CN105925649A - Preparation method of skim squid protein functional active peptide with small molecular weight - Google Patents
Preparation method of skim squid protein functional active peptide with small molecular weight Download PDFInfo
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- CN105925649A CN105925649A CN201610380480.8A CN201610380480A CN105925649A CN 105925649 A CN105925649 A CN 105925649A CN 201610380480 A CN201610380480 A CN 201610380480A CN 105925649 A CN105925649 A CN 105925649A
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- 241000238366 Cephalopoda Species 0.000 title claims abstract description 204
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 72
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 65
- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 65
- 238000002360 preparation method Methods 0.000 title claims abstract description 36
- 238000011282 treatment Methods 0.000 claims abstract description 34
- 238000000605 extraction Methods 0.000 claims abstract description 17
- 210000001835 viscera Anatomy 0.000 claims abstract description 3
- 239000003795 chemical substances by application Substances 0.000 claims description 50
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- 238000000034 method Methods 0.000 claims description 43
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- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 40
- 239000000284 extract Substances 0.000 claims description 29
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- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 claims description 15
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- MVORZMQFXBLMHM-QWRGUYRKSA-N Gly-His-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CC1=CN=CN1 MVORZMQFXBLMHM-QWRGUYRKSA-N 0.000 description 2
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 2
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical compound [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
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- 201000001320 Atherosclerosis Diseases 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 108010028690 Fish Proteins Proteins 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- 244000068988 Glycine max Species 0.000 description 1
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
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- 239000003814 drug Substances 0.000 description 1
- 239000008157 edible vegetable oil Substances 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
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- 210000000232 gallbladder Anatomy 0.000 description 1
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- 230000002607 hemopoietic effect Effects 0.000 description 1
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- 239000004531 microgranule Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
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- 238000012986 modification Methods 0.000 description 1
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- 230000000474 nursing effect Effects 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000008935 nutritious Nutrition 0.000 description 1
- 238000011017 operating method Methods 0.000 description 1
- LQRJAEQXMSMEDP-XCHBZYMASA-N peptide a Chemical compound N([C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](C)C(=O)NCCCC[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)C(\NC(=O)[C@@H](CCCCN)NC(=O)CNC(C)=O)=C/C=1C=CC=CC=1)C(N)=O)C(=O)C(\NC(=O)[C@@H](CCCCN)NC(=O)CNC(C)=O)=C\C1=CC=CC=C1 LQRJAEQXMSMEDP-XCHBZYMASA-N 0.000 description 1
- 238000011056 performance test Methods 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 230000013777 protein digestion Effects 0.000 description 1
- 230000004853 protein function Effects 0.000 description 1
- 239000003531 protein hydrolysate Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 229910052711 selenium Inorganic materials 0.000 description 1
- 239000011669 selenium Substances 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 235000017557 sodium bicarbonate Nutrition 0.000 description 1
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 229930193551 sterin Natural products 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 235000012976 tarts Nutrition 0.000 description 1
- 229960003080 taurine Drugs 0.000 description 1
- 239000002699 waste material Substances 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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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Wood Science & Technology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Toxicology (AREA)
- Gastroenterology & Hepatology (AREA)
- Biophysics (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Meat, Egg Or Seafood Products (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention relates to the field of nutrient supplements, in particular to a preparation method of a skim squid protein functional active peptide with small molecular weight. The preparation method comprises the following steps: firstly, performing viscera removal treatment on a squid to obtain a squid body and then performing cutting treatment and breaking treatment to obtain minced squid pieces; secondly, performing ultrasonic decoloring treatment, ultrasonic color fixing treatment and extraction treatment on the squid pieces to obtain a squid protein extracting solution; thirdly, performing enzymolysis treatment, pressurizing treatment and ultrafiltration treatment in sequence to obtain the active peptide. According to the preparation method provided by the invention, through pressurizing an enzymatic hydrolysate, the obtained active squid small peptide is uniform in broken chain, and the molecular weight can be controlled within 3KD; through the decoloring treatment and the color fixing treatment, the treatment time of the squid body is greatly shortened, and a fat layer in the squid body is removed, so that the quality of the small peptide is radically ensured; all steps and parameters in the preparation method provided by the invention have synergetic effects, so that the quality of an active peptide product can be further ensured.
Description
Technical field
The present invention relates to belong to nutrient prime replenisher field, be specifically related to a kind of low-molecular-weight defat squid albumen
The preparation method of functional activity peptide.
Background technology
Along with the development of China's ocean squid fishery, squid becomes the aquatic products processing raw material that China is main.It is reported,
The year processing capacity of China squid is about 30～400,000 tons of squids.Squid is rich in calcium, phosphorus, ferrum element, beneficially bone
Bone is grown and hemopoietic, can effectively treat anemia；Contain substantial amounts of taurine simultaneously, the gallbladder in blood can be suppressed
Sterin content, fatigue alleviating, recover vision, improve liver function, polypeptide and selenium contained by it have antiviral,
Anti-actinism, has simultaneously and has tonifying-Yin and nourishing-stomach, the function of qi-restoratives skin moistening.
