CN101978827A - Ningnanmycin composition and preparation method thereof - Google Patents
Ningnanmycin composition and preparation method thereof Download PDFInfo
- Publication number
- CN101978827A CN101978827A CN2010105681260A CN201010568126A CN101978827A CN 101978827 A CN101978827 A CN 101978827A CN 2010105681260 A CN2010105681260 A CN 2010105681260A CN 201010568126 A CN201010568126 A CN 201010568126A CN 101978827 A CN101978827 A CN 101978827A
- Authority
- CN
- China
- Prior art keywords
- ningnanmycin
- compounds
- composition
- formula
- condition
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Abstract
The invention provides a pesticide composition containing ningnanmycin. The pesticide composition also contains two ningnanmycin analogs which can effectively prevent and control plant virus diseases, fungus diseases and bacteria diseases separately and together. In addition, the invention also provides a preparation method for the pesticide composition, a prepared pesticide preparation and application of the ningnanmycin analogs and the like.
Description
Technical field
The invention belongs to pesticide field, particularly, the present invention relates to comprise the composition pesticide of Ningnanmycin, it is characterized in that, it also comprises two kinds of Ningnanmycin analogs.In addition, the invention still further relates to the application etc. of the preparation method of above-mentioned agriculturally useful compositions, the pesticidal preparations that is mixed with and Ningnanmycin analog.
Background technology
Ningnanmycin (ningnanmycin) be a kind of from streptomyces noursei Xichang mutation (Slreptomyces noursei var.xichangenis) CGMCC No.0194 isolated alkaline water dissolubility cytosine nucleoside antibiotic (its chemical constitution as shown in Equation 1), inhibited to Gram-negative and positive bacteria, especially the pathogen of the powdery mildews of plants such as bacterial blight of rice and wheat, cucumber, beans and flowers and tobacco mosaic virus etc. all there are good preventive and therapeutic effect (referring to No. the 93104287.9th, Chinese patent; To Gu Xi, etc. a kind of new farm antibiotics---Ningnanmycin. microorganism journal, the 35th volume, the 5th phase: 368-374).The substitute of chemistry agriculturals such as current withered peaceful as leaf, the phenazine oxide of Ningnanmycin is promoted the use of at the wide geographic area of China's the South and the North, has obtained huge economic and social benefit.
Existing literature report has related to applying of Ningnanmycin and has been mixed with the application of compound preparation with other farm antibioticss.For example, Chinese patent application relates to the preparation and the technology of purifying Ningnanmycin for No. 99113215 and No. 200310112003: Chinese patent application No. 99122552, No. 200510018042, No. 200610151121, No. 200710056845, No. 200710188014, No. 200810053081 and No. 201010118266 etc., what disclose the Ningnanmycin for preparing is made into separately that preparation uses or it is made into the compound preparation use with gibberellin, virazole and/or N-(5-methyl-1,3-thiazole-2-yl)-4-methyl-1,2,3-thiadiazole etc.These reports all continue characteristic and other tunnings of further investigation streptomyces noursei Xichang mutation fermentation, especially those Ningnanmycin analogs with farm antibiotics potentiality or antibiont synergist potentiality that ferment and produce.
The inventor reaches on the basis of the research experience in year surplus in the of ten Ningnanmycin and zymophyte thereof own, through long-term arduous experiment and the fortune composition that some is accidental, obtained a kind of composition that comprises Ningnanmycin and two kinds of Ningnanmycin analogs surprisingly, wherein Ningnanmycin only need account for about 40% of described composition weight, just can play and the identical agricultural control efficiency of Ningnanmycin that has commercially available same composition weight now, thereby reduced the consumption of Ningnanmycin in actual use, all had positive effect for cost and prevent or suspension of pesticide resistance generation etc.; Simultaneously, these two kinds of Ningnanmycin analogs of inventor's discovery provide foundation for developing new farm antibiotics or antibiotic synergist etc.In addition, the inventor also provides the preparation method of said composition, and this method only need be adjusted the technology of streptomyces noursei Xichang mutation fermentation, just the disposable above-mentioned composition of preparing of energy, proportioning components is stable, need not adding other compositions as existing bibliographical information afterwards.
Summary of the invention
The technical problem to be solved in the present invention is the composition that comprises Ningnanmycin that provides new, in fact it reduced usefulness (containing) amount of Ningnanmycin, but effect does not but reduce, and this can reduce the production cost of Ningnanmycin on the one hand, on the other hand can suspension of pesticide resistance generation.Secondly, the technical problem to be solved in the present invention is to overcome the prior art prejudice of (as, No. the 99113215th, Chinese patent application), uses the method that prolongs fermented incubation time to come disposable preparation above-mentioned composition, and it is low and do not subtract the advantage of effect to have a cost.Also have, the technical problem to be solved in the present invention is to provide the agricultural chemicals of two kinds of Ningnanmycin analogs in the above-mentioned composition to use, and can be used for being prepared into composition pesticide and pesticidal preparations, also helps to develop new agricultural chemical compound.
Particularly, in first aspect, the invention provides the composition that comprises Ningnanmycin, it is characterized in that, it also comprises formula as follows 2 compounds and formula 3 compounds
That is, the invention provides a kind of composition, it comprises formula as follows 1 compound, formula 2 compounds and formula 3 compounds
Wherein, formula 2 compounds have mass spectral analysis collection of illustrative plates shown in Figure 1; Formula 3 compounds have mass spectral analysis collection of illustrative plates shown in Figure 2.That is, the invention provides the composition that comprises Ningnanmycin, it is characterized in that, the compound that it also comprises the compound with mass spectral analysis collection of illustrative plates shown in Figure 1 and has mass spectral analysis collection of illustrative plates shown in Figure 2.
