CN107227267A - A kind of soybean small peptide and preparation method and application - Google Patents
A kind of soybean small peptide and preparation method and application Download PDFInfo
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- CN107227267A CN107227267A CN201710334372.1A CN201710334372A CN107227267A CN 107227267 A CN107227267 A CN 107227267A CN 201710334372 A CN201710334372 A CN 201710334372A CN 107227267 A CN107227267 A CN 107227267A
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Abstract
The invention discloses bacillus cereus (Bacillus cereus) bacterial strain that one plant is used for decomposition of protein, China typical culture collection center is preserved in, preserving number is CGMCC NO:14049.Public affairs of the invention have also opened the soybean small peptide of the method that soybean small peptide is prepared using the bacterial strain and gained, and the invention also discloses a kind of carbonated soda water healthy beverage.The bacillus cereus protein breakdown ability of the present invention is strong, available for the soybean small peptide and its Related product for developing edible high economic value added, with significant economic value.
Description
Technical field
The present invention relates to biological technical field, in particular it relates to which one plant is used for the bacillus cereus of decomposition of protein
(Bacillu scereus) bacterial strain, and the method for soybean small peptide and the soybean small peptide of gained are prepared using the bacterial strain, the present invention
Further relate to a kind of carbonated soda water healthy beverage.
Background technology
Soybean is most important in the world and maximum yield beans, is used as the most abundant most cheap protein that the mankind are edible
Source, its is amino acid contained relatively comprehensively, especially based on lysine, the deficiency of lysine in just complementary grain protein.It is sub-
People from continent eating soybean fermentive food, including the mold fermentation product such as fermented soya bean, fermented bean drink, beans sauce and various fermented bean curd, have not only evaded tired
The protein allergies symptom for disturbing American-European ethnic group can also be by eliminating its factor for suppressing nutrition, while producing a variety of after microbial action
Flavor peptide with permanent fragrant, thus be more easy to be received and digested and assimilated by human body.
Along with world scholar in recent years to the research and development of functional food and the progress of biotechnology, Soybean Peptide system
Product particularly soybean small peptide functional food receives the favor of global food industry.At present, soybean small peptide is soybean protein through water
Soybean small peptide below the mixture for the small-molecular peptides that solution is obtained, pentapeptide may pass through mucous membrane of small intestine and be directly absorbed by the body, its
In have effects that physiologically active small-molecular peptides are not available for former soybean protein and its amino acid constituted.Soybean is small
The various healthcare functions such as lowering blood pressure and blood fat of peptide functional food, promote fat metabolism, it is anti-oxidant, antifatigue, promote element absorption and
The functions such as low irritability are also progressively found and are widely used, so product great industrial development of the exploitation rich in soybean small peptide
Prospect.
In addition, modern life mode stresses quick, efficient and practicality, the demand pleased for instant and sense organ is opened
Send out and fed substantial amounts of polished rice smart face and for example high sugar carbonic acid soda of other convenient food etc., the single diet of these nutrition fills
Denounce salinity, sugar, energy and acid, long-term consumption and nutrient can be caused excessively to take in and internal osmotic pressure when amount of drinking water is supplied
It is unbalance.Under this unsound social diet agitation, the alkaline healthy beverage of the simple low sugar low nutrition element rich in fresh water
Demand increases severely with day.
The present invention evades the macroscopic design thinking set of the high concentration soy-bean peptide rich in nutrient in the past, is absorbed in out
The low sugar alkaline carbonic acid beverage containing a small amount of soybean small peptide of clean taste is sent out, the need for having suited market, special people has been looked after
The demand of group, is directed to process optimization and the healthy beverage of sense organ pleasure is made us in exploitation, meets the taste of consuming public and many
Fang Yaoqiu.
The content of the invention
In order to solve the above problems, above-mentioned target is realized, the present invention provides following technical scheme.
One of technical scheme is to provide one plant of bacillus cereus (Bacillus cereus) bacterial strain, and it is protected
China typical culture collection center is hidden in, preserving number is CCTCC NO：14049.
Another technical scheme of the present invention is to provide a kind of microbial inoculum, and its active component is above-mentioned bacillus cereus bacterium
Strain.
Another technical scheme of the present invention is to provide above-mentioned Bacillus cereus strain in decomposing soya-bean protein or system
Application in standby soybean small peptide.
