JPH1135578A - Compound n-0285, its production and use - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject new compound capable of exhibiting excellent herbicidal, plant disease-containing, antitumor, antiarteriosclerotic, immunosuppressive and bronchodilation activities and useful as a medicine or an agricultural chemical by culturing a specific mold. SOLUTION: The subject compound is expressed by the formula. The compound is useful as an agricultural chemical (herbicide, plant disease-controlling agent, etc.), or a medicine (antitumor agent, antiasthmatic agent, antiarteriosclerotic agent, antiallergic agent, antirheumatic agent, etc.). The compound is obtained by culturing a non-identified strain No-155 (FERM P-16083) at a temperature of 20-28 deg.C at a pH of 5.0-8.0 usually for 8-16 days under an aerobic condition and subsequently isolating the compound from the culture product. When used as a herbicide or a plant disease-controlling agent, the compound is applied at a dose of 0.001-10 kg/ha. When used as an antitumor, the compound is administered at a daily dose of 0.01-100 mg/kg body.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a compound N-028
5. It relates to its production method and its use. Since the compound N-0285 of the present invention has various physiological activities such as herbicide, it can be used not only as a herbicide but also as a pesticide, a medicine or an intermediate thereof. Therefore, the present invention is also heavily used in these technical fields.

[0002]

BACKGROUND OF THE INVENTION As a compound having a chemical structure similar to the compound N-0285 according to the present invention, Brefeldin A (Bref
eldin A) [Betina et al., Acta Chem. Scand., 19: 519 (1
965)], Brefeldin C [Hutchinso
n, et al., J. Am. Chem. Soc., 103: 5956-5959 (1981)], and the compound N-0285 is clearly distinguished from these known compounds by different molecular formulas and various absorption spectra.

[0003]

Conventionally, various physiologically active substances produced by microorganisms have been known. However, there are only a few substances that are actually used as agricultural chemicals or pharmaceuticals, and new physiologically active substances are always required. . An object of the present invention is to provide compound N-0285, a production method and a use thereof.

[0004]

The present inventors have conducted various searches for substances produced by microorganisms in order to meet the above expectations. It was found that a strong herbicidally active substance was produced in a culture of a filamentous fungus designated 155, and the active ingredient was successfully isolated as Compound N-0285. The present inventors have studied in detail the physiological activity of the isolated compound N-0285, and have found usefulness as an agricultural chemical such as herbicidal activity and plant disease control activity. Furthermore, it was confirmed that the compound of the present invention has antitumor activity, antiatherosclerotic activity, immunosuppressive activity, and bronchodilator activity, and found usefulness as a medicament, thereby completing the present invention.

The present invention provides the following formula:

Embedded image Which provides compound N-0285 represented by the formula:

Further, the present invention relates to a compound N-
0285-producing bacterium was cultured and the compound N-
0285 is collected in the method for producing compound N-0285. As an example of the compound N-0285-producing bacterium used in the present invention, No. 1 newly isolated from the soil of Tsukiyono-cho, Gunma Prefecture is used. There are 155 strains.

Further, the present invention provides a pesticide, particularly a herbicide, a plant disease controlling agent or a medicament, particularly an antitumor agent, an anti-asthmatic agent, an anti-atherosclerotic agent, which comprises the compound N-0285 as an active ingredient. And anti-asthmatic, anti-allergic and anti-rheumatic drugs.

BEST MODE FOR CARRYING OUT THE INVENTION The microorganism of the present invention has the following properties.

1) No. Mycological properties of 155 strains Growth state in various media When the 155 strain was cultured on an oatmeal agar medium at 25 ° C. for 10 days, a colony having a white or pale yellow fluffy shape with a diameter of about 20 to 22 mm and showing a smooth but moderate bulge was formed. The back is pale yellow. The malt extract agar medium has a diameter of 12 to 15 mm (at 25 ° C. for 10 days), forms a white fluffy, smooth colony, and has a white back surface. 2 diameters on potato dextrose agar
0 to 25 mm (25 ° C., 10 days), the front and back surfaces showed the same properties as the oatmeal agar medium. On a Tzapek yeast agar medium, the diameter is 20 to 22 mm (at 25 ° C, 1 ° C).
(0 days), formed colonies showing white fluffy, smooth but moderate bumps. No sexual spores or conidia were formed in the media tested. At 37 ° C., it does not grow in any medium.

[0009] 2. Morphological features under a microscope The hyphae are 1-5 μm in diameter and are transparent to pale yellow, branched,
No bulkhead. No sexual spores or conidia were formed in the media tested.

[0010] No. from the above properties. Strain 155 cannot be identified as a specific genus of mold due to lack of characteristic reproductive organs and is therefore considered as an unidentified mold. This strain is identified by the Unidentified Strain No. 1
At the display of No. 55, the microfabrication laboratory No. 16083 (FERM P-
16083).

No. The 155 strain is liable to change its properties as seen in other molds. For example, no. 155 or a mutant (naturally occurring or inducible), transzygote or genetically modified from this strain,
Anything that produces compound N-0285 can be used in the present invention.

2) Cultivation of Compound N-0285 Producing Bacteria In order to produce compound N-0285 according to the present invention, first, a microorganism having the ability to produce compound N-0285 is cultured in a medium. As nutrient sources, known nutrients conventionally used for mold culture can be used. For example, as the carbon source, glucose, sucrose, starch syrup, starch, dextrin, soybean oil, and the like can be used. As a nitrogen source, peptone, meat extract, yeast extract, corn steep liquor, oatmeal, ammonium sulfate, ammonium chloride, ammonium nitrate, urea and the like can be used.
In addition, salt, magnesium sulfate, copper sulfate,
Inorganic salts such as zinc sulfate, manganese chloride, calcium carbonate, and phosphate can be added alone or in combination.In addition, organic substances that promote the growth of the strain and the production of compound N-0285, such as nucleic acids, vitamins, Trace elements such as inorganic substances can be appropriately added. When foaming during culturing is remarkable, an antifoaming agent or the like may be appropriately added.

The most suitable culture method is a culture method under aerobic conditions such as shaking or aeration and stirring culture. The culture temperature is 20 to 28 ° C, and the pH of the medium is 5.0 to 5.0.
It is preferably adjusted to 8.0. Culture time is usually 8 ~
After culturing for 16 days, compound N-0285 accumulates in the medium. Therefore, it is preferable to terminate the cultivation when the amount of accumulation in the culture reaches the maximum. In addition, these culture conditions may be selected optimally according to the characteristics of the production bacteria used or the culture method.

