CN1932027A - Double enzyme hydrolysis process for preparing soybean peptide without bitter - Google Patents
Double enzyme hydrolysis process for preparing soybean peptide without bitter Download PDFInfo
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- CN1932027A CN1932027A CN 200610017190 CN200610017190A CN1932027A CN 1932027 A CN1932027 A CN 1932027A CN 200610017190 CN200610017190 CN 200610017190 CN 200610017190 A CN200610017190 A CN 200610017190A CN 1932027 A CN1932027 A CN 1932027A
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Abstract
The double enzyme hydrolysis process for preparing soybean peptide without bitter includes material pre-treatment, enzymolysis, terminating reaction, purifying product and mathematic simulation of the hydrolysis process. Soybean separating protein material is first pretreated at high temperature to destroy the structure of soybean protein and shorten enzymolysis time, then hydrolyzed with two kinds of proteinase of different characteristics to reach proper hydrolysis degree before high temperature treatment to terminating enzyme reaction. Through further pH regulation to precipitate residual protein, centrifugal separation and desalting the supernatant, the soybean peptide product is obtained. The soybean peptide product is ivory powder and has amino acid composition the same as that of soybean protein, balanced amino acid proportion and rich nutrients.
Description
Technical field:
The invention belongs to biological technical field, especially relate to a kind of different enzyme of two specific characters that utilizes and handle soybean protein isolate, hydrolytic process is simulated and predicted, obtain the method for the soybean peptides product of lower molecular weight, no bitter taste by mathematical model.
Background technology:
Soybean peptides is meant that soy-protein is after hydrolytic action, the protein hydrolyzate that obtains through process such as separating, refining again, its multiple peptide mixt after by soybean protein hydrolysis is formed jointly, wherein also contains compositions such as small-amount free amino acid, carbohydrate and inorganic salt.The protein content of soybean peptides is about 85%, and its amino acid is formed almost completely the same with soy-protein, and the balance of indispensable amino acid is good, and content is also abundant.The soybean peptides molecular weight is usually below 10000, and some functional oligopeptide molecular weight are less.Soybean peptides water-soluble very high is even still have flowability under 50% high density; Simultaneously, the soy-protein aqueous solution of about 10% concentration will solidify once heating, but for the aqueous solution of soybean peptides, does not produce solidification phenomenon.From pH and solvability, soy-protein can form precipitation near isoelectric pH 4.3, and soybean peptides is in full pH scope, keep dissolved state, and have stronger water absorbability and moisture retention, arrestin matter form gel, adjust protein food product hardness, improve mouthfeel and characteristic such as easily digest and assimilate.Little peptide in the diet (2～3 amino acid) and big peptide (10～51 amino acid) can intactly pass through enteron aisle, are directly absorbed by enteron aisle, and little peptide absorption rate is faster than the total free aminoacids of same composition, and the protein utilization height; Biologically active peptides is being organized the biological effect that may cause body on the level.Isolated multiple little peptide has the adjusting vegetative nervous system from the enzymic hydrolysate of some food proteins, and the active cells immunologic function is improved cardiovascular function and the anti-ageing physiologically active of waiting for a long time.And the soybean peptides antigenicity is low, can not produce anaphylaxis.These find exploitation peptide class new function protective foods significant.It has been found that in recent years soybean peptides also has many physiological functions, as reducing cholesterol, inhibition elevation of blood pressure, nonallergic, raise immunity and Green Tea Extract damage (delaying senility) etc.
Soy proteinaceous hydrolysis reaction can be undertaken by acid, alkali or enzyme catalysis, and acid, base catalysis technology have bigger destruction because its reaction is violent to nutritive ingredient, and near eliminating, adopting enzyme process catalysis is the main method that present biologically active peptides is produced.The enzymic hydrolysis meeting has greatly improved the character of soybean protein, but makes reasons such as the intramolecular hydrophobic amino acid residue exposure of soybean protein make gained soybean peptides product that comparatively serious bitter taste be arranged owing to hydrolytic action, has limited the application of soybean peptides product.In order to remove the bitter taste of soybean peptides product, people have developed multiple debitterized technique and have comprised absorption, shelter or the like, and independent debitterized technique unavoidably makes complex process, and cost increases, and productive rate is descended.
