JPH0246001B2 - - Google Patents

Description

[Detailed description of the invention]

(Industrial Application Field) The present invention relates to a plant disease control agent containing antibiotic SF-2370 as an active ingredient. (Prior Art) The present inventors, while searching for substances effective against plant diseases, discovered that the novel antibiotic substance SF-2370 is effective against various plant diseases including rice leaf blight. Substance SF-2370 is a new antibiotic that was previously filed by the applicant (October 9, 1982).
(See specification of Japanese Patent Application No. 59-210524 filed in Japan)
However, the present inventors have now discovered that it is also useful as a plant disease control agent. (Problem to be solved by the invention) Since 1942, many antibiotics have been discovered and put into practical use in the fields of pharmaceuticals, veterinary drugs, preservatives, agricultural chemicals, etc., and their excellent selective activity has been reconsidered in recent years. There is. However, there are still many medical and industrial fields that remain unsolved because no effective substances have been found. For example, rice leaf blight, peach borer bacterium, and vegetable soft rot have not been adequately controlled because there are no accurate control agents. Furthermore, the emergence of more effective control agents for rice sheath blight, rice blast, etc. is expected. The present inventors focused on the above problems, and
The aim is to solve this problem by discovering and providing new pest control agents. (Means for Solving the Problems) In order to meet the above-mentioned expectations, the present inventors continued to search for substances effective against plant diseases.
Antibiotic SF-2370 substance is used against rice leaf blight and other diseases.
They discovered that it is effective against various plant diseases, and thereby completed the present invention. Therefore, the present invention provides a plant disease control agent characterized by containing the antibiotic SF-2370 substance as an active ingredient. The plant disease control agent according to the present invention takes any form or usage mode that can be adopted as an agricultural and horticultural agent, especially a fungicide, except that the active ingredient is the above-mentioned SF-2370 substance. be able to. Specifically, for example, the SF-2370 substance can be used as it is, or diluted with water, solid powder, or other suitable carrier, and if necessary, an auxiliary agent such as a spreading agent can be added, or it can be used as a pesticide. Various liquid or solid carriers are mixed according to methods commonly used in manufacturing, and if necessary, auxiliary agents such as wetting agents, spreading agents, dispersing agents, emulsifying agents, fixing agents, and lubricants are added. In addition, it can be used in various formulations such as wettable powders, solutions, emulsions, powders, granules, and fine granules. In producing these preparations, the liquid carrier used is one that acts as a solvent for the SF-2370 substance;
Alternatively, those that can be dispersed or dissolved with the aid of adjuvants are used. For example, water, aromatic hydrocarbons, aliphatic hydrocarbons, alcohols, esters, ketones, highly polar dimethylformamide, dimethyl sulfoxide, etc. Solid carriers include clay, kaolin, talc, diatomaceous earth, bentonite, Mineral powders such as calcium carbonate and silica,
Gravel, wood flour and other organic powders and granules can be used. Non-ionic as an adjuvant,
Thickening materials such as anionic, cationic, and amphoteric surfactants, ligninsulfonic acid or its salts, gums, fatty acid salts, and methylcellulose are listed. (Action of the Invention) The plant disease control agent of the present invention can be used by spraying on the foliage of crops, and can also be applied to the surface of water, in water, on the surface of broken soil, or into broken soil. In this case, compatible agricultural and horticultural chemicals or fertilizers may be used. Such agricultural and horticultural chemicals include, for example, fungicides, insecticides, herbicides, and plant growth regulators. When using the plant disease control agent of the present invention as a liquid, the spray liquid usually contains 10
It is desirable to contain it at a concentration of 1000ppm to 1000ppm (in the case of concentrated small-volume spraying, aircraft spraying, etc., it can be used as a more concentrated spraying liquid as necessary), and powders, granules, fine granules, etc. When used as a compound, it is desirable that the content be 0.1 to 30%. The amount of application varies depending on the type and severity of the target disease, type of target crop, application mode, etc.
For example, if it is applied to paddy fields, if it is a hydrate (20% active ingredient) per 10 ares, for example, 50 to 200,
If it is a water solvent (10% active ingredient), for example, 50 to 200
For granules (5% active ingredient), for example, 2 to 6
Kg, for example, 2 to 6 if it is a powder (2% active ingredient)
Application rates of the order of Kg are generally appropriate. (Example) Next, examples of formulation of the plant disease control agent of the present invention will be described. However, the present invention is not limited to the following examples, and many modifications and modification methods not exemplified here are possible. It goes without saying that it is possible to adopt Formulation example 1 (Wettable powder) Ingredients by weight Parts SF-2370 20 Clay 10 Diatomaceous earth 65 Lignosulfonic acid calcium salt 3 Polyoxyethylene alkyl allyl ether 2 If the above ingredients are uniformly ground and mixed, the active ingredient 20 Obtain a hydrating agent containing %. Formulation example 2 (granules) Ingredients by weight : SF-2370 substance 10 Clay 87 Carboxymethyl cellulose 3 Mix the above ingredients, add an appropriate amount of water, knead into a mold, and then dry to obtain 10% of the active ingredient. Obtain granules containing: Formulation Example 3 (Powder) Ingredients by Weight SF-2370 Substance 3 Calcium Stearate 1 Silicic Anhydride Powder 1 Clay 47 Talc 48 By uniformly grinding and mixing the above ingredients, a powder containing 3% of the active ingredient is obtained. Formulation example 4 (Emulsion) Ingredients by weight SF-2370 substance 20 N,N-dimethylformamide 20 Xylene 45 Polyoxyethylene alkyl phenyl ether
12 Calcium salt of alkylbenzenesulfonate 3 By uniformly mixing and dissolving the above ingredients, an emulsion containing 20% of the active ingredient is obtained. Formulation example 5 (Flowable agent) Ingredients Part by weight SF-2370 substance 25 Machine oil 67 Polyoxyethylene alkyl phenyl ether
5 Calcium salt of alkylbenzenesulfonate 3 By uniformly mixing and pulverizing the above ingredients, a flowable agent containing 25% of the active ingredient is obtained. (Effects of the Invention) The excellent efficacy of the plant disease control agent of the present invention will be demonstrated through test examples. Test Example 1 Antibacterial test against plant pathogenic bacteria Using the plant pathogenic bacteria shown in the table below as test bacteria, the presence or absence of growth of the test bacteria on an agar plate was investigated.
The minimum inhibitory concentration (MIC) was determined. That is,
SF-2370 substance was mixed into a potato decoction agar medium to create a dilution series, which was then poured into a shear dish to solidify to create an agar plate. Inoculate the test bacteria onto the agar plate, and inoculate it at 25℃ for 48 hours in the case of bacteria.
In the case of filamentous fungi, the presence or absence of growth of the test bacteria was observed after culturing for 72 hours. The MIC measurement results are shown in the table below.