3) Purification of Compound N-0285 In order to collect Compound N-0285 from the culture after completion of the culture, a separation and purification method generally used for isolating a microbial metabolite from the culture is used. Used. That is, concentration under reduced pressure, lyophilization, for example, organic solvent extraction using butanol, ethyl acetate, chloroform, benzene, etc., various ion exchange chromatography, gel filtration chromatography using Sephadex LH-20, etc., activated carbon, silica gel, etc. Compound N-028 can be obtained by adsorbing or desorbing an active substance by adsorption chromatography or thin-layer chromatography, or by high-performance liquid chromatography using a reversed-phase column, alone or in combination in any order, and repeatedly.
5 can be isolated and collected.

The compound N-0285 thus obtained is
The following physicochemical properties are shown. 1. Molecular weight: 296 [(EI-MS): m / z 296
(M) ⁺ ]; Composition formula: C ₁₆ H ₂₄ O ₅ ; 3. Properties and color: colorless transparent oily neutral substance; 4. UV absorption spectrum: shown in FIG. Infrared external absorption spectrum: ν _max ^KBr (cm ⁻¹ ) = 355
6, 2945, 2935, 1712, 1261, 106
8, 981; shown in FIG. Solubility in solvent: chloroform, ethyl acetate,
6. Easily soluble in methanol, poorly soluble in water; ¹ H nuclear magnetic resonance spectrum: The ¹ H nuclear magnetic resonance spectrum was measured in deuterated methanol is shown in Fig. 3, and the chemical shifts (.delta. value) below: δ (ppm): 7.39 ( 1H, dd ), 5.85 (1
H, m), 5.82 (1H, dd), 5.23 (1H,
dd), 4.80 (1H, m), 4.06 (1H,
m), 3.87 (1H, m), 3.50 (1H, d
d), 2.12 (1H, m), 2.09 to 2.01 (2
H, m), 1.89 to 1.70 (5H, m), 1.59
(1H, m), 1.24 (3H, d), 0.91 (1
H, m); ¹³ C nuclear magnetic resonance spectrum: The ¹³ C nuclear magnetic resonance spectrum was measured in deuterated chloroform are shown in Figure 4, show that the chemical shifts (.delta. value) below: δ (ppm): 21.1,28.0 , 33.1, 35.
0, 35.7, 48.9, 52.9, 73.2, 76.
6,77.1,83.7,118.2,134.1,1
35.2, 154.5, 168.3; Rf value: silica gel thin layer chromatography (Kies
el gel 60F254, Merck) Ethyl acetate: methanol (10: 1 V / V%) shows 0.35.

Further, from the results of the above physicochemical properties and spectrum analysis, the chemical structure of compound N-0285 is represented by the following structural formula.

Embedded image Is shown.

The amount of the compound of the present invention to be applied as a herbicide varies depending on the application scene, application time, application method, target weeds, cultivated crops, etc., but generally 0.001 per hectare (ha) as an active ingredient. About 10 to 10 kg, preferably about 0.001 to 5 kg is appropriate.

If necessary, the compound of the present invention may be mixed with other kinds of herbicides, various insecticides, fungicides, plant growth regulators, synergists and the like at the time of preparation or spraying. In particular, by mixing and applying other herbicides, it is possible to expect a reduction in cost due to a reduction in the amount of applied medicine, an increase in the herbicidal spectrum due to the synergistic action of the mixed chemical, and a higher herbicidal effect. At this time, a combination with a plurality of known herbicides is also possible at the same time.

The amount of the compound of the present invention applied as a plant disease control agent varies depending on the application scene, application time, application method, target weeds, cultivated crops and the like, but is generally 0.001 per ha as an active ingredient. About 10 kg, preferably about 0.001 to 5 kg is appropriate.

The compound of the present invention may be mixed and used with other kinds of herbicides, various insecticides, fungicides, plant growth regulators, synergists and the like at the time of preparation or spraying as required. In particular, by mixing and applying other herbicides, it is possible to expect a reduction in cost due to a reduction in the amount of applied medicine, an increase in the herbicidal spectrum due to the synergistic action of the mixed chemical, and a higher herbicidal effect. At this time, a combination with a plurality of known herbicides is also possible at the same time.

When the compound of the present invention is applied as an agricultural chemical, it is generally mixed with a suitable solid carrier or liquid carrier, and if necessary, a surfactant, a penetrant, a spreading agent, a thickener, a deicing agent, A binder, an anti-caking agent, an anti-decomposition agent, etc. are added, and the composition can be put into practical use in any dosage form such as a liquid, emulsion, wettable powder, dry flowable, flowable, powder, and granule. Also, from the viewpoint of labor saving and safety improvement,
The preparation of any of the above dosage forms can also be provided in a water-soluble package.

Examples of the solid carrier include kaolinite,
Natural minerals such as pyrophyllite, sericite, talc, bentonite, acid clay, attapulgite, zeolite and diatomaceous earth; inorganic salts such as calcium carbonate, ammonium sulfate, sodium sulfate and potassium chloride; synthetic silicic acid; and synthetic silicate. .

Examples of the liquid carrier include water, alcohols (such as ethylene glycol, propylene glycol, and isopropanol), aromatic hydrocarbons (such as xylene, alkylbenzene, and alkylnaphthalene), ethers (such as butyl cellosolve), and ketones (such as cyclohexanone). And the like, esters (γ-butyrolactone, etc.), acid amides (N-methylpyrrolidone, N-octylpyrrolidone, etc.) and vegetable oils (soybean oil, rapeseed oil, cottonseed oil, castor oil, etc.).

These solid and liquid carriers may be used alone or in combination of two or more.

Examples of the surfactant include polyoxyethylene alkyl aryl ether, polyoxyethylene styryl phenyl ether, polyoxyethylene polyoxypropylene block copolymer, polyoxyethylene fatty acid ester, sorbitan fatty acid ester and polyoxyethylene sorbitan fatty acid ester. Nonionic surfactants, alkylbenzene sulfonates, lignin sulfonates, alkyl sulfosuccinates, naphthalene sulfonates, alkyl naphthalene sulfonates, salts of formalin condensates of naphthalene sulfonic acid, and alkyl naphthalene sulfonic acid formalin Salts of condensates, polyoxyethylene alkylaryl ether sulfates and phosphates, polyoxyethylene styryl phenyl ether sulfates and phosphates and salts Ionic surfactants such as triethanolamine salts. The content of these surfactants is not particularly limited, but is preferably in the range of usually 0.05 to 20 parts by weight based on 100 parts by weight of the preparation of the present invention. These surfactants may be used alone or in combination of two or more.

