DE2329486A1 - NEW ANTIBIOTICS AND METHODS FOR THEIR PRODUCTION - Google Patents

Description

New Antibiotics and Procedures to Her;

The invention relates to new antibiotic substances B-41 and a method for producing this antibiotic substance, in which one in a medium to the Species Streptomyces that produces B-41 strains and then the obtained B-41 from the culture substance isolated. The invention also relates to an insecticidal and acaricidal agent, which as an active ingredient at least contains one of the new antibiotic substances B-41.

Many organic compounds have already been used as insecticidal and acaricidal agents. Among the antibiotic However, substances show only a few identical

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early insecticidal and acaricidal effects. In addition, these substances have not yet been put to practical use found.

A new antibiotic substance B ~ 41 has now been found, which is produced by the strain B-41-146. This new strain belongs to the species Streptomyces. The .inventive Antibiotic not only has a much higher acaricidal activity than the previously known compounds with acaricidal activity, but is also capable of causing insects harmful to agriculture and horticulture to control how aphids, larvae of insects of the species Lepdoptera etc.

The antibiotic substance B-41, which is the active ingredient of Insecticidal and acaricidal agents of the present invention could be divided into nine kinds of substances namely A ^, Ap, A ,, A., IL, Bp, B ,, C. and Cp will.

The physicochemical properties of these substances are summarized in the following table:

3 0 a 8 5 1/1 2 1 fi

Molecular formula A. A ₉ A, A /, B ₁ B ₀ B, C. C ₉ ^C 32 ^H 48 ° 7 ^C 5B ^K 56 ° O ^C 31 ^K 44 ° 7 ^C 32 ^H 46 ° 7 ^C 39 ^H 58 ° 10 ^C 32 ^H 46 ° 7 ^C 33 ^H 43 ° 7 ^C 36 ^H 4 ₇ ° 9 ^NC 37 ^H 49 ° 9 ^N

Elemental calculated C 70.56 67.83 70.43 70.82 68.19 70.82 71.19 C: 67.80 C 68.18

tarana- H: 7.43 H 7.58

lysis (%). H 8.88 8.39 8.39 8.54 8.51 8.54 8.69 N: 2.20 N: 2.15

found C 70.74 71.73 65.73 69.85 68.00 69, S6 70.72 C: 65.93 C: 68.09

H: 7 53 Hi 7 54

H 9.18 8.26 7.89 8.37 8.32 8.32 8 * 59 N: 2 ^ 14 N: 2 * 13

Molecular Osmometry

large- see me-
«·» ■ · weight method (in

"Acetone) 513.9 672.1 517.0-629.5 524.3

■ "·. Mass

- ~ spectrum

- * (M +) 544 672 528. 542 686 542 556 679 693

^ (Note 1) (Note Λ)

Melting amorphous amorphous 212 193 176 139 amorphous amorphous amorphous

kt ( ⁰ C) powder powder -215 -195 -178 -142 powder powder powder

Specific rotation ■

£ ° + 160 ° + 54 ° + 106 ° + 103 ° + 75 ° + 131 ° + 126 ° + 57 ° +54

(Concentration of the co

Sample 5 mg / 2 ml, jr-

Length of the layer oo

in acetone 10 cm) -h- CD

UV absorption spectrum (\ -max, mu) 240.5

' Fig. 1

IR absorption spectrum (Nujol method)

NMR spectrum (in (CD,) ₂ · CO in the case of FIGS. 19 to 25 and in CDCl in the case of FIGS. 26 and 27

245
Fig. 2 Fig. 3

245 245 245 245 240 240 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9

Fig. 10 Fig. 11 Fig. 12 Fig. 13 Fig. 14 Fig. 15 Fig. 16 Fig. 17 Fig. 18

60 MHz Fig. 19 Fig. 20 Fig. 21 Fig .22 leaf 23 Fig. 2 4 Fig. 25 Fig. 26 Fig. 2 Mass spectrum (main cn peaks in the Bedin 544 672 528 542 686 542 556 679 692 OD gungen 75 eV, tempera 402 181 400 414 414 400 414 568 582 CTt ture of ionization 181 153 181 195 195 181 195 400 414 - ^k room 200 C and rehearsal 153 151 • 153 167 167 153 167 181 195 temperature 120 to 151 151 151 153 167 ro 190 ⁰ C) 125 151 151 111 111 (Note 1) (Grade he·. Solubility in lo bad solvents soluble in what ser, easy dto dto dto dto dto dto dto löslüi in n-hexane, B _e nzol, Acetone, Ethanol -5 and Chlo roform

