CN106337074A - Cirrhinus molitorella bone collagen extracting method - Google Patents
Cirrhinus molitorella bone collagen extracting method Download PDFInfo
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- CN106337074A CN106337074A CN201610946819.6A CN201610946819A CN106337074A CN 106337074 A CN106337074 A CN 106337074A CN 201610946819 A CN201610946819 A CN 201610946819A CN 106337074 A CN106337074 A CN 106337074A
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- 102000008186 Collagen Human genes 0.000 title claims abstract description 51
- 108010035532 Collagen Proteins 0.000 title claims abstract description 51
- 210000000988 bone and bone Anatomy 0.000 title claims abstract description 46
- 229920001436 collagen Polymers 0.000 title claims abstract description 38
- 238000000034 method Methods 0.000 title claims abstract description 35
- 241000703788 Cirrhinus molitorella Species 0.000 title claims abstract description 28
- 239000000843 powder Substances 0.000 claims description 50
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 39
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 27
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 20
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 20
- 238000002791 soaking Methods 0.000 claims description 19
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 14
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 claims description 12
- 238000005119 centrifugation Methods 0.000 claims description 12
- 239000000463 material Substances 0.000 claims description 12
- 241000594011 Leuciscus leuciscus Species 0.000 claims description 10
- 230000029087 digestion Effects 0.000 claims description 8
- 238000000605 extraction Methods 0.000 claims description 8
- 239000003960 organic solvent Substances 0.000 claims description 7
- 238000010298 pulverizing process Methods 0.000 claims description 7
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 6
- 239000004365 Protease Substances 0.000 claims description 5
- 108091005804 Peptidases Proteins 0.000 claims description 4
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 claims description 4
- 230000003544 deproteinization Effects 0.000 claims description 4
- 238000007654 immersion Methods 0.000 claims description 4
- 238000001556 precipitation Methods 0.000 claims 1
- 241000251468 Actinopterygii Species 0.000 abstract description 27
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 abstract description 9
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 abstract description 4
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 abstract description 4
- 229910001424 calcium ion Inorganic materials 0.000 abstract description 4
- 238000002360 preparation method Methods 0.000 abstract description 4
- 235000013305 food Nutrition 0.000 abstract description 3
- 230000000717 retained effect Effects 0.000 abstract 1
- 239000007788 liquid Substances 0.000 description 26
- 239000006228 supernatant Substances 0.000 description 12
- 239000012535 impurity Substances 0.000 description 10
- 102000004169 proteins and genes Human genes 0.000 description 10
- 108090000623 proteins and genes Proteins 0.000 description 10
- 239000000284 extract Substances 0.000 description 9
- 230000008569 process Effects 0.000 description 9
- 102000004190 Enzymes Human genes 0.000 description 7
- 108090000790 Enzymes Proteins 0.000 description 7
- 238000001035 drying Methods 0.000 description 7
- 238000012545 processing Methods 0.000 description 7
- 238000012360 testing method Methods 0.000 description 7
- 239000002639 bone cement Substances 0.000 description 6
- 238000004140 cleaning Methods 0.000 description 6
- 229940088598 enzyme Drugs 0.000 description 6
- 238000001914 filtration Methods 0.000 description 6
- 238000011282 treatment Methods 0.000 description 6
- 241000252335 Acipenser Species 0.000 description 5
- 210000004712 air sac Anatomy 0.000 description 5
- 238000009835 boiling Methods 0.000 description 5
- 238000005485 electric heating Methods 0.000 description 5
- 230000002255 enzymatic effect Effects 0.000 description 5
- 239000008236 heating water Substances 0.000 description 5
- 238000000751 protein extraction Methods 0.000 description 4
- 239000000047 product Substances 0.000 description 3
- 235000019419 proteases Nutrition 0.000 description 3
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 239000011575 calcium Substances 0.000 description 2
- 229910052791 calcium Inorganic materials 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000004108 freeze drying Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- NTIZESTWPVYFNL-UHFFFAOYSA-N Methyl isobutyl ketone Chemical compound CC(C)CC(C)=O NTIZESTWPVYFNL-UHFFFAOYSA-N 0.000 description 1
- 108090000526 Papain Proteins 0.000 description 1
- 101710172711 Structural protein Proteins 0.000 description 1
- 238000009360 aquaculture Methods 0.000 description 1
- 244000144974 aquaculture Species 0.000 description 1
