CN106119326A - The method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage - Google Patents
The method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage Download PDFInfo
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- CN106119326A CN106119326A CN201610480502.8A CN201610480502A CN106119326A CN 106119326 A CN106119326 A CN 106119326A CN 201610480502 A CN201610480502 A CN 201610480502A CN 106119326 A CN106119326 A CN 106119326A
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- 108010035532 Collagen Proteins 0.000 title claims abstract description 29
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 27
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 27
- 238000000034 method Methods 0.000 title claims abstract description 25
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- 238000000926 separation method Methods 0.000 claims description 6
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- 238000010612 desalination reaction Methods 0.000 claims description 5
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 claims description 2
- 102000002322 Egg Proteins Human genes 0.000 claims description 2
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- 229910052791 calcium Inorganic materials 0.000 description 3
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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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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
- C08B37/0063—Glycosaminoglycans or mucopolysaccharides, e.g. keratan sulfate; Derivatives thereof, e.g. fucoidan
- C08B37/0069—Chondroitin-4-sulfate, i.e. chondroitin sulfate A; Dermatan sulfate, i.e. chondroitin sulfate B or beta-heparin; Chondroitin-6-sulfate, i.e. chondroitin sulfate C; Derivatives thereof
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- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/26—Preparation of nitrogen-containing carbohydrates
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Abstract
The invention discloses a kind of method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage, the method be with slaughterhouse fresh cattle cartilage as raw material, coproduction prepares collagen protein polypeptide and chrondroitin, the method process conditions are gentle, energy consumption is low, the product purity obtained is high, steady quality is reliable, achieve the high-efficiency comprehensive utilization of cattle cartilage resource, achieve cattle cartilage intensive processing and from cattle cartilage coproduction extract collagen protein polypeptide and chrondroitin, improve cattle and butcher the added value of by-product, and without newly polluting generation in production process, decrease the pollution to environment in all directions, environmental protection.
Description
Technical field
The present invention relates to technical field of bioengineering.From cattle cartilage, collagen protein is extracted many more particularly, to one
The method of peptide coproduction chrondroitin.
Background technology
Collagen protein polypeptide has the digestibility of height, except a small amount of in oral cavity, stomach near by the effect of protease
Outside one step is decomposed, the oral human body that enters passes the oral cavity of people, stomach rapidly, is directly entered small intestinal, by intestinal absorption, eventually enters into
Blood circulation of human body system, organ and cell tissue, play rapidly its physiological action and biological function.In the world to collagen egg
White research conclusion, the number-average molecular weight of collagen protein is 2000~3000 when, most useful for absorption of human body.
The source of natural biological calcium such as ocean Concha Ostreaes etc., due to marine pollution matter, so having been reported that this kind of calcium preparation is deposited
At element pollutions such as lead cadmiums.Concha Ostreae often contains each Heavy Metallic Elements, long-term taking can make these heavy metal depositions in
Internal cause chronic poisoning.
Chrondroitin has been found to reach to supplement the effect of soft tissue of joint structure via being administered orally, and takes cartilage clinically
Element can effectively alleviate arthritic pain and inflammation phenomenon, has the effect delaying joint ageing simultaneously.Deep by high-tech
Processing, can treat nervous headache, trigeminal neuralgia, coronary heart diseases and angina pectoris, myocardial ischemia, cardiovascular and cerebrovascular disease, joint
Bitterly, the disease such as atherosclerosis and hepatitis.Can there is enhancing human body constitution as the additive in health product, food and resist
Pathogenic bacteria, beauty treatment, the anti-ageing effect of waiting for a long time.Now exporting to American-European multiple country, the market demand is vigorous.
Chrondroitin is prepared with salt solution, alkaline hydrolysis and enzymatic isolation method mostly by manufacturing enterprise now, and using more method is to adopt
The method combined with enzymolysis with alkali extracts chrondroitin, uses this alkaline process enzymolysis production process to there is complex steps, cycle
Longer, in high temperature steaming, alkaline extraction, repeatedly enzymolysis and decolorization, chrondroitin is destroyed, and product yield is on the low side, produces
Cost is high.
Accordingly, it is desirable to provide a kind of extract yield is high, and method is simple, the collagen protein polypeptide of mild condition, biological calcium,
The co-production of chrondroitin.