Summary of the invention:
Technical problem to be solved by this invention provides the method that a kind of raw material is easy to get, production technique simply accurately prepares the soybean peptides of lower molecular weight, no bitter taste efficiently.
The present invention is to be raw material with the soybean protein isolate, adopts the method for double-enzyme hydrolysis to prepare the lower molecular weight soybean peptides, thereby has removed the bitter taste of soybean peptides product preferably, for road has been put down in its application.The present invention has set up the mathematical model about each controlling factor in degree of hydrolysis and the hydrolytic process, can infer the influence of the wherein arbitrary factor of adjusting to hydrolysis reaction according to this model; Also can infer corresponding reaction conditions according to required degree of hydrolysis; Reach required hydrolysis effect by adjusting other adjustings under certain condition confined condition aborning.This model provides possibility for realizing that digitizing is accurately controlled, and production is had directive significance.
The preparation method of soybean peptides of the present invention relates to steps such as raw materials pretreatment, enzymolysis, reaction terminating, purifying products, and hydrolytic process is carried out mathematical simulation.Raw materials pretreatment is about to the soybean protein isolate raw material through pyroprocessing, destroys the structure of soybean protein, shortens enzyme processing time.Adopt the different proteolytic enzyme of two specific characters to be hydrolyzed then, adopt pyroprocessing to stop enzyme reaction to suitable degree of hydrolysis, regulate the pH value and make remaining protein precipitate last centrifugation, the gained supernatant liquor is the soybean peptides product after desalination.It spraying drying is promptly got powder dress soybean peptides product.
This test has drawn the degree of hydrolysis Y of Alcalase hydrolysis by novo soybean protein isolate about concentration of substrate X by homogeneous design and quadratic polynomial stepwise regression analysis
1, enzyme-to-substrate compares X
2, reaction times X
3Mathematical model be:
Y＝-0.004216810156X
1 2-0.012570741235X
2 2
-0.008208720443X
1*X
2+0.00006524879464X
1*X
3
+0.09785141731X
1+0.18829124604X
2-0.0005398321290X
3
-0.584988919。
The method that the present invention prepares soybean peptide without bitter comprises the steps:
A) raw materials pretreatment
Be that 6～10% soybean protein isolate is 70 ℃～90 ℃ preheatings with concentration of substrate, and be aided with at the uniform velocity 200～400r/min stirring, and carry out pre-treatment, make the fats oxidn enzymatic inactivation in the soybean protein isolate, pretreatment time is 10～15min, to reach the effect of partially denaturing and homogeneous;
B) enzymic hydrolysis
Adding reactor 200～400r/min through pretreated soybean protein isolate is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 2～4M and the HCL of 2～4M to enzyme, vigor according to used enzyme, 3%～5% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip 2～4M NaOH in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches and adjusts temperature to 55 ℃ behind the 20%-24% with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 2%～5% of a protein content, keeps the constant of temperature and pH, hydrolysis time 3～5h in the enzymolysis process;
C) reaction terminating
When being reacted to predetermined degree of hydrolysis, promptly the HCL with 2～4M regulates pH to 4.3, stops to stir, and shifts out reactor, is warmed up to 80～95 ℃ rapidly, heating 5～15min, passivation proteolytic enzyme;
D) centrifugation
Centrifugal 20～the 40min of 4000～6000rpm will precipitate with mixed peptide and separate, and collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide;
E) refining
Supernatant liquor after centrifugal is crossed H
+The type cation exchange resin carries out desalting refinement;
F) concentrate
Liquid soybean peptides after the desalination is carried out vacuum concentration, make it volume and be reduced to original 1/4～1/2;
G) drying
Spraying drying promptly gets the soybean peptides product, and the product yield is 58.7%～67.8%.