Chinese patent application CN103947818A discloses the preparation method of a kind of squid bioactive peptide: by former
Material process, ultrasonic Treatment, HIGH PRESSURE TREATMENT obtain finished product.Compared to existing technology, the time shortens dramatically, tool
Have technical maturity, reliable, this method can cost-effective, reduce energy consumption, improve the features such as extraction efficiency.
Chinese patent application CN104982980A discloses a kind of processing side nursing one's health squid Protein reconstitution goods
Method: squid carries out pretreatment, deacidification processes, then chopping and mixing starch, edible oil, soybean separation protein
Wait adjuvant in vain, thus obtain squid Protein reconstitution goods.These squid Protein reconstitution goods not only do not have Peru's squid
The tart flavour of fish itself, and while remaining squid local flavor very well, also have concurrently nutritious, be prone to disappear
The advantage changed, is especially suitable for the crowd that child, old people, the digestion powers such as crowd of just recovering from a dangerous illness are more weak.
At present, from squid, obtain the method for bioactive peptide a lot of etc., but there is shortcomings, although high pressure side
Just, but the protein chain of squid not easy fracture, the molecular weight of albumen is excessive, causes the waste of resource.Therefore,
Current scientific research and practice need a kind of new method, prepare molecular weight, activated, be prone to
The method of the squid small peptide absorbed.
Summary of the invention
The present invention provides the preparation method of a kind of low-molecular-weight defat squid protein functional bioactive peptide, the method
By pretreatment, the decolouring step such as fixation, enzymolysis pressurization, prepare molecular weight and be below 3KD, be prone to
The squid protein functional bioactive peptide of absorption of human body.
The present invention is achieved by the following technical solutions:
The preparation method of a kind of low-molecular-weight defat squid protein functional bioactive peptide, comprises the following steps:
Pre-treatment step: squid is removed internal organs and processes, obtain squid plate；Again described squid plate is entered
Row scribing processes, break process, obtains the squid fragment rubbed；
Decolouring fixation step: described squid fragment is carried out ultrasonic desolventing technology, ultrasonic fixation treatment, extraction
Process, obtain squid protein extract；
Enzymolysis pressurization steps: described squid protein extract is carried out successively enzymolysis processing, pressurized treatments, surpasses
Filter processes, and obtains described bioactive peptide.
Above-mentioned preparation method, as a kind of preferred implementation, in described pre-treatment step, described squid
For Argentina squid or Peru squid；Preferably, described squid is live squid or freezing new less than 15 days
Fresh squid.
Above-mentioned preparation method, as a kind of preferred implementation, in described pre-treatment step, described squid
The thickness of plate is between 3cm～8cm；It is highly preferred that similar plastics in tripe got rid of by described squid plate
The vertebrae of sheet, white mucosa；Further, during described scribing processes, cutter cuts described squid plate about
The thickness of 1/2, is separated by 2cm～5cm between every cutter.
Above-mentioned preparation method, as a kind of preferred implementation, in described decolouring fixation step, decolorising agent is
The NaOH solution of 0.01～0.045mol/L；Color fixing agent be mass percent concentration be 0.1%～0.4%
Ca(OH)2Solution.
Above-mentioned preparation method, as a kind of preferred implementation, in described decolouring fixation step, described decolouring
The condition processed is as follows: described squid fragment is 100:2-8 with the mass ratio of decolorising agent, and ultrasonic temperature is
45-65 DEG C, power is 200-350W, and the time is 1～3h；The condition of described fixation treatment is as follows: described squid
Fish fragment is 100:1-4 with the mass ratio of color fixing agent, and ultrasonic temperature is 45-65 DEG C, and power is 200-350W,
Time is 1～3h.
Above-mentioned preparation method, as a kind of preferred implementation, in described decolouring fixation step, described extraction
The extractant used in process is pure water, and Extracting temperature is 80-95 DEG C, and extraction time is 90-150min.
Above-mentioned preparation method, as a kind of preferred implementation, in described enzymolysis pressurization steps, described enzymolysis
Process specific as follows: described squid protein extract first regulates pH to 8.0-10.0 with alkali liquor, add described
Alkaline protease, enzymolysis 1-3h, temperature is 55 DEG C-65 DEG C；Then at 90-100 DEG C of enzyme denaturing 4-10min；Again will
Temperature is down to 40-50 DEG C, obtains squid protein enzymatic hydrolyzate.It is highly preferred that described alkali liquor is food stage hydroxide
Sodium, food stage potassium hydroxide or food stage sodium bicarbonate, concentration is 0.02-0.04mol/L；Further, institute
The weight ratio stating alkaline protease and described squid fragment is (1-5): 100；Described alkaline protease is preferably
Properase E。
Above-mentioned preparation method, as a kind of preferred implementation, in described enzymolysis pressurization steps, described pressurization
Process specific as follows: at 100-160 DEG C, be first pressurized to 5kgf/cm2, and keep 15-20min, make pressure
Uniformly, pressure then is made suddenly to be down to 0kgf/cm within the 0-4 second2。
Above-mentioned preparation method, as a kind of preferred implementation, in described enzymolysis pressurization steps, described ultrafiltration
Process be use ultrafilter membrane molecular cut off be the molecule of below 3KD.
