CN108251458B - A kind of degradation agent of aflatoxin, preparation method and application - Google Patents
A kind of degradation agent of aflatoxin, preparation method and application Download PDFInfo
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- CN108251458B CN108251458B CN201810071264.4A CN201810071264A CN108251458B CN 108251458 B CN108251458 B CN 108251458B CN 201810071264 A CN201810071264 A CN 201810071264A CN 108251458 B CN108251458 B CN 108251458B
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- 229930195730 Aflatoxin Natural products 0.000 title claims abstract description 74
- XWIYFDMXXLINPU-UHFFFAOYSA-N Aflatoxin G Chemical compound O=C1OCCC2=C1C(=O)OC1=C2C(OC)=CC2=C1C1C=COC1O2 XWIYFDMXXLINPU-UHFFFAOYSA-N 0.000 title claims abstract description 74
- 239000005409 aflatoxin Substances 0.000 title claims abstract description 74
- 230000015556 catabolic process Effects 0.000 title claims abstract description 71
- 238000006731 degradation reaction Methods 0.000 title claims abstract description 71
- 239000003795 chemical substances by application Substances 0.000 title claims abstract description 50
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 238000000855 fermentation Methods 0.000 claims abstract description 30
- 230000004151 fermentation Effects 0.000 claims abstract description 30
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- 235000011399 aloe vera Nutrition 0.000 claims abstract description 23
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- 239000007788 liquid Substances 0.000 claims abstract description 19
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- 238000000034 method Methods 0.000 claims abstract description 15
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- 238000003756 stirring Methods 0.000 claims description 10
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- 239000012467 final product Substances 0.000 claims description 7
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- 239000008103 glucose Substances 0.000 claims description 6
- 229910017053 inorganic salt Inorganic materials 0.000 claims description 6
- 239000012266 salt solution Substances 0.000 claims description 6
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- 229910021580 Cobalt(II) chloride Inorganic materials 0.000 claims description 5
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- 229910052921 ammonium sulfate Inorganic materials 0.000 claims description 5
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- 229910000359 iron(II) sulfate Inorganic materials 0.000 claims description 5
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- CSNNHWWHGAXBCP-UHFFFAOYSA-L magnesium sulphate Substances [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 5
- 229910000357 manganese(II) sulfate Inorganic materials 0.000 claims description 5
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- PQMOXTJVIYEOQL-UHFFFAOYSA-N Cumarin Natural products CC(C)=CCC1=C(O)C(C(=O)C(C)CC)=C(O)C2=C1OC(=O)C=C2CCC PQMOXTJVIYEOQL-UHFFFAOYSA-N 0.000 description 2
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- 230000007423 decrease Effects 0.000 description 2
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- 125000004852 dihydrofuranyl group Chemical class O1C(CC=C1)* 0.000 description 2
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- C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
- C12P1/02—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using fungi
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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Abstract
The invention discloses a kind of degradation agent of aflatoxin, preparation method and applications, belong to aflatoxin degradation technique field.Aflatoxin degradation agent of the invention is prepared by fermentation liquid and 1% peppermint oil, 0.1~0.5% laccase, 0.2% Tween-80 of the fresh aloe of 60~100g, 75~125g lemon.It is mutually cooperateed between each component of the present invention, makes the fully degraded of aflatoxin.Aflatoxin degradation agent of the present invention is safe and efficient to the degradation of aflatoxin, reaction is mild, can degrade 96% or more aflatoxin.
Description
Technical field
The invention belongs to aflatoxin degradation technique fields, and in particular to a kind of degradation agent of aflatoxin, its system
Preparation Method and application.
Background technique
Aflatoxin with its toxicity, carcinogenicity, teratogenesis, mutagenicity title, it is true by aspergillus flavus, aspergillus parasiticus etc.
The secondary metabolite that bacterium generates, is in the nature the derivative of dihydrofuran cumarin, the source of carcinogenic nature is cumarin
Mechanism, toxicity source are dihydrofuran ring structures.Aflatoxin is seriously polluted in grain, feed, to the mankind and domestic animal
Health threat is very big.The mankind, which eat the food containing aflatoxin by mistake, can lead to Acute Aflatoxicosis, and liver by
To damage, or even cause liver cancer.The feed that domestic animal eats aflatoxin contamination can lead to feed feed intake, production performance and exempt from
The reduction of epidemic disease function.
Currently, mainly having physical method, chemical method and biological detoxication method for the poison-removing method of aflatoxin.However, by
In that there are solvent extractions is at high cost, extraction is not thorough, and effect stability is poor, safety for each method the problems such as, it is difficult to be widely applied.
