CN108094712A - Aflatoxin B in a kind of distillers ' grains1Biodegrading process - Google Patents

Aflatoxin B in a kind of distillers ' grains1Biodegrading process Download PDF

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CN108094712A
CN108094712A CN201711472042.5A CN201711472042A CN108094712A CN 108094712 A CN108094712 A CN 108094712A CN 201711472042 A CN201711472042 A CN 201711472042A CN 108094712 A CN108094712 A CN 108094712A
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distillers
grains
culture
inoculation
aflatoxin
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CN108094712B (en
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常敬华
何志明
栾雨婷
刘聪
项莹
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Liaoning Baijiali feed Sales Co.,Ltd.
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Liaoning Technical University
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K10/00—Animal feeding-stuffs
    • A23K10/30—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms
    • A23K10/37—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms from waste material
    • A23K10/38—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms from waste material from distillers' or brewers' waste
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L5/00—Preparation or treatment of foods or foodstuffs, in general; Food or foodstuffs obtained thereby; Materials therefor
    • A23L5/20—Removal of unwanted matter, e.g. deodorisation or detoxification
    • A23L5/28—Removal of unwanted matter, e.g. deodorisation or detoxification using microorganisms
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
    • Y02P60/80—Food processing, e.g. use of renewable energies or variable speed drives in handling, conveying or stacking
    • Y02P60/87—Re-use of by-products of food processing for fodder production

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Abstract

Aflatoxin B in a kind of distillers ' grains1Biodegrading process, belong to feed safety technical field, step is as follows：By straw mushroom strain by solid-liquid medium culture, formation straw mushroom culture solution is placed in distillers ' grains after cultivating and filters out the object for meeting laccase activity, spare；Aspergillus flavus culture solution is cultivated, aspergillus flavus culture solution culture is added in into distillers ' grains by proportioning, completes inoculation；The distillers ' grains that inoculation is taken to complete add cultivate ingredient thereto, after abundant mixing, by the straw mushroom culture solution mixing after proportioning inoculation screening, condition of culture are controlled to be cultivated, aflatoxin B in completion distillers ' grains1Degradation, after measured, aflatoxin B1Degradation rate reaches more than 86.02%.Aflatoxin B in the distillers ' grains of the present invention1Biodegrading process, aflatoxin B in distillers ' grains can be significantly reduced1Content, for reduce livestock and poultry to aflatoxin B1Intake, improve product potency, the health risk for reducing economic loss and humans and animals is of great significance.

Description

Aflatoxin B in a kind of distillers ' grains1Biodegrading process
Technical field：
The invention belongs to feed safety technical fields, and in particular to aflatoxin B in a kind of distillers ' grains1Degradation side Method.
Background technology：
Aflatoxin is the very similar compound of a kind of chemical constitution that the moulds such as aspergillus flavus, aspergillus parasiticus generate, It is the most important mycotoxin of pollution grain and oil crop, food and feed in China and world wide, there is strong toxicity and carcinogenic Property, most important of which is that aflatoxin B1(AFB1), I class carcinogenic substances are classified as by the World Health Organization, seriously endanger people and animals Health.And its secondary metabolite remains in animal food and can form potential hazard to human health by food chain. Such as aflatoxin MlIt is aflatoxin B1Main metabolites in mammal body.Be present in animal milk, The edible parts such as liver, eggs, are common in milk, it has very strong toxicity and carcinogenicity.In actual production, stringent control is raised AFB in material1Content to ensure safety of human and livestock, reduce economic loss have direct realistic meaning.
Distillers ' grains are with the residue for raw material after everfermentation, distillation extraction alcohol such as sorghum, wheat, corn, white wine The nutritional ingredients such as most protein, fat, calcium, phosphorus in remaining raw material in grain.The feed whole nation is directly produced using distillers ' grains Feed-use grain can be saved every year ten thousand tons at least more than 350.But fresh grain stillage moisture is big, rich in abundant nutrient, easily grows mould Bacterium, especially aflatoxin.During wine brewing, 3 times of cereal materials generate 1 times of vinasse accessory substance, if raw material is dirty Dye, aflatoxin can be concentrated to 3 times of raw material in distillers ' grains, seriously endanger fowl poultry safety.But aflatoxin property is stablized, General processing method cannot remove its toxicity.Therefore, aflatoxin in distillers ' grains is effectively controlled and eliminated, be badly in need of A kind of safe efficient and environmentally friendly poison-removing method.
The content of the invention：
The purpose of the present invention is overcoming above-mentioned the shortcomings of the prior art, aflatoxin B in a kind of distillers ' grains is provided1 Biodegrading process, by method provided by the invention, the aflatoxin B in distillers ' grains can be made1It significantly reduces, and increases vinasse Biological value.
To achieve the above object, the present invention uses following technical scheme：
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and is cultivated, completes culture, In：The condition of culture is：Cultivation temperature is 25~40 DEG C, and incubation time is 3~8d；
(2) fluid nutrient medium culture：
PD fluid nutrient mediums are cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in the training of PD liquid It supports in base, is cultivated, form straw mushroom culture solution, it is spare；Wherein：The condition of culture is：25~40 DEG C of cultivation temperature, training Support 3~8d of time, the straw mushroom culture solution spore concentration 105~107cfu/mL；
In the step 1 (2), the process for preparation of PD fluid nutrient mediums is：Peeled potatoes 250g, glucose 20g are taken, Distilled water 1000ml is mixed, and it is 6~7 to adjust pH, is configured to PD fluid nutrient mediums.