The composition of first aspect present invention is composition pesticide preferably.In this article, " agricultural chemicals " has implication known in the art, promptly uses and is used to prevent and/or treat the medicine of (that is, control) plant pest to plant.Discover through the inventor, although the composition Ningnanmycin content of first aspect present invention has only about 40%, but can replace Ningnanmycin to use fully, control efficiency and the commercially available not significant difference of Ningnanmycin, therefore the composition of first aspect present invention can be the composition pesticide that is used to prevent and treat the disease that composition pesticide, especially bacterial blight-resisting, powdery mildew and/or the tobacco mosaic virus of the viroses of plant, mycosis and/or bacteriosis that Ningnanmycin can prevent and treat cause.
The composition of first aspect present invention can comprise water, also can not comprise water, and preferably its water content is lower than 0.5% (w/w), more preferably less than 0.4% (w/w), the moisture content as shown in the specific embodiment of the invention.Usually the composition of first aspect present invention exists as former medicine, is convenient to transportation, but before the field uses, will be diluted to preparation usually and use.This low water content, when can be diluted to preparation at auxiliary materials such as composition water with first aspect present invention, make things convenient for the percentage composition of calculation composition in preparation, thus in convenient former medicine transportation, also conveniently be mixed with the preparation that former medicine has diluted.The composition of first aspect present invention preferably is made up of formula 1 compound, formula 2 compounds and formula 3 compounds and water.
The composition of first aspect present invention preferably contains the Ningnanmycin greater than 35% (w/w).Particularly, in the composition of first aspect present invention, the content of Ningnanmycin is greater than 35% (w/w), be preferably 40-50% (w/w), more preferably 40-45% (w/w) is more preferably 40-42% (w/w), the content as shown in the specific embodiment of the invention.Preparation method provided by the invention can disposablely prepare the composition of Ningnanmycin content greater than 35% (w/w) (be preferably 40-50% (w/w), more preferably 40-45% (w/w) is more preferably 40-42% (w/w)), and stable content is fine.In this article, as do not add special instruction, content is all recently represented with weight percent.
The composition of first aspect present invention preferably contains formula 2 compounds of 35-55% (w/w).Particularly, in the composition of first aspect present invention, the content of formula 2 compounds is 35-55% (w/w), is preferably 40-50% (w/w), 40-47% (w/w) more preferably, the content as shown in the specific embodiment of the invention.Preparation method provided by the invention can disposablely prepare the composition that formula 2 compounds contents are 35-55% (w/w) (being preferably 40-50% (w/w), more preferably 40-47% (w/w)), and stable content is fine.Discover that through the inventor this compound is present in the composition that contains Ningnanmycin, can play and the essentially identical control efficiency of commercially available Ningnanmycin.
The composition of first aspect present invention preferably contains formula 3 compounds of 7-23% (w/w).Particularly, in the composition of first aspect present invention, the content of formula 3 compounds is 7-23% (w/w), is preferably 10-20% (w/w), 12-17% (w/w) more preferably, the content as shown in the specific embodiment of the invention.Preparation method provided by the invention can disposablely prepare the composition that formula 3 compounds contents are 7-23% (w/w) (being preferably 10-20% (w/w), more preferably 12-17% (w/w)), and stable content is fine.Discover that through the inventor this compound is present in the composition that contains Ningnanmycin, can play and the essentially identical control efficiency of commercially available Ningnanmycin.
The composition of first aspect present invention can be also formulated together by obtaining (as synthetic or purification) formula 1 compound, formula 2 compounds and formula 3 compounds respectively, and prepare.But the composition of first aspect present invention preferably prepares by the time that prolongs fermented and cultured.Prior art (as, No. the 99113215th, Chinese patent application) prejudice is thought, prolong the fermentation after fermentation and enter dynamic stage, but the inventor discovers, although can produce dynamic stage new compound (as, formula 2 compounds and formula 3 compounds), but the control efficiency of the composition of preparing do not influenced.The composition of first aspect present invention preferably prepares greater than 70 hours (more preferably greater than 70.5 hours, as 72 hours) by the time that prolongs fermented and cultured.Particularly, preferably preparation by the following method of the composition of first aspect present invention:
(1) rejuvenation streptomyces noursei Xichang mutation;
(2) on solid culture medium, cultivate mycelium;
(3) carry out enrichment culture in the liquid medium within;
(4) carry out fermented and cultured in fermentation tank, the time of fermented and cultured is preferably greater than 70 hours, more preferably greater than 70.5 hours, as 72 hours; With
(5) purifying.
In step (1), the mutation of streptomyces noursei Xichang refers to the CGMCC No.0194 bacterial strain that is preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center and is equal to bacterial strain, as having the bacterial strain of identical 16s rRNA with CGMCC No.0194 bacterial strain.These bacterial strains can obtain easily by buying on market.Rejuvenation preferably 26-30 ℃ (as, 28 ℃) and 65%-90% (as, 80%) the condition of relative moisture under carry out, usually on solid culture medium aerobic culture 14-18 (as, 16) day.In this article, solid culture medium contains corn flour, soybean cake powder, Portugal's glucose, corn starch, dusty yeast, NaCl, CaCO
3, (NH
4)
2SO
4, KH
2PO
4, MgSO
4And agar powder, preferably its weight ratio prescription is: corn flour 3%, soybean cake powder 4%, Portugal's glucose 1%, corn starch 1%, dusty yeast 0.1%, NaCl 0.5%CaCO
30.5%, (NH
4)
2SO
40.2%, KH
2PO
40.04%, MgSO
40.35%, agar powder 2.2%, surplus is a water, regulates pH value to 6.8 with NaOH.