- 230000003796 beauty Effects 0.000 description 1
- 239000003519 biomedical and dental material Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 210000002744 extracellular matrix Anatomy 0.000 description 1
- 239000013505 freshwater Substances 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 235000012054 meals Nutrition 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 229940055729 papain Drugs 0.000 description 1
- 235000019834 papain Nutrition 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 239000013049 sediment Substances 0.000 description 1
- 238000005406 washing 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
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
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- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Biochemistry (AREA)
- Wood Science & Technology (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Toxicology (AREA)
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Abstract
The invention relates to the technical field of food industry, in particular to a cirrhinus molitorella bone collagen extracting method. In the preparation process of the extracting method, nosubstance such as acetic acid or EDTA causing calcium ion loss is not adopted, thus, the collagen content is higher in the cirrhinus molitorella bone collagen obtained by adopting the method, and the calcium ions in fish bones are also retained. It is detected that the collagen content in the cirrhinus molitorella bone collagen obtained by adopting the method is 13.9%-21.1%.
Description
Technical field
The present invention relates to food industrial technical field, more particularly, to a kind of extracting method of Cirrhina molitorella bone collagen.
Background technology
Collagen protein is a kind of structural protein of extracellular matrix, is a kind of white, opaque, unbranched fibrous type
Protein, because its good biology performance has obtained extensive concern at aspects such as biomedical material, food, beauty treatments.I
Guo Shi Aquatic product big country, the expansion with aquaculture scale and the development of processing industry, many aquatic products include useless after it is processed
Gurry contains abundant collagen protein, can become the good source of collagen protein.
Cirrhina molitorella is one of important economic fish in South China, the fishbone that the course of processing produces, and the leftover bits and pieces such as fish tail account for
The 40%～55% of raw material fish quality.If not being effectively treated, not only result in environmental pollution, and a large amount of preciousnesses can be wasted
Resource.And the protein containing in these leftover bits and pieces is close with the flesh of fish, mineral particularly calcium content is higher than the flesh of fish, in addition, under
Heel contains nutrient needed by human.Generally, these leftover bits and pieces only carry out roughing, are not fully utilized, thus
Can cause to waste.
Extract collagen protein from Cirrhina molitorella processing dead meal, comprehensive utilization, the fall of fresh-water fishes processing waste can be promoted
The processing cost of low Cirrhina molitorella, provides the added value of its processing, can reduce the pollution of environment simultaneously, obtain good economic benefit and
Social benefit.But it is in the existing method preparing collagen protein from the leftover bits and pieces such as fishbone, when extracting bone collagen, past
Toward first sloughing calcareous hydrolytic collagen again, to prepare purer collagen protein, but calcareous a large amount of losses so can be caused.
Content of the invention
In view of this, the technical problem to be solved in the present invention is to provide a kind of extracting method of Cirrhina molitorella bone collagen,
Extract collagen content in the collagen protein of acquisition in this way higher, and extraction process do not slough calcareous.
The invention provides a kind of extracting method of Cirrhina molitorella bone collagen, comprising:
Step 1: after protease digestion dace fish bone material, cleaned, dry, pulverizing, prepared fishbone coarse powder；
Step 2: described fishbone coarse powder, after defat, deproteinization, after being extracted with aqueous citric acid solution, is sunk through centrifugation reject
Form sediment, after lyophilization, obtain Cirrhina molitorella bone collagen；
Described defat adopts organic solvent to soak；
Described deproteinization adopts nacl solution soaking and/or naoh solution soaking.
Described dace fish bone material is Cirrhina molitorella fishbone, is the leftover bits and pieces that the course of processing produces.Described digestion is particularly as follows: by Cirrhina molitorella
Collagen material adds protein enzyme solution after mixing with water.The addition of water is preferred with submergence dace fish bone material.