Another technical scheme of the present invention is to provide a kind of method for preparing soybean small peptide, and it comprises the following steps：
(a) dregs of beans is weighed, is sterilized, bacillus cereus is inoculated with, and ferment, it is preferable that fermentation time is 15 days；
(b) fermented bean dregs obtained by step (a) add protease and digested；
(c) enzymolysis product obtained by filtration step (b) simultaneously stands purification, obtains de- bitter deodorization and the soybean egg of color is removed in part
White clear liquid, it is preferable that the filtering is carried out with 400 mesh filter clothes, it is preferable that the standing purification activated carbon and florisil silica
Carry out, it is preferable that activated carbon and the florisil silica consumption is 5 ‰ and is 10 ‰ mass fractions, g/kg；
(d) soybean protein clear liquid obtained by step (c) is subjected to below 10000Da ultrafiltration membrane treatments, then carry out 1000Da with
Lower NF membrane and below 500Da nanofiltration membrane treatments, collect the soybean small peptide filtered out and contain water mixed liquid, continue the soybean filtered out to this
Small peptide carries out water removal nanofiltration membrane treatment containing water mixed liquid, obtains soybean small peptide.
Further, the above-mentioned method for preparing soybean small peptide also includes：
(e) soybean small peptide obtained by step (d) is concentrated, then be freeze-dried, it is preferable that after concentration, added
Electrolysis water carries out antibacterial, it is preferable that electrolysis water consumption is 5 ‰.
Preferably, above-mentioned bacillus cereus is the Bacillus cereus strain of the present invention.
Preferably, above-mentioned protease is thermolysin, it is preferable that the thermophilic bacteria protein enzyme dosage is 5 ‰ mass
Number (g/kg).
Another other technical scheme of the present invention is to provide a kind of soybean small peptide, and it is as the method described in any of the above item
It is prepared.
Another technical scheme of the present invention is to provide a kind of beverage, and by weight, it includes following composition：
1-5 parts of soybean small peptide, it is preferable that the soybean small peptide is above-mentioned soybean small peptide；
10-12 parts of citric acid, it is preferable that the citric acid is Citric Acid Mono；
60-70 parts of xylitol；
7.5-8 parts of sodium bicarbonate；
1500 parts of water.
In one embodiment, the following composition of the beverage：
3 parts of Soybean Peptide；
10 parts of citric acid；
60 parts of xylitol；
7.5 parts of sodium bicarbonate,
1500 parts of water.
Typically, bacillus cereus is common pathogenic bacteria, in distributed in nature extensively, the inspection in various food
Extracting rate is also higher, and often storage temperature is improper before consumption for food, and standing time is long, makes waxy bud of the pollution in food
The gemma of spore bacillus or remaining is able to growth and breeding, or due to the heat-stable toxin produced containing bacillus cereus, and lead
Cause the generation of food poisoning.The incidence of disease of poisoning is higher, and generally 60%~100%.Thus, rarely have bacillus cereus to be used for
The case of food production.
However, bacillus cereus can produce bacterialprotease, the ability with effective degraded or decomposing soya-bean protein, this
Invention is based on this, and having separated one plant has the new bacillus cereus of stronger protein breakdown ability, and develops one kind
Produce the new technology of soybean small peptide.
The beneficial effects of the invention are as follows：
(1) the high activity bacillus cereus that separates of the present invention has a strong protein breakdown ability, and enzyme activity=
184.79U/mL, can be the less various small peptides of molecular weight the breaks down proteins in dregs of beans.
(2) present invention develops a kind of fermented bean dregs and extracted on the basis of high activity bacillus cereus is isolated to
The new technology of small-molecular peptides, by methods such as fermentation, hydrolysis, decolouring and deodorizing, separation, extractions, soybean oil is refined using industry
The squeezing extraction leaching dregs of beans of accessory substance two develops the soybean small peptide and its Related product of edible high economic value added, and accessory substance is also
It can continue to use as feed, substantially increase the comprehensive utilization value of soybean protein.
(3) present invention has also outputed one kind containing the small molecular weight titanium extracted in the soybean protein for soaking dregs of beans from fermentation two
Low pained, low bad smell sugar-free carbonated soda water healthy beverage, rapid on the premise of clean mouthfeel is kept can supplement human body
The moisture of missing.Can also take into account mouthfeel, the sense of taste, alkaline and micro nutrition on the basis of the intake of sugared part is not increased, without point
Powder and stabilizer, are a kind of healthy drinks.
Brief description of the drawings
With reference to accompanying drawing and to the detailed description of the specific embodiment of the invention, those skilled in the art will become more apparent that this
Above-mentioned and other purposes, the advantages and features of invention.
Fig. 1 is the visual results after the Bacillus cereus strain culture 3 days of the present invention, i.e. flat-plate bacterial colony form.
Fig. 2 is the microscopy results after the Bacillus cereus strain culture 3 days of the present invention, i.e., after Gram's staining
Form.
