CN106755282A - A kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process - Google Patents

A kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process Download PDF

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Publication number
CN106755282A
CN106755282A CN201611155437.8A CN201611155437A CN106755282A CN 106755282 A CN106755282 A CN 106755282A CN 201611155437 A CN201611155437 A CN 201611155437A CN 106755282 A CN106755282 A CN 106755282A
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grass carp
protein
prediction
microwave
hydrolysis
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CN201611155437.8A
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Chinese (zh)
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曹书华
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/34—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving hydrolase
    • C12Q1/37—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving hydrolase involving peptidase or proteinase
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/90—Enzymes; Proenzymes
    • G01N2333/914—Hydrolases (3)
    • G01N2333/948—Hydrolases (3) acting on peptide bonds (3.4)
    • G01N2333/95—Proteinases, i.e. endopeptidases (3.4.21-3.4.99)

Abstract

The invention discloses a kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process, belong to protein deep process technology field.The invention provides a kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process, methods described is that grass carp protein is added in a certain amount of distilled water, and regulation pH adds Alcalase alkali proteases, and protein digestion is carried out using microwave；According to quadratic power model：DH=235.40712+45.27037A+29.57606B+9.59297E 003C 2.58750AB+0.000206AC+0.002484BC 1.85550A2‑1.36387B2‑0.00001255C2, prediction protease grass carp protein is hydrolyzed into specific hydrolysis angle value needed for technological parameter to select suitable technological parameter, or grass carp protein is hydrolyzed reached degree of hydrolysis by prediction enzymatic hydrolysis system under given conditions；Model of the invention, the degree of accuracy is high, highly reliable.