【table】

[Table] Test Example 2 Control effect test on rice leaf blight (1) Rice seedlings at the 5-leaf stage (cultivar ``Jukoku''), 8 seedlings each grown in plastic pots with a diameter of 6.5 cm, were treated with 28
A single needle inoculated with rice leaf blight bacteria cultured on potato semi-synthetic slant medium for 72 hours at ℃. 24 hours after inoculation, the hydrating powder produced in Formulation Example 1 was prepared into a spray solution of a predetermined concentration, and sprayed onto the rice plants at a rate of 40 ml/3 pots using a spray gun of a chemical spraying device. After air-drying, it was placed in an artificial climate room at 28°C during the day and 23°C at night, and the length of the lesion was measured after 10 days, and the control value was calculated using the following formula. Control value (%) = (1 - average lesion length of treated plots/average lesion length of non-treated plots) × 100 The occurrence of chemical damage was also visually observed. The results were as shown in the table below.

[Table] Test Example 3 Test for control effect on rice white leaf blight (2) Rice seedlings grown in the same manner as in Test Example 2 were transferred with their pots to separate plastic beakers, and filled with water to form a paddy field. A predetermined amount of the granules produced in Formulation Example 2 were sprinkled on the edge of rice seedlings, and 5 days later, rice leaf blight bacteria were inoculated in the same manner as in Test Example 2. The pest control effect was as shown in the table below.

[Table] Test Example 4 Control effect test on rice sheath blight A flowable agent produced according to Formulation Example 5 was applied at a predetermined concentration to paddy rice (cultivar "Jukoku") at the heading stage grown in a 1/5000 are Wagner pot. A spray solution was prepared and sprayed at a rate of 70 ml/3 pots using a chemical spray gun (2 kg/cm ² ). Immediately after air-drying, the sheath blight bacteria obtained by culturing on a peptone-added potato decoction agar medium for 48 hours were punched out using a cork borer with a diameter of 0.5 cm, and the mycelia-containing agar piece was placed in the center of the plant, 15 cm from the ground surface. It was inserted into the area and inoculated. After inoculation, each pot was covered with a vinyl cylinder to encourage the invasion of the sheath blight fungus and left in a glass greenhouse at 30°C during the day and 24°C at night to develop the disease. Ten days after inoculation, the infected stems were examined. The lesion length was measured and the control value was calculated according to the following formula. At the same time, the occurrence of drug damage was also observed. Control value = (1 - average lesion length of treated area/average lesion length of untreated area) x 100 The results were as shown in the table below.