Above-mentioned preparation method, as a kind of preferred implementation, also includes dense after described enzymolysis pressurization steps
Contracting drying steps: by described bioactive peptide through process, spray drying treatment are concentrated in vacuo, obtain squid albumen
Functional activity Gly-His-Lys；It is highly preferred that the concrete grammar of described concentrate drying is: in vacuum 82.7～
Under 90.6KPa, by described bioactive peptide by nebulizer, dimerization becomes the 10～200 vaporific microgranules of purpose, described mist
Shape granule directly contacts with hot-air and carries out heat exchange, and the short time completes to be dried, and drying time is 10～15min.
Compared to existing technology, there is advantages that
1, because the molecular weight control of the squid protein functional bioactive peptide of the present invention is at below 3kD, so easily
Digested, there is anticancer, resisting fatigue, defying age, raising body immunity, detoxicating intestine
Etc. effect；In addition with rich in taste, the feature such as edible easy, can be used for preparing antioxidation, blood pressure lowering,
Atherosclerosis or the health food of beauty treatment or medicine；There is wide market and application prospect.
2, enzymolysis solution is pressurizeed by the present invention, the activated squid small peptide obtained, and not only chain rupture is uniform,
And molecular weight control can be within 3KD；The present invention substantially reduces squid also by ultrasonic decolouring and fixation
Plate processes the time, and eliminates the fat deposit in squid plate, fundamentally ensure that the quality of little peptide；This
Synergism between each step and parameter of bright method, can be further ensured that the quality of bioactive peptide product.
Detailed description of the invention
The preparation method of a kind of low-molecular-weight defat squid protein functional bioactive peptide, comprises the following steps successively:
Step one, pretreatment: choose thickness and reach Argentinian squid or the Peru squid of 3cm～8cm, by liver
Dirty organ such as grade removes, and draws out the vertebrae of similar plastics sheet, white mucosa in tripe, obtains squid plate；And
Removing surface blade twill scribing at this squid plate, the thickness of blade incision squid plate about 1/2, between every cutter
It is separated by 2cm～5cm, rubs after squid plate is cleaned with pure water, obtain the squid fragment of rubbing or become
Squid is rotten.Owing to having squid skin to wrap up outside squid plate, even if there is also the thickest skin and flesh adhesion when rubbing
Squid block, process so carrying out in advance cutting drawing, in order to later stage decolouring is more uniformly rapidly.
Wherein, the squid of employing is live squid or the freezing new Fresh squid less than 15 days；
Exemplarily, the thickness of above-mentioned squid can be the arbitrary value in 3cm, 4cm, 5cm, 8cm or appoint
Meaning scope therebetween；The interval of above-mentioned every cutter can be any in 2cm, 3cm, 4.5cm, 5cm
Value or scope the most therebetween.
Step 2, decolouring and fixation:
Step by step one, ultrasonic desolventing technology: the NaOH solution using 0.01～0.045mol/L is decolorising agent,
Add in the squid fragment rubbed and ultrasonic Treatment, be neutral with pure water washing to pH, obtain decolouring and produce
Thing.
Wherein, this squid fragment is 100:2-8 with the mass ratio of decolorising agent, and ultrasonic temperature is 45-65 DEG C, merit
Rate is 200-350W, and the time is 1～3h.
Step by step two, ultrasonic fixation treatment: the Ca (OH) using mass percent concentration to be 0.1%～0.4%2
Solution is color fixing agent, adds in above-mentioned Decolorization product and ultrasonic Treatment, obtains fixation product.
Wherein, squid fragment is 100:1-4 with the mass ratio of color fixing agent, and ultrasonic temperature 45-65 DEG C, power is
200-350W, the time 1～3h.
Step by step three, extraction process: after above-mentioned fixation product is washed to pH neutrality, use the pure water to be
Protein extracting agent, extracts collagen protein in 80-95 DEG C of hot pure water, and extraction times is 90-150min, mistake
Filter insoluble thing, obtain squid protein extract.
In this step, the purpose of desolventing technology is to slough squid skin and squid meat skin color；Fixation treatment
Purpose is to ensure that the fixing of color after squid decolouring, generally light color；Two steps are ultrasonic is solid in order to improve decolouring
Colour efficiency, saving processes the time.Additionally, decolouring and fixation treatment can remove the fat deposit in squid plate,
Fundamentally ensure the quality of little peptide and the yield of small active peptides.
