1 and β of collagen protein
Chain reduces, and meanwhile, generates the not specific polypeptide that the background (substrate) of gel is colored.
According to Fig. 9, the subelement composition of the collagen protein only separated so that acid solvent processes is studied, knot
Fruit, under the acetic acid of 0.01M, starts to find to be equivalent to the α 1 and β chain of collagen protein in reaction after 6 hours, after 24 hours, with
This α 1 and β chain together finds α 2 and γ chain.This phenomenon is as acetic acid concentration and uprises and significantly more manifest, along with reaction is little
Time passage and α 1, α 2, β and γ chain also amount on increase.
As a result, when carrying out ultrasonic Treatment under acid solvent, the α 1 of collagen protein, α 2, β and γ chain are equivalent to faster
Increase fastly, thus when carrying out ultrasonic Treatment under acid solvent, cause the separation of collagen protein at short notice.
It addition, according to Figure 13, can confirm that following situation: repeat ultrasonic Treatment and the collagen protein that extracts also has
Typical collagen structure.
The confirmation of embodiment 6. collagen protein
In order to confirm to manufacture in example, parallel ultrasonic Treatment under acid solvent and the collagen protein that separates is with collagen
The form fractionation of albumen is still with the form fractionation of gelatin, and research is because of pepsin (1:10,000.Yakuri pure
Chem., co..ltd., Japan) eupepsy of collagen protein that produces.The characteristic of collagen protein is as follows: its structure is the hardest
Gu, will not be decomposed by digestive enzyme, and be decomposed by collagenase (Collagenase).But, if collagen protein is because of warm
Gelatinization, then decomposed by digestive enzyme.Therefore, utilize this characteristic, if can determine whether as to the collagen separated according to ultrasound wave
Albumen carries out pepsin and causes digestion, then be the form fractionation with gelatin, if not causing digestion, then with collagen protein
Form fractionation.
After collagen concentration is adjusted to 1mg/ml, heat 5 minutes and gelatinization at 100 DEG C.Relative to collagen egg
The concentration of Pseudobulbus Bletillae (Rhizoma Bletillae) gelatin and add the pepsin of 0.5%, thus after carrying out the process of 0～30 minute at 10 DEG C, Xiang Geshi
Material add the SDS of 8M, the mercaptoethanol (mercaptoethanol) of 2%, the Tris-HCl (pH value is 8.0) of 20mM and 100
Heat 2 minutes at DEG C, so that enzymatic activity stops.Hereafter, digestion pattern is analyzed according to SDS-PAGE (Lammli method).This
Time, collagen protein standard substance uses Acid soluble Collagen (acid-soluble collagen protein) (TypeII, from
white rabbit skin.,Sigma,USA.).Its result such as Figure 11.In fig. 11, the TYPE I's on the basis of No.1 is into
Collagen protein, No.2 is under the acetic acid of 0.01M, and the amplitude (amplitude) with 80% carries out the ultrasonic Treatment of 12 hours
And the collagen protein separated, No.3,4,5 are for be processed into pepsic result by this collagen protein.No.6 be with 100 DEG C heat at
The reason collagen protein of No.2 and gelatinization, No.7,8,9 are to analyze described gelatin is processed into pepsic result.
In fig. 11, the implication of each No. is as follows.
S:molecular weight marker
No.1:I type collagen (acid soluble)
No.2:Collagen of fish skin Isolated by sonication with acetic acid.
No.3,4,5:Collagen treated with pepsin for 10,20and 30mim.
No.6:Gelatin obtained from collagen (No.2) by heating at 100 DEG C.
No.7,8,9:Gelatin (No.6) treated with pepsin for 10,20and 30mim.
According to Figure 11, under acid solvent, the collagen protein (No.2) separated by ultrasound wave is even if carrying out pepsin
Ferment treatment, also will not cause the change (No.3,4 and 5) of main component i.e. α and the β chain of collagen protein.But, if to collagen
After albumen carries out heat treatment and makes gelatinization (No.6), carry out pepsin, then find following situation: the master of collagen protein
Want composition i.e. α and β chain to be wholly absent, low molecular composition increases (No.7,8 and 9).Therefore, according to result above, confirm
Following situation: the composition separated by ultrasound wave is collagen protein, and not gelatin or the hydrolyzate of collagen protein.