TV) CD

Color iodine / chlorine reaction roform

yellow yellow yellow yellow yellow yellow yellow yellow yellow

(in the Ninhydrin nega- nega pH who 12th nega nega nega nega nega nega nega pH who 12th to Slim tiv tiv ten of tiv tiv tiv tiv tiv tiv tiv ten of GO schi cht-
chroma-
togra-
phy) spray
with sweat
fic acid
under him Brown red 2 to Brown Brown red brown brown Brown Brown 2 to OK heat lich lich ^ p purple purple OO C. Colours Colours CD cc
OD Potassium perm cn manganate solution yellow yellow yellow yellow yellow yellow yellow yellow yellow Neutraiization Ti no no no tration: pKa- dto dto dto ' dto pK dto pKa- dto Ln alcohol value value value tr » holic solution at at at colorless pH who color ten of Come on colorless 2 to 9 color (at Come on a pH value of 8.7 he followed a Kri stallisa tion where through the Measurement not possible became) color color color color-color Come on Come on Come on go Go Color of the substance

Footnote 1: The mass spectra of C. and Cp were determined for the acetylated derivative which had been obtained by substituting the hydrogen atom of the substituent FL in the given structural formula by a CH 1 CO group.

The accompanying drawings show the UV absorption spectra, the IR absorption spectra and the NMR spectra of the antibiotic substance B-41. Figures 1 through 9 show the UV spectra of A ₁ , Ap, A ^, A ,, B., Bp, B ,, C. and Cp of the antibiotic substance B-41. Figures 10 to 10 show the IR spectra of A., Ap, A ,, A., B ₁ , Bp, B ,, C ₁ and Cp of the antibiotic substance B-41 and Figures 19 to 27 show the NMR Spectra of A ₁ , Ap, A ^, A », B ₁ , Bp, B ,, C ^ and Cp of the antibiotic substance B-41.

In the table below, the Rf values of the above substances, measured by thin layer chromatography, are given. For this, a thin layer chromatography film containing a fluorescent reagent (made by Tokyo Kasei Kogyo Co. Ltd., trademark "Spotfilm fluorescent") was used. The detection was carried out according to the intensity of the emitted fluorescence when the individual substances were irradiated with UV rays with 2536 Å.

3 (J 9851/1216

Silica gel F

Solution A. A ₀ A, A, B. B ₀ B, C. C ₀ medium- ^{Ί ά} * * ^{Ί ά} * ^{Ί ά} system

Accton / n-He-

xan (30/70) 0.47 0.32 0.42 0.44 0.47 0.61 0.63 0.22 0.24

Ethyl acetate /

Benzene (50 /

50) 0.61 0.62 0.63 0.65 0.79 0.80 0.82 0.45 0.47

Ethyl acetate / chloroform

(25:75) 0.27 0.35 0.39 0.41 0.75 0.79 0.81 0.12 0.13

Acetone / ben-

zol (40:60) 0.85 0.75 0.77 0.79 0.92 0.92 0.92 0.6n 0.63

Acetone / ben-

zol (15:75) 0.39 0.32 0.39 0.39 0.57 0.51 0.53 0.11 0.13

Ethano1 / η-He-

xan (2:98) 0 0 0 00 00 00

Alumina F

Solution Α _Λ A ₀ A-. AB B ₀ B, C ₁ C ₀ medium- ' ^d * * ^{Ί d} * ^{j ά} system

Acetone / n-He-

xan (3C-.70) 0.55 0.16 0.32 0.34 0.67 0.92 0.92 0 0

Ethyl acetate /

Benzene (50:

50) 0.56 0.10 0.21 0.23 0.73 0.81 0.83 0 0

Ethyl acetate / chloroform

(25:75) 0.40 0.10 0.15 0.17 0.63 0.65 0.67 0 0

Acetone / ben-

zol (15:75) 0.27 0.05 0.09 0.11 0.35 0.45 0.47 0 0

Ethano1 / η-He-

xan (2:98) 0.17 0.03 0.07 0.09 0.20 0.42 0.44 0 0

3U y 8 S 1/1 2 1 6

From the physicochemical properties given above, in particular the mass spectra with high resolution, as well as from the results of the X-ray analysis, the following structural formula was obtained for the substances A ₁ , A 1, B, B 3, C 1 and C ₂ of B-41 :

R, -

^Λ 3 H CHj CKj C ₂ H ^Α 4 H CH ₂ C ₂ Ii ₅ ^Β 2 CHj CHj CHj ^Β 3 CHj CHj C ₂ H ₅ ^C l H -ΠΗ ^ .000 CHv j C ₂ H

Similarly, the following structural formula resulted for A ₁ of B-41:

CH.

⁰ Vl <> .A

OCH ₂ 3 U 9 H 5 i / 1 2 1 G

The structures of Ap and IL of B-41 have not yet been elucidated. From the splitting of the mass spectra, however, it can be concluded that these compounds have a similar chemical structure to the substances A ^, A ^, A ,, B ^, B ^, Ο ,, and Cp.

Since there are no known antibiotic substances, which have the specified chemical structures and the specified physico-chemical properties must the substance B-41 according to the invention is a new antibiotic.

Streptomyces strain B-41-146 which contains the antibiotic B-41 has the mycological properties given below.

1. Morphological properties:

On most laboratory media, a finely branched substrate mycelium developed into a long, airy mycelium that formed turns with spirals or loops.