Examples of Agrochemical Formulations Examples of agrochemical formulations containing the compound of the present invention as an active ingredient are shown below, but the invention is not limited thereto.

Wetting agent 0.1 to 80 parts of the compound of the present invention 5 to 98.9 parts of a solid carrier 1 to 10 parts of a surfactant Other 0 to 5 parts Others include, for example, an anti-caking agent and an anti-decomposition agent.

Emulsion 0.1 to 30 parts of the compound of the present invention 55 to 95 parts of a liquid carrier Surfactant 4.9 to 15 parts

Flowable agent Compound of the present invention 0.1 to 70 parts Liquid carrier 15 to 98.89 parts Surfactant 1 to 12 parts Others 0.01 to 30 parts Others include, for example, an antifreezing agent and a thickener.

Dry flowable compound 0.1 to 90 parts of the compound of the present invention 0 to 98.9 parts of solid carrier 0 to 20 parts of surfactant 0 to 10 parts of others Others include, for example, a binder and a decomposition inhibitor.

Solution 0.01 to 30 parts of the compound of the present invention 0.1 to 50 parts of liquid carrier 50 to 99.89 parts of water Others 0 to 10 parts Others include, for example, antifreezing agents and spreading agents.

Granules 0.01 to 10 parts of the compound of the present invention 90 to 99.99 parts of solid carrier Others 0 to 10 parts Others include, for example, a binder and a decomposition inhibitor.

[0033] The compound of the present invention cropland example paddy particularly upland, as a non-arable land herbicide, soil treatment, even in the one of the processing method of the foliage treatment, Inuhouzuki (Solanu m nigr
um), Datura (Datura stramoniu m) the Solanaceae family, which typified by (Solanaceae) weed, velvetleaf (Abutilon
theophrasti ), American deer ( Sida spinosa )
( Malvacea e) weeds and Malagaea ( Ipomoe a purpurea ) and other morning glory ( Ipomoe a sp.)
ps.) and Morning Glory acids (Calystegia spps.) Convolvulaceae family (Convolvulaceae) weeds typified by, Amaranthus lividus (Amaran
Thus lividus ), Aobille ( Amaranthus retroflexus )
( Amaranthacea e) weeds, eel fir ( Xanthium pensylvanicum ), ragweed ( Ambrosia art )
emisiaefolia), sunflower (Helianthus annuu s), galinsoga quadriradiata (Galinsog a ciliat a), western thorns thistle (Cirsiu m arvens e), Senecio vulgaris (Seneci o vulgari
s ), Compositae weeds represented by Erigeron annus, etc., and Roripp a indic
a ), Sawfish ( Sinapi s arvensis ), Nasuna ( Cap )
cruciferae ( Cruc ) represented by sellaBursapastoris
iferae weeds, Polygonum m Blumei , Polygonaceae weeds ( Polygonaceae ) weeds represented by Polygonum convolvulu s, etc., Portulac a olerac
ea ), etc. ( Portulacacea e) weeds, Shiroza ( Chenopodiu malbu m), and Kookasa ( Chenopod )
iu m ficifolium), kochia (Kochia scopari a) Chenopodiaceae represented by like (Chenopodiaceae) weeds, chickweed (St
Caryophyll ( Caryophyll ) represented by ellari a medi a)
acea e) weeds, persian speedwell (Veronica persica) Scrophulariaceae represented by, etc. (Scrophulariaceae) weeds, dayflower (Commelin a communis) commelinaceae represented by such (Commelinacea e) weeds, henbit (Lamium a
mplexicaul e), Laminaria purpureum
( Labiatae ) weeds represented by, etc., cypress ( Euphorbi a supina ), and cypress ( Euphorbi a ma)
culata ) and Euphorbiacea ( Euphorbiacea)
e) Weeds, Galium spurium , Rubi ( Rubi )
a akan e) and other Rubiacea ( Rubiacea e) weeds and violets ( Viol a mandshuric a) and other violets ( Vi
olacea e) weeds, horned horned flowers ( Sesbania exalt)
at a), sicklepod (Cassia obtusifolia) legume that typified by (Leguminosa e) broad-leaved weeds such as weeds (Broad-leav
ed weeds), wild sorghum ( Sorgha m bicolo r), eucalypt ( Panicum dichotomiflorum ), Johnsongrass ( Sorghu m halepens e), dog millet ( Echinochlo a crus-g)
alli var. crus-galli ), Echinochlo a
crus-galli var. praticol a), cultivated flies ( Echinochlo a
utilis ), crabgrass ( Digitari a adscendens ), oats ( Avenafatua ), crabgrass ( Eleusine indica ), enokorogosa ( Setari a viridi s), and sparrows ( Al )
opecurus aeguali s) and other grass weeds (Gramin
aceous weeds, daylily ( Cyperu s rotundus , Cyperu)
s esculentus ) and other Cyperaceae weeds (Cy
peraceous weeds) and various upland weeds (Cropland weeds)
It has high herbicidal activity at low dose.

In addition, as a herbicide for paddy fields, in any of soil treatment and foliage treatment under flooded conditions, a hermit mosquito ( Alisma canaliculatum ) and a mosquito ( Sagittaria ) are used.
trifoli a), Uridagawa ( Sagittaria pygmaea ) and other weeds ( Alismataceae ) weeds, Cyperu s difformi s, Cyperu s seroti
nu s), fireflies ( Scirpu s juncoide s), Krogwai ( El
eocharis kuroguwai) Cyperaceae represented by, etc. (Cyperaceae) weed, false pimpernel (Scrophulariaceae represented by Lindemia pyxidaria), etc. (Scrothulariaceae) weeds, pontederiaceae represented by Monochoria (Monochoria vaginalis), etc. (Potenderiaceae) weed, pondweed ( Pota
mogeto n distinctus) potamogetonaceae represented by such as (Po
tamogetonaceae ), Lythraceae weed represented by Rotala indica, etc., Echinochlo a oryzicol a, and Echino
chlo a crus-galli var. formosensi s), dog millet ( Echin
ochlo a crus-galli var. crus-galli )
Paddy weeds have high herbicidal activity at low dose.

