CN104059129A - Antifungal peptide and application thereof to inhibition of generation of aflatoxin - Google Patents
Antifungal peptide and application thereof to inhibition of generation of aflatoxin Download PDFInfo
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- CN104059129A CN104059129A CN201410244941.XA CN201410244941A CN104059129A CN 104059129 A CN104059129 A CN 104059129A CN 201410244941 A CN201410244941 A CN 201410244941A CN 104059129 A CN104059129 A CN 104059129A
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Abstract
The invention provides an antifungal peptide and an application thereof to inhibition of generation of aflatoxin. The antifungal peptide has the amino acid sequence as follows: Ala-Asp-Leu-Pro-Val-Lys-Ser-Gly-Asp-Thr-Pro-Glu. The antifungal peptide provided by the invention is capable of effectively inhibiting the growth of aspergillus flavus and aspergillus parasiticus and the generation of aflatoxin, favorable and durable in inhibiting effect and suitable for large-scale production and application.
Description
Technical field
The present invention relates to a peptide species, particularly relate to a kind of anti-fungus peptide and suppressing the aborning application of aflatoxin.
Background technology
Aflatoxin (Aflatoxins) is the similar compound of a class chemical structure, mainly by flavus (Aspergillus flavus) and Aspergillus parasiticus (A.parasiticus)) produce, isolation identification goes out tens kinds of aflatoxin such as B1, B2, G1, G2, M1, M2 at present, and its basic structure is dihydrofuran ring and tonka bean camphor.Aflatoxin is the extremely strong highly toxic substance of a kind of toxicity, and its hazardness is mainly that people and animal livers tissue are had to destruction, can cause liver cancer even dead when serious.
Flavus and Aspergillus parasiticus extensively exist in air and soil, and it can produce aflatoxin after the food crop such as infecting peanut, corn in these crop storages.The most common with aflatoxin B1 in the food of natural contamination, its toxicity and carinogenicity are also the strongest.Due to tremendous economic loss and Health hazard that aflatoxin contamination brings, therefore researchist is finding the method that suppresses aflatoxin generation bacteria growing and remove aflatoxin always both at home and abroad.
At present, mainly comprise physical removal method, chemical removal method and biological removal method for the method that suppresses aflatoxin generation bacteria growing and removal aflatoxin.Physical removal method directly removes or makes aflatoxin decompose and change nontoxic intermediate product into aflatoxin, it comprises extraction process, absorption method, heat treating process, radiation method etc.; Chemical removal method is mainly to use chemical reagent to remove the growth of aflatoxin or inhibition flavus, thereby reduces the synthetic of toxin, and conventional chemical reagent comprises alkali, oxygenant etc.; Biological removal method is mainly to utilize the antimicrobial substance of antagonistic microbe and meta-bolites or plant-sourced to suppress the synthetic of aflatoxin, and much research has been found that multiple-microorganism for example bacterium, actinomycetes, mould, yeast and algae can both suppress the synthetic or aflatoxin degradation of aflatoxin.
Though physical removal method and chemical removal method have certain effect aspect synthesizing suppressing aspergillus growth and toxin, but inevitably can destroy the nutritive ingredient of product, the quality of reduction product, also may produce by product and chemical agent residue simultaneously, thereby cause certain harm; And biological removal method is at present mostly in the laboratory study stage, it is comparatively wasted time and energy, and the inhibition of contratoxin is limited, is not suitable for producing on a large scale and applying.
Except aforesaid method, some researchists also utilize transgenic technology to cultivate the farm crop of some aspergillus flavus resistings both at home and abroad, as aspergillus flavus resisting peanut, aspergillus flavus resisting corn etc., on this basis in conjunction with the method for physics, chemistry, biological removal toxin, thereby aflatoxin is controlled effectively, but the security of transgenic plant there is no final conclusion, and do not approved yet.Therefore, be badly in need of a kind of method of inhibition aflatoxin of efficient and applicable scale operation and application.
Summary of the invention
The invention provides a kind of anti-fungus peptide and suppressing aflatoxin aborning application, for solve prior art limited to the inhibition of aflatoxin, be not suitable for the technological deficiency such as scale operation and application.
The invention provides a kind of anti-fungus peptide, aminoacid sequence is: Ala-Asp-Leu-Pro-Val-Lys-Ser-Gly-Asp-Thr-Pro-Glu (SEQ ID NO:1).
The present invention also provides a kind of above-mentioned anti-fungus peptide suppressing the aborning application of aflatoxin.