Hereinafter, with reference to alterations, the content of the preferred embodiment explaining the present invention is as follows.But, at this
In the explanation of invention, for the purport of the clear and definite present invention, omit for known function or the explanation of composition.
Figure 14 is the outline ideograph of the collagen protein segregation apparatus utilizing ultrasound wave of one embodiment of the invention, Figure 15
It it is the outline ideograph of the separative element of the collagen protein segregation apparatus utilizing ultrasound wave of Figure 14.
With reference to these figures, utilize ultrasound wave collagen protein segregation apparatus (100, hereinafter referred to as collagen protein segregation apparatus
100) comprising: test portion case 110, it houses fish skin；Separative element 130, its reception is supplied from the fish skin of test portion case 110 and is separated into
Collagen protein, thus the collagen protein separated is supplied again to test portion case 110；Ultrasonic wave generator unit 150, it is connected to
Separative element 130, in order to produce in order to the ultrasound wave that fish skin is separated into collagen protein；Cooling end 170, it is to intercept separation gel
The mode of the heat produced during former albumen possesses；Test portion supply pipe 180a, its from test portion case 110 to separative element 130 side supply fish
Skin；Test portion discharge pipe 180b, the collagen protein separated by separative element 130 is supplied to test portion case 110 side by again；Follow
Ring pump 191, it possesses on the prolongation path of test portion supply pipe 180a, in order to can forcibly make fish skin and collagen protein along examination
Material supply pipe 180a and test portion discharge pipe 180b circulation；And circulating load regulation controller 192, the driving of its regulation circulating pump 191
Degree.
Test portion case 110 is to house supply to be separated into the composition of the fish skin (test portion) of collagen protein to separative element 130 side,
Comprising: reservoir 111, it stores fish skin；And the 2nd cooling water inflow and outflow portion 113, the outside of its completely enclosed reservoir 111,
The space of the cooling current into and out that can make to be supplied from cooling end 170 is thus provided.
Reservoir 111 is to provide to store the composition of the hollow cylindrical in the space of the fish skin as test portion.Certainly, originally
The claim of invention is not limited because of reservoir 111 shape, and according to other embodiments of the invention, reservoir 111 also is able to
Possess with tetragon etc..
In reservoir 111, not only store fish skin, and together store when fish skin is separated into collagen protein required
Acid solution, this acid solution uses from the past to be separated from fish skin by collagen protein always, therefore omits detailed
Explanation.
The downside of reservoir 111 is to supply pipe 180a by the test portion of description to be connected to the downside of separative element 130
(specifically, for the downside in process portion 131), thus fish skin can be supplied to separative element 130 side, described reservoir 111
Upside be upside (specifically, upper for process portion 131 being connected to separative element 130 by test portion discharge pipe 180b
Side), thus the collagen protein separated again can be discharged to reservoir 111 side.
2nd cooling water inflow and outflow portion 113 is to provide the composition in following space: in order to make cooling water along the 2nd cooling current
Entering pipe 175a flows into or makes cooling water to cooling water flow out of pipe 175b to the outflow of cooling body 171 side along the 2nd from cooling body 171.
To this end, the 2nd cooling water inflow and outflow portion 113 be separate certain intervals with the outer peripheral face from reservoir 111 and can be complete
The mode of the outside of totally-enclosed reservoir 111 possesses, be exactly based on this possess reservoir the 111 and the 2nd cooling current become a mandarin
Go out the space between portion 113, make the cooling water can inflow and outflow.Certainly, identically with described reservoir 111, the right of the present invention
Require not limited because of the shape in the 2nd cooling water inflow and outflow portion 113.
On the other hand, separative element 130 is following composition: receives and is supplied from the fish skin of test portion case 110 and is divided by this fish skin
From becoming collagen protein, thus the collagen protein separated is supplied again to test portion case 110 side.
Separative element 130 comprises: process portion 131, fish skin is separated into collagen protein by it；And the 1st cooling water inflow and outflow
Portion 133, it is by the outside in completely enclosed process portion 131, it is provided that can make to be supplied from the cooling current into and out of cooling end 170
Space.