In the embodiment of the present invention, described protease is papain, the mass ratio of described protease and dace fish bone material
For 600u/g.
In the embodiment of the present invention, the temperature of described digestion is 60 DEG C, and the time is 20min～40min.Some specific embodiments
In, the time of digestion is 30min.
After digestion, also include being heated to the step of enzyme denaturing of seething with excitement, the time of ebuillition of heated is 10min
The flesh of fish of residual after digestion, is removed through over cleaning.Then it is dried, be dried by the way of drying, specially
60 DEG C, 12h is dried.
Crushed after being dried adopts high speed disintegrator to pulverize the fishbone being dried, and obtains fishbone coarse powder；Grinding time is 8min
～10min.
In the embodiment of the present invention, described organic solvent is selected from n-butyl alcohol, isopropanol or acetone.
In the embodiment of the present invention, defat is to soak after 3h～12h described fishbone coarse powder with organic solution, and reject is organic molten
Agent, with clear water rinsing.
In some specific embodiments, with the time of organic solvent immersion for 6 hours～12 hours.It is specially 6h or 12h.
Specifically, 6h is soaked or with acetone soak 12h with n-butyl alcohol or isopropyl acetone.
In the embodiment of the present invention, fishbone coarse powder is 1:(10～40 with the quality-volume ratio of organic solution).
In some embodiments, fishbone coarse powder is 1:30 with the quality-volume ratio of organic solution.
After soaking through organic solvent, in fishbone coarse powder, lipid is removed.
The removal of foreign protein is included using nacl solution soaking and/or naoh solution soaking.
The present invention does not limit to nacl solution soaking, the order of naoh solution soaking, can successively be respectively adopted two kinds molten
Immersion is steeped, also can be only with one of which solution soaking, and the present invention is not construed as limiting to this.But effect shows, only entered with nacl solution
After row soaks, the yield of collagen protein is not as good as the effect using naoh solution soaking.
The mass fraction of described nacl solution is 5%～10%；Fishbone coarse powder in described nacl solution soaking, through defat
Quality-volume ratio with described nacl solution is 1:(5～20)；Soak time is 3h～12h.
In some specific embodiments, the mass fraction of nacl solution is 7%；In described nacl solution soaking, through defat
Fishbone coarse powder is 1:20 with the quality-volume ratio of described nacl solution；Soak time is 9h.
The concentration of described naoh solution is 0.05mol/l；In described naoh solution soaking, the fishbone coarse powder through defat and institute
Quality-the volume ratio stating naoh solution is 1:(2～6)；Soak time is 3h～12h.
In some specific embodiments, the concentration of naoh solution is 0.05mol/l～0.2mol/l；Described naoh solution soaking
In, the fishbone coarse powder through defat is 1:5 with the quality-volume ratio of described naoh solution；Soak time is 3h.
After nacl solution soaking and/or naoh solution soaking, foreign protein is removed.Then carried out with citric acid solution
The extraction of collagen protein.
The concentration of described aqueous citric acid solution is 0.1mol/l；The temperature of described extraction is 0 DEG C～4 DEG C, described extraction
Mode is extraction, described, the quality-volume ratio through Deproteinated fishbone coarse powder and described aqueous citric acid solution be 1:(10～
40).
In some specific embodiments, described quality-volume through Deproteinated fishbone coarse powder and described aqueous citric acid solution
Than for 1:30.
The temperature of described centrifugation is 0～4 DEG C, and rotating speed is 8000rpm, and the time is 10min.
Supernatant after centrifugation is fish bone collagen protein extract, supernatant sucking filtration, and frozen drying obtains Cirrhina molitorella bone
Collagen protein.
The Cirrhina molitorella bone collagen that the extracting method that the present invention provides is obtained.
The present invention provide preparation method, preparation during, not using meeting lead to calcium ion loss acetic acid or
The materials such as edta, therefore, in the albumen of the Cirrhina molitorella ossein being obtained with the method provided by the present invention, not only the content of collagen protein is relatively
Height, also retains the calcium ion in fishbone.After testing, in the Cirrhina molitorella bone collagen that the method that the present invention provides is obtained, collagen
The content of albumen is 13.9%～21.1%.