Exemplarily, in above-mentioned ultrasonic desolventing technology, the concentration of above-mentioned NaOH solution can be 0.01mol/L,
Arbitrary value in 0.02mol/L, 0.03mol/L, 0.04mol/L, 0.045mol/L or model the most therebetween
Enclosing, the mass ratio of above-mentioned squid fragment and decolorising agent can be any in 7%, 6%, 5%, 4%, 3%
Value or scope the most therebetween, above-mentioned ultrasonic temperature can be 45 DEG C, 50 DEG C, 55 DEG C, 60 DEG C, 65 DEG C
In arbitrary value or scope arbitrarily the most therebetween, power can be 200W, 250W, 300W, 330W,
Arbitrary value in 350W or scope the most therebetween, the time can be in 1h, 1.5h, 2h, 3h
Arbitrary value or scope the most therebetween；In above-mentioned ultrasonic fixation treatment, above-mentioned Ca (OH)2The matter of solution
Amount percent concentration can be the arbitrary value in 0.1%, 0.2%, 0.25%, 0.3%, 0.4% or any two
Scope between person, the mass ratio of above-mentioned squid fragment and decolorising agent can be 4%, 3.5%, 3%, 2%,
Arbitrary value in 1% or scope the most therebetween, above-mentioned ultrasonic temperature can be 45 DEG C, 50 DEG C, 55 DEG C,
60 DEG C, the arbitrary value in 65 DEG C or scope the most therebetween, power can be 200W, 250W, 300
Arbitrary value in W, 330W, 350W or scope the most therebetween, the time can be 1h, 1.5h,
Arbitrary value in 2h, 3h or scope the most therebetween；During said extracted processes, Extracting temperature can be
80 DEG C, 85 DEG C, 88 DEG C, 90 DEG C, the arbitrary value in 95 DEG C or scope the most therebetween；Extraction times
For 90min, 95min, 100min, 105min, 100min, 120min, 130min, 140min,
Arbitrary value in 145min, 150min or scope the most therebetween.
Step 3, enzymolysis pressurize:
Step by step one, enzymolysis processing: this squid protein extract first regulates pH to 8.0-10.0 with alkali liquor, then
Adding alkaline protease, alkaline protease is preferably Properase E, and enzymolysis time is 1-3h, hydrolysis temperature
It it is 55 DEG C-65 DEG C；Then at 90-100 DEG C of enzyme denaturing 4-10min cool the temperature to again 40-50 DEG C (such as: 40 DEG C,
45 DEG C, 50 DEG C), obtain squid protein enzymatic hydrolyzate.
Wherein, this alkaline protease is (1-5) with the weight ratio of squid fragment: 100.
Step by step two, pressurized treatments: being placed in pressure pan by this squid protein enzymatic hydrolyzate, the requirement of pressurization is:
At 100-160 DEG C, first it is pressurized to 5kgf/cm2, and keep 15-20min, make pressure uniform, then at 0-4
Suddenly pressure is made to be down to 0kgf/cm in Miao2.The pressurized conditions of the present invention uniformly fracture and 3KD to long peptide chain
The yield of following small active peptides plays an important role.
Step by step three, hyperfiltration treatment: by the little molecule that rolling NF membrane molecular cut off is below 3KD,
Obtain squid bioactive peptide liquid.The bioactive peptide molecule amount of this squid bioactive peptide liquid is below 3KD, for oligopeptide,
Easily it is rapidly absorbed by a human.
In this step, the purpose of enzymolysis processing is that to allow protein digestion be longer polypeptide；The mesh of pressurized treatments
Be that long polypeptide chain is processed as little peptide by pressurization the method for moment blood pressure lowering by longer polypeptide.
Exemplarily, in above-mentioned enzymolysis processing, the pH value of above-mentioned squid protein extract is adjusted to 8,8.5,
9, arbitrary value or scope the most therebetween in 10, the above-mentioned pH value being used for regulating squid protein extract
Alkali liquor be food stage sodium hydroxide, food stage potassium hydroxide or food stage sodium bicarbonate, concentration is
0.02-0.04mol/L, above-mentioned enzymolysis time is arbitrary value or arbitrarily the most therebetween in 1h, 1.5h, 2h, 3h
Scope, the temperature of enzymolysis is 55 DEG C, 56 DEG C, 58 DEG C, 60 DEG C, 62 DEG C, 64 DEG C, arbitrary value in 65 DEG C
Or scope arbitrarily the most therebetween, enzyme-removal temperature is 90 DEG C, 93 DEG C, 95 DEG C, 98 DEG C, arbitrary value in 100 DEG C
Or scope arbitrarily the most therebetween, the enzyme denaturing time is arbitrary value or appoint in 4min, 5min, 8min, 10min
Meaning scope therebetween, alkaline protease and the weight ratio of squid fragment are 1:100,2:100,2.5:100,
Arbitrary value or scope the most therebetween in 3:100,4:100,5:100；In above-mentioned pressurized treatments, pressurization
Temperature be 100 DEG C, 102 DEG C, 105 DEG C, 108 DEG C, 110 DEG C, 115 DEG C, 120 DEG C, 125 DEG C, 130 DEG C,
135 DEG C, 140 DEG C, 145 DEG C, 150 DEG C, 155 DEG C, 158 DEG C, arbitrary value or the most therebetween in 160 DEG C
Scope, the time of holding be in 15min, 16min, 17min, 18min, 19min, 20min arbitrarily
Value or scope arbitrarily the most therebetween, the time of blood pressure lowering is 0 second, 1 second, 2 seconds, 3 seconds, 4 seconds, in 5 seconds
Arbitrary value or scope the most therebetween.