[Table] Test Example 5 Rice blast control efficacy test The emulsion prepared in Formulation Example 4 above was applied to four-leaf stage rice seedlings (cultivar "Jukoku") grown in plastic pots of 6.5 cm in diameter, with eight seedlings each. A spraying liquid of a predetermined concentration was prepared and sprayed using a spray gun at a rate of 40ml/3 pots. After the rice seedlings were air-dried, they were inoculated by uniformly spraying a conidial suspension of rice blast fungus, and kept in a humid room at 24°C overnight. Afterwards, they were transferred to an artificial climate room at 25℃ to develop the disease, and 7 days after inoculation,
The number of lesions was counted, and the control value was calculated using the following formula. Control value (%) = (1 - average number of lesions in treated area/average number of lesions in untreated area) x 100 The results were as shown below.

[Table] Test Example 6 Chinese cabbage soft rot control efficacy test When 10 true leaves of Chinese cabbage (cultivar "Shonan No. 2"), which were grown in three pots each in a No. Prepare the wettable powder to the specified concentration and use a spray gun to
It was sprayed at a rate of ml/3 pots. After air-drying, Chinese cabbage soft rot bacteria cultured in potato decoction liquid medium,
Multiple needles were inoculated into the midrib of Chinese cabbage. After keeping it in a humid room for 1 day and night, it was transferred to an artificial climate room to allow the disease to develop, and after 2 days, depending on the degree of disease onset, 0 for no disease, 5 for the entire plant, and 1 to 4 for the middle. The disease severity was investigated by giving an index, and the control value was calculated using the following formula. Control value (%) = (1 - average disease severity in treated plots/average disease severity in untreated plots) x 100 In addition, the occurrence of leaf damage was determined by intraocular observation at the time of disease onset investigation. The results were as shown in the table below.

【table】

[Table] Antibiotics used as active ingredients in the present invention
Since the SF-2370 substance is a new substance, it will be explained below. SF-2370 material can be summarized as follows:
It is a neutral substance that exhibits the following properties: pale yellow crystalline, molecular formula is O ₂₇ H ₂₁ N ₃ O ₅ , molecular weight is
467 (mass spectrometry, FD-MS)・The melting point is 256℃, the specific optical rotation is [α] ²⁵ _D +57゜ (c0.1・methanol),
For color reactions, Greig-Rieback
Leaback) reagent, negative for ninhydrin and ferric chloride reagents, soluble in pyridine, dimethylformamide, dimethyl sulfoxide, methyl cellosolve and chloroform, sparingly soluble in methanol and ethyl acetate, insoluble in water and n-hexane. It is an antibiotic that is characterized by: Furthermore, the physical and chemical properties of SF-2370 substance are detailed. (1) Appearance: Pale yellow crystal (2) Elemental analysis: C69.50%, H4.74%, N8.88% (3) Molecular formula: C ₂₇ H ₂₁ N ₃ O ₅ (4) Molecule Amount: 467 (Mass spectrometry. FD-MS) (5) Melting point: 256°C (6) Specific rotation: [α] ²⁵ _D +57° (c0.1.methanol) (7) Ultraviolet absorption spectrum: In methanol
228nm (E1% 1cm725), 250nm (E1% 1cm717),
265nm shoulder, 280nm shoulder, 290nm (E1% 1cm1708), 320nm shoulder, 335nm (E1% 1cm432), 350nm (E1% 1cm325) and
It exhibits characteristic absorption at 367 nm (E1% 1 cm353). These absorptions do not change significantly even when acids or alkalis are added. (8) Infrared absorption spectrum: The infrared absorption spectrum measured in the potassium bromide tablet is as shown in Figure 1 of the attached drawings. (9) Nuclear magnetic resonance spectrum: ^{The 1} H nuclear magnetic resonance spectrum at 400 MHz measured in deuterated chloroform is shown in Figure 2, and ^{the 13} C nuclear magnetic resonance spectrum at 100 MHz is shown in Figure 3. (10) Silica gel thin layer chromatography (Kieselgel 60F254): Developing solvent Rf value ethyl acetate 0.58 Chloroform-methanol (20:1) 0.63 Benzene-acetone (2:1) 0.45 (11) Color reaction: Greig-Lieback ( Positive for Greig-Leaback reagent, negative for ninhydrin and ferric chloride reagents. (12) Solubility: Soluble in pyridine, dimethylformamide, dimethyl sulfoxide, methyl cellosolve and chloroform, sparingly soluble in methanol/ethyl acetate, water, n
- Insoluble in hexane. (13) Classification of basic, acidic, and neutral: Neutral Table 7 shows another antibacterial and antifungal spectrum of SF-2370 substance obtained by the agar plate dilution method.