Summary of the invention
It is an object of the invention to provide a kind of method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage.Should
The product purity that method process conditions are gentle, energy consumption is low, obtain is high, steady quality is reliable, it is achieved that cattle cartilage resource efficient
Comprehensive utilization, it is achieved that the intensive processing of cattle cartilage and from cattle cartilage coproduction extract collagen protein polypeptide and chrondroitin, carry
High cattle butchers the added value of by-product, and without newly polluting generation in production process, decreases the pollution to environment in all directions,
Environmental protection.
For reaching above-mentioned purpose, the present invention uses following technical proposals:
The method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage, comprises the steps:
1) choose fresh cattle cartilage, clean, after 110 DEG C~120 DEG C of steaming and decoctings 50~60min, at 2.5~3.0MPa pressure
Lower separation, obtains cattle chondroprotein and cattle cartilage;
2) by step 1) the cattle cartilage that obtains cleans, is crushed to particle diameter 0.1~0.5mm, obtains cattle cartilage powder, and add it
The pure water of 1~1.5 times of quality, stirs, and inoculates plant lactobacillus, regulates pH to 5.0~6.0,35~50 DEG C of temperature
The lower ferment at constant temperature 18~48h of degree, obtains fermentation liquid;
3) by step 2) gained fermentation liquid centrifugation, separation condition is 16000~18000r/min, obtains clear liquid and precipitation
Thing;
4) by step 3) gained precipitate cleans, and adds pure water, stir, add papain, 48
~55 DEG C of enzymolysis 8~12h, obtain enzymolysis solution;Enzymolysis solution is heated to 80~100 DEG C again and keeps 40~60min, obtain the enzymolysis of inactivation
Liquid;
5) by step 4) gained inactivation enzymolysis solution through be concentrated by ultrafiltration, anion-exchange resin column carry out column chromatography, collect
Effluent;Again through deionized water eluting, obtain eluent;Effluent is mixed homogeneously with eluent, uses press atomization drying, obtain glue
Former protein polypeptide;
6) by step 5) in anion-exchange resin column salt solution eluting after deionized water eluting, by obtain
Eluent ultrafiltration desalination, adds dehydrated alcohol, obtains mixed liquor, stands 4~6h, obtains precipitation;In precipitation, add hydrogen peroxide enter
Row oxidative decoloration, then through drying under reduced pressure, obtain chrondroitin.
Preferably, step 2) in, the addition of described plant lactobacillus is 1.5~3wt% of the cattle cartilage after pulverizing.
Preferably, step 4) in, the addition of described papain is sedimentary 0.5~1wt%.
Preferably, step 6) in, in described mixed liquor, the volumetric concentration of dehydrated alcohol is 60~80%.
Preferably, step 6) in, described precipitation is 1:4~1:5 with the mass ratio of hydrogen peroxide.
Further, heretofore described plant lactobacillus includes Lactobacillus plantarum, lactobacillus casei, Lactobacillus brevis
Deng.Acidproof and the bile tolerance ability of the plant lactobacillus in the present invention is strong, and it can be survived under the adverse circumstances that nutrition is uneven,
Thus survival rate is higher, colonization ability is higher.
The papain used in the present invention, for the production of functional peptide, can make protein clear up Viability peptide and ammonia
Base acid, its decomposable asymmetric choice net is more more than pancreatin, widely protein substrate.
The plant lactobacillus, the papain that use in the present invention all can be obtained, such as plant lactobacillus by commercially available purchase
Including but not limited to buy from Jiang Yuan Chemical Co., Ltd. of Shenzhen, papain includes but not limited to that purchase is graceful from upper Hypon
Bio tech ltd.
Beneficial effects of the present invention is as follows:
1, the present invention is by using plant lactobacillus fermentation, biological enzymolysis and combining ultrafiltration, carry out column chromatography and pressure spray
The technology such as mist is dried, are continuously separated acquisition collagen protein polypeptide, chrondroitin, and the purity of the product obtained is high, this joint production process phase
Higher than per unit area yield process equipment utilization rate, save the energy and adjuvant, significantly reduce production cost.
2, the present invention uses plant lactobacillus to ferment cattle cartilage, compared with directly using enzymatic isolation method, and mild condition, energy consumption
Low, product purity is high, steady quality is reliable;Use anion-exchange resin column to carry out column chromatography, inorganic ions can be removed, increase
The degree of purity of product.
