CN1840681A - Process for producing functional soybean peptide by two-step method - Google Patents
Process for producing functional soybean peptide by two-step method Download PDFInfo
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- CN1840681A CN1840681A CNA2006100458091A CN200610045809A CN1840681A CN 1840681 A CN1840681 A CN 1840681A CN A2006100458091 A CNA2006100458091 A CN A2006100458091A CN 200610045809 A CN200610045809 A CN 200610045809A CN 1840681 A CN1840681 A CN 1840681A
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Abstract
The preparation method for a functional soybean peptide comprises: with bean pulp degreased at low temperature as material, composite hydrolyzing soyabean protein with proteinase and microbe while adding bacillus subtilis. This invention needs low cost, improves product physical and biological performances, and is benefit to health care.
Description
Technical field
The invention belongs to the soybean deep processing technology field, the technology of the definite preparation functional soybean peptide of saying so.
Background technology
Soybean peptides is a kind of deep processed product that soy-protein forms through protease hydrolysis.Soybean polypeptide is compared with traditional soy-protein, have good water solubility, molecular weight is little, solution viscosity is little, be heated and do not solidify, provide rapidly human body energy, no protein denaturation, free from beany flavor, no residue, do not produce characteristic such as precipitation under acidic conditions, and have easily digest and assimilate, low antigenicity, reducing cholesterol and significant advantage such as blood pressure, adjusting human physiological functions.Therefore, soybean peptides is a kind of active polypeptide of excellent property, as functional factor in the foodstuffs industry and high-quality protein, has very wide application prospect, and become one of focus of domestic and international soy-protein research, in fields such as food, medical science, daily-use chemical industry, demonstrated tempting DEVELOPMENT PROSPECT.Mainly contain two factors at present and restricted the industrialized development of peptide based food: the one, cost is high, and the 2nd, the mouthfeel incompatibility that bitter taste causes.
Nattokinase (Nattokinase) is the Taka-proteinase that directly decomposes thrombus.Also have the osteoporosis of preventing, antibiotic, sterilization, preventing hypertension, anticancer, suppress hyperglycemia, prevent effects such as brain aging and whole intestines.Nattokinase is a kind of serineprotein kinase with strong thrombus-dissolving effect of being found from tempeh natto (Natto) at first by Japanese scholar.Because Nattokinase derives from food, has no side effect, so very likely be developed to the liquid enzymes oral preparation with clear and definite curative effect or contain the enzyme healthy beverage.Bacillus subtilus (Bacillussubtilis (Ehrenberg) Cohn) is the main production bacterial classification of japanese traditional food natto, contain a large amount of functional ingredient---Nattokinase in the tunning, but Nattokinase has the stink of natto uniqueness, directly be used in food, on mouthfeel, have certain problem.Therefore how removing or alleviate the peculiar smell that produces in the fermenting bacillus subtilis process, and it is applied in during soybean polypeptide produces, make the functional soy peptide product that contains Nattokinase, is the problem that needs solve.
Summary of the invention
The purpose of this invention is to provide a kind of raw material and be easy to get, production technique is simple, and mouthfeel is better, contains the preparation method of the functional soy peptide of Nattokinase.
In order to achieve the above object, the technical solution used in the present invention is:
1. extraction soybean protein isolate
Raw material: defatted soybean meal
(1) alkali dissolution
Defatted soybean meal is pulverized the back and is crossed 40～60 mesh sieves, press material-water ratio 1: 8～15 water are added batch extractor, with 10～30%NaOH solution material liquid pH is regulated 7.0～9.0, at the uniform velocity stir 40～60min under 40～60 ℃ of degree, collect protein extract after the centrifugation, residue utilizes the lixiviate of above-mentioned condition secondary, and is standby after twice protein extract merged.
(2) sour precipitating
Add 10～30% hydrochloric acid soln in the soybean protein extracting solution, regulator solution pH value is 4.2～4.5, treat most of protein precipitation after, centrifugation obtains purity and is the soybean protein isolate more than 90% after the water washing and precipitating.
2. protease hydrolysis
(1) soybean protein isolate is mixed with 3～5% protein soln, under 80～90 ℃ of conditions, handles 10～15min.
(2) regulating protein soln pH value is 7.0～9.0, add the proteinic proteolytic enzyme of 1000～1500u/g (as microbial proteases such as Bacillus subtilus 1389, actinomycetes 166, aspergillus terricola 3942, aspergillus niger 3350 and bacillus licheniformis 2709), hydrolysis 3～5h under 40～55 ℃ of conditions, reaction finishes the back and heat 8～15min enzyme that goes out in 80～100 ℃ of water-bath.