Prepared bioactive peptide solution can be concentrated for increasing bioactive peptide concentration, be dried to powder；Therefore conduct
A kind of preferred implementation, the preparation of the low-molecular-weight defat squid protein functional bioactive peptide that the present invention provides
Method can also include after step 3 step 4, i.e. concentrate drying step:
By this squid bioactive peptide liquid through process, spray drying treatment are concentrated in vacuo, obtain squid protein function
Property active peptide powder.Exemplarily, in above-mentioned steps four, the concrete grammar of concentrate drying is: in vacuum 82.7～
Under 90.6KPa, by squid bioactive peptide liquid by nebulizer (spray gun), dimerization become 10～200 mesh (such as 20 mesh,
30 mesh, 50 mesh, 80 mesh, 120 mesh, 160 mesh, 180 mesh, 190 mesh) vaporific microgranule, with hot-air
Directly contact carries out heat exchange, and the short time completes to be dried, drying time be 10～15min (such as 11min,
12min、13min、14min)。
Below in conjunction with specific embodiment, the present invention is expanded on further.Should be understood that these embodiments are only used for this
Invention rather than restriction the scope of the present invention.Externally should be understood that after having read present disclosure,
The present invention is made various changes or modifications by those skilled in the art, and these equivalent form of values fall within the application institute equally
Attached claims limited range.
Embodiment 1
In the present embodiment, preparation method in turn includes the following steps:
(1) pretreatment: choose thickness and reach Argentinian squid or the Peru squid of 3cm, the organs such as liver are gone
Remove, obtain squid plate；And removing surface blade twill scribing, the thickness of blade incision squid plate about 1/2,
It is separated by 5cm between every cutter, rubs after squid plate is cleaned with pure water, obtain the squid fragment rubbed.
(2) decolouring: be decolorising agent by the NaOH solution of 0.045mol/L, by squid fragment and decolorising agent it
Mass ratio is 100:4, is joined by decolorising agent in squid fragment and ultrasonic Treatment, and temperature 45 C, power is
300W, time 2h, be neutral with pure water washing to pH, obtain Decolorization product.
(3) fixation: the Ca (OH) using concentration to be 0.4% (quality)2Solution is color fixing agent, squid fragment with
The mass ratio of color fixing agent is 100:3, is joined by color fixing agent in Decolorization product and ultrasonic Treatment, temperature 45 C,
Power is 200W, processes time 2h, obtains fixation product.
(4) extract: after fixation product is washed to pH neutrality, employing pure water is protein extracting agent,
Extracting collagen protein in 80 DEG C of hot water, extraction times is 2h, filters, removes insoluble thing, and extraction obtains
Squid protein extract.
(5) enzymolysis: be first that 0.02mol/L food stage sodium hydroxide solution regulates this squid albumen and carries by concentration
Take the pH to 8.0-10.0 of liquid, then add alkaline protease Properase E enzymolysis 2h at 60 DEG C, then exist
Enzyme denaturing 5min at 100 DEG C, is cooled to 40 DEG C, and enzyme dosage is the 2.0% of squid fragment weight, obtains squid egg
White enzymolysis solution.
(6) pressurization: again the squid protein enzymatic hydrolyzate obtained is put in pressure pan, be pressurized at 110 DEG C
5kgf/cm2, and keeping 15min so that it is pressure is uniform, then makes suddenly pressure be down to 0kgf/cm at 4 seconds2。
(7) ultrafiltration: be the little molecule within 3KD by rolling NF membrane molecular cut off, obtains squid
Bioactive peptide liquid.
(8) concentrate drying: be concentrated in vacuo by this squid bioactive peptide liquid, is spray-dried, obtains product.
Embodiment 2
In the present embodiment, preparation method in turn includes the following steps:
(1) pretreatment: choose thickness and reach Argentinian squid or the Peru squid of 5cm, the organs such as liver are gone
Remove, obtain squid plate；And removing surface blade twill scribing, the thickness of blade incision squid plate about 1/2,
It is separated by 4cm between every cutter, rubs after squid plate is cleaned with pure water, obtain the squid fragment rubbed.
(2) decolouring: be decolorising agent by the NaOH solution of 0.025mol/L, by squid fragment and decolorising agent it
Mass ratio is 100:5, is joined by decolorising agent in squid fragment and ultrasonic Treatment, and temperature 55 DEG C, power is
300W, time 1h, be neutral with pure water washing to pH, obtain Decolorization product.
(3) fixation: the Ca (OH) using concentration to be 0.4% (quality)2Solution is color fixing agent, squid fragment with
The mass ratio of color fixing agent is 100:4, is added by color fixing agent in Decolorization product and ultrasonic Treatment, temperature 55 DEG C,
Power is 300W, processes time 1h, obtains fixation product.
(4) extract: after fixation product is washed to pH neutrality, employing pure water is protein extracting agent,
In 80 DEG C of hot water, extract collagen protein, extract 2h, filter, remove insoluble thing, extract and obtain squid egg
White extracting solution.
