CN106520583B - It is a kind of for strengthening the preparation method and application of bean cotyledon post-fermentation antimicrobial composition - Google Patents
It is a kind of for strengthening the preparation method and application of bean cotyledon post-fermentation antimicrobial composition Download PDFInfo
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Abstract
The present invention relates to belong to bioengineering field, it is related to a kind of for strengthening the preparation method and application of bean cotyledon post-fermentation antimicrobial composition, antimicrobial composition includes yeast agent, aspergillus oryzae microbial inoculum and lattice spore chamber bacteria agent, three kinds of microbial inoculums are prepared respectively, are then mixed for bean cotyledon post-fermentation with post-fermentation bean cotyledon in proportion.The preparation method of present invention offer antimicrobial composition, integrated artistic is suitble to industrialize continuous production, fully utilize bean paste production by-product (capsicum campanulaceae), production cycle can save 6 months, amino nitrogen improves 20%, volatility is in that fragrant component improves 3 times or more (total ester, total acid and total aldehyde content), in aflatoxin B1Yeast agent has been accessed on the peak (fermentation 30-60 days) of generation, enhance and produce ester flavor formation, and Reverse transcriptase produces the metabolism of poison aspergillus flavus and section parasitic aspergillus, reduces the content of aflatoxin B1, aflatoxin B1Lower than 0.5ppm, food safety is improved.
Description
Technical field
The present invention relates to bioengineering field is belonged to, it is related to a kind of for strengthening the preparation side of bean cotyledon post-fermentation antimicrobial composition
Method and application.
Background technique
Bean paste category Chinese tradition fermented food, so far existing more than 300 years history, it is immaterial to be listed in China
Cultural heritage.Not only production technology is unique for bean paste, also with its taste it is peppery it is aromatic, sticky suede is real, it is reddish brown it is glossy, paste flavor is strong etc.
Feature is taken the course of its own in China's sauce products, can be rated as the soul of Sichuan cuisine.Year end 2015 of cut-off, " bean paste " brand value have reached
607.16 hundred million yuan, rank " processed food class geography symbol product " whole nation first；Current year product total output reaches 1,100,000 tons, in fact
Existing 10,200,000,000 yuan of industrial output value, exports world's overwhelming majority countries and regions, earns foreign exchange more than 40,000,000 dollars.
The production of bean paste includes two stages of prior fermentation and afterripening fermentation, and prior fermentation is primarily referred to as the mould valve of semen viciae fabae
The koji-making of son and the pretreatment of pepper blank.Afterripening fermentation mainly by mature mould valve and mature pepper blank, mix in proportion by ingredient
It closes, appropriate salt and water is added, into fermenting tank for fermentation, tedding and being aged by the regular period is that bean paste is peculiar
Weather exposure technique.Therefore, the production process of bean paste can be stated are as follows: the production of bean paste is with semen viciae fabae valve system
Premised on the complicated material energy metabolism process such as bent, pepper blank and environmental microorganism, developed by unique " weather exposure "
Zymotechnique, so that the huge microbiota inhabited in Koji, pepper blank, environment ties up to fermenting grain solid, liquid, gas three phase boundary
Complicated material conversion, energetic supersession and information transfering action occurs, and ultimately form the unique ingredient of bean paste constitute and
Flavor characteristic.
Fungi has played very important effect in bean paste fermentation process, and in starter-making stage, mould can secrete egg
A variety of enzymes such as white enzyme, amylase, carbohydrase, these enzyme effects not only generate a large amount of flavor substance in capsicum and semen viciae fabae,
Also it is created condition for the growth of other microorganisms of post-fermentation phase.But it is opened since the production in bean paste post-fermentation stage is in
Environment is put, few then half a year is more, and 2 years or more " weather exposure " post-fermentation phase, there are also a large amount of environmental microorganisms to participate in its fermentation
Process, this not only determines the unique ingredient composition of bean paste and flavor characteristic, and there is also huge food safety risks, such as
Mucor and Penicillium notatum can generate frowst, produce malicious aspergillus flavus and section parasitic aspergillus is also possible to generate aflatoxin B1, band
Carry out great food safety hazards.Therefore, the fungal succession variation of bean paste post-fermentation phase is studied, and is obtained using the objective law
Bean paste post-fermentation process must be strengthened just to be very necessary.