【table】

[Table] The new antibiotic SF-2370 substance is manufactured using antibiotics belonging to the genus Actinomadura.
This is carried out by culturing the SF-2370 strain as an SF-2370 substance-producing bacterium, and collecting the antibiotic SF-2370 substance from the culture. SF used in the production of antibiotic SF-2370 substances
Strain -2370 is a strain newly isolated from the soil of Shimizu City, Shizuoka Prefecture, and the mycological properties of this strain are as follows. Morphology The basal hyphae are well elongated and branched and do not divide under normal conditions. Aerial mycelium generally has poor adhesion, but on oatmeal agar it adheres relatively well and spore formation is abundant. Spore formation on aerial mycelium is also observed on glycerol-asparagine agar, starch agar, etc. The branches of the aerial hyphae are simple branches, and no axillary branches are observed. The spore chain at the tip of the aerial hyphae is straight;
It becomes wavy, hook-shaped, or loop-shaped. When observed using an electron microscope, the spores are elliptical to short cylindrical, and have a size of 0.4 to 0.8 x 0.8 to 1.2 μm.
The surface is smooth. There are often 10 to 20 spore chains. Growth status on various media

【table】

[Table] Physiological properties (1) Growth temperature range: 15% on oatmeal agar
It grows in a temperature range of ~37°C and grows well at 25-32°C. (2) Liquefaction of gelatin: Negative (20℃, 21 days culture) (3) Starch hydrolysis: Negative (28℃, 14 days culture) (4) Nitrate reduction: Negative (28℃, 14 days culture) ( 5) Peptonization of skim milk: positive (35℃), negative (28℃) Coagulation of skim milk: positive (35℃), negative (28℃) (6) Production of melanin-like pigment: negative. Utilization of carbon source Basal medium [yeast extract (manufactured by Difco) 0.5%,
Calcium carbonate 0.1%, agar (manufactured by Difco) 1.5%]
1% of each carbon source was added and observed. (1) Used: D-glucose, D-fructose, L-rhamnose (2) Not used: D-mannitol, i-inositol, raffinose, sucrose (3) Suspected use: D-xylose, L-arabinose Cell wall composition 2,6-diaminopimelic acid in the whole cell hydrolyzate was in the meso form, and a small amount of madeulose was observed as a sugar. This SF-2370 strain was named Actinomadura sp. SF-2370, and the SF-2370 strain has been entrusted to the Institute of Microbial Technology, Agency of Industrial Science and Technology since July 28, 1982. It has been deposited with the number FERM P-7760. In the method for producing the SF-2370 substance, the bacteria described above are cultured in a medium containing nutrients that can be utilized by common microorganisms. As the nutrient source, known nutrient sources conventionally used for culturing actinomycetes can be used. For example, glucose, sucrose, starch, glycerol, starch syrup, molasses, animal/vegetable oils, etc. can be used as carbon sources. Also, as a nitrogen source, soybean flour,
Wheat germ, meat extract, peptone, yeast extract, cornstarch liquor, ammonium sulfate, sodium nitrate, and the like may be used. Others, as necessary.
It is useful to add soluble non-salts capable of producing sodium, potassium, calcium, magnesium, cobalt, chlorine, phosphate, sulfate and other ions. In addition, organic and inorganic substances that aid the growth of bacteria and promote the production of antibiotic SF-2370 substances may be appropriately added. The most suitable culture method is a culture method under aerobic conditions, especially deep culture. The appropriate temperature for culturing is
The temperature is 15-37°C, but in most cases it is cultured at around 26-30°C. Production of the antibiotic SF-2370 substance varies depending on the medium and culture conditions, but the accumulation usually reaches its maximum within 2 to 10 days in both shaking culture and tank culture. In collecting the obtained SF-2370 substance from the culture, its extraction and purification was carried out using Amberlite XAD-2 (manufactured by Rohm and Haas, USA) and Diaion HP-20 (manufactured by Mitsubishi Chemical Corporation). synthetic adsorbents such as Sephadex LH-20 (manufactured by Pharmacia Fine Chemicals, Sweden), precipitation with hexane, solvent extraction with ethyl