In step (2), cultivate mycelium preferably 26-30 ℃ (as, 28 ℃) and 65%-90% (as, 80%) the condition of relative moisture under aerobic cultivate 12-16 (as, 14) sky.Thereafter, getting the mycelium of turning out is used to increase
In step (3), enrichment culture preferably 26-30 ℃ (as, 28 ℃) condition under cultivate (as, concussion is cultivated) 12-16 (and as, 14) day, more preferably in first liquid nutrient medium, cultivating.In this article, first liquid nutrient medium contains corn flour, soybean cake powder, glucose, corn starch, dusty yeast, NaCl, CaCO
3, (NH
4)
2SO
4, KH
2PO
4And MgSO
4, preferably its weight ratio prescription is: corn flour 3%, and soybean cake powder 4%, glucose 1%, corn starch 1%, dusty yeast 0.1%, NaCl 0.5%, CaCO
30.5%, (NH
4)
2SO
40.2%, KH
2PO
40.04%, MgSO
40.35%, surplus is a water, regulates pH value to 6.8 with NaOH.
In step (4), fermented and cultured preferably 26-30 ℃ (as, 28 ℃) and 6-10L/ hour (as, 8L/ hour) the condition of throughput under cultivate, more preferably in second liquid nutrient medium, cultivating.In this article, second liquid nutrient medium contains corn flour, soybean cake powder, glucose, corn starch, dusty yeast, NaCl, CaCO
3, (NH
4)
2SO
4, KH
2PO
4And MgSO
40.35%, preferably its weight ratio prescription is: corn flour 2%, and soybean cake powder 3%, glucose 1.5%, corn starch 1%, dusty yeast 0.2%, NaCl 0.5%, CaCO
30.5%, (NH
4)
2SO
40.2%, KH
2PO
40.04%, MgSO
40.35%, surplus is a water, regulates pH value to 6.8 with NaOH.
In step (5), purifying preferably includes acidifying, filtration and drying.The inventor finds, the purifying process (as, chromatography etc.) that composition of the present invention need not by complexity prepares, and only needs simple acidifying, filtration and dry step to get final product.Particularly, acidifying preferably add acid (as, oxalic acid) acidifying pH to 5-6 (as, 5.6); Filtration preferably include with tentatively filter the filtrate that (as, press filtration) obtain and be heated to 75-95 ℃ (as, 85 ℃), be cooled to then 0-10 ℃ (as, 4 ℃), then use membrane filtration; Drying is the negative-pressure vacuum drying preferably, and its baking temperature is no more than 60 ℃, and preferred 40-60 ℃, more preferably 15-55 ℃, as 50 ℃.
In second aspect, the invention provides the described method for compositions of preparation first aspect present invention, it comprises:
(1) rejuvenation streptomyces noursei Xichang mutation, preferably rejuvenation under the condition of 28 ℃ and 80% relative moisture;
(2) cultivate mycelium on solid culture medium, preferably aerobic was cultivated 14 days under the condition of 28 ℃ and 80% relative moisture;
(3) carry out enrichment culture in the liquid medium within, preferably under 28 ℃ condition, cultivated 14 days;
(4) carry out fermented and cultured in fermentation tank, the time of preferred fermented and cultured is preferably greater than 70 hours, more preferably greater than 70.5 hours, as 72 hours, is more preferably under the condition of throughput of 28 ℃ and 8L/ hour and cultivates; With
(5) purifying preferably includes acidifying, filtration and drying.
In step (1), the mutation of streptomyces noursei Xichang refers to the CGMCC No.0194 bacterial strain that is preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center and is equal to bacterial strain, as having the bacterial strain of identical 16s rRNA with CGMCC No.0194 bacterial strain.These bacterial strains can obtain easily by buying on market.Rejuvenation preferably 26-30 ℃ (as, 28 ℃) and 65%-90% (as, 80%) the condition of relative moisture under carry out, usually on solid culture medium aerobic culture 14-18 (as, 16) day.
In step (2), cultivate mycelium preferably 26-30 ℃ (as, 28 ℃) and 65%-90% (as, 80%) the condition of relative moisture under aerobic cultivate 12-16 (as, 14) sky.Get the mycelium of turning out and be used for enrichment culture thereafter,
In step (3), enrichment culture preferably 26-30 ℃ (as, 28 ℃) condition under cultivate (as, concussion is cultivated) 12-16 (and as, 14) day, more preferably in first liquid nutrient medium, cultivating.
In step (4), fermented and cultured preferably 26-30 ℃ (as, 28 ℃) and 6-10L/ hour (as, 8L/ hour) the condition of throughput under cultivate, more preferably in second liquid nutrient medium, cultivating.