Specific embodiment
The invention provides a kind of extracting method of Cirrhina molitorella bone collagen, those skilled in the art can use for reference interior herein
Hold, be suitably modified technological parameter and realize.Specifically, all similar replacements and change are to those skilled in the art
For be it will be apparent that they are considered as including in the present invention.Preferable enforcement has been passed through in the method for the present invention and application
Example is described, and related personnel substantially can be to methods herein and application in without departing from present invention, spirit and scope
It is modified or suitably changes and combine, to realize and to apply the technology of the present invention.
The instrument that the present invention adopts is all common commercially available product, all can buy in market.
With reference to embodiment, the present invention be expanded on further:
Embodiment 1
(1) discard fishbone pretreatment:
The impurity such as manual removing Air Bladder pseudosciaenae seu Acipenser, dace fish bone material are put in container, add water to submerge fishbone, add albumen to container
Enzymatic solution.Container is put to electric-heating water bath, temperature is set as 60 DEG C, water bath time is 30min.After water-bath terminates, boiling
Water heats 10min, enzyme denaturing.Cleaning fishbone, removes the residual flesh of fish.Cleaned fishbone is dried；
(2) pulverization process:
With high speed disintegrator, the fishbone being dried is pulverized, obtain fishbone coarse powder；
(3) fishbone coarse powder ungrease treatment:
Soak fishbone coarse powder using n-butyl alcohol 6 hours, solid-liquid ratio 1:30, then clear water rinsing.
(4) foreign protein removes:
Under room temperature, first use the naoh solution of 0.05mol/l to press solid-liquid ratio 1:5 and process and soak fishbone coarse powder 3h, then clear water
Rinsing, drains；Then soak fishbone coarse powder 9h with 7% nacl solution according to solid-liquid ratio 1:20；
(5) bone collagen extracts:
Using the citric acid of 0.1mol/l, according to 1:30 (m:v) solid-liquid ratio, soak through defat removing impurities in the environment of 4 DEG C
The fishbone powder sample of albumen.High speed frozen centrifugation (8000rmp/min, 4 DEG C, 10min), supernatant is fish bone collagen protein extraction
Liquid, supernatant sucking filtration, frozen drying, obtain Cirrhina molitorella bone collagen.After testing, wherein fish bone glue protein content:
19.2%.
Embodiment 2
(1) discard fishbone pretreatment:
The impurity such as manual removing Air Bladder pseudosciaenae seu Acipenser, dace fish bone material are put in container, add water to submerge fishbone, add albumen to container
Enzymatic solution.Container is put to electric-heating water bath, temperature is set as 60 DEG C, water bath time is 30min.After water-bath terminates, boiling
Water heats 10min, enzyme denaturing.Cleaning fishbone, removes the residual flesh of fish.Cleaned fishbone is dried；
(2) pulverization process:
With high speed disintegrator, the fishbone being dried is pulverized, obtain fishbone coarse powder；
(3) fishbone coarse powder ungrease treatment:
Soak fishbone coarse powder using isopropanol 6 hours, solid-liquid ratio 1:30, then clear water rinsing.
(4) foreign protein removes:
Under room temperature, first use the naoh solution of 0.05mol/l to press solid-liquid ratio 1:5 and process and soak fishbone coarse powder 3h, then clear water
Rinsing, drains；Then soak fishbone coarse powder 9h with 7% nacl solution according to solid-liquid ratio 1:20；
(5) bone collagen extracts:
Using the citric acid of 0.1mol/l, according to 1:30 (m:v) solid-liquid ratio, soak through defat removing impurities in the environment of 4 DEG C
The fishbone powder sample of albumen.High speed frozen centrifugation (8000rmp/min, 4 DEG C, 10min), supernatant is fish bone collagen protein extraction
Liquid, supernatant sucking filtration, frozen drying, obtain Cirrhina molitorella bone collagen.After testing, wherein fish bone glue protein content:
18.7%.