The compound of the present invention can be used as a herbicide for upland fields in any of soil treatment, soil admixture treatment and foliage treatment, and is used in sports fields other than agricultural and horticultural fields such as paddy fields, fields and orchards. It can also be applied to the control of various weeds in non-agricultural lands, such as vacant lots and track ends.

The compounds of the present invention have excellent microbicidal action.
Used as a fungicide in plant protection and substance storage
can do. Preventive and cure many plant diseases
It has pathological or systemic control effects. Next,
Rice plant blasts are plant diseases
disease(Pyriculariaoryzae), Sesame leaf blight (Cochliobolus
miyabeanus), sheath blight (Rhizoctoniasolani), Wheat
Powdery mildew(Erysiphegraminis f.sp.hordei, f.s
p.tritici), Leaf spot disease (Pyrenophoragraminea)
disease(Pyrenophora teres), Fusarium head blight(Gibberellazea
e), Rust (Puccinia striiformis, P.graminis, P.r
econdita,P.hordei), snow rot(Typhula sp., Micronectr
iella nivais), naked smut (Ustilago tritici, U.nud
a), eye spot(Pseudocercosporella herpotrichoid
es), Cloud disease (Rhynchosporium secalis), Leaf blight(Sep
toriatritici), Fusarium wilt (Leptosphaerianodorum), mosquito
Black spot disease of fox (Diaporthecitri), scab(Elsinoe
fawcetti), fruit rot (Penicilliumdigitatum, P.it
alicum), Monilia disease of apple(Sclerotinia mali),
Rot (Valsamali), Powdery mildew (Podosphaerale
ucotricha), spotted leaf disease (Alternaria mali), scab
(Venturia inaequalis) Pear scab(Venturianash
icola), black spot (Alternaria Kikuchiana), Scab(Gy
mnosporangium haraeanum), Peach ash disease(Sclerotini
a cinerea), Scab(Cladosporiumcarpophilum),
Omopsis rot(Phomopsis sp.), Grape downy mildew(P
lasmoparaviticola), Black rot (Elsinoeampelin
a), Late rot (Glomerella cingulata) Powdery mildew(Unc
inulanecator), rust (Phakopsora ampelopsidis),
Oyster anthracnose (Gloeosporium kaki) deciduous disease(Cercospo
ra kaki,Mycosphaerellanawae) Downy mildew of cucumber(Ps
eudoperenospora cubensis), Anthracnose (Colletotrichum
lagenarium), powdery mildew (Sphaerotheca fuliginea)
 Vine blight(Mycosphaerellamelonis) Tomato plague
(Phytophthorainfestans) wilt disease(Fusariumoxy
sporum), ring spot disease (Alternaria solani), leaf mold(Clad
osporiumfulvam), Eggplant brown spot(Phomopsis vexan
s), powdery mildew (Erysiphe cichoracoarum), Brassica
Black spot of family vegetablesAlternaria japonica), vitiligo disease(Ceroc
osporellabrassicae) Rust on leeks(Pucciniaalli
i), soybean purpura (Cercospora kikuchii), black rot
 (Elsinoeglycines), Sunspot (Diaporthephaseololu
m), bean anthracnose (Colletotrichum lindemuthianu
m), peanut black rot (Mycosphaerella personatu
m), brown spot(Cercosporaarachidicola), Peas
Powdery mildew (Erysiphe pisi), potato summer blight (Alte
rnaria solani), strawberry powdery mildew(Sphaerothecahu
muli), Tea net blast (Exobasidiumreticulatum),
Scab(Elsinoe leucospila), tobacco scab (Altern
aria longipes), powdery mildew (Erysiphe cichoracearu
m), Anthracnose (Colletotrichum tabacum), Of sugar beet
Brown spot(Cercosporabeticola), Rose scab (Diploca
rponrosae) powdery mildew(Sphaerothecapannosa),
Brown leaf spot (Septoria chrysanthemiindici), white rust
 (Puccinia horiana), Bentgrass brown patch
(Phizoctonia solani), picium bright(Pythium sp
p.), Dollar spot(Sclerotinia homoeocarpa), Each
Typhula spp,Fusarium nivale,Sclerotinia
borealis), Hermint sporium disease(Helminthosporiu
msorokinianum, H.erythrospilum), Noshiba, Koura
Various pysium diseases of seagrass(Pythium periplocum,P.gram
inicola,P.vanterpoolii), Leaf rot(Rhizoctonia sol
ani), gray mold on various crops (Botrytis cinerea),
Sclerotium disease (Sclerotiniasclerotiorum) And the like.
Therefore, the compound of the present invention can be used in fields, paddy fields, turfgrass, and fruit trees.
Effective for controlling plant diseases in orchards, meadows, greenhouses and other non-cultivated lands
It can be used as a component.

Furthermore, surprisingly, the compounds of the present invention are useful as pharmaceuticals because they have antitumor activity, antiatherosclerotic activity, immunosuppressive activity and bronchodilator activity.

The compound of the present invention is useful as an antitumor agent for treating acute leukemia, chronic myeloid leukemia, lymphoma, etc.
It is used for the treatment of arteriosclerosis, suppression of rejection during organ transplantation, autoimmune diseases (rheumatoid arthritis, nephritis, hepatitis, etc.), HIV, atopic dermatitis, bronchial asthma, rheumatism and the like. The dosage is about 0.01 to 1 kg / kg of patient's body weight every day.
The dose is in the range of about 100 mg, preferably in the range of about 0.01 to about 50 mg per kg of body weight every day, 1 to 3 times a day.
The dosage increases or decreases depending on age, body weight, etc.

When the compound of the present invention is used as a medicine, it is formulated for administration by conventional pharmaceutical means. That is, tablets, capsules, granules and pills for oral administration are
Excipients, for example, sucrose, lactose, glucose, starch, mannitol; binders, for example, hydroxypropylcellulose, syrup, acacia, gelatin, sorbit,
Tragacanth, methylcellulose, polyvinylpyrrolidone; disintegrants such as starch, carboxymethylcellulose or its calcium salts, microcrystalline cellulose, polyethylene glycol; lubricants such as talc, magnesium or calcium stearate, silica; lubricants such as sodium laurate; It is prepared using glycerol or the like.