Summary of the invention
The purpose of the present invention is existing in view of the above technology, provide a kind of for strengthening bean cotyledon post-fermentation microbial inoculum group
Close the preparation method and application of object.
To achieve the above object, the technical solution adopted by the present invention is that: one kind for strengthen bean cotyledon post-fermentation microbial inoculum combination
The preparation method of object, the antimicrobial composition include yeast agent, aspergillus oryzae microbial inoculum and lattice spore chamber bacteria agent, and preparation method is such as
Under:
The preparation of yeast agent: the Pichia anomala of logarithmic growth phase and torulopsis are pressed into 3-5% (v/v) inoculum concentration
It is cultivated at 5 ° 30 DEG C of Be ' brewer's wort 3 days, it is 6% porous-starch, 3% sucrose and 1% that quality is then added in fermentation liquid
Glycerol, through temperature be -40 DEG C~-45 DEG C, freeze-drying time be 24-48h vacuum freezedrying be made yeast agent；
The preparation of aspergillus oryzae microbial inoculum: the aspergillus oryzae of logarithmic growth phase is bent at 6~7 ° é meters of B by 3-5% (v/v) inoculum concentration
Juice, 28-30 DEG C culture 2-5 days, when rice song juice being made to be presented micro- yellow, after the completion of culture, adding addition in fermentation liquid is quality point
The porous-starch of number 6%, 8% sucrose and 1% glycerol, through temperature be -40 DEG C~-45 DEG C, freeze-drying time be 24-48h it is cold
Freeze to be dried in vacuo and aspergillus oryzae microbial inoculum is made；
The preparation of lattice spore chamber bacteria agent: the lattice spore chamber bacterium of logarithmic growth phase is cultivated by 4-5% (v/v) inoculum concentration in sterilizing
After the completion of culture, the porous-starch of mass fraction 6%, 8% sucrose are added into fermentation liquid for 28-30 DEG C culture 2-5 days in base
With 1% glycerol, be -40 DEG C~-45 DEG C through temperature, freeze-drying time is that 24-48h vacuum freezedrying is obtained.
Further, the sterilising medium composition are as follows: wheat bran 0.2-0.4g/L, flour 1-2g/L, dry chili campanulaceae 1-
2g/L and urea 3-6g/L, pH 3.2-4.0.
In the present invention, Pichia anomala Pichia anomala preservation number are as follows: CICC 1332, lattice spore chamber bacterium
Pleospora sp. preservation number are as follows: BNCC 179839, torulopsis Torulopsis globosa preservation number are as follows:
BNCC176445, aspergillus oryzae Aspergillus oryzae preservation number are as follows: ATCC 41416.
It is a further object of the present invention to provide a kind of application of antimicrobial composition during bean cotyledon post-fermentation.
Further, during bean cotyledon post-fermentation, when fermentation was to 15-30 days according to mass ratio 0.2:10000 addition rice
Aspergillus microbial inoculum is simultaneously uniformly mixed fermentation, and when reaching 30-60 days wait ferment, yeast agent is added simultaneously according to mass ratio 0.1:10000
It is uniformly mixed, when reaching 90 days or more wait ferment, to be added dose of fermentation lattice spore chamber bacterium bacterium is added according to mass ratio 0.05:10000
Agent.
The method have the benefit that: the present invention provides the preparation method of antimicrobial composition, and integrated artistic is suitble to work
Industry continuous production fully utilizes bean paste production by-product (capsicum campanulaceae), and the production cycle can save 6 months, amino
State nitrogen improves 20%, and volatility is in that fragrant component improves 3 times or more (total ester, total acid and total aldehyde content), in aflatoxin B1It produces
Yeast agent has been accessed on raw peak (fermentation 30-60 days), enhance and produce ester flavor formation, and Reverse transcriptase produces poison aspergillus flavus
With the metabolism of section parasitic aspergillus, the content of aflatoxin B1, aflatoxin B are reduced1Lower than 0.5ppm, food is improved
Product safety.