The gained soybean peptides of the present invention slightly yellow powder that is creamy white, viscosity is bigger, and easily the moisture absorption is fine and smooth, slightly is bitter taste, and light fragrance is arranged.Its amino acid whose composition is almost completely the same with soy-protein, also has indispensable amino acid ratio balance, the characteristics that content is abundant.Eight kinds of indispensable amino acids that contained, except that methionine(Met) is limiting amino acid, other amino acid all above or near the proposed standard of The World Health Organization (WHO), therefore have higher nutritive value.Its quality index is as follows:
(1) Oranoleptic indicator
Taste: light, no bitter taste.
Color and luster: white or little yellow.
Structural state: dried powder, reconstitute a back cup end and do not have precipitation.
(2) physical and chemical index
Crude protein content: 82%～85%
Free aminoacid content: 8%
Moisture content: 6.6%
Ash oontent: 6.2%
Sugar and other content: 4%
Soluble nitrogen index: 99.8%
Tricholroacetic Acid soluble proteins 99.8%
Molecular mass: 5000～20000
The characteristic of soybean peptides product is as follows:
(1) viscosity of soybean peptides solution
The soybean peptides soltion viscosity increases with concentration and changes little.When soy-protein concentration changed between 0%～10%, viscosity changed comparatively mild, was higher than after 10% but work as concentration, and the viscosity straight line rises.And soybean peptides liquid is mobile fabulous, and concentration is also rich mobile at 50% o'clock.
(2) solvability of soybean peptides
Soybean peptides has higher solubleness, and still keeps good dissolved state at all pH, and it is transparent that solution keeps.Under cold condition, still can keep higher solubleness (solubleness of soybean peptides is 14.44g/100g water 0 ℃ the time, and the solubleness of soybean peptides is 25.91g/100g water in the time of 20 ℃, and the solubleness of soybean peptides is 31.09g/100g water in the time of 40 ℃).
(3) the anti-gel characteristic of soybean peptides
In starch, add soybean peptides and can hinder starch gel formation, make the viscosity degradation of starch paste, and amplitude big (add 1% soybean peptides and can make the high viscosity of starch paste descend 26% in concentration is 8% starch paste, MV minium viscosity descends 73%, final viscosity descends 79%).
(4) emulsifying property of soybean peptides
Soybean peptides has certain emulsifying property, and compare its emulsifying property and emulsifying stability increases with concentration with soybean protein isolate.
(5) thermostability of soybean peptides
Soybean peptides is to thermally-stabilised, and 100 ℃ are boiled the nothing precipitation.
Below further specify the present invention by testing example.
Test example 1 soybean peptides anti-oxidant activity
Soybean peptides is obviously active after measured, be clear and definite active ingredient wherein, use sephadex chromatography (SephadexG-50) and ultra-filtration membrane (poly (ether sulfone) film, molecular weight 3000 dams) that soybean peptides is separated in this test again, separating obtained component is called chromatography component 1～9; Soybean peptides is called ultrafiltration and concentration peptide and ultrafiltration filtration peptide through poly (ether-sulfone) ultrafiltration membrane ultrafiltration gained two components.With class superoxide-dismutase (SOD) activity and oxidation inhibiting rate is index, each component anti-oxidant activity such as table 1:
Table 1 soybean peptides oxidation-resistance
Table 1 Antioxidation of soy peptide
| Soybean peptides 2 | Soybean peptides 3 | Chromatography component 1 | Chromatography component 2 | Chromatography component 3 | Chromatography component 4 | Chromatography component 5 | |
| Quasi-SOD enzymic activity (U/mL) oxidation inhibiting rate (%) molecular weight content (%) | 95.02 10.51 ＜22550 | 197.0 2 21.79 ＜11667 | 95.879 10.60 22550 14.1 | 139.71 15.455 13548 7.8 | 164.36 18.18 3517 62.5 | 432.82 47.87 970 6.3 | 180.8 20 620 9.4 |
| Chromatography component 6 | Chromatography component 7 | Chromatography component 8 | Chromatography component 9 | The ultrafiltration and concentration peptide | Ultrafiltration filters peptide | ||
| The quasi-SOD enzymic activity, (U/mL) oxidation inhibiting rate, (%) molecular weight content, (%) | 273.94 30.30 13647 12.8 | 128.75 14.24 8802 76.5 | 224.63 24.84 6237 9.8 | 197.24 21.81 4022 0.98 | 151.61 16.7713 1000- 15244 | 106.21 11.75 ＜1000 |
Wherein the resistance of oxidation of chromatography component 4 is the strongest; Chromatography component 6 is taken second place; Chromatography component 8 and chromatography component 9 are active higher; It should be noted that soybean peptides 3 resistance of oxidation are also stronger, so high activity can be arranged, have using value without separation; Other each components also have activity to detect.