Process portion 131 is the composition of following hollow cylindrical: be connected to the ultrasonic wave generator unit 150 of description,
Receive and be supplied from the fish skin of test portion case 110 and this fish skin be separated into collagen protein or the collagen protein separated is supplied again
Give to test portion case 110 side.Certainly, the claim of the present invention is not limited because of the shape in process portion 131.
As it has been described above, the downside in process portion 131 and the downside of reservoir 111 and the upside in process portion 131 and reservoir
The upside of 111 is connected by test portion supply pipe 180a and test portion discharge pipe 180b respectively.
On the other hand, the 1st cooling water inflow and outflow portion 133 is to provide the composition in following space: make cooling water along the 1st cooling
Water flows into pipe 173a and flows into or make cooling water to cooling water flow out of pipe 173b to cooling body 171 side along the 1st from cooling body 171
Flow out.
To this end, the 1st cooling water inflow and outflow portion 133 be separate certain intervals with the outer peripheral face from process portion 131 and can be complete
The mode of the outside in totally-enclosed process portion 131 possesses, be exactly based on this possess process portion the 131 and the 1st cooling current become a mandarin
Go out the space between portion 133, cooling water inflow and outflow can be made.Certainly, the claim of the present invention does not becomes a mandarin because of the 1st cooling current
Go out the shape in portion 133 and limited.
On the other hand, ultrasonic wave generator unit 150 is following composition: be connected to separative element 130, in order to produce in order to
Fish skin is separated into the ultrasound wave of collagen protein.
Ultrasonic wave generator unit 150 comprises: multiple tickers 151, and it is combined in the outer peripheral face in process portion 131；And vibration
Regulation controller 153, the output degree of its regulation ticker 151.
Ticker 151 is following to constitute: according to the control of vibration regulation controller 153, produces certain vibration, thus to
The inside in process portion 131 produces ultrasound wave.
Multiple tickers 151 are isolated from each other in the range of 5～6cm, in order to will not be affected by ultrasound wave to each other
(interference), the outer peripheral face along process portion 131 adheres to.In the present embodiment, the frequency of the ultrasound wave produced by ticker 151 is
About 20kHz, its reason is, under this frequency, it is possible to the highest effect, fish skin is separated into collagen protein.
But, the frequency of ultrasound wave can also be according to the capacity in process portion 131, the storage of the fish skin being stored in process portion 131
Storages etc. are altered to other numerical value.
Vibration regulation controller 153 is the composition possessed as follows: the output degree of controlled damping mover 151, i.e.
By the frequency of the ultrasound wave that ticker 151 produces.
Vibration regulation controller 153 and ticker 151 are connected to RF Wire (Radio Frequency Wire), thus
The signal controlled by vibration regulation controller 153 is provided to ticker 151.
In the present embodiment, vibration regulation controller 153 is output as 0.1～1000W, but vibration regulates controller 153
Output can also be corresponding and different from the changeable frequency of described ticker 151 scope more.
On the other hand, cooling end 170 is following composition: cool down water to the 1st cooling side, water inflow and outflow portion 133 supply, with
Just the heat produced when separating collagen protein can be intercepted.It addition, cooling end 170 is also to the 2nd cooling side, water inflow and outflow portion 113 supply
Cooling water, plays the effect effectively intercepting the heat produced when separating collagen protein the most in the lump.
Cooling end 170 comprises: cooling body 171, it makes cooling water flow in or out；1st cooling water flow into pipe 173a and
1st cooling water flow out of pipe 173b, and cooling body 171 is connected with each other by its grade with the 1st cooling water inflow and outflow portion 133；2nd cooling
Water flows into pipe 175a and the 2nd and cooling water flow out of pipe 175b, and its grade will cooling body 171 and the 2nd cooling water inflow and outflow portion 113 phase
Connect.
Cooling body 171 is following composition: store supply to the 1st cooling water inflow and outflow portion the 133 or the 2nd cooling water inflow
The cooling water of outflow portion 113.