In the step 1 (2), PD fluid nutrient mediums first pass through sterilization treatment in advance, and sterilization process is：By PD Liquid Cultures Base is distributed into conical flask to the 2/3 of bottle volume, is sealed nozzle, is sterilized using high-pressure steam sterilizing pan, sterilizing Temperature is 121 DEG C, sterilization time 30min.
In the step 1 (2), PD fluid nutrient mediums are positioned in superclean bench platform and are cooled down, superclean bench Pre- to first pass through disinfection processing, processing procedure is：The alcohol swab impregnated with 75% alcohol cleans superclean bench table top, and passes through Ultraviolet sterilization 30min.
In the step 1 (2), fluid nutrient medium culture carries out in isothermal vibration incubator.
(3) straw mushroom liquid medium-selection in distillers ' grains：
Straw mushroom culture solution is taken, is added in distillers ' grains after carrying out solid state fermentation culture, mixing, the mixture after culture is taken, adds Enter distilled water mixing, form the mixture after dilution, supernatant laccase activity is surveyed after centrifugation, screening laccase activity is higher than 1800U/ Mixture after the dilution of L, and then the straw mushroom culture solution for meeting laccase activity requirement after the culture dilutes is filtered out, it is spare；
In the step 1 (3), distillers ' grains solid masses is (10~50) with straw mushroom culture solution liquid volume proportioning：1, Unit is g/mL.
In the step 1 (3), the additive amount of distilled water is the mixture solid quality after culture by proportioning：Distilled water Volume=1：(1~4), unit g/mL.
In the step 1 (3), centrifugal process is：The mixture after culture is taken, is placed in 50mL centrifuge tubes, by proportioning Add distillation water dissolution, after homogenizer mixing, centrifuged.
In the step 1 (3), centrifugal condition is：Rotating speed 4000rmin-1, centrifugation time 10min.
In the step 1 (3), laccase activity test method is ABTS methods.
In the step 1 (3), a laccase unit (U) is the amount that 1min converts 1 μm of required laccase of ol ABTS.
In the step 1 (3), the condition of culture after straw mushroom culture solution addition distillers ' grains is：Cultivation temperature is 25~35 DEG C, incubation time is 3~7d.
In the step 1 (3), distillers ' grains carry out sterilization treatment, the disinfecting action before straw mushroom culture solution is inoculated with It is carried out in high-pressure steam sterilizing pan, sterilising temp is 121 DEG C, sterilization time 60min.
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is taken to prepare Aspergillus flavus suspension, Aspergillus flavus suspension is added in into PD culture solutions, culture is extremely Spore concentration is 105~107Cfu/mL obtains aspergillus flavus culture solution；
In the step 2 (1), Aspergillus flavus suspension preparation process is：The malicious aspergillus flavus strain of production is inoculated in PDA inclined-planes On Tube propagation base, 3~7d is cultivated at 25~35 DEG C, after being activated to it, aseptic water washing prepares Aspergillus flavus suspension.
In the step 2 (1), the aspergillus flavus culture solution incubation carries out in isothermal vibration incubator, training Foster condition is：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm.
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Aspergillus flavus culture solution is added in into distillers ' grains, is cultivated, and by sterilization treatment, completion is inoculated with, wherein：It is described Distillers ' grains solid masses and aspergillus flavus culture solution liquid volume addition proportioning be (5~20)：1, unit g：ml；
In the step 2 (2), the condition of culture after distillers ' grains addition aspergillus flavus culture solution is：28 DEG C, 7d；Sterilize item Part is：121 DEG C, 60min.
In the step 2 (2), distillers ' grains are before malicious aspergillus flavus is produced in inoculation by sterilization treatment, the disinfecting action It is carried out in high-pressure steam sterilizing pan, sterilising temp is 121 DEG C, sterilization time 60min.
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete add cultivate ingredient thereto, after abundant mixing, the straw mushroom culture after inoculation screening Liquid mixing, is cultivated, and completes aflatoxin B in distillers ' grains1Degradation；Wherein, the medium component includes carbon source, Nitrogen source, inorganic salts and trace element, the distillers ' grains solid masses that the inoculation is completed and the straw mushroom culture solution liquid after screening Volume proportion is (10~50)：1, unit g/mL, the condition of culture are：Temperature is 25~35 DEG C, 8~20d of time.
In the step 3, the distillers ' grains that the inoculation is completed first pass through sterilization treatment in advance.
In the step 3, the condition of culture is preferably：Temperature is 28 DEG C, time 14d.
In the step 3, the distillers ' grains for being inoculated with completion are handled by pH and water content adjusting, and pH is adjusted to 4.5~7, Water content is adjusted to 55~70%.
In the step 3, the distillers ' grains pH that the inoculation is completed is adjusted to 6, and water content is adjusted to 65%.
