CN105734103A - Preparation method of casein hydrolysis peptide - Google Patents
Preparation method of casein hydrolysis peptide Download PDFInfo
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- CN105734103A CN105734103A CN201610309219.9A CN201610309219A CN105734103A CN 105734103 A CN105734103 A CN 105734103A CN 201610309219 A CN201610309219 A CN 201610309219A CN 105734103 A CN105734103 A CN 105734103A
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- 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
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/34—Extraction; Separation; Purification by filtration, ultrafiltration or reverse osmosis
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Abstract
The invention discloses a preparation method of casein hydrolysis peptide. The preparation method comprises the following steps: (1) dissolving casein with water to prepare a casein solution of which the mass concentration is 5-15wt%, and adjusting the pH value to 5-10: (2) adding an enzyme and an enzyme activity keeping agent into the casein solution to perform hydrolysis at 45-75 DEG C for 1-5 hours; (3) performing enzyme inactivation for 5-15 minutes in a water bath of 85-100 DEG C to obtain an enzyme hydrolysis solution; (4) adjusting the pH value of the enzyme hydrolysis solution to a 4-5 isoelectric point for deposition, and performing centrifugal separation; and (5) performing ultrafiltration and freeze-drying. According to the preparation method of the casein hydrolysis peptide, disclosed by the invention, the casein hydrolysis peptide is moderate in hydrolysis degree, high in content of oligopeptide, low in content of free amino acid, low in allergenicity, and easy to digest and absorb. The raw materials used in the preparation method contain no harmful substances, cannot pollute environment, and are wide in source, the preparation process is simple, and the product has the effects of improving immunity, maintaining beauty and keeping young, promoting cell growth, and resisting aging.
Description
Technical field
The present invention relates to technical field of health care food, be specifically related to the preparation method of a kind of casein hydrolysate peptides.
Background technology
Modern nutriology research finds, mankind's protein of ingesting after gastral enzyme effect, be mostly with
Little peptide form is digested and assimilated, and another new knowledge of peptide Nutrition is that protein can produce in enzymolysis process
Give birth to some biologically active peptides with special physiological regulatory function.
Casein (casein) is a kind of protein rich in bioactive sequences, specificity restriction endonuclease (as
Trypsin etc.) and similar physiological condition under, these bioactive sequences can be released, and is had
Bioactive casein biologically active peptide.These biologically active peptides are exempted from regulation gastrointestinal movement, regulation
Epidemic disease system, anti-blood pressure raise, antibacterial, antithrombotic, antiviral, anticancer, remove free radical and promote ore deposit
The aspects such as thing element absorption play an important role.
The preparation method of biologically active peptide mainly has three kinds at present: (1) extraction method, i.e. from the life of nature
Object extracts itself intrinsic natural activity peptide；(2) Hydrolyze method, will protein be hydrolyzed with
Obtain the biologically active peptide with physiological function；(3) synthetic method, i.e. the method system of applied chemistry synthesis
Standby biologically active peptide.Selecting different preparation methoies based on different purposes, different preparation methoies respectively has
Pluses and minuses.
Protein inherently biological activity propeptide, selects these polypeptide chains of suitable protease hydrolysis,
Discharge having bioactive peptide fragment, thus the biologically active peptide with physiological function can be prepared.
Employing Production by Enzymes bioactive peptide is the focus of current research.Enzymatic hydrolysis has much physics, chemical modification
Unrivaled advantage.First it is a kind of hydrolysis incomplete, halfway, its product be mainly peptide and
It not aminoacid；Next to that the mild condition of reaction, the response time is short, and efficiency is high, does not produce racemization and makees
With, the most do not destroy aminoacid；It is that product purity is high again, the advantages such as product is easily separated, with low cost；
During additionally, utilize protease to carry out aminosal, it is also possible to by selecting the kind of enzyme, control reaction bar
The measures such as part obtain the biologically active peptide with particular physiological function.