Fig. 3 is the liquid chromatogram of the soybean small peptide mix products for the different molecular weight that different pore size milipore filter is screened
Figure.
Embodiment
Raw materials used and equipment is that those skilled in the art know in embodiment, and be in the market can buy or
It is readily available or is made.
Separation, sign, culture and the preservation of the bacillus cereus of the present invention of embodiment 1.
1.1 strain isolation
10g fermented soya bean powder is weighed in 250mL conical flasks, plus sterile saline 100mL, shake 2-4 hours, it is static heavy
Form sediment, obtain supernatant, take out 10mL and be diluted, bacterium solution is diluted to 10-7, 10 are taken respectively-1、10-3、10-3、10-7These ladders
Degree carries out flat board coating experiment, is coated on casein medium, is cultivated 24 hours at 37 DEG C, sees whether to produce transparent circle.
9 plants of single bacteriums of picking generation transparent circle, which are fallen within isolation medium, is rule, purifying bacterial strain, last 9 plants of bacterial strains of slant preservation,
Followed by enzyme activity experiment.Enzyme activity highest bacterial strain, i.e. this hair are selected according to the size of transparent circle and enzyme activity size
Bright bacterial strain.
1.2 strain characterization
(1) morphological feature
On plain agar plating medium, 37 DEG C, cultivate 24 hours, can form that circular or approximate circle, quality is soft, nothing
Pigment, slightly glossiness white colony (ceraceous candle sample color) diameter 5-7mm.More vigorous, bacterium is grown on M.S.P culture mediums
Fall diameter up to 8-10mm, it is softer, provoke to be in thread, incubation time is slightly long, bacterium colony surface is in ground-glass appearance, and produces red color
Element.Bacterium colony is canescence on the female cream flat board of albumen peptozym, and opaque, surface is rougher, like ground-glass appearance or wax melting shape, bacterium colony
It is larger.
Fig. 1 is the visual results after the Bacillus cereus strain culture 3 days of the present invention.
Fig. 2 is the microscopy results after the Bacillus cereus strain culture 3 days of the present invention.
(2) 16sRNA sequences PCR primer
First, first identification is positive bacteria or negative bacterium (our bacterial strains are positive bacteria), is carried according still further to general positive bacteria
Take DNA method to extract the DNA of bacterial strain, performing PCR is entered using 16sRNA universal primers 27F and 1492R, obtained product carries out base
Because of sequencing.
Sequencing result is as follows：
GCATGGGGGCAGCTATACATGCAGTCGAGCGAATGGATTAAGA
GCTTGCTCTTATGAAGTTAGCGGCGGACGGGTGAGTAACACGT
GGGTAACCTGCCCATAAGACTGGGATAACTCCGGGAAACCGGG
GCTAATACCGGATAACATTTTGAACCGCATGGTTCGAAATTGAA
AGGCGGCTTCGGCTGTCACTTATGGATGGACCCGCGTCGCATTA
GCTAGTTGGTGAGGTAACGGCTCACCAAGGCAACGATGCGTAG
CCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACG
GCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAA
TGGACGAAAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAG
GCTTTCGGGTCGTAAAACTCTGTTGTTAGGGAAGAACAAGTGC
TAGTTGAATAAGCTGGCACCTTGACGGTACCTAACCAGAAAGC
CACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGG
CAAGCGTTATCCGGAATTATTGGGCGTAAAGCGCGCGCAGGTG
GTTTCTTAAGTCTGATGTGAAAGCCCACGGCTCAACCGTGGAG
GGTCATTGGAAACTGGGAGACTTGAGTGCAGAAGAGGAAAGT
GGAATTCCATGTGTAGCGGTGAAATGCGTAGAGATATGGAGGAA
CACCAGTGGCGAAGGCGACTTTCTGGTCTGTAACTGACACTGA
GGCGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTA
GTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGAGGGTTTCC
GCCCTTTAGTGCTGAAGTTAACGCATTAAGCACTCCGCCTGGGG
AGTACGGCCGCAAGGCTGAAACTCAAAGGAATTGACGGGGGC
CCGCACAAGCGGTGGAGCATGTGATTTAATTCGAAGCAACGCG
AAGAACCCTTACCAGGTCTTGACATCCTCTGACACCCTTAGAGA
TAGGGCTTCTCCTTTCGGGAGCAGAGTGACAGTGCTGCATGATT
GTCGTCAGCTCGTGTCGTGAGATGTTGGGATTAGTCCTGCAACG
AGCGCACTCATGATCTTAGCTGCATCATTAGCTTGAGCACTTCTA AGTGAACTGACGGTGAACAACCGGAAG(SEQ
ID NO：1)
Secondly, GENBANK applications are carried out, strain number are obtained for KY419575.Then sequence alignment, comparison result are carried out
It is to reach 99% with bacillus cereus similarity, therefore, bacterial strain of the invention is accredited as bacillus cereus.