Description

A kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process
Technical field
The present invention relates to a kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process, belong to egg White matter deep process technology field.
Background technology
In the deep-processing process of protein, enzymolysis becomes the process technology of everybody extensive concern, and egg is also turned into gradually One of essential operation in white matter process, correlative study proves small peptide compared to high molecular weight protein and free amino acid More preferably, while also having special physiologically active, such as anti-oxidant, antifatigue and strengthen immunity etc., protein is passed through absorbability After enzymolysis, superior solubility is respectively provided with good dissolubility in the range of pH very wide；Emulsification foamability is improved, Also there is good mobility in higher concentrations；It is busy to hide stable in properties.But conventional research is to obtain big water as far as possible Xie Du, obtains the smaller polypeptide of molecular weight, or even oligopeptides as far as possible, and discovered in recent years, the peptide of different degree of hydrolysis has difference Function and local flavor, and existing enzyme solution can't quickly obtain the peptide of specific degree of hydrolysis, but in hydrolytic process Middle real-time monitoring, monitors required degree of hydrolysis and stops hydrolysis, and this method for hydrolysis can not meet production needs, and hydrolysis process is not It is easy to control, because of different human users, or different batches production to obtain product quality unstable.
The content of the invention
The invention provides a kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process, the party Method is convenient, reliable, can be by calculating the response parameter needed for directly obtaining desired specific hydrolysis angle value.Solve existing enzymolysis skill The technical problem of the polypeptide of specific degree of hydrolysis can not be quickly prepared in art.
A kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process, methods described is grass carp egg White matter is added in a certain amount of distilled water, and regulation pH adds Alcalase alkali proteases, and protease is carried out using microwave Solution；According to quadratic power model：
DH=235.40712+45.27037A+29.57606B+9.59297E-003C-2.58750AB+0.000206AC+ 0.002484BC-1.85550A2-1.36387B2-0.00001255C2, predict that grass carp protein is hydrolyzed into specific water by protease Technological parameter needed for solution angle value is to select suitable technological parameter, or prediction enzymatic hydrolysis system under given conditions by grass carp Protein hydrolyzes reached degree of hydrolysis；The A represents the pH of reaction system,；The B is enzyme concentration, unit %；The C is Microwave power, unit w；The DH is the degree of hydrolysis of protein.
The enzyme activity of the Alcalase alkali proteases is 300,000 U/g.
In one embodiment of the invention, the constant interval of the A is 8-9；The constant interval of B is 2-4%；C's Constant interval is 100-900W, and the constant interval of DH is 0-100%.
In one embodiment of the invention, in the enzyme digestion reaction system course of reaction add sodium hydroxide solution with Maintenance system acid-base value is constant.
In one embodiment of the invention, the naoh concentration is 1mol/L.
In one embodiment of the invention, should be stirred continuously during the enzyme digestion reaction, keep system concentration equal One.
In one embodiment of the invention, pH meter is immersed in hydrolyzate all the time during the enzyme digestion reaction.
In one embodiment of the invention, the enzyme digestion reaction process time is within 30min.
In one embodiment of the invention, the quadratic power model builds with Responds Surface Methodology.
Advantageous Effects
The method that the present invention digests grass carp albumen process by a kind of prediction Alcalase alkali proteases microwave for supplying, leads to Alcalase alkali proteases microwave enzymolysis grass carp albumen is crossed, while being constructed with Responds Surface Methodology principle statistically Quadratic power Optimized model
DH=235.40712+45.27037A+29.57606B+9.59297E-003C-2.58750AB+0.000206AC+ 0.002484BC-1.85550A2-1.36387B2-0.00001255C2, according to the model can by degree of hydrolysis the need for formulate work Skill parameter, model of the invention, the degree of accuracy is high, highly reliable.
Specific embodiment
Embodiment 1：Alcalase alkali proteases microwave digests the center combination design of grass carp Protein Assav
Selection pH (A), Alcalase alkali proteases addition (B), microwave power (C).Each controllable variable is assigned The consumption level code for giving three consumption levels, each variable is：High and low-level " ± 1 " and central horizontal " 0 ", corresponding reality Border consumption level is as shown in table 1：
The controllable variable and consumption level of the Optimized model of table 1
Three controllable variables and consumption level according to table 1, carry out response surface analysis, and table 2 is Alcalase alkali Property the enzymolysis grass carp albumen response surface experiments design of protease microwave.
The enzymolysis grass carp albumen response surface experiments design of table 2Alcalase alkali proteases microwave
Operation sequence A B C Y/%
1 - 1 - 1 0 25.155
2 1 - 1 0 34.455
3 - 1 1 0 28.605
4 1 1 0 32.73
5 - 1 0 - 1 24.81
6 1 0 - 1 30.15
7 - 1 0 1 28.95
8 1 0 1 34.455
9 0 - 1 - 1 25.95
10 0 1 - 1 25.665
11 0 - 1 1 29.73
12 0 1 1 33.42
13 0 0 0 31.35
14 0 0 0 31.695
15 0 0 0 32.04
16 0 0 0 32.85
17 0 0 0 32.385
Embodiment 2：Alcalase alkali proteases microwave digests grass carp protein Process
The consumption level of variable in each experimental group according to table 2, it is a certain amount of that methods described is that grass carp protein is added to Distilled water in, regulation pH add Alcalase alkali proteases, carry out protein digestion using microwave；In addition enzyme digestion reaction mistake Cheng Zhongying is stirred continuously, and keeps system concentration homogeneous；System pH is determined using pH meter, pH meter is submerged all the time during enzyme digestion reaction In hydrolyzate；Sodium hydroxide solution is added in enzyme digestion reaction system course of reaction constant with maintenance system acid-base value, hydroxide Na concn is 1mol/L.Each group experiment must be according to the experiment sequence number Stochastic Expansion shown in table 2, to eliminate possible experimental error.
Embodiment 3：The structure of quadratic power model
Quadratic power model in Response to selection surface Analysis, it is as follows：
DH=K+P1A+P2B+P3C+P4AB+P5AC+P6BC+P7A2+P8B2+P9C2；Wherein：K is constant, and A, B, C are mould Type variable, P1 ... .P9 are model term coefficient.17 groups of experimental datas in table 2 are carried out into regressing calculation to above quadratic power model And variance analysis, as a result as shown in table 3.
The response surface quadratic power model analysis of variance table of table 3
Wherein, * is represented and verified as significantly.
P value in table 3 is to carry out the result that significance test is obtained to model and model term coefficient；If p value is less than 0.05, then it is assumed that model and model terms are significant；Otherwise, not significantly.As shown in Table 3, the p value of quadratic power model is less than 0.0001, therefore the model is effective.In addition, secondary visit model lose intend inspection, with the reliability of assessment models.Such as The p value for intending inspection is really lost higher than 0.05, then shows that the mistake property intended is not notable, it is secondary to visit the preferable of modeling, can enter well Row model prediction；If losing the p value for intending inspection less than 0.05, show to lose the property intended significantly, it is secondary to visit the bad of modeling, need to Model is adjusted.In table 3, model terms A, B, C, AB, BC, B2、C2P value be respectively less than 0.05, therefore be aobvious in model Write item.It is 0.217 that the p value for intending checking is lost in table 3, and model R2=97.79%, R2 Adj=94.4% the explanation model and reality Test fitting good, linear relationship significantly, can be used to predict Alcalase alkali protease microwave enzymes between independent variable and response Solution grass carp protein hydrolysis degree.
Embodiment 4：The forecast function of quadratic power model
Common process：
Grass carp protein is added in a certain amount of distilled water, is stirred, and water-bath to Alcalase alkali proteases is most Thermophilic degree, 1mol/L sodium hydroxide solution maintenance system pH value is used in addition Alcalase alkali protease courses of reaction.Every 10min determines system hydrolysis angle value.
By response surface analysis, Alcalase alkali proteases microwave enzymolysis optimum condition parameter is obtained and has been pH=9, adds Enzyme amount is that 3%, microwave power is 752w, and 30min is digested with this understanding, and actual fish meat protein degree of hydrolysis predicted value is 35.436%, actual fish meat protein degree of hydrolysis is 35.41%.With degree of hydrolysis as 35.41% as standard, carried out with common process Hydrolysis, compares the time needed for different process method protein degree reaches preset value.As shown in table 4
The different process method protein degree of table 4 reach preset value needed for time
Microwave enzymolysis substantially increases the degree of hydrolysis of grass carp albumen, and the conventional method heating-up time is long, system reaction 120min Degree of hydrolysis reaches 32.2 and remains unchanged afterwards.
Above preferred embodiment is merely to illustrate present disclosure, and in addition, the present invention also has other embodiment, As long as those skilled in the art are because of technical inspiration involved in the present invention, and formed using equivalent or equivalent deformation mode Technical scheme is all fallen within protection scope of the present invention.

