JPS644516B2 - - Google Patents

Description

[Detailed Description of the Invention] The present invention provides the following formula [] The present invention relates to a compound A11725N ₁ represented by and a method for producing the same.

More specifically, the present inventors first cultured Micromonospora sp. A11725, a strain that produces mycinamicin, a novel macrolide antibiotic, in a medium, and extracted from the culture. discovered various new antibiotic mycinamicins (Japanese Patent Application Laid-Open No. 53-54373,
(See Japanese Patent Application Nos. 54-24788 and 31316).

As a result of further research on this culture, they discovered a new substance that should be named the novel compound A11725N ₁ .

It has been found that the compound A11725N ₁ of the present invention has the following physicochemical properties, is a new substance having antibacterial activity, and is a useful compound having a core structure that is the parent of the antibiotic mycinamicin.

First, the compound A11725N ₁ of the present invention is a compound having the following physicochemical properties.

(1) Color property: Colorless crystal (2) Molecular formula: C ₂₁ H ₃₂ O ₄ (3) Elemental analysis: Measured value C = 72.49%, H = 9.43% Theoretical value C = 72.38%, H = 9.26% (4) Molecular weight: 348 (from mass spectrum) (5) Melting point: 159-160.5℃ (6) [α] ²⁶ _D : +18.9 (C = 1.0%, methanol) (7) Ultraviolet absorption spectrum: Shown in Figure 1 λmax 214.5nm (E1% 1cm=546) λmax 281nm (E1% 1cm=601) (Measurement concentration 0.028mg/ml, in methanol) (8) Infrared absorption spectrum: As shown in Figure 2,
3530, 3410, 2960, 2930, 2875, 1705, 1675,
1650, 1625, 1580, 1460, 1380, 1350, 1325,
1280, 1230, 1175, 1150, 1125, 1085, 1015,
It has absorption bands around 990, 970, 930, 890, 860, 845, and 820 ^{cm -1} (KBr method) (9) Nuclear magnetic resonance spectrum: As shown in Figure 3 (in deuterated chloroform using internal standard TMS).
100MHz) (10) TLC silica gel (Merck No. 5714) Benzene: Ethyl acetate: Ethanol (3:3:1) Rf=0.84 Chloroform-methanol-28% ammonia (15:1:0.1) Rf=0.85 (11 ) Color reaction Potassium permanganate aqueous solution decolorization reaction + ninhydrin reaction, Sakaguchi reaction, ferric chloride reaction
- (12) Distinction between acids and bases: Neutral (13) Solubility: Soluble in methanol, acetone, ethyl acetate, benzene, insoluble in water and hexane.

Furthermore, the antibacterial activity (MIC) of the compound A11725N ₁ of the present invention is as follows.

Staphyrococcus aureus0116 100γ/ml Strestococcus pyogenes NY5 200γ/ml Sarcina lutea ATCC 9341 >200γ/ml Corynebacterium diphtheriae PW8 25γ/ml Based on the above properties, the compound of the present invention A11725N ₁
is the following formula [] It turned out to be a new compound, which is a compound represented by Furthermore, the compound of the present invention has been found to be a useful compound having a nuclear structure that is the parent structure of antibiotic mycinamines, and is useful as a structural analysis reagent for antibiotic mycinamicins and as a synthetic intermediate for antibiotic mycinamicins. It is a compound.

The novel compound A11725-producing actinomycetes of the present invention are actinomycetes isolated from the soil of a potato field in Unazuki-cho, Shimoshinkawa-gun, Toyama Prefecture, and belong to the genus Micromonospora.・P. A11725 strain [This strain attaches one spore at a time to the tip of a sporophore produced from branched basal hyphae, does not form true aerial hyphae, and is a mesophilic bacterium, so It is a strain belonging to the genus Micromonospora.
P No. 4488'' (for detailed mycological properties, see Japanese Patent Application No. 53-54373).

The novel compound A11725N ₁ of the present invention is obtained as follows. That is, for example, the Micromonospora sp. A11725 strain described above is aerobically cultured in a medium containing medium components used for the cultivation of ordinary microorganisms. As the medium, a solid medium or a liquid medium can be used, and especially for mass production, a liquid medium, especially an aqueous medium, is suitable.

As a medium component, carbon sources such as glucose, starch, glycerin, sucrose, molasses, and dextrin are suitable for use.

Suitable nitrogen sources include peptone, meat extract, soybean flour, and casein hydrolyzate, but cottonseed flour, corn steep liquor, nitrates, and ammonia salts can also be used.

Other inorganic substances include sodium, potassium,
Substances containing cations such as magnesium, calcium, cobalt, manganese, iron, etc., and/or substances containing anions such as chlorine, sulfuric acid, phosphoric acid, acetic acid, etc. can be used. Furthermore, dry yeast and yeast extract can be used as bacterial growth promoting factors. Calcium carbonate can also be added to the medium to adjust the pH of the medium.