In the side of cooling body 171, possess to regulate supply to the 1st cooling water inflow and outflow portion the 133 or the 2nd cooling
The controller 171a of the circulating load of the cooling water in water inflow and outflow portion 113, operating personnel produces when considering to separate collagen protein
Heat regulates controller 171a, thus can determine to cool down the circulating load of water.
It is following composition that 1st cooling water flows into pipe 173a: by being flowed in the downside of cooling body 171 and the 1st cooling water
The downside of outflow portion 133 is connected with each other, and cooling water can be made to supply to the 1st cooling side, water inflow and outflow portion 133 from cooling body 171
Give；1st to cooling water flow out of pipe 173b be following to constitute: by will the upside of cooling body 171 and the 1st cooling water inflow and outflow portion
The upside of 133 is connected with each other, and cooling water can be made to flow out to cooling body 171 side from the 1st cooling water inflow and outflow portion 133.
That is, cooling water can cooling water flow out of pipe 173b by the 1st cooling water inflow pipe 173a and the 1st and continue cooling down this
Body the 171 and the 1st cooling circulates between water inflow and outflow portion 133, and fish skin is separated into collagen protein by the circulation at logical supercooled water
In the case of, can effectively intercept the heat produced in process portion 131.
It is following composition that 2nd cooling water flows into pipe 175a: by being flowed in the downside of cooling body 171 and the 2nd cooling water
The downside of outflow portion 113 is connected with each other, and cooling water can be made to supply to the 2nd cooling side, water inflow and outflow portion 113 from cooling body 171
Give；2nd to cooling water flow out of pipe 175b be following to constitute: by will the upside of cooling body 171 and the 2nd cooling water inflow and outflow portion
The upside of 113 is connected with each other, and cooling water can be made to flow out to cooling body 171 side from the 2nd cooling water inflow and outflow portion 113.
That is, cooling water can cooling water flow out of pipe 175b by the 2nd cooling water inflow pipe 175a and the 2nd and continue cooling down this
Body the 171 and the 2nd cooling circulates between water inflow and outflow portion 113, and fish skin is separated into collagen protein by the circulation at logical supercooled water
In the case of, can effectively intercept the heat produced in process portion 131.
On the other hand, also it is externally supplied the former of cooling water especially to the not test portion case 110 of outside for process portion 131
Because being, the collagen protein separated in process portion 131 supplies to reservoir 111 side of test portion case 110 again.
That is, the fish skin the non-once that are housed in process portion 131 are all separated into collagen protein, the most such as by ultrasound wave
Lower described, only through for a long time, fish skin is applied ultrasound wave, a certain amount of collagen protein could be obtained.
During this, fish skin one side is separated into collagen protein constantly, one side process portion 131 and reservoir 111 it
Between circulate, therefore become to the fish skin of reservoir 111 side inflow and the admixture material of collagen protein from process portion 131 and have
The state of heat to a certain degree, it is therefore desirable to intercept this heat immediately.Thus, in the present embodiment, by respectively in process portion
The outside of 131 and the outside of reservoir 111 possess the 1st cooling water inflow and outflow portion the 133 and the 2nd and cool down water inflow and outflow portion 113,
Can effectively intercept the heat produced when fish skin is separated into collagen protein.
It addition, in the way of the downside by the downside of cooling body 171 with the 1st cooling water inflow and outflow portion 133 is connected, join
Put the 1st cooling water and flow into pipe 173a, and connect with the downside by the downside of cooling body 171 with the 2nd cooling water inflow and outflow portion 113
The mode connect, configuration the 2nd cooling water flows into when fish skin is separated into collagen protein to effectively intercept by pipe 175a and produces
Heat.
That is, the fish skin being stored in reservoir 111 (certainly, is also mixed with collagen because using protocollagen segregation apparatus 100
Albumen) it is fully filled with reservoir 111 and process portion 131 the most respectively with the supply fish skin in process portion 131, therefore it is stored in cold
But the ice-cold cooling water of body 171 supply from the downside in this reservoir 111 and process portion 131 and cool down reservoir 111 and
The heat in process portion 131, thus upwards side is flowed out, and thus can improve the cooling effect of cooling water.