Specific embodiment
Below in conjunction with the embodiment of the present invention, technical scheme in the embodiment of the invention is clearly and completely described,
Obviously, described embodiments are only a part of the embodiments of the present invention, instead of all the embodiments.Based in the present invention
Embodiment, every other embodiment obtained by those of ordinary skill in the art without making creative efforts, all
Belong to the scope of protection of the invention.
Embodiment 1
It is a kind of for strengthening the preparation method of bean cotyledon post-fermentation antimicrobial composition, which is characterized in that the described microbial inoculum combination
Object includes yeast agent, aspergillus oryzae microbial inoculum and lattice spore chamber bacteria agent, the preparation method is as follows:
The preparation of yeast agent: the Pichia anomala of logarithmic growth phase and torulopsis are pressed into 3-5% (v/v) inoculum concentration
It is cultivated at 5 ° 30 DEG C of Be ' brewer's wort 3 days, it is 6% porous-starch, 3% sucrose and 1% that quality is then added in fermentation liquid
Glycerol, through temperature be -40 DEG C~-45 DEG C, freeze-drying time be 24-48h vacuum freezedrying be made yeast agent；
The preparation of aspergillus oryzae microbial inoculum: the aspergillus oryzae of logarithmic growth phase is bent at 6~7 ° é meters of B by 3-5% (v/v) inoculum concentration
Juice, 28-30 DEG C culture 2-5 days, when rice song juice being made to be presented micro- yellow, after the completion of culture, adding addition in fermentation liquid is quality point
The porous-starch of number 6%, 8% sucrose and 1% glycerol, through temperature be -40 DEG C~-45 DEG C, freeze-drying time be 24-48h it is cold
Freeze to be dried in vacuo and aspergillus oryzae microbial inoculum is made；
The preparation of lattice spore chamber bacteria agent: the lattice spore chamber bacterium of logarithmic growth phase is cultivated by 4-5% (v/v) inoculum concentration in sterilizing
After the completion of culture, the porous-starch of mass fraction 6%, 8% sucrose are added into fermentation liquid for 28-30 DEG C culture 2-5 days in base
With 1% glycerol, be -40 DEG C~-45 DEG C through temperature, freeze-drying time is that 24-48h vacuum freezedrying is obtained.
Wherein, sterilising medium forms are as follows: wheat bran 0.2-0.4g/L, flour 1-2g/L, dry chili campanulaceae 1-2g/L and urine
Plain 3-6g/L, pH 3.2-4.0.
Embodiment 2
It is a kind of for strengthening application method of the bean cotyledon post-fermentation antimicrobial composition in bean cotyledon post-fermentation, bean cotyledon post-fermentation mistake
Cheng Zhong is added aspergillus oryzae microbial inoculum and is uniformly mixed and ferment, wait ferment when fermentation was to 15-30 days according to mass ratio 0.2:10000
When reaching 30-60 days, yeast agent is added according to mass ratio 0.1:10000 and is uniformly mixed, when reaching 90 days or more wait ferment,
To be added dose of fermentation lattice spore chamber bacteria agent is added according to mass ratio 0.05:10000.
Embodiment 3
Antimicrobial composition of the present invention is to aflatoxins B in bean cotyledon post-fermentation1Influence.
(1) sample acquires
Pichia anomala (Pichia anomala CICC 1332), lattice spore chamber bacterium are added alone or in combination
(Pleospora sp.BNCC179839), torulopsis (Torulopsis globosa BNCC176445) and aspergillus oryzae
(Aspergillus oryzae ATCC 41416), total totally 40 batch post-fermentation bean paste samples, every kind of addition do 5
Secondary parallel laboratory test.(result sees below table)
(2) instrument and reagent
Instrument: 90mm mortar (Kai Pingshengxing chemical porcelain factory, Tangshan City)；Constant temperature culture oscillator (the sincere analysis instrument of Shanghai intelligence
Manufacturing Co., Ltd)；Vortex oscillator (German IRM company)；Microplate reader (Bio-Rad company, the U.S.)；Thermostat water bath (Jintan
Fuhua Instrument and Equipment Company, city).