The invention provides the preparation method of a kind of casein hydrolysate peptides, it is easy to absorb, and can effectively drop
The content of anaphylactogen in low casein.
Summary of the invention
For the deficiencies in the prior art, the technical problem to be solved is to provide a kind of casein water
Solve the preparation method of peptide.
For achieving the above object, the adopted technical solution is that:
The preparation method of a kind of casein hydrolysate peptides, comprises the following steps:
(1) casein water dissolution is configured to the casein solution that mass concentration is 5-15wt%, and adjusts
Joint pH to 5-10；
(2) enzyme and enzyme live keeping agent are added in casein solution, hydrolyze 1-5 hour at 45-75 DEG C；
(3) under 85-100 DEG C of water-bath, enzyme denaturing is lived 5-15 minute, obtains enzymolysis solution；
(4) pH value of regulation enzymolysis solution is to 4-5 isoelectric precipitation, centrifugation；
(5) ultrafiltration, lyophilizing.
In described step (1) and step (4), regulation pH value can use NaOH solution, HCl
Solution etc..
Preferably, the 0.5-5% that addition is casein solution quality of enzyme, enzyme in described step (2)
The addition of live keeping agent is the 0.1-1% of casein solution quality.
Preferably, the enzyme in described step (2) is in papain, trypsin, bromelain
The mixture of one or more.
It is highly preferred that the enzyme in described step (2) is by papain, trypsin, bromelain
Mixing, described papain, trypsin, the mass ratio of bromelain are (1-3): (1-3):
(1-3)。
Preferably, the enzyme live keeping agent in described step (2) is shell six sugar, hydroxyl isomaltulose, chitotriose
In one or more mixture.
It is highly preferred that the enzyme live keeping agent in described step (2) is by shell six sugar, hydroxyl isomaltulose, shell three
Sugar mixes, and described shell six sugar, hydroxyl isomaltulose, the mass ratio of chitotriose are (1-3): (1-3):
(1-3)。
Preferably, the centrifugation rotating speed in described step (4) is 2000-5000 rev/min, and the time is
20-40 minute.
Preferably, the ultrafiltration in described step (5) is with the ultrafilter membrane ultrafiltration of molecular cut off 1-10kD.
The preparation method of casein hydrolysate peptides of the present invention, degree of hydrolysis is moderate, and oligopeptide content is high, free amine group
Acid is low, and sensitization is low, it is easy to digest and assimilate.Unharmful substance in raw material used by the present invention, does not pollutes
Environment, and raw material sources of the present invention are extensive, and preparation technology is simple, and product has raising immunity, U.S.
Hold skin care, promote cell growth, the effect of defying age.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is described further, the following stated, only to the present invention
Preferred embodiment, not does the restriction of other forms, any technology being familiar with this specialty to the present invention
Personnel are changed to the Equivalent embodiments changed on an equal basis possibly also with the technology contents of the disclosure above.Every
Without departing from the present invention program content, any letter following example done according to the technical spirit of the present invention
Single amendment or equivalent variations, all fall within protection scope of the present invention.
Each raw material introduction in embodiment:
Shell six sugar, No. CAS: 41708-95-6.
Hydroxyl isomaltulose, No. CAS: 64519-82-0.
Chitotriose, No. CAS: 41708-93-4.
Papain, No. CAS: 9001-73-4, Zhengzhou Kang Yuan chemical products company limited provides, enzyme
Live 200U/mg.
Trypsin, No. CAS: 9002-07-7, Zhengzhou good brother chemical products company limited provides, enzyme
Live 200U/mg.
Bromelain, No. CAS: 9001-00-7, Sheng Xie bio tech ltd, Shandong provides, enzyme
Live 200U/mg.
