CN102687800B - Preparation method of aflatoxin B1 biological degradation agent and application thereof - Google Patents
Preparation method of aflatoxin B1 biological degradation agent and application thereof Download PDFInfo
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- CN102687800B CN102687800B CN 201210133701 CN201210133701A CN102687800B CN 102687800 B CN102687800 B CN 102687800B CN 201210133701 CN201210133701 CN 201210133701 CN 201210133701 A CN201210133701 A CN 201210133701A CN 102687800 B CN102687800 B CN 102687800B
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- 239000003795 chemical substances by application Substances 0.000 title claims abstract description 18
- OQIQSTLJSLGHID-WNWIJWBNSA-N aflatoxin B1 Chemical compound C=1([C@@H]2C=CO[C@@H]2OC=1C=C(C1=2)OC)C=2OC(=O)C2=C1CCC2=O OQIQSTLJSLGHID-WNWIJWBNSA-N 0.000 title claims abstract description 14
- 229930020125 aflatoxin-B1 Natural products 0.000 title claims abstract description 14
- 238000002360 preparation method Methods 0.000 title claims abstract description 14
- 239000002115 aflatoxin B1 Substances 0.000 title claims abstract description 13
- 238000006731 degradation reaction Methods 0.000 title abstract description 18
- 230000015556 catabolic process Effects 0.000 title abstract description 15
- 238000002156 mixing Methods 0.000 claims abstract description 8
- 241000228245 Aspergillus niger Species 0.000 claims abstract description 7
- 238000000034 method Methods 0.000 claims abstract description 7
- 239000007788 liquid Substances 0.000 claims description 12
- 238000006065 biodegradation reaction Methods 0.000 claims description 9
- 238000000926 separation method Methods 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 5
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 4
- 244000061456 Solanum tuberosum Species 0.000 claims description 4
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 4
- 239000008103 glucose Substances 0.000 claims description 4
- 235000015097 nutrients Nutrition 0.000 claims description 4
- 230000001954 sterilising effect Effects 0.000 claims description 4
- 238000004659 sterilization and disinfection Methods 0.000 claims description 4
- 241000894006 Bacteria Species 0.000 claims description 3
- 238000012870 ammonium sulfate precipitation Methods 0.000 claims description 3
- 238000010612 desalination reaction Methods 0.000 claims description 3
- 239000002504 physiological saline solution Substances 0.000 claims description 3
- 210000001082 somatic cell Anatomy 0.000 claims description 3
- 238000000967 suction filtration Methods 0.000 claims description 3
- 229920001817 Agar Polymers 0.000 claims description 2
- 239000008272 agar Substances 0.000 claims description 2
- 238000000855 fermentation Methods 0.000 claims description 2
- 230000004151 fermentation Effects 0.000 claims description 2
- 239000000654 additive Substances 0.000 abstract description 6
- 244000144972 livestock Species 0.000 abstract description 5
- 239000000126 substance Substances 0.000 abstract description 3
- 102000004190 Enzymes Human genes 0.000 abstract description 2
- 108090000790 Enzymes Proteins 0.000 abstract description 2
- 230000000996 additive effect Effects 0.000 abstract description 2
- 239000003674 animal food additive Substances 0.000 abstract description 2
- 230000000593 degrading effect Effects 0.000 abstract description 2
- 101100449517 Arabidopsis thaliana GRH1 gene Proteins 0.000 abstract 1
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 abstract 1
- 101100434479 Saccharomyces cerevisiae (strain ATCC 204508 / S288c) AFB1 gene Proteins 0.000 abstract 1
- 238000001035 drying Methods 0.000 abstract 1
- 210000005253 yeast cell Anatomy 0.000 abstract 1
- 229930195730 Aflatoxin Natural products 0.000 description 4
- XWIYFDMXXLINPU-UHFFFAOYSA-N Aflatoxin G Chemical compound O=C1OCCC2=C1C(=O)OC1=C2C(OC)=CC2=C1C1C=COC1O2 XWIYFDMXXLINPU-UHFFFAOYSA-N 0.000 description 4
- 239000005409 aflatoxin Substances 0.000 description 4
- 244000144977 poultry Species 0.000 description 3
- 230000001988 toxicity Effects 0.000 description 3
- 231100000419 toxicity Toxicity 0.000 description 3
- 231100000678 Mycotoxin Toxicity 0.000 description 2
- 231100000357 carcinogen Toxicity 0.000 description 2
- 239000003183 carcinogenic agent Substances 0.000 description 2