No splitting of the mycelium was observed in the initial stage.

More or less short vicious spores with dimensions of 0.6-0.9 x 1.1-1.5 η formed in chains with 10 to 50 conidia.

There were relatively short warty protrusions on the surfaces of the spores.

Spore carriers and resinification were not observed.

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- 1ΰ

2. Growth characteristics on different media:

a) sucrose nitrate agar:

Good growth; Substrate iriycelium colorless; wrong Mycelium light brown; Aerial mycelium scanty, semi-transparent, leathery '; soluble Pigment light brown.

b) Glucose asparagine agar:

Abundant growth, substrate mycelium colorless; reverse Substrate light brown; Aerial mycelium profuse, gray in color; soluble pigment light brown.

c) Glycerine asparagine agar:

Lush growth; Substrate nycelium colorless; reverse Substrate light brown; Aerial mycelium white, being on a slant on a white The background formed many shiny gray-brown spots; soluble pigment yellow-gray.

d) Inorganic Salts Starch Agar:

Lush growth; Substrate mycelium colorless; inverted mycelium yellowish-gray; Aerial mycelium gray, with many pale yellow spots on a slant crop on a gray background formed; soluble pigment glossy olive green.

e) tyrosine agar:

lush growth; Substrate mycelium gray-yellow-brown; inverted mycelium brown; Aerial mycelium gray, with sometimes yellowish gray spots appearing on a slant on a gray background formed; soluble pigment glossy brown.

f) nutrient agar:

Poor growth; Substrate mycelium colorless; inverted mycelium light brown; Aerial mycelium sparse

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borrowed, white; no soluble pigment formed.

g) yeast extract-malt extract agar:

Lush growth; Substrate mycelium gray-yellow-brown; inverted mycelium yellowish brown; LuiCtmycelium lush, gray, being on one Slant culture on a gray background formed a lot of light yellow spots; soluble pigment. yellow.

h) oatmeal agar:

Lush growth; Substrate mycelium colorless; inverted mycelium olive green; Aerial mycelium gray, whereby light yellow spots formed on a slant culture; soluble pigment pale olive.

Physiological properties:

a) Growth temperature range: 18 to 37 ⁰ C; Optimal growth temperature: 25 to 30 ⁰ C.

b) Liquefaction of gelatin: slow but strong positive.

c) Hydrolysis of starch: Strongly positive.

d) Coagulation of skimmed milk: positive (28 ⁰ C).

e) Peptonization of skimmed milk: Positive (28 C).

f) Melanin formation: negative.

g) Reduction of nitrate: positive.

h) Utilization of various carbon sources (Pridham and Gottlieb Agar)
Recycling rate:
+++ raffinose
++ D-glucose, D-fructose, sucrose, L-rhamnose,

I-inositol, D-mannitol + L-Ar & binose, D-xylose.

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From the above characteristics, it can be seen that this strain is Streptomyces chattanoogensis (International Journal of Systematic Bacteriology, Volume 18, No. 2, page 97 (1968)). However, there are the following Differences between the two tribes:

1; The trunk B-41-146 forms lush vertebrae while S. chattanoogensis branches out monopodially.

2. The spore surface of the strain B-41-146 is warty, while that of S. chattanoogensis is thorny.

3. On yeast malt extract agar and an inorganic SaI-ze starch agar the strain B-41-146 formed pale or light yellow spots on a gray background, what was not the case with S. chattanoogensis.

4. The strain B-41-146 assimilates L-arabinose, D-xylose and I-inositol, while S. chattanoogensis these carbon swell not assimilated.

Because of the above four differences in mycological 'properties it was concluded that the strain B-41-146 is a new species of Streptomyces. The strain B-41-146 was at the Research Institute of Industrial Technology of Microorganisms, Agency of Industrial Science and Technology in Japan with the depository number of Bikokenkinki No. 1438 deposited.

As is well known, actinomycetes tend in nature and through the use of artificial measures, such as UV radiation, Radiation irritation, chemical treatment, to a mutation. This applies to that used according to the invention

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B-41-146 too. The term "strain B-41-146" as used herein is intended to include all such mutants. Thus, all strains that contain the antibiotic substance fall Generate B-41, and which are not clearly distinguishable from the strain B-41-146 and its mutants, under the term "Strain B-41-146".

In the method of the invention, the antibiotic Substance B-41 obtained by culturing the B-41-146 strain in a suitable medium, and then the obtained Substance isolated from the medium. The strain can be grown in stationary cultures. When the antibiotic is in If large quantities are to be produced, it is preferable to culture the strain in a liquid with aeration and to breed agitation.

As the culture media, all those normally used for cultivating strains of the species Streptomyces can be used. Examples of suitable carbon sources are starch, dextrin, glucose, maltose, corn steep liquor and molasses. Examples of suitable nitrogen sources are meat extract, peptones, yeast extract, Soybean meal, casein, ammonium sulfate and ammonium nitrate. If necessary, potassium, calcium, magnesium, iron, copper, zinc, manganese, cobalt and similar inorganic Salts and trace elements are added.

Conventional procedures can be used to recover antibiotic B-41 from the broth, for example an extraction with an organic solvent in the presence or absence of an adsorbent or of an aid. In this case, the cells can be separated from the broth by filtration and then

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see. Solvents such as methanol or acetone extracted will. But you can also extract the broth directly with organic solvents such as chloroform, ethyl acetate, Subject to benzene, η-hexane or cyclohexane.

For cleaning the oily crude B ~ 41, which by removal of the solvent is obtained from the extract, known purification measures such as column chromatography can be used or extraction with solvents can be used.

The invention is illustrated in the examples. In these examples the activity of the broth was determined by immersion determined by two-spotted spider mites. This was done like this proceed that leaves of white beans with the zv / egg-spotted Spider mites infest v / aren, 1 minute in a 70% acetone extract of the broth or in its aqueous Thinner and then allowed to air dry. After 24 hours, the acaricidal activity was measured, whereby the activity was determined.

example 1

30 liters of a liquid culture medium (pH about 7.2) containing 2.06 glucose, 1.0.0 soybean meal and 0.2% sodium chloride was placed in a 50-1 fermentation vessel, followed by heating sterilized. The strain B-41-146 was inoculated into the medium and subjected to aerobic stirring culture at a temperature of 28 ° C., aeration of 8 l / min and stirring of 250 rpm. After the cultivation had been carried out for 120 hours, the broth showed a brilliant yellow color. At this point, the cultivation was discontinued and the activities

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did was determined. It was found that a 300-fold dilution of the broth showed an acaricidal activity of 100%. The cells were then separated from the broth by filtration and extracted with acetone. Acetone was then distilled off, whereby A4 g of a brown substance was obtained. This substance was extracted with hot hexane, whereupon the hexane was distilled off. The residue was dissolved in a small amount of methanol. The resulting solution vrurde allowed to stand overnight at -20 ⁰ C, whereby a precipitate to precipitate came. This has been removed. Then, the methanol was distilled off, whereby 35 g of a brown oily substance was obtained. The oily substance thus obtained was subjected to chromatography on an alumina column. It was eluted with chloroform. Effective fractions were collected according to the acaricidal activity. The chloroform solution was then concentrated. This procedure was repeated several times, whereby 8.2 g of a crude substance were obtained. The acaricidal activity of the crude substance when used at a concentration of 2 µg / ml was 100%.

The crude substance was passed through a column filled with "Sephadex LH-20" (trademark for a product of Pharmacia Co.) and eluted with methanol. As a result, the substances Ap, B., Bp, A-, and A ₁ , which are the main components of B-41, were eluted in this order. However, these substances overlapped one another and were therefore obtained separately in the form of two groups of B-i + Ap and Bp + A. + A. The B. + Ap group was chromatographed on a silica gel column and then eluted with chloroform ethyl acetate. In this way, 210 mg of B ^ and 115 mg of Ap were obtained. The B ₀ +

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A ₁ , + A ^ group was subjected to silica gel column chromatography, whereby 200 mg of Bp and 30 mg of B ^ which was _{similar to B 2} were obtained. The remaining A ₁ + A-2 group was then subjected to alumina column chromatography to give 372 mg of A ₁ , 42 mg of A-. and 15 mg of A. obtained A- similar to v / ar. The alumina column through which the above-described brown oily substance had passed was further eluted several times with methanol. It was subjected to silica gel column chromatography, whereby 78 mg of C ₁ from B-41 and 52 mg of C ₂ from B-41 were obtained.

To prepare the insecticidal and acaricidal compositions of the present invention, one or more of the substances A ₁ , A ₂ , A ,, A., B ₁ , B ₂ , B ,, C ₁ and Cp obtained in this way are used from B- 41 diluted with a carrier and, if necessary, mixed with other auxiliaries. In this way, the substances can be made into dusts, granules, fine granules, wettable powders, emulsifiable concentrates, oil sprays and the like. The cleaning can of course be interrupted at a desired stage of the cleaning and the resulting, not yet completely cleaned substance can also be used as an active ingredient. In the event that such a raw substance is to be used as an active ingredient as it is, it is sufficient that the raw substance is purified so that it has an acaricidal activity of 100% at a concentration of 5 ppm. In this case, the content of B-41 in the raw substance is about 50? O. The rest are impurities from the broth.

The term "carrier" is used herein to mean a synthetic or natural, inorganic or organic substance

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which is added to an insecticide so that the active ingredient easily achieves its intended goals, for example Plants, mites, harmful insects, etc., or with it storage, transport or handling of the active ingredient is facilitated.

Examples of suitable solid carriers are inorganic substances such as clay, talc, diatomaceous earth, kaolin, bentonite, Calcium carbonate and synthetic calcium silicate, natural and synthetic resins such as coumarone resins, alkyl resins and polyvinyl chloride, waxes such as carnauba wax and paraffin wax, shells of nuts, v / ie of walnuts and Coconuts, and soybean meal.

Examples of suitable liquid carriers are water, alcohols, such as ethanol, isopropanol and ethylene glycol, glycol ethers, such as ethylene glycol monophenyl ether, and diethylene glycol monoethyl ether, Ketones, such as acetone, methyl isobutyl ketone, cyclohexanone, acetophenone and isophorone, ether, such as tetrahydrofuran and dioxane, aromatic hydrocarbons such as benzene, toluene, xylene and methylnaphthalene, chlorinated hydrocarbons, v / ie trichlorethylene and carbon tetrachloride, and low, medium and high boiling points Petroleum fractions such as kerosene, light oils or aromatic hydrocarbons.

Examples of suitable propellants are furon gases, liquefied natural gases, methyl ethers and vinyl chloride monomers.

For emulsification, dispersion, wetting or spreading, surface-active agents which are ionic or non-ionic are grounded can be used. Examples of suitable anionic surfactants are sodium and calcium salts of lignosulfonic acid, sodium and potassium salts

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of oleic acid, the sodium salt of lauryl sulfonic acid, and the sodium and calcium salts of dodecylbenzenesulfonic acid. Examples of suitable cationic surfactants are higher aliphatic amines and ethylene oxide condensates of higher aliphatic amines. Examples of suitable non-ionic surfactants are the glycerides of fatty acids, sucrose esters of fatty acids, ethylene oxide condensates of higher aliphatic Alcohols, ethylene oxide condensates of higher fatty acids, ethylene oxide condensates of alkylphenols and of Alkyl naphthols and copolymers of ethylene oxide with propylene oxide.

The insecticidal and acaricidal agents of the present Invention can use a protective colloid such as gelatin, gum arabic, casein, polyvinyl alcohol or carboxymethyl cellulose, or a thixotropic agent such as sodium polyphosphate or bentonite. The according to the invention Agents can also contain other compounds with insecticidal and acaricidal activities, such as 2- (i-methylpropyl) -4,6-dinitrophenyl-ß, ß-dimethylacrylate, Di- (p-chlorophenyl) -cyclopropylcarbinol, N'- (2-methyl ~ 4-chlorophenyl) - N, N-dimethyiformaraidin, 2,4,4 ', 5-tetrachlorodiphenyl sulfone, 1,1-bis (p-chlorophenyl) -2,2,2-trichloroethanol, O, O-diethyl-S- (2-ethylthio) ethyl-phosphorodithioate, O, O-dimethyl-S- (N-methyl-N-formylcarbamoylmethyl) phosphorodithioate, 2-sec-butylphenyl-N-methylcarbamate or m-tolyl ~ N-methylcarbamate, or contain a mineral oil, increasing the effects of the agent can be increased and in some cases synergistic effects can be expected. Naturally the agents according to the invention can be mixed with fungicides, herbicides, plant growth regulators, attractants, Fertilizers and the like can be used.

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The effects of the insecticidal and acaricidal agents of the present invention will be demonstrated by the following experimental examples explained.

Experimental example

Emulsifiable concentrates which each contained 20 μj of the substances A., Ap, A, A ^, B ^ ₁ bp, B ₇ , C ₁ -and C ₀ of B-41, which were isolated according to the procedure of Example 1 were diluted to a given concentration. White bean leaves infested with two-spotted spider mites were immersed in the resulting solution for one minute and then air-dried. After 24 hours, the acaricidal activity (50) was calculated. The results obtained are summarized in the following table:

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Concentration (ppm)

fraction

20th

10

2.5

0.63

0.31

0.16

0.08

0.04

³ 1 ³ 2 ^B 3 ^C 1 ^C 2

Comparative substance *

100 100 100 19th , 9 .) 100 100 100 67 , 6 0 100 100 100 100 100 100 100 100 100 100 98.1 81.5 69.2 20th , 0 0 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100

100

56.4

100 100 93, 2 100 ) 15.2 232! 100 100 100 100 21.3 UJ
-Γ
ΟΟ 100 80.5 16, 6th 0 100 100 74, 1 18.3 0 100 100 100 100 51.8 100 100 100 100. 97.3 1 (trademark "Kelthane";

Experimental example 2

An emulsifiable concentrate that contains 20% of the raw substance B-41 obtained by subjecting the brown oily substance of Example 1 to alumina column chromatography twice was subjected to was diluted to a given concentration. The received Solution was sprayed onto apple leaves that were infested with about 100 European red mites. 5 days later the number of mites still alive was determined. The results obtained are summarized in the following table:

200 ppm 100 ppm 50 ppm

Raw B-41 5/108 14/121 50/115 Comparison substance (Kelthane) 0/97 8/103 56/128 Not treated 88/102

(The counter indicates the number of mites that were present at the time of counting lived. The denominator indicates the number of mites before spraying).

Experimental example 3

Emulsifiable concentrates containing 20% of a raw B-41 (das obtained by purifying the brown oily substance prepared in Example 1 by silica gel column chromatography had been) and a raw A. + Ap + A, mixture (that by column chromatographic purification of the crude B-41 using a mixed solvent of 50:50 ethyl acetate and benzene was obtained) were diluted to a given concentration. The solutions were sprayed on orange leaves, which are filled with citrus-red

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mites were infested. After 24 hours the acaricidal Activity (%) determined. The results obtained are summarized in the following table:

20 ppm 10 ppm 7 ppm 3.3 ppra

Raw Β-41 100 # 85.5? £

Raw A. + A ₉ + A, -

Mixture '^ ^ 100 100

Comparative substance (Kelthane) 90.9 30.8

Experimental example 4

Emulsifiable concentrates, the 20? O of a crude Λ. + B. mixture B-41 (obtained by purifying the brown oily substance prepared in Example 1 three times by silica gel chromatography with 70:30 η-hexane: acetone) and a crude A ₂ + A ^ + Bp mixture of b-41 (obtained in the same manner as the former mixture) were diluted to a given concentration. The solution was sprayed onto Chinese cabbage infested with green peach aphids. After 24 hours, the mortality (%) of the aphids was determined. The results obtained are summarized in the following table:

250 ppm 25 ppm 2.5 ppm

Raw A ₁ + B ₁ -
mixture 100% 92 , 3% 64, 2% Raw A ^ + A ^ + B ,, -
Mixture ^ ^{D c} 100 89 ,8th 52, 8th Comparative substance ** 100 73 ,1 33, 1

** 0, O-dimethyl-0- (2,2-dichlorovinyl) phosphate.

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2329A86

Experimental example 5

Eggs of the rice stem borer of the first generation were inoculated on rice plants (variety "Kinmaze"). The plant

zen were planted in 200 cm pots. The eggs were allowed to hatch to allow the larvae to infect the strains. Then, a wettable powder containing 40% of the crude B-41 was obtained by purifying the brown oily substance of Example 1 twice by an alumina column chromatography, diluted to a given concentration and then in an amount of 100 cnr ⁵ per pot sprayed the plants. 5 days later, the stems or stems were split open to determine the number of living and dead larvae. From this, the mortality of the larvae (%) was calculated. The results obtained are summarized in the following table:

100 ppm 50 ppm

Raw B-41 100% 79.4%

Comparative substance *** 82.1 25.5

*** 0,0-diethyl-0- (2-isopropyl-4-methyl-6-pyrimidinyl) pho sphorthio at.

From the experimental examples 1 to 5 it can be seen that the antibiotic substance B-41 is excellent in insecticidal and acaricidal activity and that it is particularly exerts a stronger influence on mites than conventional chemicals.

Below are working methods for making the insecticides and acaricidal agents of the present invention are given as production examples. All parts are in it based on weight.

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2329A86

Production example 1

10 parts of a crude B-41 obtained by silica gel column chromatography twice the brown oily substance obtained in Example 1, were homogeneous with 5 parts of silica, 50 parts of talc and 35 parts of clay mixed. The obtained mixture was crushed three times by means of an impact pulverizer pulverized and homogenized to a dust.

Production example 2

40 parts of the same B-41 as in Preparation Example 1 were homogeneous with 20 parts of silica, 5 parts of sodium dodecylbenzenesulfonate, 2 parts of polyvinyl alcohol and 33 parts of clay mixed. The resulting mixture was used three times pulverized using an impact pulverizer, and then homogenized into a wettable powder.

Production example 3

20 parts of the same crude B-41 as in Preparation Example 1 were mixed homogeneously with 7 parts of polyoxyethylene nonylphenyl ether, 3 parts of calcium dodecylbenzenesulfonate and 70 parts of xylene. The resulting mixture was filtered, whereby an emulsifiable concentrate was obtained .

Production example 4

10 parts of the same crude B-41 as used in production example 1 were dissolved in 10 parts of xylene. The resulting solution was mixed with 80 parts of a machine oil and then filtered to obtain an oil spray .

-25-309851 / 12 16

Claims (14)

Claims and FU an M <=> - where FL denotes a hydrogen atom or a methyl group, R ₉ denotes a methyl group or r a pyrroyloxymethyl group of the formula -CH ₂ OOC [I. means ethyl group or ethyl group, with the proviso that when FL is a hydrogen atom, R _{2 is} a methyl group or a pyrroyloxymethyl group and R is a methyl group or ethyl group, and when R ^ is a methyl group, R _{2 is} a methyl group and R ^ is a Methyl group or ethyl group. 2. A compound according to claim 1, characterized geke η η that R ,. is a water atom, R _{2 is} a methyl group and R- is a methyl group. 3. A compound according to claim 1, characterized in that R ^ is a hydrogen atom, R _{2 is} a methyl group and R is an ethyl group. -26- 30y8b1 / 12 16 4. A compound according to claim 1, characterized in that R _{1 is} a methyl group, Rp is a methyl group and R is a methyl group .sind. 5. A compound according to claim 1, characterized in that g e k e η η indicates that R ^ is a methyl group, R "a methyl group and R ^ are an ethyl group. 6. A compound according to claim 1, characterized in that R ^ is a hydrogen atom, R _{p is} a pyrroyloxymethyl group and R _{7 is} a Kethyl group. 7. A compound according to claim 1, characterized in that R ^ is a hydrogen atom, R _? a pyrroyloxymethyl group and R- ^ are an ethyl group. 8. Compound of the following formula CH 9. Antibiotic substance B-41 -A ₂ with activity against harmful insects, characterized in that it is a colorless amorphous powder that is sparingly soluble in water and easily soluble in η-hexane, benzene, acetone, ethanol and chloroform is, a molecular formula of ^ 58 ^Η 55 ^ - | 0 ' ^a composition of 71.73% carbon and 8.26? c hydrogen and the following prop- -27- 3 Ü 9 8 S i / 1 2 1 ο shafts: molecular weights of 672.1, obtained by the osmometric method in acetone, and of 672 by the mass spectroscopic method, a specific rotation of (<yO) ß = + 54 °, obtained at a concentration of the sample of 5 mg / 2 ml and the length of the layer of 10 cm in acetone, no pKa value at a pH value in the range from 2 to 12, an absorption maximum in the ultraviolet part of the spectrum at 245 mu, characteristic absorption bands of the infrared spectrum according to FIG. 11, a nuclear magnetic resonance spectrum in (CD.,) "· CO as shown in FIG 20, the main peak of the mass spectrum at 672, 181, 153 and 151, obtained under conditions of 75 eV at a Ionisationsraumtemperatur of 200 ⁰ C and a sample temperature of 120 to 190 ° C, and the following color reactions in thin-layer chromatography: yellow with iodine / chloroform, negative with ninhydrin, reddish purple with sulfuric acid and yellow with potassium permanganate. 10. Antibiotic Subtitan :; B-41-B. with an activity against harmful insects, characterized in that it is a colorless substance, which is sparingly soluble in water and easily soluble in η-hexane, benzene, acetone, ethanol and chloroform, a Suinmen formula of C ^ gH ^ gO ^ Q, a composition of 68.00% Carbon and 8.32% hydrogen and the following properties has: a melting point of 176 to 178 ° C, molecular weights of 629.5, obtained according to the osmometric Method in acetone, and from 686 according to the mass spectroscopy method, a specific rotation of (oC)]} = +75, obtained at one concentration of the sample of 5 iag / 2 ml and a length of the layer of 10 cm in acetone, no pKa value at a pH value in the range of 2 to 12, an absorption maximum in the ultraviolet part of the spectrum at 245 mn, characteristic absorption -28- 309851/1216 bands of the infrared spectrum according to FIG. 14, a magnetic nuclear resonance spectrum in (CXOp'CO according to FIG. 23, main peaks of the mass spectrum at 686, 414, 195, 167, 151 and 125, obtained under the conditions of 75 eV at an ionization space temperature of 200 ⁰ C and a sample temperature of 120 to 190 ⁰ C, and the following color reactions in thin layer chromatography: yellow with iodine / chloroform, negative with ninhydrin, reddish purple color ben with sulfuric acid and yellow with potassium permanganate. 11. Process for the production of the antibiotic substances B-AI-A ₁ , B-Vl-A ₂ , B-41-A B-41-A ^, 3-41-B ₁ , B-41-B ₂ , 3- 41-C ₁ and B-41-C ₂ , characterized in that a B-41-producing strain of the Streptomyces species is bred and that the antibiotic substances are obtained either individually or in the form of mixtures from the fermentation broth. 12. The method according to claim 11, characterized
used. that the strain B-41-146 13. · Acaricidal mixture for agriculture, characterized in that it is used as an active ingredient at least one of the antibiotic substances of compounds of the formula -29- 309851/1216 where R ₁ denotes a hydrogen atom or a methyl group, R ₂ denotes a methyl group or a pyrroyl group oxymethyl group of the formula - stands and ■ tr R ^ means a methyl group or ethyl group, with the proviso that when R ^ is a hydrogen atom, Rp is a methyl group or a pyrroyloxy methyl group and R- is a methyl group or ethyl group, and when R ^ is a methyl group, R _{2 is} a Methyl group and R is a methyl group or ethyl group, b) the compound of formula a substance that is a colorless amorphous powder that is sparingly soluble in water and easily soluble in η-hexane, benzene, acetone, ethanol and chloroform, a molecular formula of ^ -50 ^ 56 ^ 10 » ^a composition of 71, 73% carbon and 8.26% hydrogen has the following properties: molecular weights of 672.1, obtained by the osmometric method in acetone, and of 672 by the mass spectrometric method, a specific rotation of (06) jj β +54 at a concentration of the sample of 5 mg / 2 ml and the length of the layer of 10 cm in acetone, no pKa value at a pH value -30- 309851/1216 in the range from 2 to 12, an absorption maximum in the ultraviolet part of the spectrum at 245 mn , characteristic absorption bands of the infrared spectrum according to Figure 11, a nuclear magnetic resonance spectrum in (CD,) - 'CO according to Figure 20, main peaks of the mass spectrum at 672, 181, 153 and 151, obtained under the conditions of 75 eV at an ionization room temperature of 200 C and a trial temperature of 120 to 190 C, and the following color reactions in thin layer chromatography: yellow for iodine / chloroform, negative for ninhydrin, reddish purple for sulfuric acid and yellow for Potassium permanganate, and / or d) a substance that is a colorless substance that is sparingly soluble in water and easily soluble in η-hexane, benzene, acetone, ethanol and chloroform, a molecular formula of C ^ JHLgOx a composition of 68.00% carbon and 8, 32> o hydrogen and has the following properties: a melting point of 176 to 178 ° C, molecular weights of 629.5 obtained by the osmometric method in acetone, and of 686 by the mass spectroscopic method, a specific rotation of (P ) ß + 75 °, obtained with a concentration of the sample of 5 mg / 2 ml and a length of the layer of 10 a in acetone, no pKa value at a pH value in the range from 2 to 12, an absorption maximum in the ultraviolet part of the spectrum at 245 rough, characteristic absorption bands of the infrared spectrum according to FIG. 14, a nuclear magnetic resonance spectrum in (CD,) ₂ 'CO according to FIG. 23, main peaks of the mass spectrum at 686, 414, 195, 167, 151 and 125, -31- 3 Ü d H 5 1 / j 2 1 G keep in the conditions of 75 eV at a Ionisationsraumtemperatur of 200 C and a sample temperature of 120 to 190 ° C _f and the following color reactions in thin layer chromatography: yellow with iodine / chloroform, negative in ninhydrin, reddish purple in sulfuric acid, and yellow at Kaliumperraanganat together with an agriculturally acceptable carrier. 14. Insecticidal product for agriculture, thereby characterized in that it is as an active ingredient at least one of the antibiotic substances a) of compounds of the formula where R ^ is a hydrogen atom or a methyl group, R _{2 is} a methyl group or a pyrroyloxymethyl group of the formula -CHLOOcJl ■ _N J] and R, denotes a methyl group or ethyl group, with the proviso that when R «. is a hydrogen atom, R _{2 is} a methyl group or a Pyrroyloxymethylgrup- -32- 309851/1216 pe and R, a methyl group or ethyl group, and, if R «. is a methyl group, R2 is a methyl group and R, is a methyl group or ethyl group, b) the compound of formula CH. c) a substance that is a colorless amorphous powder that is sparingly soluble in water and easily soluble in η-hexane, benzene, acetone, ethanol and chloroform, a molecular formula of C-rgHt-gO ^ Q, a composition of 71, 73% carbon and 8.26% hydrogen and has the following properties: Molecular ,. weights of 672.1, obtained by the osmometric method in acetone, and of 672 by the mass spectroscopic method, a specific rotation of (oC) p = + 54 °, obtained at a sample concentration of 5 mg / 2 ml and length the layer of. 10 cm in acetone, no pKa value at a pH value in the range from 2 to 12, an absorption maximum in the ultraviolet part of the spectrum at 245 m 11, characteristic absorption bands of the infrared spectrum according to Figure 11, a nuclear magnetic resonance spectrum in (CIU) ₂ * CO according to Figure 20, main -33- 309851/1216 peaks of the mass spectrum at 672, 181, 153 and 151, obtained under conditions of 75 eV at a Ionisationsraumtemperatur of 200 C and a sample temperature tuijvoii 120 to 190 ⁰ C, and Hen following color reactions in thin layer chromatography: yellow with iodine / chloroform, negative with ninhydrin, reddish purple with sulfuric acid and yellow with potassium permanganate, and / or d) a substance that is a colorless substance that is sparingly soluble in water and easily soluble in η-hexane, benzene, acetone, ethanol and chloroform, a sum formula of C ^ qH ^ qO ^^, a composition of 68.00 % carbon and 8.32% hydrogen and the following properties to V / eisti a melting point 176-178 ⁰ C, molecular weights of 629.5, obtained according to the method ocrnometrisehen in acetone, and 686 after the massenspektroskopisehen method, a specific rotation of + 75 °, obtained with a concentration of the sample of 5 mg / 2 ml and a length of the layer of 10 cm in acetone, no pKa value at a pH value in the range from 2 to 12, an absorption maximum in the ultraviolet part of the spectrum at 245 mu, characteristic absorption bands of the infrared spectrum according to FIG. 14, a nuclear magnetic resonance spectrum in (CD,) ₂ 'C0 according to FIG. 23, main peaks of the mass spectrum at 686, 414, 195, 167, 151 and 125, obtained under the conditions of 75 eV at an ionization room temperature of 200 ° C and a sample temperature of 120 to 190 ⁰ C, and the following color reactions in thin layer chromatography: yellow with iodine / chloroform, negative with ninhydrin, reddish purple with sulfuric acid and yellow with potassium permanganate,
along with an agriculturally acceptable vehicle contains. 309851/1216