Embodiment 1
The preparation method of casein hydrolysate peptides, comprises the following steps:
(1) casein deionized water dissolving is configured to the casein solution that mass concentration is 10wt%
(the most every 10 grams of caseins of 100 grams of deionized water dissolving), and adjust with the sodium hydroxide solution of 2mol/L
Joint pH to 7.8；
(2) will enzyme and enzyme live keeping agent addition casein solution be uniformly mixed, the wherein addition of enzyme
For the 1.2% of casein solution quality, the addition of enzyme live keeping agent be casein solution quality 0.6% (i.e.
Every 100 grams of casein solution add enzyme 1.2 grams, enzyme live keeping agent 0.6 gram), at 60 DEG C, hydrolyze 2 little
Time, obtain hydrolyzing afterproduct；
(3) then hydrolysis afterproduct is placed in lower 10 minutes enzyme denaturing of 95 DEG C of water-baths live, obtains enzymolysis solution；
(4) with the pH value of the hydrochloric acid solution regulation enzymolysis solution of 2mol/L to 4.6 casein isoelectric point, IPs, treat
After Precipitation, remove unhydrolysed casein with 3000r/min centrifugation 30min, be centrifuged
After supernatant；
(5) take the ultrafilter membrane ultrafiltration of the supernatant molecular cut off 6kD after being centrifuged, collect filtrate,
Lyophilizing at-30 DEG C, obtains the casein hydrolysate peptides of embodiment 1 preparation.
Enzyme in described step (2) by papain, trypsin, bromelain is in mass ratio
1:1:1 is uniformly mixed and obtains.
Enzyme live keeping agent in described step (2) is by shell six sugar, hydroxyl isomaltulose, chitotriose in mass ratio
It is uniformly mixed for 1:1:1 and obtains.
Embodiment 2
Substantially the same manner as Example 1, differ only in: described enzyme is by papain, Trypsin
Enzyme is uniformly mixed for 1:1 in mass ratio and obtains.Can be prepared by the casein hydrolysate peptides of embodiment 2.
Embodiment 3
Substantially the same manner as Example 1, differ only in: described enzyme is by papain, Fructus Ananadis comosi egg
White enzyme is uniformly mixed for 1:1 in mass ratio and obtains.Can be prepared by the casein hydrolysate peptides of embodiment 3.
Embodiment 4
Substantially the same manner as Example 1, differ only in: described enzyme is by trypsin, bromelain
Enzyme is uniformly mixed for 1:1 in mass ratio and obtains.Can be prepared by the casein hydrolysate peptides of embodiment 4.
Embodiment 5
Substantially the same manner as Example 1, differ only in: described enzyme live keeping agent wheat sugared by shell six, different
Bud ketose alcohol is uniformly mixed for 1:1 in mass ratio and obtains.Can be prepared by the casein water of embodiment 5
Solve peptide.
Embodiment 6
Substantially the same manner as Example 1, differ only in: described enzyme live keeping agent is by shell six sugar, shell three
Sugar is uniformly mixed for 1:1 in mass ratio and obtains.Can be prepared by the casein hydrolysate peptides of embodiment 6.
Embodiment 7
Substantially the same manner as Example 1, differ only in: described enzyme live keeping agent by hydroxyl isomaltulose,
Chitotriose is uniformly mixed for 1:1 in mass ratio and obtains.Can be prepared by the casein hydrolysis of embodiment 7
Peptide.
Test case 1
The enzymolysis solution obtaining embodiment 1-7 step (3) tests its degree of hydrolysis, and concrete outcome is shown in Table 1:
Table 1: casein hydrolysis degree test table
| Degree of hydrolysis/% | |
| Embodiment 1 | 17.60 |
| Embodiment 2 | 8.30 |
| Embodiment 3 | 8.76 |
| Embodiment 4 | 9.89 |
| Embodiment 5 | 8.18 |
| Embodiment 6 | 7.65 |
| Embodiment 7 | 8.12 |
Comparing embodiment 1 and embodiment 2-4, embodiment 1 (papain, trypsin, Fructus Ananadis comosi egg
White enzyme) degree of hydrolysis is apparently higher than embodiment 2-4 (in papain, trypsin, bromelain
Any two kinds compound).Comparing embodiment 1 and embodiment 5-7, embodiment 1 (shell six Fructus Hordei Germinatus ketone sugared, different
Sugar alcohol, chitotriose) degree of hydrolysis is apparently higher than embodiment 2-4 (shell six sugar, hydroxyl isomaltulose, shell three
In sugar, any two kinds compound).