Test example 2 soybean peptides hypolipidemic activities
Soybean peptides has tangible hypolipidemic activity, and from the data of table 2 as seen, wherein the high dosage administration has tangible reduction effect to the rat blood serum triglyceride level.Compare significant difference (P＜0.05=), thereby have statistical significance with control group.
Table 2 soybean peptides is to the influence of rat blood serum
Tabel 2.effect of soy peptide on Wister mice，serum
| Group and dosage | Number of animals (only) | Cholesterol CHO (mmol/L) | Triglyceride level TG (mmol/L) | High-density lipoprotein (HDL) HLD (mmol/L) |
| High dose group low dose group control group | 10 10 10 | 1.044±0.071 1.138±0.060 1.145±0.097 | 0.493±0.056 a 0.627±0.048 ab 0.721±0.066 b | 0.236±0.025 0.225±0.006 0.202±0.014 |
Same row do not have identical lowercase person significant difference (P＜0.05=
Test example 3 soybean peptides anti-fatigue actives
Whether have and animal is set up rapidly improve the effect of endurance for investigating soybean peptides, we have carried out the mouse swimming test, and wherein soybean peptides 1 action effect is obvious, and the data from table 3 as can be seen, adhere to that to soybean peptides group mouse the time of swimming is long, high dose group is longer than low dose group.And the feedwater group time is the shortest, and statistical procedures shows high dose group result significantly (P＜0.05), shows that soybean peptides has the effect that animal is set up and improve endurance.
Table 3 soybean peptides is to the influence of mouse swimming
Tabel 3 Effect of soy peptide on the mice of swimming time
| Group | Number of animals (only) | Swimming time (S) |
| High dose group low dose group 2% Glucose Liquid distilled water | 10 10 10 10 | 639±34 b 511±58 ab 431±49 a 361±31 a |
Same row do not have identical lowercase person significant difference (P＜0.05=)
Test example 4 soybean peptides hypotensive activities
Whether has the effect that reduces hyperpietic's blood pressure for investigating soybean peptides, we have carried out angiotensin-converting enzyme (ACE) and have suppressed activity test, it is active that soybean peptides 2 has stronger ACE inhibition in this problem, be clear and definite active ingredient wherein, with sephadex chromatography soybean peptides 2 is separated again in this problem, 5 kinds of components of gained are called chromatography component 1-5; Suppress percentage with ACE and represent, each component suppresses activity and sees Table 4:
Table 4 soybean peptides hypotensive activity
Table 4 antihypertension of soy peptide
| Soybean peptides 2 | Chromatography component 1 | Chromatography component 2 | Chromatography component 3 | Chromatography component 4 | Chromatography component 5 | |
| Suppress percentage % | 31.4 | 7.9 | 24.7 | 39.7 | 56.9 | 58.3 |
| Content (%) | 14.1 | 7.8 | 62.5 | 6.3 | 9.4 |
Chromatography component 4, chromatography component 5 hypotensive activities are stronger, and chromatography component 1, chromatography component 2, chromatography component 3 are active less, and have and actively reduce and the trend strengthened with molecular weight.We consider that soybean peptides 3 is to get at the basic enterprising one-step hydrolysis of soybean peptides 2 thus, and molecular weight is littler, and its activity may be stronger, but after testing, it suppresses percentage only has 12.5%, half of still not enough soybean peptides 2.Branch factorial ACE belongs to excision enzyme, and the structure of the C of its inhibitor end is bigger to suppressing activity influence, and soybean peptides 3 produce in because of adding excision enzyme, destroyed the structure of the C end of inhibitor, cause active decline.