(5) enzymolysis: by this squid protein extract, is first 0.02mol/L food stage potassium hydroxide by concentration
Solution regulation pH to 8.0, then add alkaline protease Properase E enzymolysis 2h at 65 DEG C, then at 100 DEG C
Lower enzyme denaturing 4min, is cooled to 50 DEG C, and enzyme dosage is the 2.0% of squid fragment weight, obtains squid protease
Solve liquid.
(6) pressurization: again the squid protein enzymatic hydrolyzate obtained is put in pressure pan, be pressurized at 120 DEG C
5kgf/cm2, and keeping 15min so that it is pressure is uniform, then makes suddenly pressure be down to 0kgf/cm at 4 seconds2。
(7) ultrafiltration: be the little molecule within 3KD by rolling NF membrane molecular cut off, obtains squid
Bioactive peptide liquid.
(8) concentrate drying: be concentrated in vacuo by this squid bioactive peptide liquid, is spray-dried, obtains product.
Embodiment 3
In the present embodiment, preparation method in turn includes the following steps:
(1) pretreatment: choose thickness and reach Argentinian squid or the Peru squid of 8cm, the organs such as liver are gone
Remove, obtain squid plate；And removing surface blade twill scribing, the thickness of blade incision squid plate about 1/2,
It is separated by 3cm between every cutter, rubs after squid plate is cleaned with pure water, obtain the squid fragment rubbed.
(2) decolouring: be decolorising agent by the NaOH solution of 0.025mol/L, by squid fragment and decolorising agent it
Mass ratio is 100:6, is joined by decolorising agent in squid fragment and ultrasonic Treatment, and temperature 55 DEG C, power is
300W, time 1h, be neutral with pure water washing to pH, obtain Decolorization product.
(3) fixation: the Ca (OH) using concentration to be 0.4% (quality)2Solution is color fixing agent, squid fragment with
The mass ratio of color fixing agent is 100:4, is added by color fixing agent in Decolorization product and ultrasonic Treatment, temperature 55 DEG C,
Power is 300W, processes time 1h, obtains fixation product.
(4) extract: after fixation product is washed to pH neutrality, employing pure water is protein extracting agent,
In 80 DEG C of hot water, extract collagen protein, extraction times 2h, filter, remove insoluble thing, extract and obtain squid
Fish protein extracting solution.
(5) enzymolysis: by this squid protein extract, is first 0.02mol/L food stage sodium hydroxide by concentration
Solution regulation pH to 8.0, then add alkaline protease Properase E enzymolysis 1h at 60 DEG C, then at 100 DEG C
Lower enzyme denaturing 5min, is cooled to 45 DEG C, and enzyme dosage is the 4.0% of squid fragment weight, obtains squid protease
Solve liquid.
(6) pressurization: again the squid protein enzymatic hydrolyzate obtained is put in pressure pan, be pressurized at 135 DEG C
5kgf/cm2, and keeping 20min so that it is pressure is uniform, then makes suddenly pressure be down to 0kgf/cm at 4 seconds2。
(7) ultrafiltration: be the little molecule within 3KD by rolling NF membrane molecular cut off, obtains squid
Bioactive peptide liquid.
(8) concentrate drying: be concentrated in vacuo by this squid bioactive peptide liquid, is spray-dried, obtains product.
Embodiment 4
In the present embodiment, preparation method in turn includes the following steps:
(1) pretreatment: choose thickness and reach Argentinian squid or the Peru squid of 7cm, the organs such as liver are gone
Remove, obtain squid plate；And removing surface blade twill scribing, the thickness of blade incision squid plate about 1/2,
It is separated by 2cm between every cutter, rubs after squid plate is cleaned with pure water, obtain the squid fragment rubbed.
(2) decolouring: be decolorising agent by the NaOH solution of 0.045mol/L, by squid fragment and decolorising agent it
Mass ratio is 100:7, is joined by decolorising agent in squid fragment and ultrasonic Treatment, and temperature 55 DEG C, power is
300W, time 1h, be neutral with pure water washing to pH, obtain Decolorization product.
(3) fixation: the Ca (OH) using concentration to be 0.4% (quality)2Solution is color fixing agent, squid fragment with
The mass ratio of color fixing agent is 100:3, is added by color fixing agent in Decolorization product and ultrasonic Treatment, temperature 55 DEG C,
Power is 300W, processes time 1h, obtains fixation product.
(4) extract: after fixation product is washed to pH neutrality, employing pure water is protein extracting agent,
Extracting collagen protein in 80 DEG C of hot water, extraction times is 2h, filters, removes insoluble thing, and extraction obtains
Squid protein extract.
(5) enzymolysis: by this squid protein extract, is first 0.02mol/L food stage potassium hydroxide by concentration
Solution regulation pH to 8.0, then add alkaline protease Properase E enzymolysis 2h at 65 DEG C, then at 100 DEG C
Lower enzyme denaturing 8min, is cooled to 50 DEG C, and enzyme dosage is the 1.0% of squid fragment weight, obtains squid protease
Solve liquid.