Test case 2
Enzymolysis solution prepared by casein solution embodiment 1-7 step (1) prepared and step (3) enters
Row allergen content is tested, and calculates anaphylactogen reduction rate, and computing formula is: anaphylactogen reduction rate (%)
In=(allergen content in allergen content-enzymolysis solution in casein solution)/casein solution, anaphylactogen contains
Amount × 100%, concrete outcome is shown in Table 2.
Table 2: anaphylactogen reduction rate
| Anaphylactogen reduction rate/% | |
| Embodiment 1 | 99.92 |
| Embodiment 2 | 93.03 |
| Embodiment 3 | 93.32 |
| Embodiment 4 | 94.45 |
| Embodiment 5 | 92.67 |
| Embodiment 6 | 92.83 |
| Embodiment 7 | 93.89 |
Comparing embodiment 1 and embodiment 2-4, embodiment 1 (papain, trypsin, Fructus Ananadis comosi egg
White enzyme) anaphylactogen reduction rate is apparently higher than embodiment 2-4 (papain, trypsin, Fructus Ananadis comosi egg
In white enzyme, any two kinds compound).Comparing embodiment 1 and embodiment 5-7, (shell six is sugared, different for embodiment 1
Maltulose alcohol, chitotriose) anaphylactogen reduction rate is apparently higher than embodiment 2-4 (shell six Fructus Hordei Germinatus sugared, different
In ketose alcohol, chitotriose, any two kinds compound).
Test case 3
Casein hydrolysate peptides embodiment 1-7 prepared, is placed in 25 DEG C, protects under relative humidity 85% environment
Hide half a year, use " GB/T 4789.2-2010 microbiological test of food hygiene total plate count mensuration "
Carry out total plate count test.Concrete test result is shown in Table 3.
Table 3: total plate count test table cfu/g
| Total plate count | |
| Embodiment 1 | 1.6×103 |
| Embodiment 2 | 3.2×103 |
| Embodiment 3 | 4.1×103 |
| Embodiment 4 | 3.5×103 |
| Embodiment 5 | 4.3×103 |
| Embodiment 6 | 4.7×103 |
| Embodiment 7 | 4.5×103 |
Inventor is found surprisingly that, after adding enzyme and enzyme live keeping agent, total plate count is decreased obviously, product
Anti-microbial property significantly improve, the shelf-life is significantly extended.Comparing embodiment 1 and embodiment 2-4,
Embodiment 1 (papain, trypsin, bromelain) anti-microbial property is apparently higher than embodiment
2-4 (in papain, trypsin, bromelain, any two kinds compound).Comparing embodiment 1
With embodiment 5-7, embodiment 1 (shell six sugar, hydroxyl isomaltulose, chitotriose) anti-microbial property is the highest
In embodiment 2-4 (in shell six sugar, hydroxyl isomaltulose, chitotriose, any two kinds compound).
Claims (8)
1. the preparation method of a casein hydrolysate peptides, it is characterised in that comprise the following steps:
(1) casein water dissolution is configured to the casein solution that mass concentration is 5-15wt%, and adjusts
Joint pH to 5-10；
(2) enzyme and enzyme live keeping agent are added in casein solution, hydrolyze 1-5 hour at 45-75 DEG C；
(3) under 85-100 DEG C of water-bath, enzyme denaturing is lived 5-15 minute, obtains enzymolysis solution；
(4) pH value of regulation enzymolysis solution is to 4-5 isoelectric precipitation, centrifugation；
(5) ultrafiltration, lyophilizing.