In the step 3, the carbon source is glucose, soluble starch, and one kind in sucrose or maltose is described Carbon source addition quality for inoculation complete distillers ' grains quality 1~4%；
The nitrogen source be peptone, beef extract, KNO3, dusty yeast, NH4NO3In one kind, the nitrogen source addition matter Measure complete distillers ' grains quality for inoculation 1~2.5%；
The inorganic salts are KH2PO4, CaCl2Or MgSO4In one kind, inorganic salts addition quality is has been inoculated with Into the 0.05~0.35% of distillers ' grains quality；
The trace element is CuSO4, ZnSO4, MnSO4In one kind, described trace element addition quality is inoculation Complete 0.1~0.4mg/kg of distillers ' grains quality.
In the step 3, culture medium addition quality is preferably：
Carbon source addition quality completes the 3% of distillers ' grains quality for inoculation；
Nitrogen source addition quality completes the 1.5% of distillers ' grains quality for inoculation；
Inorganic salts addition quality completes the 0.15% of distillers ' grains quality for inoculation；
Trace element addition quality completes the 0.2mg/kg of distillers ' grains quality for inoculation.
Straw mushroom aflatoxin B in degradation of white spirit grain1The application in field.
Beneficial effects of the present invention：
Aflatoxin B in distillers ' grains provided by the invention1Biodegrading process, aspergillus flavus poison can be significantly reduced in distillers ' grains Plain B1Content, for reduce livestock and poultry to aflatoxin B1Intake, improve product potency, reduce economic loss and people and The health risk of animal is of great significance.
Description of the drawings：
Fig. 1 is 50 μ g/L AFB1Standard items HPLC collection of illustrative plates；
Fig. 2 is 20 μ g/L AFB1Standard items HPLC collection of illustrative plates；
Fig. 3 is 10 μ g/L AFB1Standard items HPLC collection of illustrative plates；
Fig. 4 is 1 μ g/L AFB1Standard items HPLC collection of illustrative plates；
Fig. 5 is 0.5 μ g/L AFB1Standard items HPLC collection of illustrative plates；
Fig. 6 is aflatoxin B in the distillers ' grains of embodiment 21Biodegrading process in be inoculated with AFB in the distillers ' grains of completion1's HPLC collection of illustrative plates；
Fig. 7 is aflatoxin B in the distillers ' grains of embodiment 21Biodegrading process in degrade after AFB in distillers ' grains1HPLC Collection of illustrative plates.
Specific embodiment：
With reference to embodiment, the present invention is described in further detail.
Following previous experiments are conventional method unless otherwise specified with the experimental method used in embodiment.
Material, reagent used etc. in following previous experiments and embodiment, unless otherwise specified, commercially It arrives.
Following previous experiments and aflatoxin B used in embodiment1(AFB1) standard items：It is purchased from SIGMA companies, 50 μ g/LAFB1Standard items HPLC collection of illustrative plates is as shown in Figure 1,20 μ g/L AFB1Standard items HPLC collection of illustrative plates is as shown in Fig. 2, 10 μ g/L AFB1 Standard items HPLC collection of illustrative plates is as shown in figure 3,1 μ g/L AFB1Standard items HPLC collection of illustrative plates is as shown in figure 4,0.5 μ g/L AFB1Standard items HPLC collection of illustrative plates is as shown in Figure 5；
Following previous experiments are with vinasse buying in embodiment in Fuxin San Gou breweries；
Following previous experiments and the AFB in Examples 1 to 81Content assaying method is as follows：
The distillers ' grains being inoculated with after aspergillus flavus and the distillers ' grains for completing aflatoxin degradation take about 20g in 60 DEG C of dry 12h Vinasse are crushed with pulverizer after drying, measure distillers ' grains moisture and AFB1Content, Toxic extraction processing method refer to GB/ T30955-2014。
Chromatographic condition：Chromatographic column is Venusil MP C18 (5 μm, 4.6mm × 250mm)；Column temperature is 40 DEG C；Mobile phase is Methanol:Water (V:V=53:47)；Flow velocity is 0.8mL/min；Using Post-column photochemical derivatization method：Photochemical derivatization device 254nm；With Fluorescence detector detects, excitation wavelength 360nm, launch wavelength 450nm, 20 μ L of sample size, calculates aflatoxin B1Content.