Injections, solutions, emulsions, suspensions, syrups and aerosols can be used as a solvent for the active ingredient, for example, water, ethyl alcohol, isopropyl alcohol, propylene glycol, 1,3-butylene glycol, polyethylene glycol; Agents such as sorbitan fatty acid esters, polyoxyethylene sorbitan fatty acid esters,
Polyoxyethylene fatty acid esters, polyoxyethylene ethers of hydrogenated castor oil, lecithin; suspending agents, for example, cellulose derivatives such as carboxymethyl sodium salt and methylcellulose; natural gums, such as tragacanth and gum arabic; preservatives, for example, paraoxybenzoate It is prepared using acid esters, benzalkonium chloride, sorbate and the like.

As the ointment which is a transdermal absorption preparation, for example, white petrolatum, liquid paraffin, higher alcohol, macrogol ointment, hydrophilic ointment, aqueous gel base and the like are used.
Suppositories are prepared using, for example, cocoa butter, polyethylene glycol, lanolin, fatty acid triglycerides, coconut oil, polysorbate and the like.

[0042]

The production examples of the compound of the present invention are shown below. Various methods for producing the compound N-0285 can be devised based on the properties revealed by the present invention. Therefore,
This example does not limit the scope of the invention in any way.

Production Example In a 500 ml Erlenmeyer flask, 0.1% of polypeptone, 0.1% of yeast extract, and 10% of tomato juice (manufactured by Kagome Co., Ltd.) were added to a PD (potato dextrose) medium (manufactured by Difco). 100 ml of liquid medium consisting of composition
And autoclaved at 120 ° C. for 20 minutes. The unidentified strain No. One platinum loop of 155 was inoculated, and cultivation was performed at 25 ° C. at 140 rpm for 3 days. The culture solution obtained here was dispensed into a 5 liter Erlenmeyer flask (1
0, autoclaved) and transferred to 25 ° C, 14
Rotating culture was performed at 0 rotation / minute for 12 days. After cultivation,
The resulting culture was filtered to obtain a filtrate and cells.

[0044] This culture filtrate was used to prepare Diaion HP-20.
It was adsorbed on 500 ml (registered trademark, manufactured by Mitsubishi Chemical Corporation) and eluted with 2 liters of 75% methanol / water. The eluate was concentrated under reduced pressure to remove methanol, extracted three times with 500 ml of ethyl acetate, and the combined ethyl acetate solution was concentrated under reduced pressure to obtain 1.3 g of a brown oily substance. This was subjected to silica gel chromatography using chloroform-methanol (100: 0 to 20) as a developing solvent, and the fraction containing Compound N-0285 was concentrated to dryness to obtain about 400 mg of a light brown oily substance. Next, this fraction was subjected to thin layer chromatography (Kieselgel 60F254, manufactured by Merck) using ethyl acetate-methanol (10: 1) as a developing solvent, and an active fraction having an Rf value of 0.35 was collected. Is concentrated to dryness to give a colorless oily compound N-
Approximately 160 mg of 0285 was obtained.

The compound N-0285 thus obtained is
The following physicochemical properties are shown. 1. Molecular weight: 296 [(EI-MS): m / z 296
(M) ⁺ ] 2. Composition formula: C ₁₆ H ₂₄ O ₅ 3. Property and color: neutral substance with clear and colorless oil Ultraviolet absorption spectrum: shown in FIG. 5. Infrared external absorption spectrum: ν _max ^KBr (cm ⁻¹ ) = 355
6, 2945, 2935, 1712, 1261, 106
8, 981 shown in FIG. 6. Solubility in solvent: chloroform, ethyl acetate,
6. Easily soluble in methanol, poorly soluble in water ¹ H nuclear magnetic resonance spectrum: FIG. 3 shows the ¹ H nuclear magnetic resonance spectrum measured in deuterated methanol. The chemical shift (δ-value) is shown below. δ (ppm): 7.39 (1H, dd), 5.85 (1
H, m), 5.82 (1H, dd), 5.23 (1H,
dd), 4.80 (1H, m), 4.06 (1H,
m), 3.87 (1H, m), 3.50 (1H, d
d), 2.12 (1H, m), 2.09 to 2.01 (2
H, m), 1.89 to 1.70 (5H, m), 1.59
(1H, m), 1.24 (3H, d), 0.91 (1
H, m) 8. ¹³ C nuclear magnetic resonance spectrum: FIG. 4 shows a ¹³ C nuclear magnetic resonance spectrum measured in deuterated chloroform. The chemical shift (δ-value) is shown below. δ (ppm): 21.1, 28.0, 33.1, 35.
0, 35.7, 48.9, 52.9, 73.2, 76.
6,77.1,83.7,118.2,134.1,1
35.2, 154.5, 168.3 9. Rf value: silica gel thin layer chromatography (Kies
el gel 60F254, Merck) Ethyl acetate: methanol (10: 1 V / V%) shows 0.35.

Further, from the results of the above physicochemical properties and spectrum analysis, the chemical structure of Compound N-0285 was

Embedded image It is.

Next, the biological properties of Compound N-0285 will be specifically described with reference to Test Examples. However, it is not limited only to these. Test example Herbicidal activity test of compound N-0285 Put sterilized flood soil in a plastic box of 15 cm in length, 23 cm in width, and 8 cm in depth, and mixed seedlings of firgrass, morning glory, ibis, bluegrass, and ragweed, and covered about 1.5 cm. The plant was grown in a greenhouse at 25 to 30 ° C. for 14 days, and compound N-0285 prepared in the following emulsion was uniformly sprayed on the foliage so as to have a predetermined dose. Seven days after the application of the chemical, the herbicidal effect on the above-mentioned various weeds was investigated according to the following criteria. The results are shown in Table 1.

The symbols in the table have the following meanings. A: sea fir, B: morning glory, C: strawberry, D: blue vine, E: ragweed

The composition of the emulsion is shown below. Compound N-0285 20 parts Xylene 59 Parts Isophorone 15 parts Solpol 3005X 6 parts (non-ionic Mixture of anionic surfactant and anionic surfactant; trade name of Toho Chemical Industry Co., Ltd.) In the above blending examples, "parts" means parts by weight.