1.3 condition of culture
The culture medium of screening production protease：
Skimmed milk power 4g, KH2PO40.36g、Na2HPO41.07g、ZnCL20.04g、 CaCL20.002g、MgSO40.5g、
FeSO40.02g, NaCL 0.16g, tryptone 0.04g, agar 10g, distilled water 1000mL, 121 DEG C sterilize 20 minutes.
Storage medium (LB culture mediums)：
Beef extract 2.5g, peptone 5g, sodium chloride 2.5g, agar 10g, the mL of distilled water 500,121 DEG C sterilize 20 minutes.
Cultivation temperature：28~35 DEG C.
1.4 preservation explanations
Strain name：Bacillus cereus；
Latin name：Bacillus cereus；
Preservation mechanism：China General Microbiological culture presevation administrative center, abbreviation CGMCC；
Address：Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3；
Preservation date：On 04 19th, 2017；
Collection is registered on the books numbering：CGMCC NO：14049.
The preparation of the soybean small peptide of embodiment 2.
Embodiment 2.1
Soybean small peptide is prepared from according to following steps：
(a) two leaching dregs of beans are weighed, water is added, high pressure steam sterilization is inoculated with the bacillus cereus of the present invention in fermentation tank
Interior sealed fermenting 72 hours；
(b) fermented bean dregs obtained by step (a) are added into water, thermolysin is added after fully infiltrating
(thermolysin) digested, rear ultrasound extraction obtains enzymolysis liquid, thermophilic bacteria protein enzyme dosage is 5 ‰ mass fraction (g/
kg)；
(c) enzymolysis liquid obtained by step (b) is crossed into 400 mesh filter clothes to remove after particulate matter, adds activated carbon and florisil silica
Half an hour after-purification is stood, then activated carbon and florisil silica are filtered to remove through plate and frame filter press, de- bitter deodorization and part is obtained
Except the soybean protein clear liquid of color, activated carbon and florisil silica consumption are 5 ‰ and are 5 ‰ mass fractions (g/kg)；
(d) soybean protein clear liquid obtained by step (c) is carried out after 10000Da ultrafiltration membrane treatments, it is sterile big that collection is filtered out
Beans small peptide slightly contains water mixed liquid, then the soybean small peptide is slightly carried out at 1000Da NF membranes and 500Da NF membranes containing water mixed liquid
After reason, collection filters out soybean small peptide containing water mixed liquid, continues to filter out soybean small peptide containing water mixed liquid progress water removal NF membrane to this
After processing, collection does not filter out component solution for soybean small peptide mixed concentrated liquid, filters out component and crosses CSM RE2540-TE filter membranes receipts
Collect aqueous phase filtrate, then through 121 DEG C of high pressure steam sterilizations 30 minutes；
(e) it will add that electrolysis water is antibacterial after soybean small peptide mixed concentrated liquid concentration obtained by step (d), electrolysis water consumption is
5 ‰ mass fraction (g/kg), then be freeze-dried, the powdery soybean small peptide for obtaining 1000Da, 500Da and ＜ 500Da is mixed
Compound, the mixture separates base amino acid containing soybean simultaneously.
Embodiment 2.2
Soybean small peptide is prepared from according to following steps：
(a) two leaching dregs of beans are weighed, water is added, high pressure steam sterilization is inoculated with the bacillus cereus of the present invention in fermentation tank
Interior sealed fermenting 36 hours；
(b) fermented bean dregs obtained by step (a) are added into water, thermolysin is added after fully infiltrating
(thermolysin) digested, rear ultrasound extraction obtains enzymolysis liquid, thermophilic bacteria protein enzyme dosage is 3 ‰ mass fraction (g/
kg)；
(c) enzymolysis liquid obtained by step (b) is crossed into 400 mesh filter clothes to remove after particulate matter, adds activated carbon and florisil silica
Half an hour after-purification is stood, then activated carbon and florisil silica are filtered to remove through plate and frame filter press, de- bitter deodorization and part is obtained
Except the soybean protein clear liquid of color, activated carbon and florisil silica consumption are 7 ‰ and are 10 ‰ mass fractions (g/kg)；
(d) soybean protein clear liquid obtained by step (c) is carried out after 10000Da ultrafiltration membrane treatments, it is sterile big that collection is filtered out
Beans small peptide slightly contains water mixed liquid, then the soybean small peptide is slightly carried out at 1000Da NF membranes and 500Da NF membranes containing water mixed liquid
After reason, collection filters out soybean small peptide containing water mixed liquid, continues to filter out soybean small peptide containing water mixed liquid progress water removal NF membrane to this
After processing, collection does not filter out component solution for soybean small peptide mixed concentrated liquid, filters out component and crosses CSM RE2540-TE filter membranes receipts
Collect aqueous phase filtrate, then through 121 DEG C of high pressure steam sterilizations 30 minutes；
(e) it will add that electrolysis water is antibacterial after soybean small peptide mixed concentrated liquid concentration obtained by step (d), electrolysis water consumption is
10 ‰ mass fraction (g/kg), then be freeze-dried, obtain 1000Da, 500Da and ＜ 500Da powdery soybean small peptide
Mixture, the mixture separates base amino acid containing soybean simultaneously.