acetate, columns with silica gel, etc. Although chromatography etc. can be used, the following collection method is efficient. That is, bacterial cells and other solid substances are separated from the culture solution using a supernatant such as diatomaceous earth, and then the active ingredients in the bacterial cells are extracted with 70% acetone water. After the extract is concentrated under reduced pressure to remove acetone, the concentrated solution is extracted with ethyl acetate. The extract is dehydrated over anhydrous sodium sulfate, concentrated under reduced pressure to remove ethyl acetate, and dried. This was further subjected to silica gel column chromatography using chloroform-ethyl acetate (10:1) as a developing solvent, and Sephadex LH using methanol as a developing solvent.
High purity SF-2370 substance can be obtained by appropriately combining column chromatography such as 20. Next, an example of manufacturing the new antibiotic SF-2370 substance is shown as a reference example. Reference example: glucose 2.0%, wheat germ 1.0, peptone 0.5
%, yeast extract 0.5%, and calcium carbonate 0.1% (PH 7.0) was dispensed into a 100 ml Erlenmeyer flask and sterilized at 120° C. for 15 minutes. This was inoculated with Actinomadilla sp. SF-2370 (Feikoken Bacterial Serial No. 7760), and was heated at 28°C for 7 days at a rate of 220% per minute.
Culture was performed by rotation. 20 ml of this culture was placed in a 1 volume jar fermentor containing 600 ml of production medium (PH 7.0) consisting of 1.5% glucose, 1.0% wheat germ, 1.0% cornstarch liquor, 0.5% firma media, and 0.3% calcium carbonate. The cells were inoculated and cultured with aeration at 28° C. for 5 days (aeration volume: 600 ml/min, rotation speed: 500 revolutions/min). After the culture was completed, diatomaceous earth was used as an auxiliary agent to obtain cultured bacterial cells. 500 ml of 70% acetone water was added to the bacterial cells to extract the active ingredient, and the bacterial cells were separated. Then, the bacterial cell extract was concentrated under reduced pressure to remove acetone, and the resulting concentrated liquid was
250 ml of ethyl acetate was added to 250 ml and shaken to extract the active ingredient. Repeat this extraction operation twice,
After the obtained ethyl acetate extraction, 500 ml of the extract was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain an oily substance. N-hexane was added to this oily substance, and the resulting precipitate was collected to obtain 286 mg of a crude substance. This crude substance was dissolved in a mixture of chloroform and ethyl acetate (10:1), and silica gel C-200 (manufactured by Wako Pure Chemical Industries, Ltd.) 60
ml column and developed with 600 ml of a chloroform-ethyl acetate (10:1) mixture. The developing solution is silica gel thin layer chromatography (Merck, Kieselgel, 60F254.5714; developing solvent: ethyl acetate)
and irradiated with ultraviolet light (254nm) to obtain an Rf value of 0.58.
The fractions showing antibacterial activity in a paper disk bioassay using Sarcina lutea as the test bacterium were collected and concentrated to dryness under reduced pressure to obtain 216 mg of pale yellow powder. This crude powder was dissolved in ethyl acetate and subjected to preparative silica gel thin layer chromatography (Merck Kieselgel 60F254,
5744, developing solvent: ethyl acetate), the active portion (Rf value 0.58) was scraped off, and extracted with ethyl acetate. After concentrating the extract under reduced pressure, methanol was added and left overnight to obtain 79mg of pale yellow crystals of SF-2370 substance.
was gotten.

[Brief explanation of the drawing]

Figure 1 shows the infrared absorption spectrum of SF-2370 substance in potassium bromide tablets, and Figure 2 shows the SF-2370 substance in potassium bromide tablets.
400MHz measured in deuterated chloroform with 2370 substances
¹ H nuclear magnetic resonance spectrum is shown, and Figure 3 shows SF-
100M measured in deuterated chloroform of 2370 substances
Hz ¹³ C nuclear magnetic resonance spectrum is shown.

Claims (1)

[Claims] 1. A plant disease control agent characterized by containing an antibiotic SF-2370 substance as an active ingredient.