Criteria 5: Herbicidal rate 90% or more (almost completely killed) 4: Herbicidal rate 70% to 90% 3: Herbicidal rate 40% to 70% 2: Herbicidal rate 20% ~ 40% 1 ... Weed kill rate 5% ~ 20% 0 ... Weed kill rate 5% or less (almost no effect)

The above-mentioned herbicidal rate is obtained by measuring the amount of above-ground grass in the chemical-treated area and the above-ground grass rate in the untreated area according to the following formula. Herbicidal rate = {1- (amount of aboveground grass in treated area / amount of aboveground grass in untreated area)} × 100

[0052]

[Table 1] Table 1 量 Dose (g / a) A BCDE {10 5 5 3 3 3} ──────────────────────────────

As described above, by foliage treatment of Compound N-0285, each kind of broadleaf weeds caused significant growth inhibition on the 7th day after the treatment, and it is clear that the herbicidal effect is apparent. .

2. (A) Cucumber downy mildew controlling effect test Compound N-0285 is contained in 1.5-leaf stage cucumber (cultivar: Sagamihanjiro) grown in a pot having a diameter of 7 cm. 20 ml of a chemical solution prepared by diluting the emulsion described in Test Example 1 above with water to a predetermined concentration was sprayed per pot using a spray gun. The day after spraying, a spore suspension of Pseudoperonospora cubensis (2 × 1
0 ⁵ / ml) was sprayed, 25 ° C. cucumbers inoculated,
They were placed in an inoculation box with a humidity of 95% or more all day and night. Then, it was placed in a greenhouse and the ratio of the lesion area formed 7 days after inoculation to the inoculated leaves was measured, and the control value was calculated according to the following formula. Table 2 shows the test results. Control value = {1- (diameter of lesion treated with drug / diameter of lesion not treated with drug)} x 100

(B) Cucumber Powdery Mildew Control Effect Test The above-mentioned Test Example 1 containing the compound N-0285 in the 1.5-leaf stage cucumber (variety: Sagami Hanshiro) grown in a pot having a diameter of 7 cm. Was diluted with water and adjusted to a predetermined concentration, and a spray gun was used to spray 20 ml per pot using a spray gun. The day after spraying, a spore suspension of cucumber downy mildew (Sphaerotheca fuliginea) (3 × 10 ⁵
Per ml) and inoculated. Then put it in the greenhouse,
The ratio of the lesion area formed 10 days after the inoculation to the inoculated leaves was measured, and the control value was calculated according to the following formula. Table 2 shows the test results. Control value = {1- (diameter of lesion treated with drug / diameter of lesion not treated with drug)} x 100

(C) Rice Blast Control Effect Test The three-leaf stage rice (variety: Nipponbare) grown in a pot having a diameter of 7 cm was prepared by adding the emulsion described in the above Test Example 1 containing the compound N-0285 with water. Using a spray gun, 20 ml of the drug solution diluted to a predetermined concentration was used for each pot.
Sprayed. The day after spraying, the rice blast fungus (Pyricularia or
sprayed with a spore suspension (2 × 10 ⁵ cells / ml)
The inoculated rice was placed in an inoculation box at 25 ° C. and a humidity of 95% or more all day and night. Then, it was placed in a greenhouse and the ratio of the lesion area formed 7 days after inoculation to the inoculated leaves was measured, and the control value was calculated according to the following formula. Table 2 shows the test results. Control value = {1- (diameter of lesion treated with drug / diameter of lesion not treated with drug)} x 100

(D) Barley powdery mildew control effect test The above-mentioned test containing compound N-0285 in 1.5 to 2-leaf stage barley (cultivar: Norin 41) grown in a pot having a diameter of 5.5 cm. A drug solution prepared by diluting the emulsion described in Example 1 with water to a predetermined concentration was sprayed using a spray gun at a rate of 20 ml per pot. The day after spraying, barley leaves affected by barley powdery mildew (Erysiphe graminis f. Sp. Hordei) were applied and inoculated. Then, it was placed in a greenhouse and the ratio of the lesion area formed 7 days after inoculation to the inoculated leaves was measured, and the control value was calculated according to the following formula.
Table 2 shows the test results. Control value = {1- (diameter of lesion treated with drug / diameter of lesion not treated with drug)} x 100

(E) Wheat Leaf Rust Control Effect Test The above test example containing compound N-0285 in 1.5 to 2 leaf stage wheat (cultivar: Norin 61) grown in a pot having a diameter of 5.5 cm. A drug solution prepared by diluting the emulsion described in 1 with water to a predetermined concentration was sprayed using a spray gun to spray 20 ml per pot. The day after spraying, a spore suspension (2 × 10 ⁵ cells / ml) of wheat rust (Erysiphe graminis f. Sp. Hordei) was sprayed, and the inoculated wheat was placed in an inoculation box at 25 ° C. and a humidity of 95% or more for 24 hours. Was. Then, it was placed in a greenhouse and the ratio of the lesion area formed 7 days after inoculation to the inoculated leaves was measured, and the control value was calculated according to the following formula. Table 2 shows the test results. Control value = {1- (diameter of lesion treated with drug / diameter of lesion not treated with drug)} x 100

The symbols in the table have the following meanings. A: Cucumber downy mildew, B: Cucumber powdery mildew, C: Rice blast, D: Barley powdery mildew, E: Wheat red rust

[0060]

[Table 2] Table II 除 Control value (%) Spray concentration (ppm) ) ───────────────────────── ABCDE ─────────────────── {500 100 40 70 50 30 100 100 90 20 70 50 20} ───────────

As described above, by spraying compound N-0285, the lesions of various plant pathogenic fungi are remarkably suppressed, and it is clear that the disease control effect is exhibited. No phytotoxicity to useful crops due to spraying was observed.