Embodiment 2.3
Soybean small peptide is prepared from according to following steps：
(a) two leaching dregs of beans are weighed, water is added, high pressure steam sterilization is inoculated with the bacillus cereus of the present invention in fermentation tank
Interior sealed fermenting 72 hours；
(b) fermented bean dregs obtained by step (a) are added into water, thermolysin is added after fully infiltrating
(thermolysin) digested, rear ultrasound extraction obtains enzymolysis liquid, thermophilic bacteria protein enzyme dosage is 4 ‰ mass fraction (g/
kg)；
(c) enzymolysis liquid obtained by step (b) is crossed into 400 mesh filter clothes to remove after particulate matter, adds activated carbon and florisil silica
Half an hour after-purification is stood, then activated carbon and florisil silica are filtered to remove through plate and frame filter press, de- bitter deodorization and part is obtained
Except the soybean protein clear liquid of color, activated carbon and florisil silica consumption are 5 ‰ and are 2 ‰ mass fractions (g/kg)；
(d) soybean protein clear liquid obtained by step (c) is carried out after 10000Da ultrafiltration membrane treatments, it is sterile big that collection is filtered out
Beans small peptide slightly contains water mixed liquid, then the soybean small peptide is slightly carried out at 1000Da NF membranes and 500Da NF membranes containing water mixed liquid
After reason, collection filters out soybean small peptide containing water mixed liquid, continues to filter out soybean small peptide containing water mixed liquid progress water removal NF membrane to this
After processing, collection does not filter out component solution for soybean small peptide mixed concentrated liquid, filters out component and crosses CSM RE2540-TE filter membranes receipts
Collect aqueous phase filtrate, then through 121 DEG C of high pressure steam sterilizations 30 minutes；
(e) it will add that electrolysis water is antibacterial after soybean small peptide mixed concentrated liquid concentration obtained by step (d), electrolysis water consumption is
7 ‰ mass fraction (g/kg), then be freeze-dried, the powdery soybean small peptide for obtaining 1000Da, 500Da and ＜ 500Da is mixed
Compound, the mixture separates base amino acid containing soybean simultaneously.
Embodiment 2.4
Soybean small peptide is prepared from according to following steps：
(a) two leaching dregs of beans are weighed, water is added, high pressure steam sterilization is inoculated with the bacillus cereus of the present invention in fermentation tank
Interior sealed fermenting 48 hours；
(b) fermented bean dregs obtained by step (a) are added into water, thermolysin is added after fully infiltrating
(thermolysin) digested, rear ultrasound extraction obtains enzymolysis liquid, thermophilic bacteria protein enzyme dosage is 5 ‰ mass fraction (g/
kg)；
(c) enzymolysis liquid obtained by step (b) is crossed into 400 mesh filter clothes to remove after particulate matter, adds activated carbon and florisil silica
Half an hour after-purification is stood, then activated carbon and florisil silica are filtered to remove through plate and frame filter press, de- bitter deodorization and part is obtained
Except the soybean protein clear liquid of color, activated carbon and florisil silica consumption are 2 ‰ and are 7 ‰ mass fractions (g/kg)；
(d) soybean protein clear liquid obtained by step (c) is carried out after 10000Da ultrafiltration membrane treatments, it is sterile big that collection is filtered out
Beans small peptide slightly contains water mixed liquid, then the soybean small peptide is slightly carried out at 1000Da NF membranes and 500Da NF membranes containing water mixed liquid
After reason, collection filters out soybean small peptide containing water mixed liquid, continues to filter out soybean small peptide containing water mixed liquid progress water removal NF membrane to this
After processing, collection does not filter out component solution for soybean small peptide mixed concentrated liquid, filters out component and crosses CSM RE2540-TE filter membranes receipts
Collect aqueous phase filtrate, then through 121 DEG C of high pressure steam sterilizations 30 minutes；
(e) it will add that electrolysis water is antibacterial after soybean small peptide mixed concentrated liquid concentration obtained by step (d), electrolysis water consumption is
5 ‰ mass fraction (g/kg), then be freeze-dried, the powdery soybean small peptide for obtaining 1000Da, 500Da and ＜ 500Da is mixed
Compound, the mixture separates base amino acid containing soybean simultaneously.