3. microbial fermentation
Add 1.0～2.5% sucrose in hydrolysate of soybean protein liquid, its pH value is adjusted to 6.5～7.5,20～30min sterilizes under 121 ℃ of conditions.Sterilization, cooling, insert 4～6% the subtilis that can produce Nattokinase (as subtilis 1.107, subtilis B.S.Hezilll), in culture temperature is that 25～35 ℃, fermentor tank mixing speed are that 80～120rpm, air flow are condition bottom fermentation 32～40h of 4～6L/min, centrifugation 5～20min removes thalline under 5000～10000rpm, obtains fermented supernatant fluid.
4. refining
(1) debitterize, decolouring
Add 10～30g/L gac (gac/protein is w/v) in fermented supernatant fluid, regulating the pH value is 7.0～8.0, reacts 1～3h under 25～35 ℃ of conditions.The solid matters such as gac in the raw material are removed in centrifugation.
(2) desalination
Utilize acidic cation-exchange resin and basic anion exchange resin to carry out desalination, the temperature of soybean peptides is 30～40 ℃ during desalination, and flow velocity is 12～20m
3Protein liquid/(m
3Resin h).
5. canned
Adopt aseptic condition to carry out canned.
It is raw material that the present invention adopts defatted soybean meal, utilize proteolytic enzyme and microbial fermentation composite hydrolysis soybean protein, and add Bacillus subtilus in process of production, a kind of preparation technology who contains the functional polypeptide product of Nattokinase is provided, not only can reduce production costs, technology is simple, the bitter taste of product obviously reduces, and products obtained therefrom has not only kept original easily the digesting and assimilating of soybean peptides, low antigenicity, reducing cholesterol and blood pressure, regulate significant advantages such as human physiological functions, also possessed Nattokinase simultaneously and prevented osteoporosis, antibiotic, sterilization, preventing hypertension, anticancer, suppress hyperglycemia, prevent the effect of brain aging and whole intestines, at food, medical science, fields such as daily-use chemical industry have DEVELOPMENT PROSPECT.
Embodiment
Embodiment 1
1. extraction soybean protein isolate
Raw material: defatted soybean meal
(1) alkali dissolution
Defatted soybean meal is pulverized the back and is crossed 60 mesh sieves, press material-water ratio 1: 10 water is added batch extractor, with 15%NaOH solution material liquid pH is regulated 7.2, at the uniform velocity stir 50min under 50 ℃ of degree, collect protein extract after the centrifugation, residue utilizes the lixiviate of above-mentioned condition secondary, and is standby after twice protein extract merged.
(2) sour precipitating
Add 20% hydrochloric acid soln in the soybean protein extracting solution, regulator solution pH value is 4.3, treat most of protein precipitation after, centrifugation obtains purity and is the soybean protein isolate more than 90% after the water washing and precipitating.
2. protease hydrolysis
(1) soybean protein isolate is mixed with 4% protein soln, at 90 ℃ of thermal pretreatment 10min.
(2) regulating protein soln pH value is 8.5, adds proteinic Bacillus subtilus 1389 proteolytic enzyme of 1200u/g, hydrolysis 4h under 50 ℃ of conditions, and reaction finishes the back and heat the 10min enzyme that goes out in boiling water bath.
3. microbial fermentation
In hydrolysate of soybean protein liquid, add 2% sucrose, its pH value is adjusted to 7.0, sterilization, cooling, insert 5% subtilis 1.107, in culture temperature is that 30 ℃, fermentor tank mixing speed are that 100rpm, air flow are the condition bottom fermentation 36h of 5L/min, and centrifugation 10min removes thalline under the 8000rpm, obtains fermented supernatant fluid.
4. refining
(1) debitterize, decolouring
Add 20g/L gac (gac/protein is w/v) in fermented supernatant fluid, regulating the pH value is 7.5, reacts 2h under 30 ℃ of conditions.The solid matters such as gac in the raw material are removed in centrifugation.
(2) desalination
Utilize storng-acid cation exchange resin 732 and strongly basic anion exchange resin 717 to carry out desalination, the temperature of soybean peptides is 40 ℃ during desalination, and flow velocity is 20m
3Protein liquid/(m
3Resin h).
5. canned
Adopt aseptic condition to carry out canned.