3. Cell growth inhibitory activity test of compound N-0285 (A) Growth inhibitory effect on THP-1 cells Human leukemia Cellline THP-1 cells maintained in RPMI1640 medium containing 10% FBS
About 500000 in RPMI1640 medium containing 0% FBS
Cells / ml, and 96 well pla
100 μl of cell suspension per well was added to te. Compound N- in dimethyl sulfoxide (DMSO)
0285 is 500 times the processing density (5, 0.5, 0.
(0.5 mg / ml), and the drug solution diluted 2.5-fold with the above culture solution was added in an amount of 0.5 μl per well.
Was added. After 5 days of culture at 5% CO ₂ , 37 ° C., 5 m
50 μl of a g / ml MTT solution was added per well, and the culture was continued for another 4 hours. 0.01N hydrochloric acid / 2
-Add 50 μl of propanol solution per well,
After dissolving the MTT reaction, the absorbance was measured at 550 nm using a microplate reader. By comparing the absorbance of untreated cells and cells treated with a known concentration of the drug, the concentration of the drug that inhibits cell growth by 50% (IC
₅₀ , μg / ml). Test result IC ₅₀ = 0.08 μg / ml

(B) Inhibition of proliferation on intimal smooth muscle cells: An aortic intimal hypertrophy caused by arteriosclerosis caused by a high cholesterol diet load was prepared by an oxygen dispersion method, and 10% FBS was prepared.
Intimal smooth muscle cells subcultured in the containing DME medium were used. After treating the cells with 0.125% trypsin and 50% versin, the cells were suspended in 10% FBS-DME medium to a cell concentration of about 130,000 cells / ml. 4
50 μl of the cell suspension was added to an 8-well plate, and 200 μl of a 10% FBS-DME medium was further added. After culturing for 6 hours, the number of initially adherent cells was counted, and the medium after culturing was replaced with a 10% FBS-DME medium to which compound N-0285 diluted with DMSO was added. After culturing for 5 days, the cells were treated with trypsin, and the number of cells was counted using a Coulter counter. The drug concentration that inhibits cell proliferation by 50% (IC ₅₀ , μg / ml) by comparing the counts of untreated cells and cells treated with a known concentration of drug
Was calculated. Test result IC ₅₀ = 0.28 μg / ml

(C) Proliferation inhibitory effect on T and B lymphocyte cells The spleen was excised from BALB / c mice (male, 6 weeks old).
Spleen cells were prepared. A mononuclear cell fraction was collected by specific gravity centrifugation using Histopaque 1083 (d = 1.083) and used as a lymphocyte suspension (survival rate, purity> 90%). After centrifugation, the sedimentation was performed using RPMI-1640 medium (10% FBS,
0 μM 2-ME) in a 96-well plate.
Plated at a concentration of 0000 cells / well. Concanavalin A (con A) or lipopolysaccharide (LPS) was added thereto at a final concentration of 3, 20 μg, respectively.
/ Well, compound N-0285 diluted with DMSO was added to each concentration, and the cells were cultured in a 5% CO ₂ incubator for 3 days. After 3 days, add 10 μl of cell counting kit reagent to each well.
After adding / well and culturing again for 2 hours, a wavelength of 450
The absorbance at nm was measured. The proliferation caused by con A stimulation was defined as T lymphocyte proliferation, and the proliferation stimulated by LPS was defined as B lymphocyte proliferation, and the absorbance was compared to calculate the concentration of a drug that inhibits each proliferation by 50% (IC ₅₀ , μg / ml). . Test result T lymphocyte proliferation inhibition IC ₅₀ = 5.54 μg / ml B lymphocyte proliferation inhibition IC ₅₀ = 6.27 μg / ml

As described above, the inhibition of the growth of various animal cells by the treatment with Compound N-0285 was clarified, and it was confirmed that the compound of the present invention has antitumor activity, antiatherosclerotic activity and immunosuppressive activity.

4. Bronchodilator activity test of compound N-0285 300 to 400 g of Hartley male guinea pigs (Japan SLC) were killed by exsanguination and the trachea was removed. After removing fat and connective tissue, the trachea was cut into a spiral having a width of about 2 mm, and divided into three pieces containing 4 to 5 smooth muscle tissues. Specimens 37 ° C., and suspended in organ bath of 20ml containing modified-Tyrode solution aerated with a gas containing 95% O ₂ and 5% CO _2, was added a load of 1g. Muscle relaxation is an isonic transducer (Nihon Kohden, TD-112S)
Pen recorder (Yokogawa Hokushin Electric, Type 3066)
Recorded. After resting the sample for 60-90 minutes, 0.1 M of isoproterenol was added to relax. After washing, this was repeated at intervals of 30 to 40 minutes until a certain relaxation reaction was obtained. The compound of the present invention was cumulatively added to relax, and finally 1 mM aminophylline was added to obtain a maximum relaxation reaction. The intensity of the relaxation response is defined as the contraction height before drug application is 0.
And when the relaxation of aminophylline is 100%, 5
It was expressed as the concentration of the drug that relaxes by 0% (EC ₅₀ , μg / ml). Compound N-0285 was used after dissolving in DMSO and diluting with distilled water. Test result IC ₅₀ = 10 μg / ml

As described above, the relaxing action of the tracheal muscle isolated from guinea pigs by the treatment with Compound N-0285 was clarified, and it was confirmed that Compound N-0285 had bronchodilator activity.

Examples of Agrochemical Formulations Examples of agrochemical formulations containing the compound of the present invention as an active ingredient are shown below, but the invention is not limited thereto.

Wetting agent 20 parts of the compound of the present invention 76 parts of pyrophyllite 2 parts of sorbol 5039 2 parts (mixture of nonionic surfactant and anionic surfactant: trade name of Toho Chemical Industry Co., Ltd.) Carplex # 80 2 parts (Synthetic hydrated silica: trade name of Shionogi & Co., Ltd.) The above is uniformly mixed and pulverized to obtain a wettable powder.

Emulsion 5 parts of the compound of the present invention 75 parts of xylene 15 parts of N-methylpyrrolidone 15 parts of sorbol 2680 5 parts (mixture of nonionic surfactant and anionic surfactant: trade name of Toho Chemical Industry Co., Ltd.) Are uniformly mixed to form an emulsion.

[0071]

TABLE 3 Flowable agent 25 parts of the compound of the present invention 10 parts of agrizole S-710 (nonionic surfactant: trade name of Kao Corporation) Lunox 1000C 0.5 part (anionic surfactant: Toho Chemical Industry Co., Ltd.) ) Trade name) Xanthan gum 0.02 parts Water 64.48 parts After mixing above uniformly, wet pulverize to make flowable.