On the other hand, test portion supply pipe 180a is to possess to side, process portion 131 supply fish skin from reservoir 111
Composition, test portion discharge pipe 180b be in order to can to reservoir 111 side discharge by process portion 131 separate collagen protein and have
Standby composition.It is of course also possible, as described before, here, fish skin and collagen protein refer to because use protocollagen segregation apparatus 100
Each other according to the material of state of a certain amount of mixing.
It addition, on the prolongation path of test portion supply pipe 180a, possess circulating pump 191, this circulating pump 191 is by following
Circular rector regulates controller 192 and controls its driver.
Operating personnel regulates the driver of circulating pump 191 by regulation circulating load regulation controller 192, according to circulation
The driver of pump 191, supplies a certain amount of fish skin from reservoir 111 to side, process portion 131, meanwhile, is stored in process
The collagen protein in portion 131 is also discharged to reservoir 111 side, thus fish skin and collagen protein circulate sustainably.
It addition, make circulating pump 191 possess test portion supply pipe 180a side and not test portion discharge pipe 180b side be because examining
Consider and be fully filled with reservoir 111 and process portion 131 the most respectively to the fish skin supplied in process portion 131.
The collagen protein segregation apparatus 100 of the present embodiment obtains collagen egg merely with acid solution the most as before
In vain, but ultrasound wave used along and obtain collagen protein, therefore reduce the making of acid (acid) used when separating collagen protein
Consumption, thus has the advantage that the method separation collagen protein not only by environmental protection, but also can improve glue in the lump
The output capacity of former albumen.
Meanwhile, the collagen protein segregation apparatus 100 of the present embodiment uses when being so that fish skin to be separated into collagen protein
Situation be limited and be illustrated, it is also possible to for gelatin is separated into collagen protein polypeptide.That is, the right of the present invention is wanted
Ask and not only comprise the situation that fish skin is separated into collagen protein, but also comprise and replace fish skin with gelatin, and obtain collagen protein
Polypeptide replaces the situation of collagen protein.
Figure 16 is the collagen protein segregation apparatus utilizing ultrasound wave representing and using Figure 14, separates the method for collagen protein
Precedence diagram, Figure 17 is to represent to use Figure 14's to utilize the collagen protein segregation apparatus of ultrasound wave to separate collagen during collagen protein
The chart of albumen output capacity, Figure 18 is to represent by SDS-electrophoresis method, utilizes the collagen protein of ultrasound wave to divide to using Figure 14
The chart of the result that the collagen protein separated from device is analyzed.
With reference to these figures, in order to use the collagen protein segregation apparatus 100 of the present embodiment, fish skin is separated into collagen egg
In vain, should be through following process: first, fish skin and acid solution are stored in test portion case (S1 step)；Use is connected to receive and supplies
Give from the fish skin of test portion case and the separative element of acid solution and separative element, and produce the ultrasonic of ultrasound wave within a certain period of time
Ripple generation unit, is separated into collagen protein (S2 step) by fish skin；By the collagen protein separated with unsegregated fish skin from dividing
Again it is sent to from unit try bin side (S3 step)；(S4 walks until reaching certain output capacity to repeat S2 step and S3 step
Suddenly).
By described collagen protein separation method, at certain treatment conditions (acetic acid of acid solution: 0.01M；Cooling water
Temperature: 4 DEG C；Test portion feed speed: 0.5L/ divides；The frequency of ultrasound wave: 20kHz) under, when fish skin is separated into collagen protein
Collagen protein output capacity as shown in figure 17.
That is, after carrying out the process of 2 hours, the fish skin of about substantially 15% is separated into collagen protein, after 4 hours, greatly
The fish skin causing about 33% is separated into collagen protein, and after 12h, the fish skin of about substantially 53% is separated into collagen protein.Separately
Outward, after 15 hours, even if constantly fish skin being applied ultrasound wave, also the output capacity of collagen protein will not be caused bigger shadow
Ring.
On the other hand, by SDS-electrophoresis method, being analyzed the collagen protein utilizing as above method and separate, result is such as
Figure 18 equally maintains typical collagen structure, processes separated collagen egg even with the enzyme beyond removing glue protoenzyme
In vain, also will not cause decomposition, thus can confirm that separated composition is collagen protein.