Particularly, described aflatoxin can produce bacterium by aflatoxin and be produced, for example can be by least one generation in flavus and Aspergillus parasiticus.
The application mode of anti-fungus peptide of the present invention is versatile and flexible, both can be sprayed directly on the material of easy generation aflatoxin, thereby efficiently and enduringly suppress the generation of aflatoxin at its duration of storage; In addition, can also during arable farming, be sprayed at the blade of crop or for cultivating the soil of described crop, thereby after harvesting crops, suppress the generation of the aflatoxin in crop at its duration of storage.
The present invention also provides a kind of method that aflatoxin produces that suppresses, and commute produces the material of aflatoxin and bestows above-mentioned anti-fungus peptide.
Further, by water-soluble described anti-fungus peptide, be mixed with mass percent concentration and be after 0.01～0.05% anti-fungus peptide solution, be sprayed on the material of described easy generation aflatoxin.
Further, the material of described easy generation aflatoxin is the material of easy infection flavus and/or Aspergillus parasiticus, such as rice, wheat, corn, peanut, soybean, walnut, cottonseed etc., particularly corn and peanut; It includes but not limited to grain, food, feed or Chinese medicine; And the moisture content of controlling in the material that sprays the described easy generation aflatoxin after described anti-fungus peptide solution is not more than 25%.
Anti-fungus peptide of the present invention is good to the inhibition of aflatoxin, and therefore in the time using, required relative concentration is lower; On the other hand, when concentration is too low, can may cause sprinkling amount to strengthen, thereby make spray after material in moisture content excessive, the growth of the easy like this mould (for example flavus and Aspergillus parasiticus) that causes producing aflatoxin, thereby bring disadvantageous effect, therefore preferably concentration range is 0.01～0.05%.
Further, described grain is rice, wheat or corn, and described food is peanut.
The present invention also provides a kind of method that aflatoxin produces that suppresses, and during arable farming, bestows above-mentioned anti-fungus peptide to described crop or for cultivating the soil of described crop.
Particularly, described in carrying out during crop is easy to infect flavus or Aspergillus parasiticus, bestow, for example, during peanut cultivation, can be before its results within 30-50 days, bestow described anti-fungus peptide.And, in the time bestowing described in carrying out, can within interval 10-30 days, carry out.In addition, say from inhibition, can be 10 days interval time; From cost angle, can be 20 days interval time.
Further, by water-soluble described anti-fungus peptide, be mixed with mass percent concentration and be after 0.01～0.05% anti-fungus peptide solution, be sprayed at the blade surface of described crop or for cultivating the soil of described crop.
Further, the consumption of described anti-fungus peptide solution is every mu of 8～12L/, for example every mu of 10L/.
Anti-fungus peptide provided by the invention is simple in structure, easy to prepare, and easily carries out scale operation, and this anti-fungus peptide can effectively suppress the growth of flavus and Aspergillus parasiticus and the generation of aflatoxin, and inhibition is good and lasting.In addition, anti-fungus peptide provided by the invention is in the time being applied to the generation of inhibition aflatoxin, and application mode is simple and diversified, and application cost is cheap, is particularly suitable for large-scale application.
Embodiment
For making the object, technical solutions and advantages of the present invention clearer, below in conjunction with embodiments of the invention, the technical scheme in the embodiment of the present invention is clearly and completely described, obviously, described embodiment is the present invention's part embodiment, instead of whole embodiment.Based on the embodiment in the present invention, those of ordinary skill in the art, not making the every other embodiment obtaining under creative work prerequisite, belong to the scope of protection of the invention.
Embodiment 1
The preparation of anti-fungus peptide:
Adopt the anti-fungus peptide that ABI433A full-automatic polypeptide synthetic instrument synthetic amino acid array is Ala-Asp-Leu-Pro-Val-Lys-Ser-Gly-Asp-Thr-Pro-Glu, after the desalination of HPLC reversed phase column chromatography, the molecular weight of measuring this anti-fungus peptide by Tricine-SDS-PAGE method is 1380Da.
Embodiment 2
Anti-fungus peptide prepared by embodiment 1 is dissolved in sterilized water, is mixed with respectively mass percent concentration and is 0.01% and 0.03% anti-fungus peptide solution, carries out bacteriostatic test.
Flavus through cultivating and Aspergillus parasiticus are mixed with respectively to spore concentration and are 10
5cFUmL
-1the spore suspension of left and right, for example join respectively, in the plate of solid medium (PDA substratum) that sterilizing is housed, every ware adds 20mL left and right, in each plate, place subsequently the Oxford cup of sterilizing, to adding respectively in the Oxford cup of each plate the above-mentioned concentration of 100 μ L to be respectively 0.01% and 0.03% anti-fungus peptide solution, as test example 2-1 and 2-2, simultaneously with the sterilized water 2-1 as a rule that opposes.
Each plate is placed in to 28 DEG C of constant incubators and cultivates after 72h, measure antibacterial circle diameter, wherein each test repeats 3 times, averages, and the results are shown in Table 1.
The bacteriostatic test of table 1 anti-fungus peptide
Table 1 result shows:
Anti-fungus peptide prepared by the present invention can significantly suppress the growth of flavus and Aspergillus parasiticus, and then can effectively suppress the generation of its meta-bolites aflatoxin; And the concentration of the fungistatic effect of this anti-fungus peptide and anti-fungus peptide solution has certain positive correlation, therefore in the time of practical application, can prepare according to demand the anti-fungus peptide solution of suitable concentration.
Embodiment 3
Anti-fungus peptide prepared by embodiment 1 is dissolved in sterilized water, is mixed with respectively mass percent concentration and is 0.02% and 0.05% anti-fungus peptide solution.
Get common commercially available corn grain (moisture content is about 15%), after detecting its aflatoxin content, divide four batches, work, be respectively that 0.02% and 0.04% anti-fungus peptide solution and sterilized water are evenly sprayed on three batches of corn grain wherein by above-mentioned concentration, as test example 3-1,3-2 and reference examples 3-1, and the moisture content of the rear three batches of corn grain of control sprinkling is 22% left and right; Another batch do not make the corn grain routine 3-2 in contrast of any processing.
After adopting wrapping machine to pack above-mentioned four batches of corn grain, be placed in storage room and carry out normal temperature storage, after storage the 10th day respectively, 30 days, within 60 days and 90 days, sample, adopt enzyme linked immunological kit to measure the wherein content of aflatoxin B1, measuring method is specially: accurately take 10g and be placed in centrifuge tube through the sample of pulverizing, add 2.5g sodium-chlor and 100mL methanol aqueous solution (volume ratio of methyl alcohol and water is 7:3), carry out high-speed stirring with homogenizer, then centrifugal 3min under 4000r/min, centrifuged supernatant is placed in to separating funnel, add 100mL pure water, and with twice of the dichloromethane extraction of 50mL, collect after whole extraction liquids, at 40 DEG C, do near with rotatory evaporator reduction vaporization, dry up by nitrogen gas stream again, finally accurately add 2.0mL methyl alcohol with dissolved residue, and measure the content of aflatoxin B1 with enzyme linked immunological kit, the results are shown in Table 2.
The inhibition of table 2 anti-fungus peptide to aflatoxin in corn
Table 2 result shows:
With respect to the corn grain that adopts sterilized water processing, the content of not making aflatoxin in the corn grain of any processing is relatively low, the growth that easily impels the bacterial strain of easy generation aflatoxin is described, thereby has a negative impact when moisture content is higher; And generation and the accumulation of corn grain aflatoxin in duration of storage that the anti-fungus peptide solution that adopts the present invention to prepare sprays are significantly suppressed, and inhibition continuous and effective in a long time, thereby the storage life that is conducive to improve corn grain; In addition the concentration that, improves anti-fungus peptide solution can further improve the inhibition to aflatoxin.
Embodiment 4
Anti-fungus peptide prepared by embodiment 1 is dissolved in the water, and is mixed with mass percent concentration and is 0.01% anti-fungus peptide solution.
Adopt ordinary method to cultivate peanut, and in harvesting peanut first 60 days, interval 10 days, 20 days and 30 days are sprayed on peanut leaf surface by 0.01% anti-fungus peptide solution of above-mentioned preparation in the mode of spraying respectively, the consumption of the anti-fungus peptide solution of using when each sprinkling is every mu of 10L/, it is as test example 4-1, simultaneously not spray the peanut plant routine 4-1 in contrast of anti-fungus peptide solution.
Adopt ordinary method results peanut, and be stored in storage room under room temperature, after 30 days, adopt aforesaid method to measure the content of the aflatoxin B1 in peanut in peanut storage, the results are shown in Table 3.
The inhibition of table 3 anti-fungus peptide to Aflatoxin in Peanut byHigh
Table 3 result shows:
The peanut of process anti-fungus peptide processing before results, the content of the lay up period aflatoxin after results obtains certain inhibition, and without peanut prolongation along with storage time at duration of storage of any processing, the content of aflatoxin significantly raises, the generation that can effectively suppress crop duration of storage aflatoxin when anti-fungus peptide of the present invention is used during arable farming is described, the method is simple, easy handling not only, and is conducive to the large-scale practical application of anti-fungus peptide.
Finally it should be noted that: above each embodiment, only in order to technical scheme of the present invention to be described, is not intended to limit; Although the present invention is had been described in detail with reference to aforementioned each embodiment, those of ordinary skill in the art is to be understood that: its technical scheme that still can record aforementioned each embodiment is modified, or some or all of technical characterictic is wherein equal to replacement; And these amendments or replacement do not make the essence of appropriate technical solution depart from the scope of various embodiments of the present invention technical scheme.
Claims (10)
1. an anti-fungus peptide, is characterized in that, aminoacid sequence is: Ala-Asp-Leu-Pro-Val-Lys-Ser-Gly-Asp-Thr-Pro-Glu.
2. anti-fungus peptide claimed in claim 1 is suppressing the aborning application of aflatoxin.
3. application according to claim 2, is characterized in that, described aflatoxin is by least one generation in flavus and Aspergillus parasiticus.
4. suppress the method that aflatoxin produces, it is characterized in that, commute produces the material of aflatoxin and bestows anti-fungus peptide claimed in claim 1.
5. method according to claim 4, is characterized in that, by water-soluble described anti-fungus peptide, be mixed with mass percent concentration and be after 0.01～0.05% anti-fungus peptide solution, is sprayed on the material of described easy generation aflatoxin.
6. method according to claim 5, it is characterized in that, the material of described easy generation aflatoxin is grain, food, feed or Chinese medicine, and the moisture content of controlling in the material that sprays the described easy generation aflatoxin after described anti-fungus peptide solution is not more than 25%.
7. method according to claim 6, is characterized in that, described grain is rice, wheat or corn, and described food is peanut.
8. suppress the method that aflatoxin produces, it is characterized in that, during arable farming, bestow anti-fungus peptide claimed in claim 1 to described crop or for cultivating the soil of described crop.
9. method according to claim 8, it is characterized in that, by water-soluble described anti-fungus peptide, be mixed with mass percent concentration and be after 0.01～0.05% anti-fungus peptide solution, be sprayed at the blade surface of described crop or for cultivating the soil of described crop.
10. method according to claim 9, is characterized in that, the consumption of described anti-fungus peptide solution is every mu of 8～12L/.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107021996A (en) * | 2017-05-24 | 2017-08-08 | 中国海洋大学 | A kind of small peptide, its application and bactericidal composition therefrom |
| CN107646790A (en) * | 2017-10-18 | 2018-02-02 | 浙江青莲食品股份有限公司 | The cultural method of children phase panda pig |
| CN107646791A (en) * | 2017-10-18 | 2018-02-02 | 浙江青莲食品股份有限公司 | Cherish the cultural method of son phase panda pig |
| CN107736300A (en) * | 2017-10-18 | 2018-02-27 | 浙江青莲食品股份有限公司 | The cultural method of nursing period panda pig |
| CN107787911A (en) * | 2017-10-18 | 2018-03-13 | 浙江青莲食品股份有限公司 | The cultural method of adult panda pig |
| CN108660120A (en) * | 2017-03-27 | 2018-10-16 | 中国科学院微生物研究所 | Anti-fungus peptide and application thereof |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108660120A (en) * | 2017-03-27 | 2018-10-16 | 中国科学院微生物研究所 | Anti-fungus peptide and application thereof |
| CN107021996A (en) * | 2017-05-24 | 2017-08-08 | 中国海洋大学 | A kind of small peptide, its application and bactericidal composition therefrom |
| CN107021996B (en) * | 2017-05-24 | 2020-02-14 | 中国海洋大学 | Short peptide, application thereof and antibacterial composition obtained from short peptide |
| CN107646790A (en) * | 2017-10-18 | 2018-02-02 | 浙江青莲食品股份有限公司 | The cultural method of children phase panda pig |
| CN107646791A (en) * | 2017-10-18 | 2018-02-02 | 浙江青莲食品股份有限公司 | Cherish the cultural method of son phase panda pig |
| CN107736300A (en) * | 2017-10-18 | 2018-02-27 | 浙江青莲食品股份有限公司 | The cultural method of nursing period panda pig |
| CN107787911A (en) * | 2017-10-18 | 2018-03-13 | 浙江青莲食品股份有限公司 | The cultural method of adult panda pig |
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