In addition, appropriate amounts of silicone resin, animal and vegetable oils, etc. can be added to the culture medium to suppress foaming during culture.

A particularly suitable medium for carrying out the method of the invention is a medium containing glucose, cottonseed flour, corn steep liquor, calcium carbonate as medium components.

As the culture conditions, known culture conditions used for the production of known antibiotics can be used. The culture temperature ranges from 20°C to 37°C, with temperatures of 26°C to 30°C being particularly suitable. The number of culture days varies depending on the culture conditions, but
Usually it takes 4 to 5 days.

Any known culture method can be used, but the aerated agitation culture method in a fermentation tank is particularly suitable for mass production. In order to isolate and collect the compound A11725N ₁ of the present invention from a culture, first, bacterial cells and other solid substances are removed by filtration or centrifugation, and then extracted from the liquid by an extraction method using an organic solvent. is the most suitable method. Organic solvents used for extraction include chlorinated hydrocarbons such as chloroform, dichloroethylene, and trichlorethylene, and fatty acid esters such as ethyl acetate, butyl acetate, and amyl acetate.
Any organic solvent that dissolves A11725N ₁ well and is difficult to mix with water can be used.

The organic solvent extract containing A11725N ₁ is distilled off under reduced pressure to obtain a crude substance containing A11725N ₁ . The crude substance is fractionated using methods such as column chromatography using silica gel or countercurrent distribution, and each fraction is subjected to silica gel thin layer chromatography to confirm its components.
Fractions containing pure A11725N ₁ are collected and the solvent is distilled off under reduced pressure to dryness to obtain a white powder.

The present invention will be explained below with reference to examples, but it is not limited thereto.

Example [A] 100 ml of a medium (PH7.0) containing 1% dextrin, 1% glucose, 0.5% casein hydrolyzate, 0.5% yeast extract, and 0.1% calcium carbonate was dispensed into a 500 ml Erlenmeyer flask, 120℃, 20
Heat sterilized for minutes. One platinum loop from a slant culture of Micromonospora sp. A11725 strain was inoculated into 10 bottles of this medium, and cultured with shaking at 30°C for 120 hours. This was then transferred to a 30-volume jar fermenter containing heat-sterilized medium 20 with the same composition as above, and incubated at 30°C and 300 rpm for 72 hours.
The culture was carried out with aeration and stirring under conditions of sterile air at 20 m/min. Next, 5% glucose, 2% cottonseed flour, 0.5% corn steep liquor, and 0.1 dipotassium phosphate.
%, magnesium sulfate 0.1%, calcium carbonate
Heat-sterilized medium containing 0.5% (PH7.2)
Transfer the above culture to a 250 volume tank containing 200
Transplant 10, 30℃, 120 hours, 250r.pm/min
Culture was carried out with aeration under sterile air conditions of 100°C to obtain a culture of 190°C.

[B] A liquid 160 was obtained by filtering the culture 190 to remove bacterial cells and other solid substances.

This liquid was extracted with the same amount of ethyl acetate to obtain 160 ml of an ethyl acetate solution containing Nippon. This was concentrated under reduced pressure to a concentration of 50%, and then the ethyl acetate layer was washed with 20% of an aqueous hydrochloric acid solution of pH 2.5, and the ethyl acetate layer containing the target compound was concentrated to dryness to obtain 73g of a brown syrupy crude product. Ta.

[C] Dissolve 73g of the above crude product in 500ml of benzene,
It was adsorbed onto a silica gel column (5 cm x 55 cm) that had been filled with benzene in advance. Then benzene-ethyl acetate-ethanol (170:30:2)
The mixture was developed with different solvents and fractionated into 50 ml portions. For each fraction, benzene-ethyl acetate-
The target product was confirmed by thin layer chromatography using ethanol (3:3:1) as a developing solvent, and fractions containing it were collected. The 100th fraction from the 75th fraction contains only the substance identified as A11725N ₁ , and this fraction is concentrated and dried to obtain 1.3 g of A11725N ₁ .
I got it.

[Brief explanation of drawings]

Figure 1 shows the ultraviolet absorption spectrum of the novel compound A11725N ₁ of the present invention, and Figure 2 shows the novel compound of the present invention.
FIG. 3 shows the infrared absorption spectrum of A11725N ₁ and the nuclear magnetic resonance spectrum of A11725N ₁ , the novel compound of the present invention.

Claims (1)

[Claims] 1 Compound A11725N represented by the following formula [] ₁ 2 Compounds belonging to the genus Micromonospora
A method for producing a novel compound A11725N ₁ , which comprises culturing A11725N ₁ producing bacteria in a medium and collecting compound A11725N ₁ from the culture. 3 Compounds belonging to the genus Micromonospora
A11725N ₁ The producing bacterium is Micromonospora S.
A method for producing the novel compound A11725N ₁ according to claim 2, which is P. A11725 strain.