Test example 5 soybean peptides fermentable
Soybean peptides has the effect that promotes fermentation, has the effect of tangible promotion yeast growth, and is higher than peptone.And can promote the secretion of useful metabolite.Soybean polypeptide can promote the proliferation function of milk-acid bacteria, bifidus bacillus, yeast, mould and other mushrooms.
The soybean peptides group increases by 8.9 times with respect to the blank group, is equal to protein group, is higher than 2.5 times of peptones.
5 pairs of yeast-leavened influences of table 5 soybean peptides
Tabel.5 Effect of soy peptide on the feament of yeast
| Water | Soybean protein | Soybean peptides | Peptone | |
| Yeast number 106/ml yeasts grow multiple | 0.28 0 | 2.5 8.9 | 2.5 8.9 | 1.0 3.57 |
Grow and metabolism because soybean polypeptide has the microorganism growth of promotion, can use it in the food such as producing sour milk, cheese, vinegar, bread, soy sauce, ham.Enhance productivity, improve stability, trophicity and the local flavor of quality; Can also be used for production of enzyme preparation.
Embodiment:
Embodiment 1
Accurately take by weighing concentration of substrate and be 8.38% soybean protein isolate, with soybean protein isolate 75 ℃ of preheatings, and be aided with at the uniform velocity 400r/min and stir, adding reactor 400r/min is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature after carrying out pre-treatment 15min, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 4M and the HCL of 4M to enzyme, vigor according to used enzyme, 4.5% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip the HCL of 4M NaOH and 4M in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches 20% back and adjusts temperature to 55 ℃ with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 4% of a protein content, maintenance temperature and pH's is constant in the enzymolysis process, hydrolysis time 4h, hydrolysis finishes the back and transfers pH to 4.3 precipitation enzyme and remaining protein, 90 ℃ of heating 15min make the thorough passivation of enzyme, the centrifugal 40min of 4000rpm will precipitate with mixed peptide and separate, collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide, it desalination and concentration to volume is reduced to 1/2 original back spraying drying, promptly get the soybean peptides product, productive rate is 64.6%.
Embodiment 2
Accurately take by weighing concentration of substrate and be 6% soybean protein isolate, with soybean protein isolate 90 ℃ of preheatings, and be aided with at the uniform velocity 200r/min and stir, adding reactor 200r/min is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature after carrying out pre-treatment 10min, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 2M and the HCL of 2M to enzyme, vigor according to used enzyme, 3% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip the NaOH of 2M in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches 24% back and adjusts temperature to 55 ℃ with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 5% of a protein content, maintenance temperature and pH's is constant in the enzymolysis process, hydrolysis time 3h, hydrolysis finishes the back and transfers pH to 4.3 precipitation enzyme and remaining protein, 80 ℃ of heating 10min make the thorough passivation of enzyme, the centrifugal 40min of 5000rpm will precipitate with mixed peptide and separate, collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide, it desalination and concentration to volume is reduced to 1/4 original back spraying drying, promptly get the soybean peptides product, productive rate is 67.8%.
Embodiment 3
Accurately take by weighing concentration of substrate and be 8% soybean protein isolate, with soybean protein isolate 85 ℃ of preheatings, and be aided with at the uniform velocity 300r/min and stir, adding reactor 300r/min is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature after carrying out pre-treatment 10min, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 4M and the HCL of 4M to enzyme, vigor according to used enzyme, 4% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip the NaOH of 4M and the HCL of 4M in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches 22% back and adjusts temperature to 55 ℃ with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 2.5% of a protein content, maintenance temperature and pH's is constant in the enzymolysis process, hydrolysis time 5h, hydrolysis finishes the back and transfers pH to 4.3 precipitation enzyme and remaining protein, 95 ℃ of heating 15min make the thorough passivation of enzyme, the centrifugal 30min of 4000rpm will precipitate with mixed peptide and separate, collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide, it desalination and concentration to volume is reduced to 1/2 original back spraying drying, promptly get the soybean peptides product, productive rate is 62.7%.