Reagent: methanol, n-hexane, chloroform (being to analyze pure, Chengdu Ke Long chemical reagent factory), aflatoxin
B1 (AFB1) ELISA kit (German R-Biopharm company).
(3) method
1. sample pretreatment: taking post-fermentation bean paste sample 20g, homogeneous paste is ground into mortar, envelope saves
It is spare afterwards.
2. aflatoxin B1Extract: extracting method uses improved national standard method, specifically: weigh 2.5 ± 0.05g
Bean cotyledon sample after grinding, is placed in 250mL stuffed conical flask, 5mL n-hexane and 12.5mL60% methanol aqueous solution is added, by force
Forced oscillation 5min is centrifuged 5min in 4000r/min after standing.It takes 5mL middle layer in separatory funnel, tri- chloromethane of 10mL is added
Alkane, stratification after concussion release and collect lower layer's chloroform.The chloroform for taking 1mL to collect will evaporate in evaporating dish
Ware is put into draught cupboard after 65 DEG C of water-baths volatilize, and 100 μ L60% methanol aqueous solutions are added and redissolve, and adds the dilution of 400 μ L samples
Liquid (appended by kit) is diluted, and 4 DEG C of preservations are to be measured.
3. detection method: sample measurement is carried out according to the method on kit operation instructions, it is specific as follows: to take suitable
Hole item is fixed on micropore grillage, is ready to be placed at room temperature for the required reagent of 2h or more and sample extracting solution；By standard items and
Sample to be tested is added in corresponding aperture, then 50 μ L antibody enzyme conjugates are added in every hole, is put in 37 DEG C of warm bath after slight oscillatory
30min；Liquid in hole is outwelled after the water bath is over, is washed microwell plate 5 times with washing lotion in kit, it for the last time should be on blotting paper
It pats dry；Be added 50 μ L developing solution A after washing in every hole, then plus 50 μ L developing solution B, in 37 DEG C of warm bath 10min after slight oscillatory；
50 μ L terminate liquids are added in every hole, detects absorbance in 450nm after mixing, is as a result read in 5min.
4. data processing: with aflatoxin B1The logarithm of standard concentration is X-axis, percentage absorbance value (each hole
Light absorption value is measured divided by 0 concentration standards light absorption value) it is Y-axis, draw standard curve.Then AFB1 content in sample are as follows:
X=10^c × k
AFB1 content, ug/kg in X- sample；
Log concentration value corresponding to the percentage absorbance value of each sample in c- standard curve；
K- extension rate.
(4) result and analysis
1. different microbial inoculums strengthen AFB1 content situation in bean paste sample
The content situation (μ g/kg) of AFB1 in the bean paste sample that the different microbial inoculums of table 1 are strengthened
Note: " ND " refers to lower than detection limit (0.2 μ g/kg).
Those skilled in the art of the present technique are appreciated that unless otherwise defined, all terms used herein (including technology art
Language and scientific term) there is meaning identical with the general understanding of those of ordinary skill in fields of the present invention.Should also
Understand, those terms such as defined in the general dictionary, which should be understood that, to be had and the meaning in the context of the prior art
The consistent meaning of justice, and unless defined as here, it will not be explained in an idealized or overly formal meaning.
It should be noted last that: the above embodiments are only used to illustrate and not limit the technical solutions of the present invention, although ginseng
It is described the invention in detail according to above-described embodiment, it will be apparent to an ordinarily skilled person in the art that: it still can be to this
Invention is modified or replaced equivalently, without departing from the spirit or scope of the invention, or any substitutions,
It is intended to be within the scope of the claims of the invention.