Previous experiments 1, straw mushroom Spawn incubation optimum condition determine：
Straw mushroom strain is taken to be inoculated in PDA plate culture medium to be cultivated, temperature is respectively set as 25 DEG C, 28 DEG C and 40 DEG C, the time is respectively set as 3d, 6d, 7d and 8d, is cultivated；Take peeled potatoes 250g, glucose 20g, distilled water 1000ml is mixed, and it is 6~7 to adjust pH, is configured to PD fluid nutrient mediums, PD fluid nutrient mediums are distributed into conical flask In at the 2/3 of bottle volume, seal nozzle, sterilized using high-pressure steam sterilizing pan, sterilising temp is 121 DEG C, sterilizing Time is 30min, and the alcohol swab impregnated with 75% alcohol cleans superclean bench table top, and through ultraviolet sterilization 30min, will go out PD fluid nutrient mediums after bacterium are positioned in superclean bench platform, are cooled to room temperature, by the straw mushroom Jing Guo solid medium culture Strain is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration incubator respectively at 25 DEG C, 28 DEG C and 40 DEG C, cultivates 3d, 6d, 7d and 8d form straw mushroom culture solution；
Distillers ' grains are taken, it is respectively 4.5,6 and 7 to adjust pH, and it is respectively 55%, 65% and 70% to adjust water content, inoculation grass Mushroom culture solution at 25 DEG C, 28 DEG C and 35 DEG C, cultivates 8d, 14d and 20d, completes culture, observes bacterium in straw mushroom culture solution respectively Silk upgrowth situation shows that the optimal culture condition of straw mushroom strain is：Solid culture temperature is 28 DEG C in PDA plate culture medium, training It is 6d to support the time, and Liquid Culture temperature is adjusted to 6 for 28 DEG C, incubation time 6d, distillers ' grains pH in PD fluid nutrient mediums, aqueous Amount is adjusted to 65%, and incubation time of the straw mushroom culture solution in distillers ' grains is 14d, when cultivation temperature is 28 DEG C, in straw mushroom strain Mycelial growth it is the most luxuriant；
Previous experiments 2, optimum carbon source, optimum nitrogen source, optimal trace element and corresponding addition quality determine：
(1) the malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, cultivates 3d at 28 DEG C, live to it After change, sterile water is rinsed, prepares Aspergillus flavus suspension, Aspergillus flavus suspension is added in into PD culture solutions, is put in isothermal vibration training It is carried out in foster case, 200rpm, 7d is cultivated at 30 DEG C, it is 10 to cultivate to spore concentration5Cfu/mL obtains aspergillus flavus culture solution；
(2) distillers ' grains are taken, after sterilized, add in aspergillus flavus culture solution, the two proportioning is bent with Huang for distillers ' grains solid masses Mould culture solution liquid volume addition proportioning is 10：1, unit g：Ml will cultivate 7d at 28 DEG C of distillers ' grains after inoculation aspergillus flavus, AFB is measured after sterilizing1Content；
(3) distillers ' grains water content is adjusted to 65%, pH and is adjusted to 6.0 after sterilizing, and adds glucose, soluble starch, sugarcane respectively Sugar, maltose are as unique additional carbon, after determining optimum carbon source, set 4 additive amounts, be respectively vinasse quality 1%, 2%th, 3%, 4%；Select dusty yeast, peptone, beef extract, KNO3、NH4NO3As unique additional nitrogen source, optimum nitrogen source is determined Afterwards, 4 additive amounts are set, are respectively 1%, 1.5%, 2%, the 2.5% of vinasse quality；It is separately added into CaCl2、MgSO4、 KH2PO4Three kinds of inorganic salts, after determining optimal inorganic salts, set 4 additive amounts, be respectively vinasse quality 0.05%, 0.15%, 0.25%th, 0.35%；Select zinc (ZnSO4), manganese (MnSO4), copper (CuSO4) three kinds of additional trace elements, it determines optimal additional micro- After secondary element, 4 additive amounts are set, is respectively 0.1mg/kg, 0.2mg/kg, 0.3mg/kg, 0.4mg/kg of vinasse quality, fills Straw mushroom culture solution mixing is inoculated with after point mixing respectively, 28 DEG C of culture 14d measure AFB after sterilizing1Content, calculate degradation rate, really Determine optimum carbon source, nitrogen source, inorganic salts and trace element and corresponding optimum additive amount.
Experimental result：
Under different condition of culture, straw mushroom is measured to aflatoxin B1Degradation rate find, optimum carbon source is grape Sugar can make straw mushroom to AFB1Degradation rate be 82.07%；Optimum nitrogen source is peptone, degradation rate 80.94%；It is optimal inorganic Salt is KH2PO4, degradation rate 79.35%；Optimal trace element is copper (CuSO4), degradation rate 77.16%, specific experiment number According to as shown in the following table 1~4；
1 carbon source of table is to AFB1The influence of degradation rate
2 nitrogen source of table is to AFB1The influence of degradation rate
3 inorganic salts of table are to AFB1The influence of degradation rate
4 trace element of table is to AFB1The influence of degradation rate
Optimization obtains glucose and most preferably adds quality completes distillers ' grains quality for inoculation 3%, and straw mushroom is to AFB at this time1's Average degradation rate is up to 82.06%；The optimal addition quality of peptone completes the 1.5% of distillers ' grains quality for inoculation, drops at this time Solution rate is 85.74%；KH2PO4Optimal addition quality the 0.15% of distillers ' grains quality is completed for inoculation, degradation rate is at this time 80.08%；Add the CuSO that quality is 0.2mg/kg4Most beneficial for straw mushroom to AFB1Degradation, degradation rate 77.05%, finally Optimal conditions be：3% glucose, 1.5% peptone, 0.15%KH2PO4, the CuSO of 0.2mg/kg428 DEG C, pH 6, training When supporting 28 DEG C of temperature, incubation time 14 days, straw mushroom can be by 53.52 μ gkg in distillers ' grains-1AFB1It is down to 7.48 μ gkg-1, drop Solution rate is 86.02%, and specific experiment data are as shown in the following table 5~8；
Table 5 adds the glucose of different quality to AFB1The influence of degradation rate
Table 6 adds the peptone of different quality to AFB1The influence of degradation rate
Table 7 adds the KH of different quality2PO4To AFB1The influence of degradation rate
Table 8 adds the CuSO of different quality4To AFB1The influence of degradation rate
Example 1 below~8 are carried out on the basis of the previous experiments：
Embodiment 1