3, present invention achieves the co-production obtaining collagen protein polypeptide, chrondroitin from cattle cartilage continuously in a large number, skill
Art is reliable, constant product quality, it is achieved that the efficient utilization of cattle cartilage resource.
4, the fresh cattle cartilage of slaughter hall is immediately placed between processingization system by the present invention, has stopped Os Bovis seu Bubali in freezing, transport
Go mouldy, maintain the nutritional labeling of cattle cartilage to greatest extent;Shorten extraction time.Achieve the efficient of cattle cartilage resource
Comprehensive utilization and intensive processing, improve cattle and butcher the added value of by-product.In the whole course of processing, all in enclosed environment
Complete, each link is strictly separated, it is to avoid secondary pollution.
Detailed description of the invention
In order to be illustrated more clearly that the present invention, below in conjunction with preferred embodiment, the present invention is described further.Ability
Field technique personnel should be appreciated that following specifically described content is illustrative and be not restrictive, and should not limit this with this
The protection domain of invention.
Embodiment 1
The method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage, step is as follows:
1) steaming and decocting
Choosing fresh slaughterhouse fresh cattle cartilage is raw material, cleans, chooses fresh cattle cartilage, cleans, and steams at a temperature of 110 DEG C
After boiling 55min, separate with high pressure 2.5MPa, obtain cattle chondroprotein and cattle cartilage;
2) fermentation is pulverized
By step 1) the cattle cartilage that obtains is cleaned, is crushed to mean diameter is 0.2mm, obtains cattle cartilage powder;To cattle cartilage
Powder add the pure water of its equivalent and stirs, adding the plant lactobacillus accounting for cattle cartilage quality 1.5%, regulating pH
To 5.0, at a temperature of 40 DEG C, ferment at constant temperature 28h, obtains fermentation liquid;
3) centrifugation
By step 2) the centrifugal separator separation of gained fermentation liquid, rotating speed is 16000r/min, obtains clear liquid and precipitate;
4) enzymolysis
By step 3) gained precipitate cleans, and adds pure water, stir, add precipitate quality 0.5%
Papain, at 50 DEG C, enzymolysis 10h, obtains enzymolysis solution;Again enzymolysis solution is heated to 90 DEG C, enzyme-deactivating 50min, obtain inactivation
Enzymolysis solution;
5) collagen protein polypeptide is prepared
By step 4) enzymolysis solution of gained inactivation through being concentrated by ultrafiltration, anion-exchange resin column carries out column chromatography, collects stream
Go out liquid;Again through deionized water eluting, obtain eluent;Effluent is mixed homogeneously with eluent, with pressure spray dryer, obtains collagen
Protein polypeptide;
6) chrondroitin is prepared
By step 5) in anion-exchange resin column salt solution eluting after deionized water eluting, take eluent and carry out
Ultrafiltration desalination, adding dehydrated alcohol to the volumetric concentration of dehydrated alcohol in gained solution is 75%, the solution left standstill that will obtain
5h, obtains precipitation;In precipitation, add the hydrogen peroxide of its 3 times of quality, carry out oxidative decoloration, drying under reduced pressure, obtain chrondroitin.
The yield utilizing the collagen protein polypeptide that preparation method of the present invention produces is 95.3%, and chrondroitin purity reaches 99%.