Above, specific embodiment is described and illustrated, but the present invention is not limited to described enforcement
Example, those skilled in the art are it will be appreciated that various correction and change can be carried out without departing from the thought of the present invention and scope
Shape.Therefore, these fixed cases or variation individually can not understand according to the technological thought of the present invention or viewpoint, the reality deformed
Execute example should be subordinated in scope of the presently claimed invention.
Claims (10)
1. the collagen protein segregation apparatus utilizing ultrasound wave, it is characterised in that comprise:
Test portion case, it houses fish skin；
Separative element, its reception is supplied from the described fish skin of described test portion case and is separated into collagen protein, thus will be separated
Described collagen protein supplies again to described test portion case；And
Ultrasonic wave generator unit, it is connected to described separative element, in order to produce in order to described fish skin is separated into described collagen
The ultrasound wave of albumen.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 1, it is characterised in that further include cooling
Portion, it is to described separative element side supply cooling water, in order to can intercept the heat produced when separating described collagen protein；
Described separative element comprises:
Process portion, its reception is supplied from the fish skin of described test portion case and this fish skin is separated into collagen protein；And
1st cooling water inflow and outflow portion, its outer peripheral face from described process portion separates certain intervals and closes described process portion
Outside, thus provides and makes to be supplied from the space of the described cooling current into and out of described cooling end.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 2, it is characterised in that described test portion case and bag
Contain:
Reservoir, it stores described fish skin；And
2nd cooling water inflow and outflow portion, it separates certain intervals from the outer peripheral face of described reservoir and closes described reservoir
Outside, thus provides and makes to be supplied from the space of the described cooling current into and out of described cooling end.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 2, it is characterised in that described ultrasound wave produces
Raw unit comprises:
At least one ticker, it is combined in the outer peripheral face in described process portion and produces ultrasound wave；And
Vibration regulation controller, it regulates the output degree of described ticker.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 4, it is characterised in that: described ticker is
It is isolated from each other in the range of 5～6, in order to the multiple tickers that will not be affected by ultrasound wave each other.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 4, it is characterised in that: by described vibration
The frequency of the ultrasound wave that son produces is 20 kHz.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 4, it is characterised in that: described vibration regulation
Controller is output as 0.1～1000 W.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 3, it is characterised in that described cooling end bag
Contain:
Cooling body；
1st cooling water flows into pipe, and it prolongs from the downside in the lower the most described 1st cooling water inflow and outflow portion of described cooling body
Long, in order to cooling water cools down water inflow and outflow portion side inflow from described cooling body to the described 1st；
1st cooling water flow out of pipe, and it prolongs from the upside of the upper lateral described cooling body in described 1st cooling water inflow and outflow portion
Long, in order to cooling water flows out to described cooling body side from described 1st cooling water inflow and outflow portion；
2nd cooling water flows into pipe, and it prolongs from the downside in the lower the most described 2nd cooling water inflow and outflow portion of described cooling body
Long, in order to cooling water cools down water inflow and outflow portion side inflow from described cooling body to the described 2nd；And
2nd cooling water flow out of pipe, and it prolongs from the upside of the upper lateral described cooling body in described 2nd cooling water inflow and outflow portion
Long, in order to cooling water flows out to described cooling body side from described 2nd cooling water inflow and outflow portion.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 8, it is characterised in that further include:
Test portion supply pipe, the downside of the downside of described reservoir with described process portion is connected with each other by it, in order to can be from described storage
The portion that deposits supplies fish skin to side, described process portion；And
Test portion discharge pipe, the upside in described process portion is connected with each other by it with the upside of described reservoir, in order to can be to described examination
Bin side discharges the collagen protein separated by described separative element.
The collagen protein segregation apparatus utilizing ultrasound wave the most according to claim 9, it is characterised in that further include:
Circulating pump, it possesses on the prolongation path of described test portion supply pipe, in order to can forcibly make described fish skin and described
Collagen protein is along described test portion supply pipe and the circulation of described test portion discharge pipe；And
Circulating load regulation controller, it regulates the driver of described circulating pump, in order to fish skin described in scalable and described collagen
The circulating load of albumen.