6. products obtained therefrom detect parameters
Degree of hydrolysis is (%): 32.0
Molecular weight (Da): to be lower than 1000
NSI value (%): 〉=95
Nattokinase (urokinase unit/mL fermented liquid): 〉=200
Embodiment 2
1. extraction soybean protein isolate
Raw material: defatted soybean meal
(1) alkali dissolution
Defatted soybean meal is pulverized the back and is crossed 60 mesh sieves, press material-water ratio 1: 10 water is added batch extractor, with 15%NaOH solution material liquid pH is regulated 7.2, at the uniform velocity stir 50min under 50 ℃ of degree, collect protein extract after the centrifugation, residue utilizes the lixiviate of above-mentioned condition secondary, and is standby after twice protein extract merged.
(2) sour precipitating
Add 20% hydrochloric acid soln in the soybean protein extracting solution, regulator solution pH value is 4.3, treat most of protein precipitation after, centrifugation obtains purity and is the soybean protein isolate more than 90% after the water washing and precipitating.
2. protease hydrolysis
(1) soybean protein isolate is mixed with 5% protein soln, at 80 ℃ of thermal pretreatment 15min.
(2) regulating protein soln pH value is 7.0, adds proteinic Bacillus subtilus 1389 proteolytic enzyme of 1500u/g, hydrolysis 3.5h under 40 ℃ of conditions, and reaction finishes the back and heat the 10min enzyme that goes out in boiling water bath.
3. microbial fermentation
In hydrolysate of soybean protein liquid, add 2.5% sucrose, its pH value is adjusted to 7.5, sterilization, cooling, insert 4% subtilis 1.107, in culture temperature is that 35 ℃, fermentor tank mixing speed are that 80rpm, air flow are the condition bottom fermentation 40h of 6L/min, and centrifugation 5min removes thalline under the 10000rpm, obtains fermented supernatant fluid.
4. refining
(1) debitterize, decolouring
Add 10g/L gac (gac/protein is w/v) in fermented supernatant fluid, regulating the pH value is 7.0, reacts 3h under 25 ℃ of conditions.The solid matters such as gac in the raw material are removed in centrifugation.
(2) desalination
Utilize storng-acid cation exchange resin 732 and strongly basic anion exchange resin 717 to carry out desalination, the temperature of soybean peptides is 30 ℃ during desalination, and flow velocity is 12m
3Protein liquid/(m
3Resin h).
5. canned
Adopt aseptic condition to carry out canned.
6. products obtained therefrom detect parameters
Degree of hydrolysis is (%): 30.2
Molecular weight (Da): to be lower than 1000
NSI value (%): 〉=95
Nattokinase (urokinase unit/mL fermented liquid): 〉=200
Embodiment 3
1. extraction soybean protein isolate
Defatted soybean meal
(1) alkali dissolution
Defatted soybean meal is pulverized the back and is crossed 60 mesh sieves, press material-water ratio 1: 10 water is added batch extractor, with 15%NaOH solution material liquid pH is regulated 7.2, at the uniform velocity stir 50min under 50 ℃ of degree, collect protein extract after the centrifugation, residue utilizes the lixiviate of above-mentioned condition secondary, and is standby after twice protein extract merged.
(2) sour precipitating
Add 20% hydrochloric acid soln in the soybean protein extracting solution, regulator solution pH value is 4.3, treat most of protein precipitation after, centrifugation obtains purity and is the soybean protein isolate more than 90% after the water washing and precipitating.
2. protease hydrolysis
(1) soybean protein isolate is mixed with 3% protein soln, at 90 ℃ of thermal pretreatment 10min.
(2) regulating protein soln pH value is 9.0, adds proteinic Bacillus subtilus 1389 proteolytic enzyme of 1000u/g, hydrolysis 5h under 55 ℃ of conditions, and reaction finishes the back and heat the 8min enzyme that goes out in boiling water bath.
3. microbial fermentation
In hydrolysate of soybean protein liquid, add 1% sucrose, its pH value is adjusted to 6.5, sterilization, cooling, insert 6% subtilis 1.107, in culture temperature is that 25 ℃, fermentor tank mixing speed are that 120rpm, air flow are the condition bottom fermentation 32h of 4L/min, and centrifugation 20min removes thalline under the 5000rpm, obtains fermented supernatant fluid.
4. refining
(1) debitterize, decolouring
Add 30g/L gac (gac/protein is w/v) in fermented supernatant fluid, regulating the pH value is 8.0, reacts 1h under 35 ℃ of conditions.The solid matters such as gac in the raw material are removed in centrifugation.