Embodiment 2.5
Soybean small peptide is prepared from according to following steps：
(a) two leaching dregs of beans are weighed, water is added, high pressure steam sterilization is inoculated with the bacillus cereus of the present invention in fermentation tank
Interior sealed fermenting 60 hours；
(b) fermented bean dregs obtained by step (a) are added into water, thermolysin is added after fully infiltrating
(thermolysin) digested, rear ultrasound extraction obtains enzymolysis liquid, thermophilic bacteria protein enzyme dosage is 2 ‰ mass fraction (g/
kg)；
(c) enzymolysis liquid obtained by step (b) is crossed into 400 mesh filter clothes to remove after particulate matter, adds activated carbon and florisil silica
Half an hour after-purification is stood, then activated carbon and florisil silica are filtered to remove through plate and frame filter press, de- bitter deodorization and part is obtained
Except the soybean protein clear liquid of color, activated carbon and florisil silica consumption are 10 ‰ and are 5 ‰ mass fractions (g/kg)；
(d) soybean protein clear liquid obtained by step (c) is carried out after 10000Da ultrafiltration membrane treatments, it is sterile big that collection is filtered out
Beans small peptide slightly contains water mixed liquid, then the soybean small peptide is slightly carried out at 1000Da NF membranes and 500Da NF membranes containing water mixed liquid
After reason, collection filters out soybean small peptide containing water mixed liquid, continues to filter out soybean small peptide containing water mixed liquid progress water removal NF membrane to this
After processing, collection does not filter out component solution for soybean small peptide mixed concentrated liquid, filters out component and crosses CSM RE2540-TE filter membranes receipts
Collect aqueous phase filtrate, then through 121 DEG C of high pressure steam sterilizations 30 minutes；
(e) it will add that electrolysis water is antibacterial after soybean small peptide mixed concentrated liquid concentration obtained by step (d), electrolysis water consumption is
2 ‰ mass fraction (g/kg), then be freeze-dried, the powdery soybean small peptide for obtaining 1000Da, 500Da and ＜ 500Da is mixed
Compound, the mixture separates base amino acid containing soybean simultaneously.
The liquid chromatogram of the gained soybean small peptide of embodiment 2.1 is as shown in figure 3, this is filtered out with different pore size milipore filter
The liquid chromatogram of the soybean small peptide mixture of the different molecular weight come.(A), the liquid chromatogram of 10000Da filter liquors and
The liquid chromatogram of 5000Da filter liquors；(B), the liquid chromatogram of the liquid chromatogram of 1000Da filter liquors and 500Da filter liquors
Figure.High-efficient liquid phase chromatogram condition is：Cosmosil 5C18- AR II chromatographic columns (4.6mm × 250 mm), gradient type elution (0%
~80% acetonitrile %, 60 minutes), mobile phase A is addition 0.1%TFA MilliQ ultra-pure waters, and Mobile phase B is pure acetonitrile, flow velocity
For 0.5mL/min, Detection wavelength is 220nm, and column temperature is 35 DEG C.
Spectrogram fine structure illustrates the smaller filter liquor of molecular weight, and chromatographic peak information is abundanter, and the quantity of small peptide is fewer, dense
Degree is higher.
The liquid chromatogram of soybean small peptide product obtained by embodiment 2.2-2.5 is similar with Fig. 3.
The preparation of the carbonated soda water drinks of embodiment 3.
The preparation method of carbonated soda water drinks of the present invention is mainly included the following steps that：
(a) raw material is weighed by weight：Soybean small peptide 1-5 parts made from embodiment 2.1,1500 parts of water, citric acid 10-
12 parts, 60-70 parts of xylitol, 7.5-8 parts of sodium bicarbonate；
(b) carbon dioxide is filled with after above-mentioned raw materials are mixed to saturation, filling and capping is simultaneously stored.
The preparation method is further elaborated below by following embodiments.
Embodiment 3.1
The present embodiment provides the parts by weight of beverage raw material：1 part of soybean small peptide (below 500Da), 1500 parts of water, citric acid
12 parts, the citric acid be Citric Acid Mono, 70 parts of xylitol, 8 parts of sodium bicarbonate, the sodium bicarbonate be sodium acid carbonate, be filled with two
Carbonoxide to saturation, filling and capping is simultaneously stored.