In step (5), purifying preferably includes acidifying, filtration and drying.The inventor finds, the purifying process (as, chromatography etc.) that composition of the present invention need not by complexity prepares, and only needs simple acidifying, filtration and dry step to get final product.Particularly, acidifying preferably add acid (as, oxalic acid) acidifying pH to 5-6 (as, 5.6); Filtration preferably include with tentatively filter the filtrate that (as, press filtration) obtain and be heated to 75-95 ℃ (as, 85 ℃), be cooled to then 0-10 ℃ (as, 1 ℃), then use membrane filtration; Drying is the negative-pressure vacuum drying preferably, and its baking temperature is no more than 60 ℃, and preferred 10-60 ℃, more preferably 45-55 ℃, as 50 ℃.
Prior art (as, No. the 99113215th, Chinese patent application) prejudice is thought, prolong the fermentation after fermentation and enter dynamic stage, thereby think and to adopt the tunning before dynamic stage to come the purifying Ningnanmycin, but the inventor discovers, although can produce dynamic stage new compound (as, formula 2 compounds and formula 3 compounds), but do not influence the control efficiency of the composition of preparing, and this fermentation all there is positive role for reducing the Ningnanmycin consumption and slowing down drug-fast generation.Can predict, other can quicken the fermentation process that arrives dynamic stage, promptly can produce the described method for compositions of first aspect present invention and also contain in the present invention, are not subjected to concrete fermentation time and limit.The method of second aspect present invention has been prepared the described composition of first aspect present invention, comprises wherein institute's preferred compositions.
In the third aspect, the invention provides pesticidal preparations, it comprises acceptable assistant on described composition pesticide of first aspect present invention and the agricultural chemicals.Pesticidal preparations is tangible product, can be formulations such as aqua, emulsion, pulvis.In this article, " acceptable assistant on the agricultural chemicals " refers to can add in agricultural chemicals and prepares synergist and/or the auxiliary material that forms preparation.Synergist is other agricultural chemical compounds or composition pesticide normally, can with the composite formation pesticidal preparations of the described composition pesticide of first aspect present invention.Those that typical synergist such as the cited document of this paper background technology are reported comprise pest-resistant dose, antibacterial agent and antivirotic, as gibberellin, virazole and N-(5-methyl-1,3-thiazole-2-yl)-4-methyl-1,2,3-thiadiazole etc.In the specific embodiment of the present invention, described composition pesticide of first aspect present invention and Imidacloprid can be re-dubbed pesticidal preparations, so the preferred pesticidal preparations of the present invention comprises described composition of first aspect present invention and Imidacloprid.
Auxiliary material does not normally have the reagent of disease-resistant worm activity, comprises carrier (as, water), emulsifier, stabilizing agent, filler etc.Typical auxiliary material comprises water, neopelex, diatomite, caproic acid lignocaine alcohol ester etc.The present invention preferably is mixed with pesticidal preparations No. 02144510 with reference to Chinese patent application, and therefore inventing preferred pesticidal preparations comprises the described composition of first aspect present invention and neopelex and diatomite.
In fourth aspect, the invention provides the application in the viroses of plant, mycosis and/or bacteriosis that the control Ningnanmycin can be prevented and treated of described composition of first aspect present invention or the described pesticidal preparations of third aspect present invention.The inventor finds that the control efficiency of described composition of first aspect present invention and commercially available Ningnanmycin do not have remarkable difference, and can reduce the actual amount of Ningnanmycin, therefore can reduce or suspension of pesticide resistance generation, thereby be used to prevent and treat the viroses of plant, mycosis and/or the bacteriosis that Ningnanmycin can be prevented and treated.Described composition of first aspect present invention or the described pesticidal preparations of third aspect present invention can independent control or with other composition pesticides or the composite control of pesticidal preparations.In the specific embodiment of the present invention, separately control and composite control can realize the effect of preventing and treating.The present invention preferably provides the application in preventing and treating the disease that bacterial leaf-blight, powdery mildew and/or tobacco mosaic virus cause of the described composition of first aspect present invention or the described pesticidal preparations of third aspect present invention.
Aspect the 5th, the invention provides formula 2 compounds and be used for preventing and treating the composition pesticide of the disease that bacterial leaf-blight, powdery mildew and/or tobacco mosaic virus cause or the application of pesticidal preparations in preparation.Formula 2 compounds exist in the described composition of first aspect present invention, but do not reduce the control efficiency of Ningnanmycin, therefore can add in composition pesticide or the pesticidal preparations, produce composition pesticide or the pesticidal preparations that makes new advances thus.The present invention preferably provides the preparation that is combined in of formula 2 compound cotypes, 3 compounds and/or Ningnanmycin to be used to prevent and treat bacterial leaf-blight, the composition pesticide of the disease that powdery mildew and/or tobacco mosaic virus cause or the application in the pesticidal preparations, be used to prevent and treat bacterial leaf-blight suc as formula 2 compounds in preparation, the composition pesticide that contains formula 3 compounds and/or Ningnanmycin of the disease that powdery mildew and/or tobacco mosaic virus cause (as, the described composition of first aspect present invention) or contain application in the pesticidal preparations (as, the described pesticidal preparations of third aspect present invention) of formula 3 compounds and/or Ningnanmycin.In preparation process, formula 2 compounds can initiatively add in composition pesticide or the pesticidal preparations, also can produce in the production process of composition pesticide, as preparation produces in the described method of second aspect present invention
Aspect the 6th, the invention provides formula 3 compounds and be used for preventing and treating the composition pesticide of the disease that bacterial leaf-blight, powdery mildew and/or tobacco mosaic virus cause or the application of pesticidal preparations in preparation.Formula 3 compounds exist in the described composition of first aspect present invention, but do not reduce the control efficiency of Ningnanmycin, therefore can add in composition pesticide or the pesticidal preparations, produce composition pesticide or the pesticidal preparations that makes new advances thus.The present invention preferably provides the preparation that is combined in of formula 3 compound cotypes, 2 compounds and/or Ningnanmycin to be used to prevent and treat bacterial leaf-blight, the composition pesticide of the disease that powdery mildew and/or tobacco mosaic virus cause or the application in the pesticidal preparations, be used to prevent and treat bacterial leaf-blight suc as formula 3 compounds in preparation, the composition pesticide that contains formula 2 compounds and/or Ningnanmycin of the disease that powdery mildew and/or tobacco mosaic virus cause (as, the described composition of first aspect present invention) or contain application in the pesticidal preparations (as, the described pesticidal preparations of third aspect present invention) of formula 2 compounds and/or Ningnanmycin.In preparation process, formula 3 compounds can initiatively add in composition pesticide or the pesticidal preparations, also can produce in the production process of composition pesticide, as preparation produces in the described method of second aspect present invention.