- 239000000287 crude extract Substances 0.000 description 2
- 238000000502 dialysis Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000007670 refining Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 241000228197 Aspergillus flavus Species 0.000 description 1
- 241000228230 Aspergillus parasiticus Species 0.000 description 1
- 208000005623 Carcinogenesis Diseases 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 101710182223 Toxin B Proteins 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000036952 cancer formation Effects 0.000 description 1
- 230000000711 cancerogenic effect Effects 0.000 description 1
- 231100000504 carcinogenesis Toxicity 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 238000001784 detoxification Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- -1 flavus Natural products 0.000 description 1
- 235000012631 food intake Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 239000002636 mycotoxin Substances 0.000 description 1
- 230000000050 nutritive effect Effects 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 231100000614 poison Toxicity 0.000 description 1
- 239000002574 poison Substances 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 229930000044 secondary metabolite Natural products 0.000 description 1
- 238000007669 thermal treatment Methods 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
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- Fodder In General (AREA)
Abstract
The invention provides a preparation method of aflatoxin B1 biological degradation agent and an application thereof. The method uses aspergillusniger capable of degrading aflatoxin B1 and the produced degradation enzyme thereof to provide a preparation method of the aflatoxin B1 biological degradation agent, and an application thereof, wherein the preparation method comprises the steps of mixing the degradation agent and substances such as brewer yeast cells according to 5-10% (V/W); after drying, preparing the aflatoxin B1 biological degradation feed additive; and mixing the additive and complete feed according to the weight ratio of 1-2%; thus, the degradation agent can degrade most of AFB1 in the feed and reduce or eliminate the aflatoxicosis phenomenon of the livestock.
Description
One) technical field
The present invention relates to AFB
lThe preparation method of biodegradation agent and application thereof belong to feed additive field.
(2) background technology
Aflatoxin (Aflatoxin, AF) is by several mycetogenetic secondary metabolites such as flavus, Aspergillus parasiticuses, is the class formation molecule mycotoxins similar with physico-chemical property, is a kind of violent in toxicity and strong carcinogen, wherein AFB
1For the most stable in the various mycotoxinss of finding so far, that toxicity is the strongest is a kind of.
Because the Aspergillus flavus that produces poison produces toxin B
1Amount maximum, and toxicity is the strongest, so generally measure B
1Content represent the content of aflatoxin in food.A large amount of epidemiology surveys and research all confirms, livestock and poultry eat contaminated feed can reduce animal production performance, cause a series of diseases, can indirectly enter human body by food chain and have extremely strong carcinogenesis, the serious threat human health.1993, AFB
1Delimited by the cancer research mechanism of the World Health Organization and be the category-A carcinogens.Traditional removal AFB
1Method mainly contains absorption, extraction, thermal treatment, Irradiation, acid-alkali treatment, redox and processes, but have waste time and energy, the detoxification rate is not high, the shortcoming that easily causes nutritive substance to run off.Therefore, eliminate this toxoid to the harm of human and livestock health in the urgent need to a kind of poison-removing method of highly effective and safe.
(3) summary of the invention:
In order to address the above problem, utilization of the present invention can aflatoxin degradation B
1Aspergillus niger (aspergillus niger) and the degrading enzyme that produces thereof a kind of AFB is provided
lThe preparation method of biodegradation agent and application thereof.
Aspergillus niger used in the present invention (aspergillus niger), available from Chinese common micro-organisms DSMZ, its preservation is numbered CGMCC NO.3.2915.
AFB
lThe preparation method of biodegradation agent is as follows:
Aspergillus niger (CGMCC NO.3.2915) is rule on PDA solid medium flat board separation, cultivate 48h for 37 ℃, transfer single bacterium colony, in access 100mlPDB liquid nutrient medium, at 37 ℃ of lower shaking culture 3～5d, get fermented liquid: with fermented liquid at 4 ℃, centrifugal 20 min under 8000 r/min conditions, or the method for employing suction filtration, separation of supernatant and somatic cells; Getting supernatant liquor, is 30～90% ammonium sulfate precipitation with concentration; With the physiological saline solution throw out of fermented liquid 1/10 volume, the desalination in 6～24 hours of dialysing namely gets the aflatoxin B1 biodegradation agent, and in gained aflatoxin B1 biodegradation agent, protein content is at 20～40mg/ml.
Described PDA solid medium (g/L) preparation method is: potato 200, and glucose 20, agar 20, after mixing, 121 ℃ of high pressure steam sterilization 15min.
Described PDB substratum (g/L) preparation method is: potato 200, and glucose 20, after mixing, 121 ℃ of high pressure steam sterilization 15min.
The condition of described shaking culture is: leavening temperature is 20～40 ℃, fermentation time 24～120h, and pH value 5.0～9.0, rotating speed is 120～200r/min.
This degradation agents is mixed with the material such as cerevisiae by 5～10%(V/W), be prepared into aflatoxin B1 biological degradation fodder additives after oven dry, additive application is 1～2%(weight ratio in the addition of complete feed).
(4) embodiment
Example 1
Aspergillus niger (CGMCC NO.3.2915) is rule on PDA solid medium flat board separation is cultivated 48h for 37 ℃, transfers single bacterium colony, in access 100mlPDB liquid nutrient medium;
At 37 ℃ of lower shaking culture 3～5d, get fermented liquid; At 4 ℃, centrifugal 20 min under 8000 r/min conditions, or the method for employing suction filtration, separation of supernatant and somatic cells are got supernatant liquor, namely get crude extract;
Ammonium sulfate precipitation crude extract with 30～90% is with 10 ml physiological saline solution throw outs, the dialysis tubing of packing into.With clear water dialysis desalination in 6～24 hours, namely get refining AFB
1(protein content is 20～40mg/ml) for degradation agents.
This degradation agents mixes with cerevisiae by 5～10%(V/W), is prepared into aflatoxin B1 biological degradation fodder additives, by 1～2%(weight ratio) mix use with complete feed.
The result of use checking:
1. get and contain AFB
1Liquid sample 100ml(AFB
1Content is 40mg/L), add refining AFB
1Degradation agents 1ml, mixing, after 37 ℃ of effect 2d, AFB in sample
1Degradation rate reach 80～90% for qualified.
2. get feed 500g(AFB
1Content is 20～100mg/kg), adds the 1% aflatoxin B1 biological degradation fodder additives according to the present invention's preparation of its weight, adds simultaneously the 300ml deionized water, mixing.After 37 ℃ of effect 3d of temperature, AFB in feed
1Degradation rate be 70～80% for qualified.
At present, due to the corn that has used mould contamination, more or less all contain AFB in commercially available complete feed
1(AFB
1).Livestock and poultry life-time service AFB
1Content surpasses the feed of 20mg/kg, namely aflatoxicosis can occur, show as that food consumption descends, weightening finish slowly, become thin, even dead.The aflatoxin B1 biological degradation fodder additives that utilizes the present invention to prepare is by 1～2%(weight ratio) mix with complete feed, can be with the most of AFB in feed
1Degraded, less or elimination livestock and poultry aflatoxicosis phenomenon.
Claims (1)
1. the preparation method of an AFB l biodegradation agent, it is characterized in that first aspergillus niger CGMCC NO.3.2915 being rule separation on PDA solid medium flat board, cultivate 48h for 37 ℃, transfer single bacterium colony, in access 100mlPDB liquid nutrient medium, at 37 ℃ of lower shaking culture 3～5d, get fermented liquid; With fermented liquid at 4 ℃, centrifugal 20min under the 8000r/min condition, or adopt the method for suction filtration, separation of supernatant and somatic cells; Getting supernatant liquor, is 30～90% ammonium sulfate precipitation with concentration; With the physiological saline solution throw out of fermented liquid 1/10 volume, the desalination in 6～24 hours of dialysing namely gets the aflatoxin B1 biodegradation agent, and in gained aflatoxin B1 biodegradation agent, protein content is at 20～40mg/ml;
Described PDA solid medium g/L preparation method is: potato 200, and glucose 20, agar 20, after mixing, 121 ℃ of high pressure steam sterilization 15min;
Described PDB liquid nutrient medium g/L preparation method is: potato 200, and glucose 20, after mixing, 121 ℃ of high pressure steam sterilization 15min;
The condition of described shaking culture is: leavening temperature is 20～40 ℃, fermentation time 24～120h, and pH value 5.0～9.0, rotating speed is 120～200r/min.
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| CN 201210133701 CN102687800B (en) | 2012-05-03 | 2012-05-03 | Preparation method of aflatoxin B1 biological degradation agent and application thereof |
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| CN 201210133701 CN102687800B (en) | 2012-05-03 | 2012-05-03 | Preparation method of aflatoxin B1 biological degradation agent and application thereof |
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| CN102687800B true CN102687800B (en) | 2013-06-19 |
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Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104099251B (en) * | 2014-04-11 | 2016-11-23 | 江南大学 | A kind of Aspergillus niger strain and the application in multiple mycotoxin is degraded thereof |
| CN104498378A (en) * | 2014-06-23 | 2015-04-08 | 湖北工业大学 | Strain producing aflatoxin B1 degrading enzyme and application thereof |
| CN104789479B (en) * | 2014-07-25 | 2018-01-16 | 江南大学 | A kind of preparation of aspergillus niger immobilized cell and its degradation to aflatoxin B1 |
| CN104789537A (en) * | 2015-04-03 | 2015-07-22 | 陕西科技大学 | Biological degradation agent for aflatoxin B1 and preparation method of degradation agent |
| CN107513512B (en) * | 2017-10-16 | 2019-12-13 | 宜宾学院 | Rhodobacter capsulatus strain for degrading aflatoxin B1 and application thereof, and degrading agent and application thereof |
| CN108034598B (en) * | 2017-11-24 | 2020-04-28 | 河南德邻生物制品有限公司 | Composition capable of simultaneously degrading aflatoxin B1 and zearalenone |
| CN110583964A (en) * | 2019-09-23 | 2019-12-20 | 江南大学 | Biological removal method for efficiently removing four aflatoxins in peanut meal |
| CN114504072A (en) * | 2022-02-21 | 2022-05-17 | 大连吉祥缘农业科技有限公司 | Application of negative ion liquid in removing aflatoxin and method for removing feed aflatoxin |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1240828A (en) * | 1998-06-30 | 2000-01-12 | 广东药学院 | Process for extracting detoxicating enzyme for aflatoxin B1 |
| CN1364421A (en) * | 2001-01-12 | 2002-08-21 | 暨南大学 | Detoxicaction method for aflatoxin in fodder |
| CN101735992A (en) * | 2008-11-18 | 2010-06-16 | 国龙科技饲料(上海)有限公司 | Exogenous composite enzyme preparation special for feed industry and preparation method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2252163A2 (en) * | 2008-03-05 | 2010-11-24 | Novozymes A/S | Detoxification of aflatoxin in feed products |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1240828A (en) * | 1998-06-30 | 2000-01-12 | 广东药学院 | Process for extracting detoxicating enzyme for aflatoxin B1 |
| CN1364421A (en) * | 2001-01-12 | 2002-08-21 | 暨南大学 | Detoxicaction method for aflatoxin in fodder |
| CN101735992A (en) * | 2008-11-18 | 2010-06-16 | 国龙科技饲料(上海)有限公司 | Exogenous composite enzyme preparation special for feed industry and preparation method and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| Degradation Products from Aflatoxin B1;RENATE MANN, et al.;《European Journal Applied Microbiology》;19761231(第2期);第297-306页 * |
| RENATE MANN, et al..Degradation Products from Aflatoxin B1.《European Journal Applied Microbiology》.1976,(第2期),第297-306页. |
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