Embodiment 3
(1) discard fishbone pretreatment:
The impurity such as manual removing Air Bladder pseudosciaenae seu Acipenser, dace fish bone material are put in container, add water to submerge fishbone, add albumen to container
Enzymatic solution.Container is put to electric-heating water bath, temperature is set as 60 DEG C, water bath time is 30min.After water-bath terminates, boiling
Water heats 10min, enzyme denaturing.Cleaning fishbone, removes the residual flesh of fish.Cleaned fishbone is dried；
(2) pulverization process:
With high speed disintegrator, the fishbone being dried is pulverized, obtain fishbone coarse powder；
(3) fishbone coarse powder ungrease treatment:
Using acetone soak fishbone coarse powder 12 hours, solid-liquid ratio 1:30, then clear water rinsing.
(4) foreign protein removes:
Under room temperature,
Pressed solid-liquid ratio 1:5 and processed with the naoh solution of 0.05mol/l and soak fishbone coarse powder 3h；
(5) bone collagen extracts:
Using the citric acid of 0.1mol/l, according to 1:30 (m:v) solid-liquid ratio, soak through defat removing impurities in the environment of 4 DEG C
The fishbone powder sample of albumen.High speed frozen centrifugation (8000rmp/min, 4 DEG C, 10min), supernatant is fish bone collagen protein extraction
Liquid, supernatant sucking filtration, frozen drying, obtain Cirrhina molitorella bone collagen.After testing, wherein fish bone glue protein content:
19.8%.
Embodiment 4
(1) discard fishbone pretreatment:
The impurity such as manual removing Air Bladder pseudosciaenae seu Acipenser, dace fish bone material are put in container, add water to submerge fishbone, add albumen to container
Enzymatic solution.Container is put to electric-heating water bath, temperature is set as 60 DEG C, water bath time is 30min.After water-bath terminates, boiling
Water heats 10min, enzyme denaturing.Cleaning fishbone, removes the residual flesh of fish.Cleaned fishbone is dried；
(2) pulverization process:
With high speed disintegrator, the fishbone being dried is pulverized, obtain fishbone coarse powder；
(3) fishbone coarse powder ungrease treatment:
Using acetone soak fishbone coarse powder 12 hours, solid-liquid ratio 1:30, then clear water rinsing.
(4) foreign protein removes:
Under room temperature, the nacl solution with 2.5% soaks fishbone coarse powder 3h according to solid-liquid ratio 1:20；
(5) bone collagen extracts:
Using the citric acid of 0.1mol/l, according to 1:30 (m:v) solid-liquid ratio, soak through defat removing impurities in the environment of 4 DEG C
The fishbone powder sample of albumen.High speed frozen centrifugation (8000rmp/min, 4 DEG C, 10min), supernatant is fish bone collagen protein extraction
Liquid, supernatant sucking filtration, frozen drying, obtain Cirrhina molitorella bone collagen.After testing, wherein fish bone glue protein content:
13.9%.
Embodiment 5
(1) discard fishbone pretreatment:
The impurity such as manual removing Air Bladder pseudosciaenae seu Acipenser, dace fish bone material are put in container, add water to submerge fishbone, add albumen to container
Enzymatic solution.Container is put to electric-heating water bath, temperature is set as 60 DEG C, water bath time is 30min.After water-bath terminates, boiling
Water heats 10min, enzyme denaturing.Cleaning fishbone, removes the residual flesh of fish.Cleaned fishbone is dried；
(2) pulverization process:
With high speed disintegrator, the fishbone being dried is pulverized, obtain fishbone coarse powder；
(3) fishbone coarse powder ungrease treatment:
Using acetone soak fishbone coarse powder 12 hours), solid-liquid ratio 1:30, then clear water rinsing.