Embodiment 2
The method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage, step is as follows:
1) steaming and decocting
Choosing fresh slaughterhouse fresh cattle cartilage is raw material, cleans, and sends in cook vat steaming and decocting 55min at a temperature of 120 DEG C
After, separate with high pressure 2.8MPa, obtain cattle chondroprotein and cattle cartilage;
2) fermentation is pulverized
By step 1) the cattle cartilage that obtains is cleaned, is crushed to mean diameter is 0.2mm, obtains cattle cartilage powder;To cattle cartilage
Powder add pure water and stirs, adding the plant lactobacillus accounting for cattle cartilage quality 2.5%, regulating pH to 6.0,
At a temperature of 45 DEG C, ferment at constant temperature 48h, obtains fermentation liquid;
3) centrifugation
By step 2) the centrifugal separator separation of gained fermentation liquid, rotating speed is 17000r/min, obtains clear liquid and precipitate;
4) enzymolysis
By step 3) gained precipitate cleans, and adds pure water, and cattle cartilage powder is stirring with the mass ratio of pure water
Uniformly, adding the papain of precipitate quality 0.8%, at 55 DEG C, enzymolysis 12h, obtains enzymolysis solution;Again enzymolysis solution is added
Hot to 100 DEG C, enzyme-deactivating 60min, obtain the enzymolysis solution of inactivation;
5) collagen protein polypeptide is prepared
By step 4) enzymolysis solution of gained inactivation through being concentrated by ultrafiltration, anion-exchange resin column carries out column chromatography, collects stream
Go out liquid;Again through deionized water eluting, obtain eluent;Effluent is mixed homogeneously with eluent, with pressure spray dryer, obtains collagen
Protein polypeptide;
6) chrondroitin is prepared
By step 5) in anion-exchange resin column salt solution eluting after deionized water eluting, take eluent and carry out
Ultrafiltration desalination, adding dehydrated alcohol to the volumetric concentration of dehydrated alcohol in gained solution is 80%, the solution left standstill that will obtain
6h, obtains precipitation;The hydrogen peroxide adding its 3 times of quality in precipitation carries out oxidative decoloration, drying under reduced pressure, obtains chrondroitin.
The yield utilizing the collagen protein polypeptide that preparation method of the present invention produces is 95.8%, and chrondroitin purity reaches
99.3%.
Embodiment 3
The method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage, step is as follows:
1) steaming and decocting
Choosing fresh slaughterhouse fresh cattle cartilage is raw material, cleans, and sends in cook vat steaming and decocting 60min at a temperature of 115 DEG C
After, separate with high pressure 3MPa, obtain cattle chondroprotein and cattle cartilage;
2) fermentation is pulverized
By step 1) the cattle cartilage that obtains is cleaned, is crushed to mean diameter is 0.5mm, obtains cattle cartilage powder;To cattle cartilage
Powder add pure water and stirs, adding the plant lactobacillus accounting for cattle cartilage quality 3%, regulating pH to 5.5,35
Ferment at constant temperature 38h at a temperature of DEG C, obtains fermentation liquid;
3) centrifugation
By step 2) the centrifugal separator separation of gained fermentation liquid, rotating speed is 18000r/min, obtains clear liquid and precipitate;
4) enzymolysis
By step 3) gained precipitate cleans, and adds pure water, and cattle cartilage powder is stirring with the mass ratio of pure water
Uniformly, adding the papain of precipitate quality 1%, at 48 DEG C, enzymolysis 8h, obtains enzymolysis solution;Again enzymolysis solution is heated to
80 DEG C, enzyme-deactivating 40min, obtain the enzymolysis solution of inactivation;
5) collagen protein polypeptide is prepared
By step 4) enzymolysis solution of gained inactivation through being concentrated by ultrafiltration, anion-exchange resin column carries out column chromatography, collects stream
Go out liquid;Again through deionized water eluting, obtain eluent;Effluent is mixed homogeneously with eluent, with pressure spray dryer, obtains collagen
Protein polypeptide;
6) chrondroitin is prepared
By step 5) in anion-exchange resin column salt solution eluting after deionized water eluting, take eluent and carry out
Ultrafiltration desalination, adding dehydrated alcohol to the volumetric concentration of dehydrated alcohol in gained solution is 80%, the solution left standstill that will obtain
4h, obtains precipitation;In precipitation, add the hydrogen peroxide of its 3 times of quality, drying under reduced pressure, obtain chrondroitin.
The yield utilizing the collagen protein polypeptide that preparation method of the present invention produces is 95.1%, and chrondroitin purity reaches
99.1%.
Obviously, the above embodiment of the present invention is only for clearly demonstrating example of the present invention, and is not right
The restriction of embodiments of the present invention, for those of ordinary skill in the field, the most also may be used
To make other changes in different forms, cannot all of embodiment be given exhaustive here, every belong to this
What bright technical scheme was extended out obviously changes or changes the row still in protection scope of the present invention.