(2) desalination
Utilize storng-acid cation exchange resin 732 and strongly basic anion exchange resin 717 to carry out desalination, the temperature of soybean peptides is 35 ℃ during desalination, and flow velocity is 15m
3Protein liquid/(m
3Resin h).
5. canned
Adopt aseptic condition to carry out canned.
6. products obtained therefrom detect parameters
Degree of hydrolysis is (%): 28.7
Molecular weight (Da): to be lower than 1000
NSI value (%): 〉=95
Nattokinase (urokinase unit/mL fermented liquid): 〉=200.
Claims (7)
1, a kind of technology of producing functional soybean peptide by two-step method is characterized in that: is raw material with the defatted soybean meal, extracts soybean protein isolate, adopt proteolytic enzyme and microbial fermentation composite hydrolysis, make the functional soybean peptide stoste that contains Nattokinase, wherein:
A. protease hydrolysis operation:
(1) will separate the soybean protein isolate that obtains and be mixed with 3～5% protein soln, under 80～90 ℃ of conditions, handle 10～15 minutes;
(2) regulating protein soln pH value is 7.0～9.0, adds the proteinic proteolytic enzyme of 1000～1500u/g, and hydrolysis is 3～5 hours under 40～55 ℃ of conditions, heats 8～15min enzyme that goes out after reaction finishes in 80～100 ℃ of water-baths;
B. microbial fermentation operation:
In the hydrolysate of soybean protein liquid that previous step obtains, add 1.0～2.5% sucrose, its pH value is adjusted to 6.5～7.5, sterilization, cooling, insert 4～6% subtilis, in culture temperature is that 25～35 ℃, fermentor tank mixing speed are that 80～120rpm, air flow are condition bottom fermentation 32～40h of 4～6L/min, and centrifugation 5～20min removes thalline under 5000～10000rpm condition, obtains fermented supernatant fluid.
2, the technology of a kind of producing functional soybean peptide by two-step method as claimed in claim 1 is characterized in that the described soybean protein isolate that extracts from defatted soybean meal, adopt following steps:
(1) alkali dissolution
Defatted soybean meal is pulverized the back and is crossed 40～60 mesh sieves, press material-water ratio 1: 8～15 water are added batch extractor, with 10～30%NaOH solution material liquid pH is regulated 7.0～9.0, at the uniform velocity stir 40～60min under 40～60 ℃ of degree, collect protein extract after the centrifugation, residue utilizes the lixiviate of above-mentioned condition secondary, and is standby after twice protein extract merged;
(2) sour precipitating
Add 10～30% hydrochloric acid soln in above-mentioned soybean protein extracting solution, regulator solution pH value is 4.2～4.5, treat most of protein precipitation after, centrifugation obtains purity and is the soybean protein isolate more than 90% after the water washing and precipitating.
3, the technology of a kind of producing functional soybean peptide by two-step method as claimed in claim 1 or 2 is characterized in that: the fermented supernatant fluid that described microbial fermentation operation obtains, proceed debitterize, decolouring processing;
Add 10～30g/L gac (gac/protein is w/v) in fermented supernatant fluid, regulating the pH value is 7.0～8.0, reacts 1～3h under 25～35 ℃ of conditions.The solid matters such as gac in the raw material are removed in centrifugation.
4, the technology of a kind of producing functional soybean peptide by two-step method as claimed in claim 3, it is characterized in that: described fermented supernatant fluid, utilize acidic cation-exchange resin and basic anion exchange resin to carry out desalination, the temperature of soybean peptides is 30～40 ℃ during desalination, and flow velocity is 12～20m
3Protein liquid/(m
3Resin h).
5, the technology of a kind of producing functional soybean peptide by two-step method as claimed in claim 1 is characterized in that: described proteolytic enzyme can be Bacillus subtilus 1389, actinomycetes 166, aspergillus terricola 3942, aspergillus niger 3350 and bacillus licheniformis 270%
6, the technology of a kind of producing functional soybean peptide by two-step method as claimed in claim 1, it is characterized in that: described microorganism is to be used for natto production, and the subtilis that can produce Nattokinase comprises subtilis 1.107, subtilis B.S.Hezilll.
7, the technology of a kind of producing functional soybean peptide by two-step method as claimed in claim 1 is characterized in that: the centrifugal rotational speed in described (b) step is that 5000～10000rpm, disengaging time are 5～20min.
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