(6) pressurization: again the squid protein enzymatic hydrolyzate obtained is put in pressure pan, be pressurized at 160 DEG C
5kgf/cm2, and keeping 20min so that it is pressure is uniform, then makes suddenly pressure be down to 0kgf/cm at 4 seconds2。
(7) ultrafiltration: be the little molecule within 3KD by rolling NF membrane molecular cut off, obtains squid
Bioactive peptide liquid.
(8) concentrate drying: be concentrated in vacuo by this squid bioactive peptide liquid, is spray-dried, obtains product.
Embodiment 5
In the present embodiment, preparation method in turn includes the following steps:
(1) pretreatment: choose thickness and reach Argentinian squid or the Peru squid of 5cm, the organs such as liver are gone
Remove, obtain squid plate；And removing surface blade twill scribing, the thickness of blade incision squid plate about 1/2,
It is separated by 4cm between every cutter, rubs after squid plate is cleaned with pure water, obtain the squid fragment rubbed.
(2) decolouring: be decolorising agent by the NaOH solution of 0.045mol/L, by squid fragment and decolorising agent it
Mass ratio is 100:4, is joined by decolorising agent in squid fragment and ultrasonic Treatment, and temperature 55 DEG C, power is
300W, time 1h, be neutral with pure water washing to pH, obtain Decolorization product.
(3) fixation: the Ca (OH) using concentration to be 0.4% (quality)2Solution is color fixing agent, squid fragment with
The mass ratio of color fixing agent is 100:2, is added by color fixing agent in Decolorization product and ultrasonic Treatment, temperature 55 DEG C,
Power is 300W, processes time 2h, obtains fixation product.
(4) extract: after fixation product is washed to pH neutrality, employing pure water is protein extracting agent,
Extracting collagen protein in 80 DEG C of hot water, extraction times is 2h, filters, removes insoluble thing, and extraction obtains
Squid protein extract.
(5) enzymolysis: by this squid protein extract, is first 0.04mol/L food stage sodium bicarbonate by concentration
Solution regulation pH to 8.0, then add alkaline protease Properase E enzymolysis 2h at 60 DEG C, then at 100 DEG C
Lower enzyme denaturing 5min, is cooled to 45 DEG C, and enzyme dosage is the 1.0% of squid fragment weight, obtains squid protease
Solve liquid.
(6) pressurization: again the squid protein enzymatic hydrolyzate obtained is put in pressure pan, be pressurized at 100 DEG C
5kgf/cm2, and keeping 20min so that it is pressure is uniform, then makes suddenly pressure be down to 0kgf/cm at 4 seconds2。
(7) ultrafiltration: be the little molecule within 3KD by rolling NF membrane molecular cut off, obtains squid
Bioactive peptide liquid.
(8) concentrate drying: be concentrated in vacuo by this squid bioactive peptide liquid, is spray-dried, obtains active peptide powder.
Embodiment 6
The present embodiment is in addition to pressurization steps is different from embodiment 5, and other operating procedures are same as in Example 5,
The pressurized conditions of this comparative example is as follows: be pressurized to 5kgf/cm at 160 DEG C2, and keep 15min so that it is pressure
Uniformly, then pressure was made suddenly to be down to 0kgf/cm at 4 seconds2。
The performance test of the product that each embodiment above-mentioned prepares and data are as follows:
Trichloroacetic acid is utilized to make protein precipitant, by the protein in enzymolysis protein powder (i.e. sample) and peptide
The peptide precipitation that chain is longer, and is dissolved out little for short chain therein peptide acid, by centrifugation, filter, digest,
Distillation, measures its albumen quality.This method is with reference to the People's Republic of China's oligomeric Gly-His-Lys of light marine fish
(QB/T2879-2007) revise on the basis of.
The protein hydrolysate (wherein comprising peptides and free amino acid) of lower molecular weight, dissolves in trichlorine
Acetic acid solution, hmw protein is susceptible to precipitation in solution of trichloroacetic acid.Sample is molten through trichloroacetic acid
Xie Hou, centrifugation goes out deposit, and in clear liquid, sour molten protein content deducts free aminoacid content and is
Oligopeptide content.
Concrete operations for take 2.00g sample (squid protein functional active peptide powder prepared by embodiment 1-6),
Add 10ml15wt%TCA (trichloroacetic acid), mix homogeneously, stand 5min.Solution is quantitatively shifted,
Under 4000rpm, centrifugal 10min, takes whole supernatant.Soluble protein is measured by GB/T 5009.5 method
Matter, protein conversion coefficient is 6.25, presses GB/T 5009.5 method simultaneously and measures free amino acid.Each sample
Product parallel assay three times.Oligopeptide content=content of soluble protein-free aminoacid content.
The active small peptide content of squid protein functional active peptide powder prepared by embodiment 1-6 is shown in Table 1.