Claims (5)
1. the method extracting collagen protein polypeptide coproduction chrondroitin from cattle cartilage, it is characterised in that comprise the steps:
1) choose fresh cattle cartilage, clean, after 110 DEG C~120 DEG C of steaming and decoctings 50~60min, divide under 2.5~3.0MPa pressure
From, obtain cattle chondroprotein and cattle cartilage;
2) by step 1) the cattle cartilage that obtains cleans, and is crushed to particle diameter 0.1~0.5mm, obtains cattle cartilage powder, and add its 1~
The pure water of 1.5 times of quality, stirs, and inoculates plant lactobacillus, regulates pH to 5.0~6.0, at a temperature of 35~50 DEG C
Ferment at constant temperature 18~48h, obtains fermentation liquid;
3) by step 2) gained fermentation liquid centrifugation, separation condition is 16000~18000r/min, obtains clear liquid and precipitate;
4) by step 3) gained precipitate cleans, and adds pure water, stir, add papain, 48~55
DEG C enzymolysis 8~12h, obtains enzymolysis solution;Enzymolysis solution is heated to 80~100 DEG C again and keeps 40~60min, obtain the enzymolysis solution of inactivation;
5) by step 4) enzymolysis solution of gained inactivation through being concentrated by ultrafiltration, anion-exchange resin column carries out column chromatography, collect and flow out
Liquid;Again through deionized water eluting, obtain eluent;Effluent is mixed homogeneously with eluent, uses press atomization drying, obtain collagen egg
White polypeptide;
6) by step 5) in anion-exchange resin column salt solution eluting after deionized water eluting, the eluting that will obtain
Liquid ultrafiltration desalination, adds dehydrated alcohol, obtains mixed liquor, stands 4~6h, obtains precipitation;In precipitation, add hydrogen peroxide carry out oxygen
Change decolouring, then through drying under reduced pressure, obtain chrondroitin.
Method the most according to claim 1, it is characterised in that step 2) in, the addition of described plant lactobacillus is powder
1.5~3wt% of cattle cartilage after broken.
Method the most according to claim 1, it is characterised in that step 4) in, the addition of described papain is heavy
The 0.5~1wt% of shallow lake thing.
Method the most according to claim 1, it is characterised in that step 6) in, the volume of dehydrated alcohol in described mixed liquor
Concentration is 60~80%.
Method the most according to claim 1, it is characterised in that step 6) in, described precipitation with the mass ratio of hydrogen peroxide is
1:4~1:5.
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CN107177658A (en) * | 2017-07-28 | 2017-09-19 | 美泰科技(青岛)股份有限公司 | A kind of preparation method of Cartilage collagen peptide |
CN108913745A (en) * | 2018-08-16 | 2018-11-30 | 余碧芝 | The method for extracting active collagen and elastin laminin using pigs and cattle cartilage |
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CN102533915A (en) * | 2011-12-31 | 2012-07-04 | 浙江工业大学 | Method for preparing chondroitin sulfate and collagen polypeptide from animal cartilages |
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CN107177658A (en) * | 2017-07-28 | 2017-09-19 | 美泰科技(青岛)股份有限公司 | A kind of preparation method of Cartilage collagen peptide |
CN108913745A (en) * | 2018-08-16 | 2018-11-30 | 余碧芝 | The method for extracting active collagen and elastin laminin using pigs and cattle cartilage |
CN110713557A (en) * | 2019-12-10 | 2020-01-21 | 无棣县兴亚生物科技有限公司 | Process for extracting chondroitin by using meat and bone meal waste pulp through biological method |
CN110713557B (en) * | 2019-12-10 | 2021-10-26 | 无棣县兴亚生物科技有限公司 | Process for extracting chondroitin by using meat and bone meal waste pulp through biological method |
CN111296755A (en) * | 2019-12-17 | 2020-06-19 | 福州宏东食品有限公司 | A nutrition-enriched oatmeal food containing marine chondroitin sulfate and its preparation method |
CN115340595A (en) * | 2022-05-09 | 2022-11-15 | 湖南伍星生物科技有限公司 | Production method for preparing bovine cartilage collagen peptide by novel degreasing process |
CN115736135A (en) * | 2022-11-18 | 2023-03-07 | 广东雅琪生物科技股份有限公司 | Micropterus salmoides feed containing compound enzymolysis protein powder and preparation method thereof |
CN117551187A (en) * | 2024-01-12 | 2024-02-13 | 烟台融科生物科技有限公司 | Method for producing bone collagen peptide by using sheep bone |
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