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 8d at 25 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 6 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 8d is cultivated in incubator at 25 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 106Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 10：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 25 DEG C After cultivating 7d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 2.5mL added to distill water dissolution, with homogenate Device mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant paint Enzymatic activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase after the culture dilutes The straw mushroom culture solution of Active pharmaceutical, it is spare, wherein, a laccase unit (U) is that 1min converts 1 μm of required paint of ol ABTS The amount of enzyme；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 7d is cultivated at 25 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration6Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 5 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml cultivates 7d at 28 DEG C, And pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 91.03 μ g·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 4.5 to adjust pH, and water content is adjusted to 70%, adds grape thereto Sugar completes the 1% of distillers ' grains quality for inoculation as carbon source, addition quality；Peptone is to be inoculated with as nitrogen source, addition quality Into the 1% of distillers ' grains quality；KH2PO4The 0.05% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality；CuSO4Make For trace element, addition quality is to be inoculated with the 0.1mg/kg for completing distillers ' grains quality, and after abundant mixing, by proportioning, inoculation is completed Distillers ' grains solid masses with the straw mushroom culture solution liquid volume proportioning after screening for 10：1, unit g/mL, after inoculation screening Straw mushroom culture solution mixing, at 25 DEG C, cultivate 20d, complete distillers ' grains in aflatoxin B1Degradation, after measured, degradation AFB in distillers ' grains afterwards1Content is 46.32 μ gkg-1, aflatoxin B1Degradation rate is 49.12%.
Embodiment 2
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 6d at 28 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 6 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 6d is cultivated in incubator at 28 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 105Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 30：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 28 DEG C After cultivating 7d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 10mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 3d is cultivated at 28 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration6Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 10 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 53.52 μg·kg-1, it is inoculated with AFB in the distillers ' grains of completion1HPLC collection of illustrative plates it is as shown in Figure 6；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 6 to adjust pH, and water content is adjusted to 65%, adds glucose thereto As carbon source, addition quality completes the 3% of distillers ' grains quality for inoculation；Peptone is completed as nitrogen source, addition quality for inoculation The 1.5% of distillers ' grains quality；KH2PO4The 0.15% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality；CuSO4Make For trace element, addition quality is to be inoculated with the 0.2mg/kg for completing distillers ' grains quality, and after abundant mixing, by proportioning, inoculation is completed Distillers ' grains solid masses with the straw mushroom culture solution liquid volume proportioning after screening for 30：1, unit g/mL, after inoculation screening Straw mushroom culture solution mixing, at 28 DEG C, cultivate 14d, complete distillers ' grains in aflatoxin B1Degradation, distillers ' grains after degradation Middle AFB1HPLC collection of illustrative plates as shown in fig. 7, after measured, AFB in distillers ' grains after degradation1Content is 7.48 μ gkg-1, aspergillus flavus poison Plain B1Degradation rate is 86.02%.
Embodiment 3
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 5d at 28 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 6 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 7d is cultivated in incubator at 28 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 105Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 30：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 28 DEG C After cultivating 7d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 10mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 3d is cultivated at 28 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration5Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 10 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 53.52 μg·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 6 to adjust pH, and water content is adjusted to 65%, adds glucose thereto As carbon source, addition quality completes the 2% of distillers ' grains quality for inoculation；Peptone is completed as nitrogen source, addition quality for inoculation The 2% of distillers ' grains quality；KH2PO4The 0.25% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality；CuSO4As Trace element, addition quality are to be inoculated with the 0.3mg/kg for completing distillers ' grains quality, after abundant mixing, by proportioning, are inoculated with completion Distillers ' grains solid masses is 30 with the straw mushroom culture solution liquid volume proportioning after screening：1, unit g/mL, after inoculation screening Straw mushroom culture solution mixing at 28 DEG C, cultivates 14d, completes aflatoxin B in distillers ' grains1Degradation, after measured, after degradation AFB in distillers ' grains1Content is 10.07 μ gkg-1, aflatoxin B1Degradation rate is 81.18%.
Embodiment 4
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 6d at 28 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 6 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 6d is cultivated in incubator at 28 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 105Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 30：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 28 DEG C After cultivating 7d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 10mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 3d is cultivated at 28 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration5Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 10 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 53.52 μg·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 6 to adjust pH, and water content is adjusted to 65%, adds glucose thereto As carbon source, addition quality completes the 4% of distillers ' grains quality for inoculation；Peptone is completed as nitrogen source, addition quality for inoculation The 2.5% of distillers ' grains quality；KH2PO4The 0.35% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality；CuSO4Make For trace element, addition quality is to be inoculated with the 0.4mg/kg for completing distillers ' grains quality, and after abundant mixing, by proportioning, inoculation is completed Distillers ' grains solid masses with the straw mushroom culture solution liquid volume proportioning after screening for 30：1, unit g/mL, after inoculation screening Straw mushroom culture solution mixing, at 28 DEG C, cultivate 14d, complete distillers ' grains in aflatoxin B1Degradation, after measured, degradation AFB in distillers ' grains afterwards1Content is 10.55 μ gkg-1, aspertoxin B1Degradation rate is 80.29%.