(4) foreign protein removes:
Under room temperature, first use the naoh solution of 0.05mol/l to press solid-liquid ratio 1:5 and process and soak fishbone coarse powder 3h, then clear water
Rinsing, drains；Then soak fishbone coarse powder 9h with 7% nacl solution according to solid-liquid ratio 1:20；
(5) bone collagen extracts:
Using the citric acid of 0.1mol/l, according to 1:30 solid-liquid ratio, soak in the environment of 4 DEG C through defat removing impurities albumen
Fishbone powder sample.High speed frozen centrifugation (8000rmp/min, 4 DEG C, 10min), supernatant is fish bone collagen protein extract, on
Clear liquid sucking filtration, frozen drying, obtain Cirrhina molitorella bone collagen.After testing, wherein fish bone glue protein content: 21.1%.
Comparative example
1) pretreatment of Cirrhina molitorella fishbone: in fishbone plus 0.1mol/l naoh solution is in 4 DEG C according to solid-liquid ratio 1g:15ml
Lower immersion 5h, removes noncollagen protein；After process fishbone distilled water cyclic washing to ph 7.0, drain, according to solid-liquid ratio 1g:
5ml adds 0.5mol/l edta solution (ph 7.4), soaks 3 days at 4 DEG C, changes 1 edta solution daily, removes fishbone
In calcium, be dried, pulverize, obtain decalcification fishbone powder.
2) preparation of field fish bone collagen albumen: add 0.5mol/l acetic acid to decalcification fishbone powder according to solid-liquid ratio 1g:3ml
Solution simultaneously soaks 3d at 4 DEG C, takes supernatant, add nacl to the final concentration of 0.9mol/ of solution after 12 000g centrifugation 30min
L, stands 60min, and 15 000g centrifugation 30min obtain precipitate, and lyophilization is fish bone collagen egg.After testing, wherein fish bone glue
Protein content: 13.9%.
The above is only the preferred embodiment of the present invention it is noted that coming for those skilled in the art
Say, under the premise without departing from the principles of the invention, some improvements and modifications can also be made, these improvements and modifications also should be regarded as
Protection scope of the present invention.
Claims (10)
1. a kind of extracting method of Cirrhina molitorella bone collagen is it is characterised in that include:
Step 1: after protease digestion dace fish bone material, cleaned, dry, pulverizing, prepared fishbone coarse powder；
Step 2: described fishbone coarse powder is after defat, deproteinization, after being extracted with aqueous citric acid solution, through centrifugation reject precipitation, cold
Cirrhina molitorella bone collagen is obtained after lyophilizing is dry；
Described defat adopts organic solvent to soak；
Described deproteinization adopts nacl solution soaking and/or naoh solution soaking.
2. it is characterised in that the temperature of described digestion is 60 DEG C, the time is 20min to extracting method according to claim 1
～40min.
3. extracting method according to claim 1 it is characterised in that described organic solvent be selected from n-butyl alcohol, isopropanol or
Acetone.
4. extracting method according to claim 1 is it is characterised in that described defat is with organic molten by described fishbone coarse powder
Immersion bubble 3h～12h after, reject organic solvent, with clear water rinsing.
5. extracting method according to claim 4 is it is characterised in that the quality-body of described fishbone coarse powder and organic solution
Long-pending ratio is 1:(10～40).
6. extracting method according to claim 1 it is characterised in that described nacl solution mass fraction be 5%～
10%；In described nacl solution soaking, the quality-volume ratio of fishbone coarse powder through defat and described nacl solution be 1:(5～
20)；Soak time is 3h～12h.
7. extracting method according to claim 1 it is characterised in that described naoh solution concentration be 0.05mol/l～
0.2mol/l；In described naoh solution soaking, the fishbone coarse powder through defat is 1:(2 with the quality-volume ratio of described naoh solution
～6)；Soak time is 3h～12h.
8. extracting method according to claim 1 is it is characterised in that the concentration of described aqueous citric acid solution is 0.1mol/
l；The temperature of described extraction is 0 DEG C～4 DEG C, and the mode of described extraction is extraction, described, through Deproteinated fishbone coarse powder with described
Quality-the volume ratio of aqueous citric acid solution is 1:(10～40).
9. it is characterised in that the temperature of described centrifugation is 0～4 DEG C, rotating speed is extracting method according to claim 8
8000rpm, the time is 10min.
10. the Cirrhina molitorella bone collagen that extracting method described in any one of claim 1～9 is obtained.
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