2. the preparation method of casein hydrolysate peptides as claimed in claim 1, it is characterised in that: described
Enzyme is the mixture of one or more in papain, trypsin, bromelain.
3. the preparation method of casein hydrolysate peptides as claimed in claim 2, it is characterised in that: described
Enzyme is mixed by papain, trypsin, bromelain, described papain, pancreas egg
White enzyme, the mass ratio of bromelain are (1-3): (1-3): (1-3).
4. the preparation method of casein hydrolysate peptides as claimed in claim 1, it is characterised in that: described
Enzyme live keeping agent is the mixture of one or more in shell six sugar, hydroxyl isomaltulose, chitotriose.
5. the preparation method of casein hydrolysate peptides as claimed in claim 4, it is characterised in that: described
Enzyme live keeping agent is mixed by shell six sugar, hydroxyl isomaltulose, chitotriose, described shell six Fructus Hordei Germinatus sugared, different
Ketose alcohol, the mass ratio of chitotriose are (1-3): (1-3): (1-3).
6. the preparation method of the casein hydrolysate peptides as according to any one of claim 1-5, its feature exists
In: described centrifugation rotating speed is 2000-5000 rev/min, and the time is 20-40 minute.
7. the preparation method of the casein hydrolysate peptides as according to any one of claim 1-5, its feature exists
In: described ultrafiltration is with the ultrafilter membrane ultrafiltration of molecular cut off 1-10kD.
8. the preparation method of the casein hydrolysate peptides as according to any one of claim 1-5, its feature exists
In: the 0.5-5% that addition is casein solution quality of enzyme in described step (2), enzyme live keeping agent
Addition is the 0.1-1% of casein solution quality.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108467487A (en) * | 2018-03-30 | 2018-08-31 | 合肥工业大学 | The dextrin modified casein polypeptide conjugate of starch base, preparation method and application |
| CN108486200A (en) * | 2018-03-30 | 2018-09-04 | 合肥工业大学 | Acylation modification casein polypeptide, nanoemulsions, preparation method and application |
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| CN102250998A (en) * | 2011-05-27 | 2011-11-23 | 山东省花生研究所 | Preparation method for peanut bioactive peptide |
| CN104059127A (en) * | 2014-05-29 | 2014-09-24 | 无限极（中国）有限公司 | Natural high-activity antihypertensive peptide, and preparation method and application thereof |
| CN105077261A (en) * | 2015-09-24 | 2015-11-25 | 上海韬鸿化工科技有限公司 | Composite fruit and vegetable ferment and preparing method thereof |
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2016
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Patent Citations (4)
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| CN101748170A (en) * | 2010-01-22 | 2010-06-23 | 西北大学 | Method for preparing 1-deoxy-D-xylulose with chemical method-enzymatic method |
| CN102250998A (en) * | 2011-05-27 | 2011-11-23 | 山东省花生研究所 | Preparation method for peanut bioactive peptide |
| CN104059127A (en) * | 2014-05-29 | 2014-09-24 | 无限极（中国）有限公司 | Natural high-activity antihypertensive peptide, and preparation method and application thereof |
| CN105077261A (en) * | 2015-09-24 | 2015-11-25 | 上海韬鸿化工科技有限公司 | Composite fruit and vegetable ferment and preparing method thereof |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108467487A (en) * | 2018-03-30 | 2018-08-31 | 合肥工业大学 | The dextrin modified casein polypeptide conjugate of starch base, preparation method and application |
| CN108486200A (en) * | 2018-03-30 | 2018-09-04 | 合肥工业大学 | Acylation modification casein polypeptide, nanoemulsions, preparation method and application |
| CN108486200B (en) * | 2018-03-30 | 2021-08-24 | 合肥工业大学 | Acylation modified casein polypeptide, nano emulsion, preparation method and application thereof |
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