Embodiment 5
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 6d at 28 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 7 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 6d is cultivated in incubator at 28 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 105Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 30：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 28 DEG C After cultivating 7d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 5mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 3d is cultivated at 28 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration5Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 10 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 53.52 μg·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 6 to adjust pH, and water content is adjusted to 65%, adds maltose thereto As carbon source, addition quality completes the 3% of distillers ' grains quality for inoculation；KNO3As nitrogen source, addition quality is completed white for inoculation The 1.5% of vinasse quality；MgSO4·7H2O completes the 0.15% of distillers ' grains quality for inoculation as inorganic salts addition quality； ZnSO4As trace element, addition quality is to be inoculated with the 0.2mg/kg for completing distillers ' grains quality, after abundant mixing, by proportioning, is connect The distillers ' grains solid masses that kind is completed is 30 with the straw mushroom culture solution liquid volume proportioning after screening：1, unit g/mL, inoculation Straw mushroom culture solution mixing after screening at 28 DEG C, cultivates 14d, completes aflatoxin B in distillers ' grains1Degradation, through survey It is fixed, AFB in distillers ' grains after degradation1Content is 29.22 μ gkg-1, aflatoxin B1Degradation rate is 45.40%.
Embodiment 6
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 6d at 28 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 7 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 6d is cultivated in incubator at 28 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 105Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 30：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 28 DEG C 7d is cultivated, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 7.5mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 3d is cultivated at 28 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration5Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 10 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 53.52 μg·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 6 to adjust pH, and water content is adjusted to 65%, adds sucrose thereto and makees For carbon source, addition quality completes the 3% of distillers ' grains quality for inoculation；Beef extract is completed white as nitrogen source, addition quality for inoculation The 1.5% of vinasse quality；CaCl2The 0.15% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality；MnSO4As Trace element, addition quality are to be inoculated with the 0.2mg/kg for completing distillers ' grains quality, after abundant mixing, by proportioning, are inoculated with completion Distillers ' grains solid masses is 30 with the straw mushroom culture solution liquid volume proportioning after screening：1, unit g/mL, after inoculation screening Straw mushroom culture solution mixing at 28 DEG C, cultivates 14d, completes aflatoxin B in distillers ' grains1Degradation, after measured, after degradation AFB in distillers ' grains1Content is 24.28 μ gkg-1, aflatoxin B1Degradation rate is 54.63%.
Embodiment 7
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 6d at 28 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 7 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 6d is cultivated in incubator at 28 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 105Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 30：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 28 DEG C After cultivating 7d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 10mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 3d is cultivated at 28 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration5Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 10 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 53.52 μg·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 6 to adjust pH, and water content is adjusted to 65%, and addition is soluble thereto Starch completes the 3% of distillers ' grains quality for inoculation as carbon source, addition quality；For dusty yeast as nitrogen source, addition quality is inoculation Complete the 1.5% of distillers ' grains quality；KH2PO4The 0.15% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality； CuSO4As trace element, addition quality is to be inoculated with the 0.2mg/kg for completing distillers ' grains quality, after abundant mixing, by proportioning, is connect The distillers ' grains solid masses that kind is completed is 30 with the straw mushroom culture solution liquid volume proportioning after screening：1, unit g/mL, inoculation Straw mushroom culture solution mixing after screening at 28 DEG C, cultivates 14d, completes aflatoxin B in distillers ' grains1Degradation, through survey It is fixed, AFB in distillers ' grains after degradation1Content is 15.63 μ gkg-1, aflatoxin B1Degradation rate is 70.79%.
Embodiment 8
Aflatoxin B in a kind of distillers ' grains1Biodegrading process, include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and cultivates 3d at 40 DEG C, is completed Culture；
(2) fluid nutrient medium culture：
Peeled potatoes 250g, glucose 20g are taken, distilled water 1000ml is mixed, and it is 7 to adjust pH, is configured to PD PD fluid nutrient mediums are distributed into conical flask to the 2/3 of bottle volume, nozzle are sealed, using high pressure by fluid nutrient medium Steam sterilization pan sterilizes, and sterilising temp is 121 DEG C, sterilization time 30min, and the alcohol swab impregnated with 75% alcohol is cleaned Superclean bench table top, and through ultraviolet sterilization 30min, the PD fluid nutrient mediums after sterilizing are positioned over superclean bench platform It is interior, it is cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums, is placed in isothermal vibration 3d is cultivated in incubator at 40 DEG C, forms straw mushroom culture solution, straw mushroom culture solution spore concentration 107Cfu/mL, it is spare；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Distillers ' grains are 121 DEG C in high-pressure steam sterilizing pan, after the 60min that sterilizes, by proportioning, and distillers ' grains solid masses and straw mushroom Culture solution liquid volume ratio is 50：1, unit g/mL take straw mushroom culture solution, add in distillers ' grains and carry out solid state fermentation at 35 DEG C After cultivating 3d, mixing takes the mixture after 2.5g cultures, is placed in 50mL centrifuge tubes, 10mL is added to distill water dissolution, uses homogenizer Mixing forms the mixture after dilution, is placed in 4000rmin-1Under the conditions of centrifuge 10min, using ABTS methods survey supernatant laccase Activity, the mixture after dilution of the screening laccase activity higher than 1800U/L, and then filter out and meet laccase work after the culture dilutes Property requirement straw mushroom culture solution, it is spare, wherein, a laccase unit (U) be 1min convert 1 μm of required laccase of ol ABTS Amount；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is inoculated on PDA slant tube culture mediums, 7d is cultivated at 25 DEG C, it is activated Afterwards, 4mL sterile waters are added on slant tube culture medium, rinsed, prepared Aspergillus flavus suspension, added in into 150mL triangular flasks The PD culture solutions of 50mL add in Aspergillus flavus suspension into PD culture solutions, are put in isothermal vibration incubator and carry out, condition of culture For：30 DEG C, incubation time 7d of cultivation temperature, concussion revolution are 200rpm, and it is 10 to cultivate to spore concentration7Cfu/mL is obtained yellow Aspergillus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Distillers ' grains are sterilized in high-pressure steam sterilizing pan, at 121 DEG C after 60min, and aspergillus flavus culture is added in into distillers ' grains Liquid, distillers ' grains solid masses are 20 with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：Ml, in 28 DEG C of cultures 7d, and pass through 121 DEG C, the sterilization treatment of 60min completes inoculation, measures AFB in the distillers ' grains that inoculation is completed1Content is 46.59 μg·kg-1；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete, after sterilizing, it is 7 to adjust pH, and water content is adjusted to 55%, adds glucose thereto As carbon source, addition quality completes the 3% of distillers ' grains quality for inoculation；NH4NO3As nitrogen source, addition quality is completed white for inoculation The 1.5% of vinasse quality；KH2PO4The 0.15% of distillers ' grains quality is completed for inoculation as inorganic salts addition quality；CuSO4As Trace element, addition quality are to be inoculated with the 0.2mg/kg for completing distillers ' grains quality, after abundant mixing, by proportioning, are inoculated with completion Distillers ' grains solid masses is 50 with the straw mushroom culture solution liquid volume proportioning after screening：1, unit g/mL, after inoculation screening Straw mushroom culture solution mixing at 35 DEG C, cultivates 8d, completes aflatoxin B in distillers ' grains1Degradation, it is after measured, white after degradation AFB in vinasse1Content is 16.88 μ gkg-1, aflatoxin B1Degradation rate is 63.76%.
Finally, it should also be noted that it is listed above be only the present invention several specific embodiments.Obviously, this hair It is bright to be not limited to above example, there can also be many variations, if the straw mushroom using other kinds is to distillers ' grains and other vinasse Degradation.The above all of deformation in this field falls within protection scope of the present invention.

Claims (10)

1. a kind of aflatoxin B in distillers ' grains1Biodegrading process, which is characterized in that include the following steps：
Step 1, straw mushroom Spawn incubation
(1) solid medium culture：Straw mushroom strain is inoculated in PDA plate culture medium and is cultivated, completes culture, wherein： The condition of culture is：Cultivation temperature is 25~40 DEG C, and incubation time is 3~8d；
(2) fluid nutrient medium culture：
PD fluid nutrient mediums are cooled to room temperature, the straw mushroom strain Jing Guo solid medium culture is inoculated in PD fluid nutrient mediums It is interior, it is cultivated, forms straw mushroom culture solution, it is spare；Wherein：The condition of culture is：25~40 DEG C of cultivation temperature, during culture Between 3~8d, the straw mushroom culture solution spore concentration 105~107cfu/mL；
(3) straw mushroom liquid medium-selection in distillers ' grains：
Straw mushroom culture solution is taken, is added in distillers ' grains after carrying out solid state fermentation culture, mixing, takes the mixture after culture, adds in and steams Distilled water mixing forms the mixture after dilution, supernatant laccase activity is surveyed after centrifugation, screening laccase activity is higher than 1800U/L's Mixture after dilution, and then the straw mushroom culture solution for meeting laccase activity requirement after the culture dilutes is filtered out, it is spare；
Step 2, the malicious aspergillus flavus of distillers ' grains inoculation production
(1) malicious aspergillus flavus actication of culture and culture are produced：
The malicious aspergillus flavus strain of production is taken to prepare Aspergillus flavus suspension, Aspergillus flavus suspension is added in into PD culture solutions, is cultivated to spore Concentration is 105~107Cfu/mL obtains aspergillus flavus culture solution；
(2) the malicious aspergillus flavus of distillers ' grains inoculation production：
Aspergillus flavus culture solution is added in into distillers ' grains, is cultivated, and by sterilization treatment, completion is inoculated with, wherein：Described is white Vinasse solid masses is (5~20) with aspergillus flavus culture solution liquid volume addition proportioning：1, unit g：ml；
Step 3, aflatoxin B in distillers ' grains1Degradation
The distillers ' grains that inoculation is taken to complete add cultivate ingredient thereto, and after abundant mixing, the straw mushroom culture solution after inoculation screening mixes It is even, it is cultivated, completes aflatoxin B in distillers ' grains1Degradation；Wherein, the medium component includes carbon source, nitrogen Source, inorganic salts and trace element, the distillers ' grains solid masses that the inoculation is completed and the straw mushroom culture solution liquid bulk after screening Product proportioning is (10~50)：1, unit g/mL, the condition of culture are：Temperature is 25~35 DEG C, 8~20d of time.
2. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 1 (3) in, distillers ' grains solid masses is (10~50) with straw mushroom culture solution liquid volume proportioning：1, unit g/mL.
3. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 1 (3) in, the condition of culture after straw mushroom culture solution addition distillers ' grains is：Cultivation temperature is 25~35 DEG C, and incubation time is 3~7d.
4. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 2 (1) in, the aspergillus flavus culture solution incubation carries out in isothermal vibration incubator, and condition of culture is：Cultivation temperature 30 DEG C, incubation time 7d, concussion revolution is 200rpm.
5. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 2 (2) in, the condition of culture after distillers ' grains addition aspergillus flavus culture solution is：28 DEG C, 7d.
6. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 3 In, the condition of culture is：Temperature is 28 DEG C, time 14d.
7. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 3 In, it being inoculated with the distillers ' grains of completion and is handled by pH and water content adjusting, pH is adjusted to 4.5~7, and water content is adjusted to 55~ 70%.
8. aflatoxin B in distillers ' grains according to claim 71Biodegrading process, which is characterized in that the step 3 In, the distillers ' grains pH that the inoculation is completed is adjusted to 6, and water content is adjusted to 65%.
9. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step 3 In, the carbon source is glucose, soluble starch, one kind in sucrose or maltose, and carbon source addition quality is connects Kind completes the 1~4% of distillers ' grains quality；
The nitrogen source be peptone, beef extract, KNO3, dusty yeast, NH4NO3In one kind, the nitrogen source addition quality be The 1~2.5% of distillers ' grains quality is completed in inoculation；
The inorganic salts are KH2PO4, CaCl2Or MgSO4In one kind, inorganic salts addition quality is completed white for inoculation The 0.05~0.35% of vinasse quality；
The trace element is CuSO4, ZnSO4, MnSO4In one kind, described trace element addition quality completes for inoculation 0.1~0.4mg/kg of distillers ' grains quality.
10. aflatoxin B in distillers ' grains according to claim 11Biodegrading process, which is characterized in that the step In 3, culture medium addition quality is：
Carbon source addition quality completes the 3% of distillers ' grains quality for inoculation；
Nitrogen source addition quality completes the 1.5% of distillers ' grains quality for inoculation；
Inorganic salts addition quality completes the 0.15% of distillers ' grains quality for inoculation；
Trace element addition quality completes the 0.2mg/kg of distillers ' grains quality for inoculation.
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108991227A (en) * 2018-08-02 2018-12-14 合肥工业大学 A kind of grain in storage for years fermentation liquid and preparation method thereof removing aflatoxin

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103215191A (en) * 2012-01-19 2013-07-24 中国农业科学院农产品加工研究所 Pleurotus ostreatus and its application in aflatoxin degradation
CN104824493A (en) * 2015-05-29 2015-08-12 东北农业大学 Degradation method of aflatoxin B1 using pleurotus citrinopileatus or shiitake mushroom
CN104988183A (en) * 2015-08-06 2015-10-21 河北省微生物研究所 Preparation method of pleurotus ostreatus fermentation broth and application of fermentation broth in degradation of aflatoxin B1

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103215191A (en) * 2012-01-19 2013-07-24 中国农业科学院农产品加工研究所 Pleurotus ostreatus and its application in aflatoxin degradation
CN104824493A (en) * 2015-05-29 2015-08-12 东北农业大学 Degradation method of aflatoxin B1 using pleurotus citrinopileatus or shiitake mushroom
CN104988183A (en) * 2015-08-06 2015-10-21 河北省微生物研究所 Preparation method of pleurotus ostreatus fermentation broth and application of fermentation broth in degradation of aflatoxin B1

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
DAPENG BAO，ETL: "Sequencing and Comparative Analysis of the Straw Mushroom (Volvariella volvacea) Genome", 《PLOS ONE》 *
周露 等: "平菇P1培养条件优化及其黄曲霉毒素降解酶的初步分离", 《核农学报》 *
王会娟: "平菇降解黄曲霉毒素的研究", 《中国优秀硕士学位论文全文数据库 农业科技辑》 *
王海云: "降解白酒糟中黄曲霉毒素B1的菌株的筛选及发酵工艺的研究", 《中国优秀硕士学位论文全文数据库 工程科技Ⅰ辑》 *
雷德柱 等: "产漆酶的草菇菌株的筛选", 《工业微生物》 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108991227A (en) * 2018-08-02 2018-12-14 合肥工业大学 A kind of grain in storage for years fermentation liquid and preparation method thereof removing aflatoxin

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