Squid protein functional active peptide powder content results prepared by table 1 embodiment 1-6
Claims (10)
1. the preparation method of a low-molecular-weight defat squid protein functional bioactive peptide, it is characterised in that:
Comprise the following steps:
Pre-treatment step: squid is removed internal organs and processes, obtain squid plate；Again by described squid plate
Carry out scribing process, break process, obtain the squid fragment rubbed；
Decolouring fixation step: described squid fragment is carried out ultrasonic desolventing technology, ultrasonic fixation treatment, carries
Take process, obtain squid protein extract；
Enzymolysis pressurization steps: described squid protein extract is carried out successively enzymolysis processing, pressurized treatments,
Hyperfiltration treatment, obtains described bioactive peptide.
The preparation method of low-molecular-weight defat squid protein functional bioactive peptide the most according to claim 1,
It is characterized in that:
In described pre-treatment step, described squid is Argentina squid or Peru squid；Preferably, described
Squid is live squid or the freezing new Fresh squid less than 15 days.
The system of low-molecular-weight defat squid protein functional bioactive peptide the most according to claim 1 and 2
Preparation Method, it is characterised in that；
In described pre-treatment step, the thickness of described squid plate is between 3cm～8cm；
Preferably, the vertebrae of similar plastics sheet, white mucosa in tripe got rid of by described squid plate；
Preferably, during described scribing processes, the thickness of described squid plate about 1/2 cut by cutter, between every cutter
It is separated by 2cm～5cm.
The preparation method of low-molecular-weight defat squid protein functional bioactive peptide the most according to claim 1,
It is characterized in that:
In described decolouring fixation step, decolorising agent is the NaOH solution of 0.01～0.045mol/L；Fixation
Agent be mass percent concentration be the Ca (OH) of 0.1%～0.4%2Solution.
5. according to the preparation of low-molecular-weight defat squid protein functional bioactive peptide described in claim 1 or 4
Method, it is characterised in that:
In described decolouring fixation step, the condition of described desolventing technology is as follows: described squid fragment and decolouring
The mass ratio of agent is 100:2-8, and ultrasonic temperature is 45-65 DEG C, and power is 200-350W, the time be 1～
3h；
The condition of described fixation treatment is as follows: described squid fragment is 100:1-4 with the mass ratio of color fixing agent,
Ultrasonic temperature is 45-65 DEG C, and power is 200-350W, and the time is 1～3h.
6. according to the preparation of low-molecular-weight defat squid protein functional bioactive peptide described in claim 1 or 4
Method, it is characterised in that:
In described decolouring fixation step, the extractant used in described extraction process is pure water, extracts temperature
Degree is for 80-95 DEG C, and extraction time is 90-150min.
The preparation method of low-molecular-weight defat squid protein functional bioactive peptide the most according to claim 1,
It is characterized in that:
In described enzymolysis pressurization steps, described enzymolysis processing is specific as follows: described squid protein extract is first
Regulating pH to 8.0-10.0 with alkali liquor, add described alkaline protease, enzymolysis 1-3h, temperature is 55 DEG C
-65℃；Then at 90-100 DEG C of enzyme denaturing 4-10min；Cool the temperature to 40-50 DEG C again, obtain squid protease
Solve liquid.Preferably, described alkali liquor is food stage sodium hydroxide, food stage potassium hydroxide or food stage carbonic acid
Hydrogen sodium, concentration is 0.02-0.04mol/L；It is highly preferred that described alkaline protease and described squid fragment
Weight ratio is (1-5): 100；Described alkaline protease is preferably Properase E.
8. according to the preparation of low-molecular-weight defat squid protein functional bioactive peptide described in claim 1 or 7
Method, it is characterised in that:
In described enzymolysis pressurization steps, described pressurized treatments is specific as follows: at 100-160 DEG C, first pressurize
To 5kgf/cm2, and keep 15-20min, make pressure uniform, then make suddenly pressure be down within the 0-4 second
0kgf/cm2。
The preparation method of low-molecular-weight defat squid protein functional bioactive peptide the most according to claim 1,
It is characterized in that:
In described enzymolysis pressurization steps, described hyperfiltration treatment be use ultrafilter membrane molecular cut off be 3KD with
Under molecule.
The preparation side of low-molecular-weight defat squid protein functional bioactive peptide the most according to claim 1
Method, it is characterised in that also include concentrate drying step after described enzymolysis pressurization steps: by described activity
Peptide, through process, spray drying treatment are concentrated in vacuo, obtains squid protein functional active peptide powder；Preferably
Ground, the concrete grammar of described concentrate drying is: under vacuum 82.7～90.6KPa, by described bioactive peptide
By nebulizer, dimerization becomes the 10～200 vaporific microgranules of purpose, and described mist particles directly contacts with hot-air
Carrying out heat exchange, the short time completes to be dried, and drying time is 10～15min.
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| CN107183652A (en) * | 2017-06-05 | 2017-09-22 | 舟山市齐晟水产有限公司 | A kind of cherry boils squid flavouring material formula |
| CN107365818A (en) * | 2017-07-07 | 2017-11-21 | 浙江海润达科技有限公司 | A kind of preparation method of mussel protein small peptide and its preparation method of respective combination chewable tablets |
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