Aspect the 7th, the invention provides the method for identifying described composition of first aspect present invention or the described pesticidal preparations of third aspect present invention, it comprises the step of carrying out the liquid chromatogram separation and carrying out Mass Spectrometer Method.Contain Ningnanmycin if detect, and detect and contain formula 2 compounds (promptly, have mass spectral analysis collection of illustrative plates shown in Figure 1) and formula 3 compounds are (promptly, have mass spectral analysis collection of illustrative plates shown in Figure 2), then can assert it is described composition of first aspect present invention or the described pesticidal preparations of third aspect present invention.Preferably can further detect the content of each composition.
The present invention has following beneficial effect: can be effectively separately and combination prevent and treat the viroses of plant, mycosis, bacteriosis; Alleviate or suspension of pesticide resistance generation; Constant product quality; The preparation method can disposablely prepare composition of the present invention; Formula 2 compounds and formula 3 compounds can be further used for the exploitation of agricultural chemical compound, composition, etc.
For the ease of understanding, below will describe in detail the present invention by concrete accompanying drawing, embodiment.It needs to be noted that these descriptions only are exemplary descriptions, do not constitute limitation of the scope of the invention.According to the description of this specification, many variations of the present invention, change all are conspicuous concerning one of ordinary skill in the art.In addition, the present invention has quoted open source literature, and these documents are in order more clearly to describe the present invention, and their full text content is all included this paper in and carried out reference, just looks like that repeated description is the same excessively in this article for their full text.
Description of drawings
Fig. 1 is the mass spectral analysis collection of illustrative plates of formula 2 compounds, and wherein last figure is its mass-spectrogram, and figure below is the second order ms collection of illustrative plates of its fragment ion.
Fig. 2 is the mass spectral analysis collection of illustrative plates of formula 3 compounds, and wherein last figure is its mass-spectrogram, and figure below is the second order ms collection of illustrative plates of its fragment ion.
Embodiment
Further specify content of the present invention by the following examples.As do not specialize, conventional means that used technological means is well known to those skilled in the art among the embodiment and commercially available common instrument, reagent, can be referring to the references such as manufacturers instruction of " microbiology experiment (the 4th edition) " (Higher Education Publishing House, Beijing) and corresponding instrument and reagent.
Frozen streptomyces noursei Xichang mutation (available from Heilongjiang Province's biopesticide Engineering Technical Research Centre) is inoculated on the freshly prepared solid culture medium, in 28 ℃, 80% relative moisture, under the condition of filtrated air circulation, cultivate and carried out rejuvenation in 16 days, the bacterial classification spore that picking produces.Aerial mycelium then is about to the bacterial classification spore inoculating on freshly prepared solid culture medium, and in 28 ℃, 80% relative moisture, aerobic was cultivated 14 days under the condition of filtrated air circulation, high-visible have mycelium to produce.Carry out enrichment culture then, promptly mycelium washed from the solid culture basal plane and enter the 1L conical flask with freshly prepared first liquid nutrient medium, adding freshly prepared first liquid nutrient medium, to be supplemented to culture volume be 250ml, in 28 ℃, cultivated 14 days with the 250rpm shaking table.Through the thalline that microscope sampling detects each bottle cultivation have all have normal morphology after, carry out fermented and cultured, the culture fluid that promptly merges 4 conical flasks, inject 10L fermentation tank (can available from German Bei Lang company), with second liquid culture and be settled to 5L, in 28 ℃ of fermented and cultured 72 hours, condition of culture is: the blowing air amount was 8L/ hour, and mixing speed is 250rpm.
Wherein, the weight ratio of solid culture medium prescription is as follows: corn flour 3%, and soybean cake powder 4%, glucose 1%, corn starch 1%, dusty yeast 0.1%, NaCl 0.5%, CaCO
30.5%, (NH
4)
2SO
40.2%, KH
2PO
40.04%, MgSO
40.35%, agar powder 2.2%, surplus is a water, regulates pH value to 6.8 with NaOH; The weight ratio prescription of first liquid nutrient medium is as follows: corn flour 3%, and soybean cake powder 4%, glucose 1%, corn starch 1%, dusty yeast 0.1%, NaCl 0.5%, CaCO
30.5%, (NH
4)
2SO
40.2%, KH
2PO
40.04%, MgSO
40.35%, surplus is a water, regulates pH value to 6.8 with NaOH; The weight ratio prescription of second liquid nutrient medium is as follows: corn flour 2%, soybean cake powder 3%, glucose 1.5%, corn starch 1.Dusty yeast 0.2%, NaCl 0.5%, CaCO
30.5%, (NH
4)
2SO
40.2%, KH
2PO
40.04%, MgSO
40.35%, surplus is a water, regulates pH value to 6.8 with NaOH.