Embodiment 3.2
The present embodiment provides the parts by weight of beverage raw material：3 parts of soybean small peptide (below 1000Da), 1500 parts of water, lemon
Acid 10 parts, the citric acid be Citric Acid Mono, 60 parts of xylitol, 7.5 parts of sodium bicarbonate, the sodium bicarbonate be sodium acid carbonate, fill
Enter carbon dioxide to saturation, filling and capping is simultaneously stored.
Embodiment 3.3
The present embodiment provides the parts by weight of beverage raw material：5 parts of soybean small peptide (below 1000Da), 1500 parts of water, lemon
Acid 11 parts, the citric acid be Citric Acid Mono, 65 parts of xylitol, 7.5 parts of sodium bicarbonate, the sodium bicarbonate be sodium acid carbonate, fill
Enter carbon dioxide to saturation, filling and capping is simultaneously stored.
Embodiment 3.4
The present embodiment provides the parts by weight of beverage raw material：5 parts of soybean small peptide (crosses CSM RE2540-TE filter membranes and collects water
The xeraphium of phase filtrate, below 500Da), 1500 parts of water, 11 parts of citric acid, the citric acid is Citric Acid Mono, xylitol 60
Part, 8 parts of sodium bicarbonate, the sodium bicarbonate is sodium acid carbonate, is filled with carbon dioxide to saturation, filling and capping is simultaneously stored.
Sensory evaluation test example
3.1-3.4 of the embodiment of the present invention is respectively labeled as No. 1-4, to soybean small peptide carbonated soda water drinks local flavor, mouthfeel,
Color and luster, sugariness, acidity, clarity and overall acceptable degree carry out subjective appreciation, obtain effective sample 51, experimental result data
It is as shown in the table：
As can be known from the above table, the subjective appreciation person of rawness type, i.e. ordinary consumer are to embodiments of the invention 3.1-3.4's
Calcareous type soybean small peptide sparkling water fancy grade is likes, and only part is somewhat liked, and peptide taste is not also felt significantly.
In addition, milipore filter and NF membrane that the filter membrane used in the present invention is used in being produced for food industry, are filled with two
The machine of carbonoxide is commercially available Soda water machine；Citric acid, xylitol, the sodium bicarbonate used in the present invention is commercially available prod, lemon
Lemon acid can be replaced by fresh squeezing lemon juice.
Described above is only the specific embodiment of the present invention, not does any formal limitation to the present invention, though
So the present invention is disclosed as above with specific embodiment, but is not limited to the present invention, any to be familiar with this professional technology people
Member, in the range of technical solution of the present invention is not departed from, when using technology contents disclosed above make it is a little change or repair
The equivalent embodiment for equivalent variations is adornd, as long as being the content without departing from technical solution of the present invention, the technology according to the present invention is real
Any simple modification, equivalent variations and modification that confrontation above example is made, still fall within the scope of technical solution of the present invention
It is interior.
Sequence table
<110>Shanghai Ocean University
<120>A kind of soybean small peptide and preparation method and application
<130>
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1150
<212> DNA
<213>Bacillus cereus（Bacillus cereus）
<400> 1
gcatgggggc agctatacat gcagtcgagc gaatggatta agagcttgct cttatgaagt 60
tagcggcgga cgggtgagta acacgtgggt aacctgccca taagactggg ataactccgg 120
gaaaccgggg ctaataccgg ataacatttt gaaccgcatg gttcgaaatt gaaaggcggc 180
ttcggctgtc acttatggat ggacccgcgt cgcattagct agttggtgag gtaacggctc 240
accaaggcaa cgatgcgtag ccgacctgag agggtgatcg gccacactgg gactgagaca 300
cggcccagac tcctacggga ggcagcagta gggaatcttc cgcaatggac gaaagtctga 360
cggagcaacg ccgcgtgagt gatgaaggct ttcgggtcgt aaaactctgt tgttagggaa 420
gaacaagtgc tagttgaata agctggcacc ttgacggtac ctaaccagaa agccacggct 480
aactacgtgc cagcagccgc ggtaatacgt aggtggcaag cgttatccgg aattattggg 540
cgtaaagcgc gcgcaggtgg tttcttaagt ctgatgtgaa agcccacggc tcaaccgtgg 600
agggtcattg gaaactggga gacttgagtg cagaagagga aagtggaatt ccatgtgtag 660
cggtgaaatg cgtagagata tggaggaaca ccagtggcga aggcgacttt ctggtctgta 720
actgacactg aggcgcgaaa gcgtggggag caaacaggat tagataccct ggtagtccac 780
gccgtaaacg atgagtgcta agtgttagag ggtttccgcc ctttagtgct gaagttaacg 840
cattaagcac tccgcctggg gagtacggcc gcaaggctga aactcaaagg aattgacggg 900
ggcccgcaca agcggtggag catgtgattt aattcgaagc aacgcgaaga acccttacca 960
ggtcttgaca tcctctgaca cccttagaga tagggcttct cctttcggga gcagagtgac 1020
agtgctgcat gattgtcgtc agctcgtgtc gtgagatgtt gggattagtc ctgcaacgag 1080
cgcactcatg atcttagctg catcattagc ttgagcactt ctaagtgaac tgacggtgaa 1140
caaccggaag 1150
Claims (10)
1. one plant of bacillus cereus (Bacillus cereus) bacterial strain, it is preserved in China General Microbiological culture presevation pipe
Reason center, preserving number is CGMCC NO:14049.