Embodiment 4
Accurately take by weighing concentration of substrate and be 10% soybean protein isolate, with soybean protein isolate 80 ℃ of preheatings, and be aided with at the uniform velocity 200r/min and stir, adding reactor 400r/min is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature after carrying out pre-treatment 15min, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 4M and the HCL of 4M to enzyme, vigor according to used enzyme, 5% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip the NaOH of 4M in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches 20% back and adjusts temperature to 55 ℃ with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 2% of a protein content, maintenance temperature and pH's is constant in the enzymolysis process, hydrolysis time 5h, hydrolysis finishes the back and transfers pH to 4.3 precipitation enzyme and remaining protein, 95 ℃ of heating 5min make the thorough passivation of enzyme, the centrifugal 20min of 6000rpm will precipitate with mixed peptide and separate, collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide, it desalination and concentration to volume is reduced to 1/2 original back spraying drying, promptly get the soybean peptides product, productive rate is 58.7%.
Embodiment 5
Accurately take by weighing concentration of substrate and be 7% soybean protein isolate, with soybean protein isolate 70 ℃ of preheatings, and be aided with at the uniform velocity 300r/min and stir, adding reactor 300r/min is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature after carrying out pre-treatment 10min, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 3M and the HCL of 3M to enzyme, vigor according to used enzyme, 3.5% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip the NaOH of 3M and the HCL of 3M in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches 20% back and adjusts temperature to 55 ℃ with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 3% of a protein content, maintenance temperature and pH's is constant in the enzymolysis process, hydrolysis time 4h, hydrolysis finishes the back and transfers pH to 4.3 precipitation enzyme and remaining protein, 90 ℃ of heating 15min make the thorough passivation of enzyme, the centrifugal 30min of 4000rpm will precipitate with mixed peptide and separate, collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide, it desalination and concentration to volume is reduced to 1/4 original back spraying drying, promptly get the soybean peptides product, productive rate is 60.2%.
Claims (1)
1, a kind of double-enzyme hydrolysis prepares the method for soybean peptide without bitter, comprises the following steps:
A) raw materials pretreatment
Be that 6～10% soybean protein isolate is 70 ℃～90 ℃ preheatings with concentration of substrate, and be aided with at the uniform velocity 200～400r/min stirring, and carry out pre-treatment, make the fats oxidn enzymatic inactivation in the soybean protein isolate, pretreatment time is 10～15min, to reach the effect of partially denaturing and homogeneous;
B) enzymic hydrolysis
Adding reactor 200～400r/min through pretreated soybean protein isolate is stirred to homogeneous and is cooled to 60 ℃ of enzyme digestion reaction temperature, regulate the optimal pH 8.0 of potential of hydrogen with the NaOH of 2～4M and the HCL of 2～4M to enzyme, vigor according to used enzyme, 3%～5% the Alcalase Sumizyme MP of accurately measuring protein content adds in the hydrolysis reactor, drip 2～4M NaOH in the reaction process, make pH be stabilized in the optimal pH 8.0 of enzyme, and make temperature remain at 60 ℃ of the optimum temperutures of enzymolysis, the alkali lye amount that day part consumes in the monitoring hydrolytic process, calculate and the monitoring degree of hydrolysis reaches and adjusts temperature to 55 ℃ behind the 20%-24% with the pH-stat method, pH value to 7.5, add the Flavourzyme compound protease, consumption is 2%～5% of a protein content, keeps the constant of temperature and pH, hydrolysis time 3～5h in the enzymolysis process;
C) reaction terminating
When being reacted to predetermined degree of hydrolysis, promptly the HCL with 2～4M regulates pH to 4.3, stops to stir, and shifts out reactor, is warmed up to 80～95 ℃ rapidly, heating 5～15min, passivation proteolytic enzyme;
D) centrifugation
Centrifugal 20～the 40min of 4000～6000rpm will precipitate with mixed peptide and separate, and collect supernatant liquor, be double-enzyme hydrolysis mixing soya beans peptide;
E) refining
Supernatant liquor after centrifugal is crossed H
+The type cation exchange resin carries out desalting refinement;
F) concentrate
Liquid soybean peptides after the desalination is carried out vacuum concentration, make it volume and be reduced to original 1/4～1/2;
G) drying
Spraying drying promptly gets the soybean peptides product, and the product yield is 58.7%～67.8%.
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