Get the zymotic fluid after above-mentioned 72 hours, under condition of stirring, add oxalic acid at once and regulate pH to 5.6, inject BYJ650 template frame filter press (can available from Jin Hai Yongchang, Hangzhou filter press Co., Ltd) press filtration then.Collect filtrate and be heated to 85 ℃, keep moving into the cryogenic freezing case at once the temperature of filtrate being reduced to 4 ℃ after 2 minutes.Cross 0.45 micron filter membrane then, collect filtrate, place negative-pressure vacuum drying box (can step butterfly heating equipment Co., Ltd) to be dried to solid in 50 ℃, promptly get the composition that the present invention contains Ningnanmycin with the pressure of 0.05MPa available from Nanjing.
Method according to embodiment 1 has prepared 5 batches of compositions that contain Ningnanmycin respectively, carries out constituent analysis.
(1) mensuration of water content
Each batch respectively takes by weighing two duplicate samples, about every part of 0.1g (that is, sample size), places SFY-3A type micro-water analyzer (can available from Shandong Zibo Hai Nuo Instr Ltd.) to detect, and is converted into moisture content according to the water content of instrument gained.The result is shown in table 2-1, all less than 0.5%.
Table 2-1 contains the sample determination of moisture result of the composition of Ningnanmycin
(2) Ningnanmycin Determination on content
With reference to manufacturers instruction, Agilent 1100HP LC liquid chromatographic detection in the use Agilent 1100HP LC-MSD liquid chromatograph-mass spectrometer contains the composition and the Ningnanmycin standard sample of Ningnanmycin, with the Ningnanmycin standard sample of concentration known (can available from China Agricultural University) is the calibration curve that contrast obtains peak area and weight relationships, and the area at the pairing peak of Ningnanmycin in the high performance liquid chromatography in each two duplicate samples of each batch is converted into mass percent concentration.The result is shown in table 2-2, and the stable content of Ningnanmycin in composition is about 40%.
Table 2-2 contains the content analysis result of Ningnanmycin in the composition of Ningnanmycin
(3) qualitative and quantitative analysis of other compositions
With reference to manufacturers instruction, use Agilent 1100HP LC-MSD liquid chromatograph-mass spectrometer (ion volume), pairing two peaks of non-Ningnanmycin are carried out mass spectral analysis respectively, wherein, retention time be the mass spectrum of composition of 2.5min and fragment ion thereof second order ms collection of illustrative plates as shown in Figure 1, instrument is confirmed its chemical constitution as shown in Equation 2; Retention time be the mass spectrum of composition of 3.3min and fragment ion thereof second order ms collection of illustrative plates as shown in Figure 2, instrument is confirmed its chemical constitution as shown in Equation 3
Calculate content according to the shown peak area of aforementioned Agilent 1100HP LC liquid chromatogram by the method for calibration curve, the mass percent concentration of composition Chinese style 2 of each batch and formula 3 compounds is respectively shown in table 2-3, the stable content of its Chinese style 2 compounds is in the scope of 40-50%, and the stable content of the compound of formula 3 is in the scope of 10-20%.
Table 2-3 contains the composition Chinese style 2 of Ningnanmycin and the content analysis result of formula 3 compounds
| Mission Number | The content (%) of |
The content of the compound of formula 3 (%) |
| 1 | ?46.18 | ?12.62 |
| 2 | ?46.52 | ?12.03 |
| 3 | ?46.08 | ?13.02 |
| 4 | ?41.28 | ?17.07 |
| 5 | ?44.09 | ?14.15 |
Embodiment 3 contains the control efficiency of the composition of Ningnanmycin
We use according to the composition that contains Ningnanmycin (A) the dilution back of embodiment 1 preparation and (B) do field trial relatively with commercially available 2% Ningnanmycin (available from Seoul Heilongjiang Qianger biochemical technology development Co., Ltd) of equivalent, in order to show its control efficiency to each crops.
(1) field control effectiveness test of stripe disease
In the Wuzhi County, Henan Province, (A, B respectively test half area to 1 mu of rice test field, prevent and treat on every side as blank (C)) in plantation paddy rice (No. 6, Henan round-grained rice) in July 7 and each dispenser on July 14 once, prevent and treat the effect of stripe disease respectively at the 7th day and observation in the 14th day after the last medication.Shown in table 3-1 and 3-2, A and B do not have evident difference to the result, can replace use fully respectively.
Table 3-1 contains the composition of Ningnanmycin and the control efficiency table of Ningnanmycin after the 7th day of equivalent
| Processed group | The rice strain number of sampling | Diseased plant rate (%) | Prevent and treat efficient (%) |
| A | 692 | 9.0 | 51.5 |
| B | 649 | 8.9 | 52.2 |
| |
700 | 18.6 | - |
Table 3-2 contains the composition of Ningnanmycin and the control efficiency table of Ningnanmycin after the 14th day of equivalent
| Processed group | The rice strain number of sampling | Diseased plant rate (%) | Prevent and treat efficient (%) |
| A | 683 | 10.0 | 58.2 |
| B | 632 | 10.1 | 57.7 |
| C | 712 | 23.9 | - |
Observed in one month after the medication, each processed group does not have any bad reaction to paddy growth, does not observe the drug toxicity incident of injury yet.This shows that the composition that contains Ningnanmycin can replace Ningnanmycin to be used to prevent and treat stripe disease separately.