[0072]

[Table 4] Dry flowable agent 75 parts of the compound of the present invention 5 parts of Hytenol NE-15 (anionic surfactant: trade name of Dai-ichi Kogyo Seiyaku Co., Ltd.) 10 parts of Vanillex N (anionic surfactant: Nippon Paper Industries, Ltd.) (Trade name) Carplex # 80 10 parts (Synthetic hydrated silicic acid: Shionogi & Co., Ltd.) After uniformly mixing and pulverizing the above, adding a small amount of water, stirring, mixing and kneading, and extruding granulation. Granulate with a machine and dry to obtain a dry flowable agent.

Granules Compound of the present invention 1 part Bentonite 55 parts Talc 44 parts After uniformly mixing and pulverizing the above, a small amount of water is added, and the mixture is kneaded by stirring, kneaded, granulated by an extrusion granulator, and dried. Granules. When used, the above preparation is used as it is or diluted 1 to 10,000 times with water so that the active ingredient is contained in an amount of 0.001 to 50 kg, preferably 0.01 to 1 kg / ha.
Sprinkle to 10kg.

Pharmaceutical Preparation Examples Next, pharmaceutical preparations containing the compound of the present invention as an active ingredient will be described, but the present invention is not limited thereto.

[0075] After mixing the above components by a conventional method, a tablet containing 50 mg of the active ingredient in one tablet was prepared.

[0076] After mixing the above components in a conventional manner, the mixture was filled into gelatin capsules to give capsules containing 50 mg of the active ingredient per capsule.

[0077] After mixing the above components, a soft capsule was prepared by a conventional method.

[0078] The above components were mixed by a conventional method to obtain a 1% ointment.

Aerosol suspension (A) Compound of the present invention 0.25% Isopropyl myristate 0.10% Ethanol 26.4% (B) 60 of 1,2-dichlorotetrafluoroethane and 1-chloropentafluoroethane ~ 40% mixture. 73.25% The above composition (A) was mixed, and the resulting mixture was charged into a container equipped with a valve. The propellant (B) was added at 20 ° C to about 2.4.
An aerosol suspension was prepared by press-fitting with a bubble nozzle to a cage pressure of 6 to 2.81 mg / cm ² .

[0080]

As described above, since the compound of the present invention has herbicidal activity against various broadleaf weeds, it is expected to be used as a foliar treatment type herbicide. Further, the compound of the present invention
It has a controlling effect on various plant disease fungi and is expected to be used as a plant disease controlling agent. Further, the compound of the present invention has antitumor activity, antiatherosclerotic activity, immunosuppressive activity and bronchodilator activity, and is therefore useful as a medicine.

[Brief description of the drawings]

FIG. 1 shows an ultraviolet absorption spectrum of compound N-0285.

FIG. 2 shows an infrared absorption spectrum of compound N-0285.

FIG. 3 shows a ¹ H nuclear magnetic resonance spectrum of compound N-0285.

FIG. 4 shows a ¹³ C nuclear magnetic resonance spectrum of compound N-0285.

──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. ⁶ Identification code FI A61K 31/365 ACD A61K 31/365 ACD ADU ADU C12N 1/14 C12N 1/14 A C12P 17/08 C12P 17/08 // ( C12N 1/14 C12R 1: 645) (C12P 17/08 C12R 1: 645) (72) Inventor Kunimitsu Nakahira 1470 Shirooka, Shirooka-cho, Minamisaitama-gun, Saitama Prefecture Nissan Chemical Industry Co., Ltd. Biological Science Research Institute (72) Inventor Tatsuya Seki 722-1, Tsuboi-cho, Funabashi-shi, Chiba Pref. Nissan Chemical Industry Co., Ltd.

Claims (13)

[Claims] 1. The following formula: A compound N-0285 represented by the formula: 2. A compound N- having the following physicochemical properties:
0285: Molecular weight: 296; Composition formula: C ₁₆ H ₂₄ O ₅ ; 3. Properties and color: colorless transparent oily neutral substance; 4. UV absorption spectrum: shown in FIG. Infrared external absorption spectrum: ν _max ^KBr (cm ⁻¹ ) = 355
6, 2945, 2935, 1712, 1261, 106
8, 981; shown in FIG. Solubility in solvent: chloroform, ethyl acetate,
6. Easily soluble in methanol, poorly soluble in water; ¹ H nuclear magnetic resonance spectrum: The ¹ H nuclear magnetic resonance spectrum was measured in deuterated methanol is shown in Fig. 3, and the chemical shifts (.delta. value) below: δ (ppm): 7.39 ( 1H, dd ), 5.85 (1
H, m), 5.82 (1H, dd), 5.23 (1H,
dd), 4.80 (1H, m), 4.06 (1H,
m), 3.87 (1H, m), 3.50 (1H, d
d), 2.12 (1H, m), 2.09 to 2.01 (2
H, m), 1.89 to 1.70 (5H, m), 1.59
(1H, m), 1.24 (3H, d), 0.91 (1
H, m); ¹³ C nuclear magnetic resonance spectrum: The ¹³ C nuclear magnetic resonance spectrum was measured in deuterated chloroform are shown in Figure 4, show that the chemical shifts (.delta. value) below: δ (ppm): 21.1,28.0 , 33.1, 35.
0, 35.7, 48.9, 52.9, 73.2, 76.
6,77.1,83.7,118.2,134.1,1
35.2, 154.5, 168.3; Rf value: silica gel thin layer chromatography (Kies
el gel 60F254, Merck) Ethyl acetate: methanol (10: 1, V / V%) shows 0.35. 3. The name of the microorganism “unidentified strain No. 15
5 and a microorganism deposited under Microorganism Deposit No. 16083 (FERM P-16083). 4. A method for producing a compound according to claim 3, wherein the microorganism according to claim 3 is cultured, and the compound according to claim 1 is collected from the culture. 5. An agricultural chemical comprising the compound according to claim 1 as an active ingredient. 6. A herbicide comprising the compound according to claim 1 as an active ingredient. 7. A fungicide for agricultural and horticultural use comprising the compound according to claim 1 as an active ingredient. 8. A medicament comprising the compound according to claim 1 as an active ingredient. 9. An antitumor agent comprising the compound according to claim 1 as an active ingredient. 10. An anti-atherosclerotic agent comprising the compound according to claim 1 as an active ingredient. An anti-asthmatic agent comprising the compound according to claim 1 as an active ingredient. 12. An antiallergic agent comprising the compound according to claim 1 as an active ingredient. 13. An antirheumatic agent containing the compound according to claim 1 as an active ingredient.