Summary of the invention
Aiming at the problems existing in the prior art, it is an object of the present invention to provide a kind of safe and efficient aflatoxin to drop
Solve agent.
In order to achieve the above object, the technical solution of the present invention is as follows:
A kind of aflatoxin degradation agent is thin by the fresh aloe of 60~100g, the fermentation liquid of 75~125g lemon and 1%
Lotus oil, 0.1~0.5% laccase, 0.2% Tween-80 are prepared.
It is by the fresh aloe of 60g, the fermentation liquid of 75g lemon and 1% peppermint oil, 0.5% paint on the basis of above scheme
Enzyme, 0.2% Tween-80 are prepared.
It is by the fermentation liquid and 1% peppermint oil, 0.3% of the fresh aloe of 80g, 100g lemon on the basis of above scheme
Laccase, 0.2% Tween-80 are prepared.
It is by the fermentation liquid and 1% peppermint oil, 0.1% of the fresh aloe of 100g, 125g lemon on the basis of above scheme
Laccase, 0.2% Tween-80 are prepared.
On the basis of above scheme, the fresh aloe, lemon fermentation liquid the preparation method comprises the following steps: fresh aloe, lemon
Juicing, is added the fermentation of bacillus amyloliquefaciens bacterium solution after juice ground-slag is broken, juice is added the fermentation of S. cervisiae bacterium solution, merges fermentation
Liquid to obtain the final product.
On the basis of above scheme, bacillus amyloliquefaciens bacterial concentration >=1010cfu/mL；The saccharomyces cerevisiae
Bacterium bacterial concentration >=108cfu/mL。
On the basis of above scheme, the preparation method of the degradation agent of the aflatoxin, step are as follows:
(1) fresh aloe, lemon, juicing, filtering；Juice ground-slag is broken to 40 mesh, adds 1.5 times of weight inorganic salt solutions, stirring
Uniformly；1.5% glucose is added in juice, stirs evenly；Juice slag and juice are cooled to room temperature in 100 DEG C of steam sterilizing 15min；
(2) bacillus amyloliquefaciens bacterium solution is added into juice slag after cooling, reaches bacillus amyloliquefaciens concentration
109Cfu/mL is placed in 37 DEG C of culture 48h；
(3) saccharomyces cerevisiae bacterium solution is added into juice after cooling, saccharomyces cerevisiae bacteria concentration is made to reach 107Cfu/mL, 28
DEG C, 150rpm, ferment 48h；
(4) by after fermentation juice slag filter waste, will filtrate be added fermentation after juice in, add peppermint oil, laccase,
Tween-80, after mixing to obtain the final product.
On the basis of above scheme, the inorganic salt solution is (g/L): (NH4)2SO42.0、KH2PO42.0、
MgSO40.3、NaCl 0.5、FeSO40.005、MnSO40.016、ZnCl20.017、CoCl20.002。
Application of the aflatoxin degradation agent of above method preparation in aflatoxin degradation.
A kind of aflatoxin biodegrading process, the degradation agent prepared using the above method, step are as follows: in wait sample of degrading
The degradation agent of above method preparation is added, after mixing in 28 DEG C of processing 2.5h, 100 DEG C of destroy the enzyme treatment 15min.
The laccase is purchased from Xia Sheng Industry Group Co., Ltd；
Peppermint oil is purchased from Jishui County poly- source hall natural perfume material oil Co., Ltd；
The bacillus amyloliquefaciens are purchased from Shandong Jun De Biotechnology Co., Ltd；
The saccharomyces cerevisiae is purchased from Shandong Kang Qin Biotechnology Co., Ltd.
Beneficial effects of the present invention
Aflatoxin degradation agent of the present invention is safe and efficient to the degradation of aflatoxin, reaction is mild, can degrade
96% or more aflatoxin.
Containing there are many effective active composition, the effective aflatoxin degradations of energy in aloe and lemon of the invention；Xie Dian
Sorodium spore bar and fermentation by saccharomyces cerevisiae can not only be such that the effective component in aloe and lemon adequately releases, they
Metabolite and a variety of enzymatic activity ingredients also have degradation to aflatoxin；Laccase itself has aflatoxin degradation
Effect, the fermentation liquid synergistic effect of aloe and lemon makes the fully degraded of aflatoxin.
Detailed description of the invention
Degradation of Fig. 1 difference aflatoxin degradation agent to aflatoxin；
Degradation of Fig. 2 aflatoxin degradation agent to different amounts of aflatoxin.
Specific embodiment
Term as used in the present invention generally has those of ordinary skill in the art usual unless otherwise specified
The meaning of understanding.
Combined with specific embodiments below, and referring to the data further detailed description present invention.Following embodiment only be
It illustrates the present invention, rather than limits the scope of the invention in any way.
Embodiment 1
Actication of culture
Bacillus amyloliquefaciens are inoculated in activation medium to cultivate, 30 DEG C of cultivation temperature, the time is for 24 hours；It will activation
Good strain is inoculated in fermentor, and 30 DEG C of temperature of control, 180rpm cultivate 18h, and thalline were collected by centrifugation, adds sterile water that bacterium is resuspended
Body makes concentration reach 1010cfu/mL。
Above-mentioned activation and fermentation medium are NB culture medium (g/L): beef extract 5.0, peptone 10.0, sodium chloride 5.0,
pH7.0。
The bacillus amyloliquefaciens are purchased from Shandong Jun De Biotechnology Co., Ltd.
Saccharomyces cerevisiae is inoculated in activation medium to cultivate, 28 DEG C, cultivates 20h；Activated strain is inoculated in
In fermentor, 28 DEG C, 150rpm, ferment 48h, and thalline were collected by centrifugation, adds sterile water that thallus is resuspended, concentration is made to reach 108cfu/
mL。
Above-mentioned activation and fermentation medium are YPD culture medium (g/L): glucose 20, peptone 20, yeast powder 10,
pH6.5。
The saccharomyces cerevisiae is purchased from Shandong Kang Qin Biotechnology Co., Ltd.
Embodiment 2
A kind of degradation agent of aflatoxin is by the peppermint of the fresh aloe of 60g, 75g lemon fermentation liquid and 1% weight
Oil, the laccase of 0.5% weight, 0.2% weight Tween-80 be prepared.
(1) fresh aloe 60g, lemon 75g, juicing, filtering；Juice ground-slag is broken to 40 mesh, adds 1.5 times of weight inorganic salts molten
Liquid stirs evenly；1.5% glucose is added in juice, stirs evenly；Juice slag and juice are cooling in 100 DEG C of steam sterilizing 15min
To room temperature；
The inorganic salt solution (g/L): (NH4)2SO42.0、KH2PO42.0、MgSO40.3、NaCl 0.5、
FeSO40.005、MnSO40.016、ZnCl20.017、CoCl20.002。
(2) into juice slag after cooling be added bacillus amyloliquefaciens bacterium solution, make bacillus amyloliquefaciens concentration >=
109Cfu/mL is placed in 37 DEG C of culture 48h；
(3) saccharomyces cerevisiae bacterium solution is added into juice after cooling, makes saccharomyces cerevisiae bacteria concentration >=107Cfu/mL, 28 DEG C,
150rpm, ferment 48h；
(4) by after fermentation juice slag filter waste, will filtrate be added fermentation after juice in, add 1% peppermint oil,
0.5% laccase, 0.2% Tween-80, after mixing to obtain the final product.
Embodiment 3
A kind of degradation agent of aflatoxin, the fresh aloe of 80g, 100g lemon fermentation liquid and 1% weight peppermint oil,
The laccase of 0.3% weight, 0.2% weight Tween-80 be prepared.
(1) fresh aloe 80g, lemon 100g, juicing, filtering；Juice ground-slag is broken to 40 mesh, adds 1.5 times of weight inorganic salts molten
Liquid stirs evenly；1.5% glucose is added in juice, stirs evenly；Juice slag and juice are cooling in 100 DEG C of steam sterilizing 15min
To room temperature；
The inorganic salt solution (g/L): (NH4)2SO42.0、KH2PO42.0、MgSO40.3、NaCl 0.5、
FeSO40.005、MnSO40.016、ZnCl20.017、CoCl20.002。
(2) into juice slag after cooling be added bacillus amyloliquefaciens bacterium solution, make bacillus amyloliquefaciens concentration >=
109Cfu/mL is placed in 37 DEG C of culture 48h；
(3) saccharomyces cerevisiae bacterium solution is added into juice after cooling, makes saccharomyces cerevisiae bacteria concentration >=107Cfu/mL, 28 DEG C,
150rpm, ferment 48h；
(4) by after fermentation juice slag filter waste, will filtrate be added fermentation after juice in, add 1% peppermint oil,
0.3% laccase, 0.2% Tween-80, after mixing to obtain the final product.
Embodiment 4
A kind of degradation agent of aflatoxin, the fresh aloe of 100g, 125g lemon fermentation liquid and 1% weight peppermint oil,
The laccase of 0.1% weight, 0.2% weight Tween-80 be prepared.
(1) fresh aloe 100g, lemon 125g, juicing, filtering；Juice ground-slag is broken to 40 mesh, adds 1.5 times of weight inorganic salts molten
Liquid stirs evenly；1.5% glucose is added in juice, stirs evenly；Juice slag and juice are cooling in 100 DEG C of steam sterilizing 15min
To room temperature；
The inorganic salt solution (g/L): (NH4)2SO42.0、KH2PO42.0、MgSO40.3、NaCl 0.5、
FeSO40.005、MnSO40.016、ZnCl20.017、CoCl20.002。
(2) into juice slag after cooling be added bacillus amyloliquefaciens bacterium solution, make bacillus amyloliquefaciens concentration >=
109Cfu/mL is placed in 37 DEG C of culture 48h；
(3) saccharomyces cerevisiae bacterium solution is added into juice after cooling, makes saccharomyces cerevisiae bacteria concentration >=107Cfu/mL, 28 DEG C,
150rpm, ferment 48h；
(4) by after fermentation juice slag filter waste, will filtrate be added fermentation after juice in, add 1% peppermint oil,
0.1% laccase, 0.2% Tween-80, after mixing to obtain the final product.
One, effect test
Illustrate aflatoxin degradation agent of the invention to aflatoxin degradation effect by taking corn flour as an example below.
1.1 test groupings
1.1.1 the aflatoxin degradation agent of embodiment 2；
1.1.2 the aflatoxin degradation agent of embodiment 3；
1.1.3 the aflatoxin degradation agent of embodiment 4；
1.1.4 embodiment 3 does not contain the aflatoxin degradation agent of aloe；
1.1.5 the aflatoxin degradation agent that embodiment 3 is fermented without bacillus amyloliquefaciens；
1.1.6 the plant component of embodiment 3 is not fermented, but decocting mode effective component extracting, the aspergillus flavus of preparation
Toxin degradation agent (the decocting mode are as follows: add the ultrapure water of 1.5 times of weight, small fire decocts 0.5h, cooled after the completion of decoction
It filters to obtain the final product)；
1.1.7 laccase control group.
1.2 experimental design
1.2.1 degradation of the different aflatoxin degradation agents to aflatoxin
The corn flour (not detecting aflatoxin) not polluted by aspergillus flavus for taking 7kg, is equally divided into 7 parts, every part adds
Add 100 μ g of aflatoxin, is uniformly mixed；Wherein the 1st~3 portion of corn flour adds the aspergillus flavus of the embodiment of the present invention 2~4 respectively
Toxin degradation agent is denoted as 2~4 groups of embodiment, additive amount 10mL respectively；The degradation agent of 4th part of corn flour addition 1.1.4, the 5th
The degradation agent of part corn flour addition 1.1.5, the degradation agent of the 6th part of corn flour addition 1.1.6 are denoted as control group 1, control group respectively
2, control group 3, additive amount 10mL；7th portion of corn flour only adds laccase, and additive amount 0.03g is denoted as laccase group；It will be above-mentioned
Degradation agent is uniformly mixed using spray mode with corn flour；After mixing after 28 DEG C of processing 2.5h, at 100 DEG C of enzyme deactivations
Manage 15min；It is as shown in Figure 1 using highly effective liquid phase chromatography detection method measurement aflatoxin content after processing.
1.2.2 degradation of the aflatoxin degradation agent to different amounts of aflatoxin
The corn flour (not detecting aflatoxin) not polluted by aspergillus flavus for taking 6kg, is equally divided into 6 parts, to 6 parts of jade
Rice flour adds 50 μ g of aflatoxin, 100 μ g, 250 μ g, 500 μ g, 750 μ g, 1000 μ g respectively, is uniformly mixed；Every portion of corn flour
The middle aflatoxin degradation agent 10mL for adding embodiment 3 respectively, is uniformly mixed using spray mode with corn flour；Mixing
After uniformly after 28 DEG C of 2.5h, 100 DEG C of destroy the enzyme treatment 15min；Aspergillus flavus is measured using highly effective liquid phase chromatography detection method after processing
Content of toxins is as shown in Figure 2.
1.3 test result
1.3.1 degradation of the different aflatoxin degradation agents to aflatoxin
As shown in Figure 1, control group 1,2,3 and the aflatoxin content of laccase group are significantly higher than the embodiment of the present invention 2~4
Group, this illustrates degradation rate significant effect of the degradation agent to aflatoxin of the embodiment of the present invention 2~4, wherein the drop of embodiment 2
Solving agent is 94.8% to the degradation rate of aflatoxin, and the degradation agent of embodiment 3 is 96.4% to the degradation rate of aflatoxin,
The degradation agent of embodiment 4 is 95.1% to the degradation rate of aflatoxin, it can be seen that, the degradation agent effect of the embodiment of the present invention 3
Fruit is best.
1.3.2 degradation of the aflatoxin degradation agent to different amounts of aflatoxin
As shown in Figure 2, the degradation rate of aflatoxin is reduced with the increase of aflatoxin content.Wherein, from 50 μ
The μ of g~250 g decline is slow；When content is 50 μ g, degradation rate reaches 98% or more；When content is 250 μ g, degradation rate also exists
90 or more；Decline from 250 μ g to 1000 μ g most fast.
The above described is only a preferred embodiment of the present invention, being not that the invention has other forms of limitations, appoint
What those skilled in the art changed or be modified as possibly also with the technology contents of the disclosure above equivalent variations etc.
Imitate embodiment.But without departing from the technical solutions of the present invention, according to the technical essence of the invention to above embodiments institute
Any simple modification, equivalent variations and the remodeling made, still fall within the protection scope of technical solution of the present invention.
Claims (8)
1. a kind of aflatoxin degradation agent, it is characterised in that: be by the fermentation of 60~100g fresh aloe, 75~125g lemon
Liquid is prepared with 1% peppermint oil, 0.1~0.5% laccase, 0.2% Tween-80；
The preparation method of the degradation agent of the aflatoxin, step are as follows:
(1) fresh aloe, lemon, juicing, filtering；Juice ground-slag is broken to 40 mesh, adds 1.5 times of weight inorganic salt solutions, stirs evenly；
1.5% glucose is added in juice, stirs evenly；Juice slag and juice are cooled to room temperature in 100 DEG C of steam sterilizing 15min；
(2) bacillus amyloliquefaciens bacterium solution is added into juice slag after cooling, makes bacillus amyloliquefaciens concentration >=109Cfu/mL,
It is placed in 37 DEG C of culture 48h；
(3) saccharomyces cerevisiae bacterium solution is added into juice after cooling, makes saccharomyces cerevisiae bacteria concentration >=107Cfu/mL, 28 DEG C,
150rpm, ferment 48h；
(4) the juice slag after fermentation is filtered into waste, filtrate is added in the juice after fermentation, peppermint oil is added, laccase, spits
Temperature -80, after mixing to obtain the final product.
2. aflatoxin degradation agent according to claim 1, it is characterised in that: be by the fresh aloe of 60g, 75g lemon
Fermentation liquid is prepared with 1% peppermint oil, 0.5% laccase, 0.2% Tween-80.
3. aflatoxin degradation agent according to claim 1, it is characterised in that: be by the fresh aloe of 80g, 100g lemon
Fermentation liquid is prepared with 1% peppermint oil, 0.3% laccase, 0.2% Tween-80.
4. aflatoxin degradation agent according to claim 1, it is characterised in that: be by the fresh aloe of 100g, 125g lemon
Fermentation liquid be prepared with 1% peppermint oil, 0.1% laccase, 0.2% Tween-80.
5. aflatoxin degradation agent according to claim 1, it is characterised in that: the bacillus amyloliquefaciens bacterium solution is dense
Degree >=1010cfu/mL；S. cervisiae bacterial concentration >=108cfu/mL。
6. the preparation method of the degradation agent of aflatoxin according to claim 1, it is characterised in that: the inorganic salts are molten
Liquid is (g/L): (NH4)2SO42.0、KH2PO42.0、MgSO40.3、NaCl 0.5、FeSO40.005、MnSO40.016、
ZnCl20.017、CoCl20.002。
7. application of the aflatoxin degradation agent of claim 6 the method preparation in aflatoxin degradation.
8. a kind of aflatoxin biodegrading process, it is characterised in that: the degradation agent prepared using claim 6 method, step are as follows:
To the degradation agent prepared wait which claim 6 method is added in sample of degrading, after mixing in 28 DEG C of processing 2.5h, 100 DEG C go out
Enzymatic treatment 15min.
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| "Bio-fungicidal activity of Aloe vera sap against mycotoxigenic seed-borne Fungi";AbdEl-Rahim M.A. El-Samawaty et al.;《Fresenius Environmental Bulletin》;20110930;第20卷(第6期);摘要，3.4，图4, 4 |
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