(2) field control effectiveness test of MRDV
In Xiuqian City, Jiangsu Province, (A, B respectively test half area to 2 mu of corn experimental field, all spray simultaneously Imidacloprid WP, prevent and treat on every side as blank (C)) in plantation corn (face difficult to understand No. 1) in dispenser on July 4 once, respectively at the effect of the 15th day and the 30th day observation control of maize rough dwarf disease after the medication.Shown in table 3-3 and 3-4, A and B do not have evident difference to the result, can replace use fully respectively.
The Ningnanmycin that table 3-1 contains the composition of Ningnanmycin and equivalent control efficiency table composite after the 15th day with Imidacloprid WP
| Processed group | The rice strain number of sampling | Diseased plant rate (%) | Prevent and treat efficient (%) |
| A | 372 | 1.1 | 83.6 |
| B | 339 | 1.2 | 82.1 |
| C | 299 | 6.7 | - |
Table 3-2 contains the control efficiency table of Ningnanmycin Imidacloprid WP after the 30th day of the composition of Ningnanmycin and equivalent
| Processed group | The rice strain number of sampling | Diseased plant rate (%) | Prevent and treat efficient (%) |
| A | 323 | 1.2 | 83.1 |
| B | 332 | 1.2 | 83.1 |
| C | 310 | 7.1 | - |
One and a half months is observed after the medication, and each processed group does not have any bad reaction to corn growth, does not observe the drug toxicity incident of injury yet.This shows that the composition that contains Ningnanmycin can replace Ningnanmycin with the composite control of maize rough dwarf disease that is used for of other agricultural chemicals.
Claims (10)
1. comprise the composition of Ningnanmycin, it is characterized in that, it also comprises formula as follows 2 compounds and formula 3 compounds
Be preferably composition pesticide, more preferably it is the composition pesticide of the disease that causes of bacterial blight-resisting, powdery mildew and/or tobacco mosaic virus.
2. the described composition pesticide of claim 1, its water content is lower than 0.5% (w/w), preferably is lower than 0.4% (w/w)
3. the described composition pesticide of claim 1, wherein the content of Ningnanmycin is preferably 40-50% (w/w) greater than 35% (w/w), more preferably 40-45% (w/w).
4. claim 1 or 3 described composition pesticides, the content of its Chinese style 2 compounds is 35-55% (w/w), the content of formula 3 compounds is 7-23% (w/w).
5. the described composition pesticide of claim 1, it is to prepare by the time that prolongs fermented and cultured, more preferably it prepares by the following method:
(1) rejuvenation streptomyces noursei Xichang mutation, preferably rejuvenation under the condition of 28 ℃ and 80% relative moisture;
(2) cultivate mycelium on solid culture medium, preferably aerobic was cultivated 14 days under the condition of 28 ℃ and 80% relative moisture;
(3) carry out enrichment culture in the liquid medium within, preferably under 28 ℃ condition, cultivated 14 days;
(4) carry out fermented and cultured in fermentation tank, the time of preferred fermented and cultured is preferably greater than 70 hours, more preferably greater than 70.5 hours, as 72 hours, is more preferably under the condition of throughput of 28 ℃ and 8L/ hour and cultivates; With
(5) purifying preferably includes acidifying, filtration and drying.
6. prepare the described method for compositions of claim 1, it comprises:
(1) rejuvenation streptomyces noursei Xichang mutation, preferably rejuvenation under the condition of 28 ℃ and 80% relative moisture;
(2) cultivate mycelium on solid culture medium, preferably aerobic was cultivated 14 days under the condition of 28 ℃ and 80% relative moisture;
(3) carry out enrichment culture in the liquid medium within, preferably under 28 ℃ condition, cultivated 14 days;
(4) carry out fermented and cultured in fermentation tank, the time of preferred fermented and cultured is preferably greater than 70 hours, more preferably greater than 70.5 hours, as 72 hours, is more preferably under the condition of throughput of 28 ℃ and 8L/ hour and cultivates; With
(5) purifying preferably includes acidifying, filtration and drying.
7. pesticidal preparations, it comprises acceptable assistant on described composition pesticide of claim 1 and the agricultural chemicals, preferred described auxiliary agent is neopelex and diatomite.
8. described composition of claim 1 or the described pesticidal preparations of claim 7 application in the viroses of plant, mycosis and/or bacteriosis that the control Ningnanmycin can be prevented and treated preferably prevents and treats the disease that bacterial leaf-blight, powdery mildew and/or tobacco mosaic virus cause.
9. formula 2 compounds or be used for preventing and treating the composition pesticide of the disease that bacterial leaf-blight, powdery mildew and/or tobacco mosaic virus cause or the application of pesticidal preparations, preferred formula 2 compound cotypes, 3 compounds and/or Ningnanmycin applied in any combination in preparation.