Claims (8)

1. a kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process, it is characterised in that the side Method is that grass carp protein is added in a certain amount of distilled water, and regulation pH adds Alcalase alkali proteases, is entered using microwave Row protein digestion；According to quadratic power model：
DH=235.40712+45.27037A+29.57606B+9.59297E-003C-2.58750AB+0.000206AC+ 0.002484BC-1.85550A2-1.36387B2-0.00001255C2, predict that grass carp protein is hydrolyzed into specific water by protease Technological parameter needed for solution angle value is to select suitable technological parameter, or prediction enzymatic hydrolysis system system under given conditions will Grass carp protein hydrolyzes reached degree of hydrolysis；The A represents the pH of reaction system；The B is enzyme concentration, unit %；The C It is microwave power, unit w；The DH is the degree of hydrolysis of protein.
2. method according to claim 1, it is characterised in that the constant interval of the A is 8-9, and the constant interval of B is 2- The constant interval of 4%, C is 100-900W, and the constant interval of DH is 0-100%.
3. method according to claim 1, it is characterised in that add hydroxide in the enzyme digestion reaction system course of reaction Sodium solution is constant with maintenance system acid-base value.
4. method according to claim 3, it is characterised in that the naoh concentration is 1mol/L.
5. method according to claim 1, it is characterised in that should be stirred continuously during enzyme digestion reaction.
6. method according to claim 1, it is characterised in that pH meter is immersed in hydrolyzate all the time during enzyme digestion reaction In.
7. method according to claim 1 and 2, it is characterised in that enzyme digestion reaction process time is 0~30min.
8. method according to claim 1, it is characterised in that the quadratic power model is built with Responds Surface Methodology 's.
CN201611155437.8A 2016-12-14 2016-12-14 A kind of method that prediction Alcalase alkali proteases microwave digests grass carp albumen process Pending CN106755282A (en)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20070031953A1 (en) * 2005-04-12 2007-02-08 Dunson James B Jr Treatment of biomass to obtain ethanol
CN102613387A (en) * 2012-03-27 2012-08-01 东北农业大学 Method for preparing soybean peptide by immobilized alkaline protease
CN102796795A (en) * 2012-09-13 2012-11-28 吉林大学 Process method for quickly preparing soybean antioxidant peptide

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20070031953A1 (en) * 2005-04-12 2007-02-08 Dunson James B Jr Treatment of biomass to obtain ethanol
CN102613387A (en) * 2012-03-27 2012-08-01 东北农业大学 Method for preparing soybean peptide by immobilized alkaline protease
CN102796795A (en) * 2012-09-13 2012-11-28 吉林大学 Process method for quickly preparing soybean antioxidant peptide

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
吴谋成: "《油菜籽加工与综合利用》", 30 June 2009, 中国轻工业出版社 *
石岭等: "响应面法优化碱性蛋白酶酶解草鱼蛋白质", 《食品科学》 *

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Application publication date: 20170531