2. a kind of microbial inoculum, its active component is the Bacillus cereus strain described in claim 1.
3. Bacillus cereus strain described in claim 1 protein degradation, decomposition of protein or prepare in small-molecular peptides should
With.
4. a kind of method for preparing soybean small peptide, it comprises the following steps：
(a) dregs of beans is weighed, is sterilized, bacillus cereus is inoculated with, and ferment, it is preferable that fermentation time is 15 days；
(b) fermented bean dregs obtained by step (a) add protease, are digested；
(c) enzymolysis product obtained by filtration step (b) simultaneously stands purification, obtains de- bitter deodorization and part except the soybean protein of color is clear
Liquid, it is preferable that the filtering is carried out with 400 mesh filter clothes, it is preferable that the standing purification activated carbon and florisil silica enter
OK, it is preferable that activated carbon and the florisil silica consumption is 5 ‰ and is 10 ‰ mass fractions, g/kg；
(d) soybean protein clear liquid obtained by step (c) is subjected to below 10000Da ultrafiltration membrane treatments, then carries out below 1000Da to receive
Filter membrane and below 500Da nanofiltration membrane treatments, collect the soybean small peptide filtered out and contain water mixed liquid, continue the soybean small peptide filtered out to this
Water removal nanofiltration membrane treatment is carried out containing water mixed liquid, soybean small peptide is obtained.
5. the method for preparing soybean small peptide described in claim 4, it further comprises：
(e) soybean small peptide obtained by step (d) is concentrated, then be freeze-dried, it is preferable that after concentration, freeze-drying
Before, electrolysis water progress is added antibacterial, it is preferable that electrolysis water consumption is 5 ‰.
6. the method for preparing soybean small peptide described in claim 4 or 5, wherein described bacillus cereus is claim 1
Described Bacillus cereus strain.
7. the method for preparing soybean small peptide described in claim 4 or 5, wherein the protease is thermolysin, preferably
Ground, the thermophilic bacteria protein enzyme dosage is 5 ‰ mass fractions (g/kg).
8. a kind of soybean small peptide, it is prepared as the method any one of claim 4-7.
9. a kind of beverage, by weight, it includes following composition：
1-5 parts of soybean small peptide, it is preferable that the soybean small peptide is the soybean small peptide described in claim 8；
10-12 parts of citric acid, it is preferable that the citric acid is Citric Acid Mono；
60-70 parts of xylitol；
7.5-8 parts of sodium bicarbonate；
1500 parts of water.
10. beverage as claimed in claim 9, it includes following composition：
3 parts of Soybean Peptide；
10 parts of citric acid；
60 parts of xylitol；
7.5 parts of sodium bicarbonate,
1500 parts of water.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111758881A (en) * | 2020-07-13 | 2020-10-13 | 上海海洋大学 | Landscaping jelly containing soybean small peptides and preparation method thereof |
| CN113243471A (en) * | 2021-06-21 | 2021-08-13 | 平凉市食品检验检测中心 | Preparation method of fermented apple juice for relieving intestinal dysbacteriosis |
| CN117683074A (en) * | 2023-12-13 | 2024-03-12 | 山东绿邦生物科技有限公司 | Bacillus cereus, protease inhibitor and method for improving yield of protein residues |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111758881A (en) * | 2020-07-13 | 2020-10-13 | 上海海洋大学 | Landscaping jelly containing soybean small peptides and preparation method thereof |
| CN113243471A (en) * | 2021-06-21 | 2021-08-13 | 平凉市食品检验检测中心 | Preparation method of fermented apple juice for relieving intestinal dysbacteriosis |
| CN117683074A (en) * | 2023-12-13 | 2024-03-12 | 山东绿邦生物科技有限公司 | Bacillus cereus, protease inhibitor and method for improving yield of protein residues |
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