10. formula 3 compounds are used for preventing and treating the composition pesticide of the disease that bacterial leaf-blight, powdery mildew and/or tobacco mosaic virus cause or the application of pesticidal preparations, preferred formula 3 compound cotypes, 2 compounds and/or Ningnanmycin applied in any combination in preparation.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010105681260A CN101978827B (en) | 2010-12-01 | 2010-12-01 | Ningnanmycin composition and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010105681260A CN101978827B (en) | 2010-12-01 | 2010-12-01 | Ningnanmycin composition and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101978827A true CN101978827A (en) | 2011-02-23 |
| CN101978827B CN101978827B (en) | 2012-11-28 |
Family
ID=43599391
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2010105681260A Active CN101978827B (en) | 2010-12-01 | 2010-12-01 | Ningnanmycin composition and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101978827B (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102515971A (en) * | 2011-12-11 | 2012-06-27 | 湖北仙隆化工股份有限公司 | Rice seed dressing agent composition used for preventing and treating black-streaked dwarf disease and its preparation method |
| WO2015160620A1 (en) | 2014-04-16 | 2015-10-22 | Bayer Cropscience Lp | Compositions comprising ningnanmycin and an insecticide |
| WO2015160619A1 (en) | 2014-04-16 | 2015-10-22 | Bayer Cropscience Lp | Compositions comprising ningnanmycin and a fungicide |
| WO2015160618A1 (en) | 2014-04-16 | 2015-10-22 | Bayer Cropscience Lp | Compositions comprising ningnanmycin and a biological control agent |
| US9192166B2 (en) | 2013-02-11 | 2015-11-24 | Bayer Cropscience Lp | Compositions comprising a streptomyces-based biological control agent and an insecticide |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106635835A (en) * | 2016-12-20 | 2017-05-10 | 陕西麦可罗生物科技有限公司 | Medium for ningnanmycin producing strain and preparation method of medium |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1093869A (en) * | 1993-04-23 | 1994-10-26 | 中国科学院成都生物研究所 | A kind of new antibiotic pesticides-Ningnan Meisu |
-
2010
- 2010-12-01 CN CN2010105681260A patent/CN101978827B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1093869A (en) * | 1993-04-23 | 1994-10-26 | 中国科学院成都生物研究所 | A kind of new antibiotic pesticides-Ningnan Meisu |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102515971A (en) * | 2011-12-11 | 2012-06-27 | 湖北仙隆化工股份有限公司 | Rice seed dressing agent composition used for preventing and treating black-streaked dwarf disease and its preparation method |
| CN102515971B (en) * | 2011-12-11 | 2013-09-04 | 湖北仙隆化工股份有限公司 | Rice seed dressing agent composition used for preventing and treating black-streaked dwarf disease and its preparation method |
| US9192166B2 (en) | 2013-02-11 | 2015-11-24 | Bayer Cropscience Lp | Compositions comprising a streptomyces-based biological control agent and an insecticide |
| WO2015160620A1 (en) | 2014-04-16 | 2015-10-22 | Bayer Cropscience Lp | Compositions comprising ningnanmycin and an insecticide |
| WO2015160619A1 (en) | 2014-04-16 | 2015-10-22 | Bayer Cropscience Lp | Compositions comprising ningnanmycin and a fungicide |
| WO2015160618A1 (en) | 2014-04-16 | 2015-10-22 | Bayer Cropscience Lp | Compositions comprising ningnanmycin and a biological control agent |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101978827B (en) | 2012-11-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106947725B (en) | Microbial agent and application thereof in saline-alkali soil improvement | |
| CN101978827B (en) | Ningnanmycin composition and preparation method thereof | |
| CN112481167A (en) | Compound microbial agent for improving soil fertility and preparation method thereof | |
| CN103992184B (en) | A kind of biological organic fertilizer | |
| CN103184184B (en) | Streptomyces albidoflavus and applications thereof | |
| CN103396971B (en) | Burkholderia cepacia and application thereof | |
| CN107937326B (en) | Paenibacillus polymyxa for preventing and treating root rot of American ginseng and application thereof | |
| CN106007959A (en) | Method for producing biomass fertilizer by virtue of livestock and poultry manures and straws | |
| CN106635903A (en) | Growth-promoting bacteria combination for enhancing salt tolerance of crops in moderate-severe saline and alkaline lands | |
| CN109504622A (en) | A kind of lysine bacillus and the composite bacteria agent of bacillus subtilis and preparation method thereof | |
| CN109796279A (en) | A kind of multi-element biological organic fertilizer and preparation method thereof | |
| CN108587636A (en) | Adjust soil conditioner and preparation method that soil acidity or alkalinity alleviates the soil salinization | |
| CN105002121A (en) | Bacillus simplex and application thereof | |
| CN102067840B (en) | Compound pesticide containing triflumizole and ningnanmycin | |
| CN107509744B (en) | Compound biopesticide for preventing and treating wheat stem basal rot and preparation method and application thereof | |
| CN102067855B (en) | Polyoxin and ningnan mycin compounded pesticide | |
| CN110157641B (en) | Biocontrol bacterium BV23 for preventing and treating corn stem-based rot and application thereof | |
| CN102067839B (en) | Composite pesticide of azoxystrobin and ningnan mycin | |
| CN102719363B (en) | Preparation method of antibacterial fermentation liquid of Solidago canadesis endophytic fungi | |
| CN102067843A (en) | Compound pesticide of tebuconazole and ningnan mycin | |
| CN106007958A (en) | Disease-resistant organic fertilizer microorganism inoculant | |
| CN102160549B (en) | Compound pesticide containing cyprodinil and ningnanmycin | |
| CN105018393A (en) | Bacillus megaterium strain and application thereof | |
| CN105936879B (en) | Bacillus subtilis K13 and its cultural method and application | |
| CN114946885B (en) | Microbial pesticide for resisting leaf spot disease of polygonatum odoratum |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |