JPS61282087A - Novel substance no.8345-a - Google Patents

Abstract

PURPOSE:To collect a novel substance from a culture of a strain of the genus Pereniporia and utilize the substance in the fields of agricultural chemicals and medicines in searching for substances having the plant growth inhibitory action and antibiotic substance action. CONSTITUTION:A fungus producing a novel antibiotic substance No.8345-A, e.g. Pereniporia medullaepanis AJ8345 (FERM-P No.8181), of the genus Pereniporia is cultivated in a culture medium containing a carbon source, nitrogen source, inorganic salt, etc. in which general basidiomycetes are grown, under aerobic conditions. The substance No.8345-A is separated from the resultant culture by using the extraction method, chromatographic method, etc. The properties of the collected substance No.8345-A are as follows; White amorphous powder. Molecular formula: C15H24O4. Melting point; 150-158 deg.C. Soluble in methanol and ethyl acetate and slightly soluble in water. Exhibiting color reactions, e.g. positive to potassium permanganate and negative to ninhydrin.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Application Field) The present invention relates to a novel substance 48345-A. This substance has plant growth inhibitory activity and antibiotic activity.
5-22754), anisomycin, toyokamayi y y
(Agria, Biol, Chem, #36 p.
2013 (1970), ), Herbisidden A,
B (J, AnLlblot,.

29.863 (1976), herbimycin (J.

Anttblot, 32, 255 (1979),
Cytovaricin (Abstract of the 5th International Pesticide Conference, l1
lb-3, 1982).

Bialajos (5th International Pesticide Conference Lecture Abstracts + fVa-1
9, 1982), among which bialaphos has been put into practical use as a herbicide.

In addition, antibiotics produced by basidiomycetes include polyacetylene compounds, terpenoid compounds, aromatic compounds, and nucleic acid analogs (Fermentation and Industry Vol-34 + All P
843) and other non-protein amino acids are known (Chemistry and Biology 14, 205 (1976)), but there are no reports on the plant growth inhibitory effects of these substances.

(Problems to be Solved by the Present Invention) The problems to be solved by the present invention are to provide a novel and useful substance that has both plant growth inhibiting action and antibiotic action.

(Means for Solving the Problems) In order to solve the above-mentioned problems, the present inventors cultivated a large number of basidiomycetes for the purpose of searching for a novel substance that has both plant growth inhibitory action and antibiotic action. As a result of searching for the liquid, Bacillus subtilis (Bacillus aubtill @
It was discovered that A3345-A, a substance that has a growth inhibitory effect on AJ1316 and at the same time suppresses plant growth, was produced. After isolating this effective substance from the culture solution, examining its physical and chemical properties, and comparing it with known substances, we found that 48345-A differs from known substances in its physical and chemical properties. Ta.

The present invention has been made based on these findings.

48345-A of the present invention is Pereniporia (Parent).
It is produced by basidiomycetes belonging to the genus -porim).

An example of this is Pereniporia medullaep.
anLs) AJ 8345 rFERM-P 81
81 (hereinafter referred to as AJ 8345 bacterium) has characteristics that advantageously produce 48345-A of the present invention.

In order to obtain A8345-A of the present invention, A3345-A producing bacteria belonging to the genus Illeniporia can be cultured by a conventional method for producing this substance. For industrially advantageous production, A3345-A producing bacteria may be cultured with aeration under aerobic conditions in a medium containing various nutrients.

The culture conditions and the composition of the medium may be those that allow general basidiomycetes to grow. In other words, the medium is basically a carbon source,
It contains a nitrogen source and inorganic salts, and if necessary, vitamins, precursors, etc. may be added.

Examples of carbon sources include glucose, arabinose,
Oxygen, starch, dextrin, glycerin, mannitol, organic acids, molasses, potato, etc. are used alone or in mixtures, and nitrogen sources include, for example, peptone, soy flour, corn cheese liquor, etc. , malt extract, amino sauce, rice bran, malt,
Urea, ammonium salts, etc. or mixtures thereof are used. Further, if necessary, an antifoaming agent such as silicone oil, soybean oil, or surfactant may be added.

The medium is preferably a liquid medium, and - of the medium is about 6.0 to about 8.
．． The culture temperature is preferably adjusted to about 20 to about 35°C.

After completion of the culture, 48345-A can be separated and collected from the culture in accordance with a conventional method for separating and collecting fermentation products from the culture. That is, Mu8345-A is collected by appropriately combining extraction methods using various organic solvents, Chroma Dog 2 Feeding using various active adsorbents, and the like.

Next, an example of manufacturing 48345-A will be shown as an example, but this example is not intended to limit the scope of the present invention in any way.

Example I Production and structural analysis of A3345-A AJ8345 grown on potato dextrose agar slope
The bacteria were mixed with 1% soluble starch, 2% glucose, 0.5% ammonium sulfate, potassium phosphate O, OSS, magnesium sulfate 0.05%, sodium chloride 0.05%,
Fermentation medium containing 70 g/l of potato extract and 4 parts of trace metal*1 (50Qm/100T in Yosaka flask)
R1 portions] and cultured with shaking or standing at 27° C. for 20 to 30 days.

*Trace metal CuSO4"5H200,64F Fe SO4・7H200,11g MnC12"4H200,7917 ZnSO4H'7I (2o O-15g distilled water
10 (1/ After removing the bacterial cells from the obtained culture solution 61 by filtration, the sterilized solution was extracted twice with ethyl acetate from 61, and the ethyl acetate layer was concentrated under reduced pressure and dried, and then added with a small amount of methanol. Next, it was purified by silica column chromatography (Roper column 8160, size B, manufactured by Merck & Co.).The developing solvent was methanol-ethyl acetate (5:95).
) was used. The active fraction was collected, concentrated to dryness under reduced pressure, dissolved in a small amount of methanol, and dissolved in 5sphadex LH-20.
The product was purified by gluchromatography using (column volume: 300 m). Methanol was used for development. After collecting the active fraction and concentrating it under reduced pressure, transfer it to semi-preparative μBondap.
The active ingredient was purified and isolated by HPLC using an aKC18 (registered trademark 2 manufactured by Nippon Waters) column.

35 timeethanol was used as the mobile phase. As shown in FIG. 4, A8345-A was fractionated at a retention time of about 58 minutes. By this method, we finally obtained 48345-
A was isolated at 19 mF.

Antibiotic 48345-A thus obtained is a novel antibiotic having the physicochemical and biological properties described below.

(1) Appearance: White amorphous powder.

(2) Elemental analysis values: carbon 63.6%, hydrogen 8.4%, nitrogen 0.1% (weight ratio) (3) Molecular weight: FD-MS Vz: 26 B (M
) (4) Molecular formula 15H2404 (5) Specific rotation: [α) −181,1(C0,2
5, methanol) (6) Melting point: 156-158°C (7) Solubility: Soluble in methanol and ethyl acetate. Slightly soluble in water.

(8) Ultraviolet absorption spectrum: Does not show characteristic absorption maximum in methanol solution.

(9) Infrared absorption spectrum: The spectrum measured in potassium bromide tablets is as shown in FIG.

αQ Hydrogen Nuclear Magnetic Resonance Spectrum The spectrum measured in double methanol is shown in Figure 2.

α carbon nuclear magnetic resonance spectrum The spectrum measured in double methanol is shown in FIG.

(6) Single value of silicadal thin layer chromatogram: methanol-ethyl acetate (5:95) 0.73 (6) Color reaction Potassium dipermanganate positive, ninhydrin negative Antibiotic M8345- with the above physicochemical properties There are no other applicable substances for A, and it is a new substance.

The biologically active antibiotic 48345-A of Example 248345-A has a Da of the composition shown in Table 1.
A minimal medium of Bacillus genus Bacillus was shown to inhibit the growth of bacteria, whereas a medium to which polybegtone and yeast extract were added did not show any growth inhibiting effect. The minimum inhibitory concentrations (MIC) for typical bacterial strains are shown in Table 2.

Table 1 - pH 7.0 It had an effect, and particularly inhibited root elongation during germination of lettuce and edible beetroot seeds. Details are as shown in Table 3.

Biological activity of 48345-A 5arclna 1utea AJ1217
50) 50Xanthomonas oryza
e AJ2783) 50, >50Baci
llus 5ubtilis AJ1316 6
．． 25) 50 medium 1. Davis minimal medium 2. Dama 18 minimal medium + 0.2% polypezotone.

0.11 yeast extract Tertiary coat Plant growth inhibitory effect

[Brief explanation of drawings]

FIG. 1 is an infrared absorption spectrum measured in a potassium bromide tablet of A3345-A. FIG. 2 is a hydrogen nuclear magnetic resonance spectrum of &8345-A measured in heavy methanol. Figure 3 is the carbon nuclear magnetic resonance spectrum of A3345-A measured in heavy methanol. Figure 4 is the carbon nuclear magnetic resonance spectrum of A3345-A.
It is a preparative sample diagram of A using a high performance liquid chroma dog 2 fee.

Claims (1)

[Claims] 1. New substance No. 1 characterized by having the following physical and chemical properties. 8345-A (1) Appearance: White amorphous powder. (2) Elemental analysis values: 63.6% carbon, 8.4% hydrogen, 0.1% nitrogen (weight ratio) (3) Molecular weight: FD-MSm/z; 268 (M^+) (
4) Molecular formula: C_1_5H_2_4O_4 (5) Specific optical rotation: [α]^2^4_D-181.1° (C0.25,
Methanol) (6) Melting point: 156-158°C (7) Solubility: Soluble in methanol and ethyl acetate, slightly soluble in water. (8) Ultraviolet absorption spectrum: Does not show characteristic absorption maximum in methanol solution. (9) Infrared absorption spectrum: The spectrum measured in potassium bromide tablets is as shown in FIG. (10) Hydrogen nuclear magnetic resonance spectrum: The spectrum measured in heavy methanol is shown in FIG. (11) Carbon nuclear magnetic resonance spectrum: The spectrum measured in heavy methanol is as shown in FIG. (12) R_f value of silica gel thin layer chromatogram: methanol-ethyl acetate (5:95) 0.73 (13) Color reaction: Potassium permanganate positive, ninhydrin negative.