Claims (4)
1. a kind of preparation method for bean cotyledon post-fermentation microbial inoculum, which is characterized in that the microbial inoculum is by yeast agent, aspergillus oryzae
Microbial inoculum and lattice spore chamber bacterium composition, the preparation method is as follows:
The preparation of yeast agent: by the Pichia anomala of logarithmic growth phase and torulopsis by 3-5% (v/v) inoculum concentration at 5 °
30 DEG C of Be ' brewer's wort cultivate 3 days, then in fermentation liquid add quality be 6% porous-starch, 3% sucrose and 1% glycerol,
It is -40 DEG C~-45 DEG C through temperature, freeze-drying time is that yeast agent is made in 24-48h vacuum freezedrying；
The preparation of aspergillus oryzae microbial inoculum: by the aspergillus oryzae of logarithmic growth phase by 3-5% (v/v) inoculum concentration in 6~7 ° of é meters of B bent juice, 28-
30 DEG C are cultivated 2-5 days, and when rice song juice being made to be presented micro- yellow, after the completion of culture, adding addition in fermentation liquid is mass fraction 6%
Porous-starch, 8% sucrose and 1% glycerol are -40 DEG C~-45 DEG C through temperature, and freeze-drying time is dry for 24-48h freezing vacuum
Dry obtained aspergillus oryzae microbial inoculum；
The preparation of lattice spore chamber bacteria agent: by the lattice spore chamber bacterium of logarithmic growth phase by 4-5% (v/v) inoculum concentration in sterilising medium
28-30 DEG C culture 2-5 days, after the completion of culture, the porous-starch of mass fraction 6%, 8% sucrose and 1% are added into fermentation liquid
Glycerol is -40 DEG C~-45 DEG C through temperature, and freeze-drying time is made for 24-48h vacuum freezedrying.
2. being used for the preparation method of bean cotyledon post-fermentation microbial inoculum according to claim 1, which is characterized in that the sterilising medium
Composition are as follows: wheat bran 0.2-0.4g/L, flour 1-2 g/L, dry chili campanulaceae 1-2 g/L and urea 3-6 g/L, pH 3.2-
4.0。
3. purposes of the microbial inoculum that preparation method according to claim 1 or claim 2 obtains in bean cotyledon post-fermentation.
4. purposes according to claim 3, which is characterized in that during bean cotyledon post-fermentation, when fermentation was to 15-30 days according to
Mass ratio 0.2:10000 is added aspergillus oryzae microbial inoculum and is uniformly mixed fermentation, when reaching 30-60 days wait ferment, according to mass ratio
0.1:10000 is added yeast agent and is uniformly mixed, and when reaching 90 days or more wait ferment, is added according to mass ratio 0.05:10000
To be added dose of fermentation lattice spore chamber bacteria agent.
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| CN107259367B (en) * | 2017-07-20 | 2020-12-22 | 四川省食品发酵工业研究设计院 | Method for improving amino acid nitrogen content of thick broad-bean sauce |
| CN108094905B (en) * | 2017-12-28 | 2021-06-18 | 西华大学 | Production method of sauced Pi county broad bean paste based on 4-ethylguaiacol fermentation strengthening |
| CN111996142B (en) * | 2020-08-20 | 2023-06-06 | 四川饭扫光食品集团股份有限公司 | Freeze dryer for preparing reinforced bean cotyledon post-fermentation microbial inoculant composition |
| CN111990601A (en) * | 2020-08-28 | 2020-11-27 | 西华大学 | Special koji for Pixian bean paste and preparation method thereof |
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| CN102948748A (en) * | 2012-11-02 | 2013-03-06 | 江苏吉贝尔药业有限公司 | Anti-fatigue health care product |
| CN105192596A (en) * | 2015-10-10 | 2015-12-30 | 西华大学 | Segmented temperature control bean paste multi-microbe co-brewing and quick fermentation method |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102948748A (en) * | 2012-11-02 | 2013-03-06 | 江苏吉贝尔药业有限公司 | Anti-fatigue health care product |
| CN105192596A (en) * | 2015-10-10 | 2015-12-30 | 西华大学 | Segmented temperature control bean paste multi-microbe co-brewing and quick fermentation method |
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