Applications Claiming Priority (5)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR10-2010-0139655 | 2010-12-30 | ||
| KR1020100139655A KR101248617B1 (en) | 2010-12-30 | 2010-12-30 | A method of extracting collagen by ultrasonication |
| KR1020110017444A KR101246595B1 (en) | 2011-02-25 | 2011-02-25 | Extracting collagen device and method for extracting collagen using the same |
| KR10-2011-0017444 | 2011-02-25 | ||
| CN201180063095.6A CN104220586B (en) | 2010-12-30 | 2011-12-30 | Method for extracting collagen using ultrasonic waves, and apparatus therefor |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201180063095.6A Division CN104220586B (en) | 2010-12-30 | 2011-12-30 | Method for extracting collagen using ultrasonic waves, and apparatus therefor |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106046150A true CN106046150A (en) | 2016-10-26 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201180063095.6A Active CN104220586B (en) | 2010-12-30 | 2011-12-30 | Method for extracting collagen using ultrasonic waves, and apparatus therefor |
| CN201610421284.0A Pending CN106046150A (en) | 2010-12-30 | 2011-12-30 | Apparatus for isolating collagen by ultrasonic wave |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201180063095.6A Active CN104220586B (en) | 2010-12-30 | 2011-12-30 | Method for extracting collagen using ultrasonic waves, and apparatus therefor |
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| Country | Link |
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| JP (1) | JP5946843B2 (en) |
| CN (2) | CN104220586B (en) |
| WO (1) | WO2012091505A2 (en) |
Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2015012682A2 (en) * | 2013-07-22 | 2015-01-29 | Universiti Putra Malaysia | A method for extracting collagen from aquatic animals, collagen and products containing it |
| CN105061587A (en) * | 2015-08-26 | 2015-11-18 | 华南理工大学 | Ultrasonic wave aided extraction method suitable for high-viscosity collagen |
| CN106220727A (en) * | 2016-08-31 | 2016-12-14 | 江苏省农业科学院 | Response phase method is utilized to optimize the ultrasonic wave added acid extracting method of collagen protein |
| CN106632667A (en) * | 2017-01-12 | 2017-05-10 | 佛山科学技术学院 | Domestic integrated collagen preparation device |
| CN109430883A (en) * | 2018-10-31 | 2019-03-08 | 广东兴亿海洋生物工程股份有限公司 | Free peptide extract of marine fishes and its preparation method and application |
| CN110669813A (en) * | 2019-10-28 | 2020-01-10 | 山西原生肽科技有限公司 | Yak rib small molecule peptide and extraction method thereof |
| CN112790372B (en) * | 2021-02-03 | 2022-04-19 | 南昌大学 | Preparation method of constant-temperature high-stability fish protein glue |
| CN112760351A (en) * | 2021-03-17 | 2021-05-07 | 华中农业大学 | Extraction method and application of fish skin collagen |
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| JP5274964B2 (en) * | 2007-10-04 | 2013-08-28 | 共栄化学工業株式会社 | Cosmetics |
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2011
- 2011-12-30 CN CN201180063095.6A patent/CN104220586B/en active Active
- 2011-12-30 WO PCT/KR2011/010352 patent/WO2012091505A2/en active Application Filing
- 2011-12-30 CN CN201610421284.0A patent/CN106046150A/en active Pending
- 2011-12-30 JP JP2013547360A patent/JP5946843B2/en not_active Expired - Fee Related
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| CN101230088A (en) * | 2008-02-22 | 2008-07-30 | 四川大学 | Method for extracting native natural collagen from animal skin or/and tendon |
| CN101676297A (en) * | 2008-09-19 | 2010-03-24 | 大连正大海洋生物有限公司 | Preparation method of sea cucumber active polypeptide under ultrasonic extraction |
| CN101735304A (en) * | 2008-11-26 | 2010-06-16 | 大连隆合海洋生物工程技术有限公司 | Method for preparing water-based collagen of codfish skin by ultrasound wave extraction |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2012091505A3 (en) | 2012-11-08 |
| CN104220586A (en) | 2014-12-17 |
| JP2014505684A (en) | 2014-03-06 |
| WO2012091505A2 (en) | 2012-07-05 |
| CN104220586B (en) | 2017-05-17 |
| JP5946843B2 (en) | 2016-07-06 |
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