JPH0523266B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Application Field] The present invention provides a novel antibiotic MG802-AF1-B,
Antibiotics belonging to it MG802-AF1-B1,
The present invention relates to a method for manufacturing MG802-AF1-B2 and MG802-AF1-B3. [Conventional technology] Until now, antibiotics produced by microorganisms have been
Over 5,000 types have been reported, and they are widely used to treat infectious diseases. Of these, Chai-nia
The production of 18 types of antibiotics has been reported from microorganisms belonging to the genus [Index of Actinomycetes Antibiotics, published by Yusako Co., Ltd.
Actinomycetes Antibiotics)]. [Problems to be Solved by the Invention] When using antibiotics, the appearance of bacteria resistant to the antibiotics is inevitable, and new antibiotics that are effective against resistant bacteria are constantly desired. [Means for Solving the Problems] To summarize the present invention, the first invention of the present invention relates to a novel antibiotic MG802-AF1-B, which has the following general formula: (In the formula, R ₁ represents hydrogen, R ₂ represents halogen, or
or R ₁ and R ₂ together indicate a conjugate, and R ₃
is a halogen). Moreover, the second invention of the present invention is based on the following formula: The antibiotic MG802− is a compound represented by
AF1−B1, the following formula: The antibiotic MG802− is a compound represented by
AF1−B2, the following formula: The antibiotic MG802− is a compound represented by
This invention relates to a method for producing AF1-B3, in which an MG802-AF1-B producing bacterium belonging to the genus Cainia is cultured, and the antibiotic MG802-AF1- is produced from the culture.
B1, MG802−AF1−B2 and MG802−AF1−B3
at least one antibiotic selected from the group consisting of
It is characterized by separating and collecting seeds. Since MG802-AF1-B exhibits antibacterial activity equivalent to Gram-positive bacteria and multidrug-resistant Gram-positive bacteria, it can be used as an antibiotic against Gram-positive bacteria that is also effective against resistant bacteria. An example of a bacterial strain producing MG802-AF1-B belonging to the genus Cainia is Chainia rubra, which was isolated from a soil sample collected by the present inventors in February 1982 in Sawada-cho, Sado-gun, Niigata Prefecture.
rubra) MG802-AF1 (Feikoken Bibori No. 8022). The mycological properties of this strain are as follows. [Mycological properties of the MG802-AF1 strain] 1. Morphology The MG802-AF1 strain forms hook-shaped to spiral-shaped aerial hyphae from branched basal hyphae under a microscope.
Whorled branches are not allowed. A mature spore chain is a chain of 10 or more spores, and the size of the spores is 0.4~
It is about 0.5 x 0.6 to 0.7 μm. The surface of the spore has short thorn-like protrusions. A sclerotia-like structure was also observed, and its diameter was approximately 5 to
It is 10μm. 2. Growth status in various media The standards shown in parentheses for describing colors are the Color Harmony Manual of Container Corporation of America.
manual) was used. (1) Sucrose/nitrate agar medium (cultured at 27℃) On the growth of light yellow daisies [2gc, Bamboo] to light brown [3io, Lt Amber], white to brownish white [3ba,
Aerial mycelia of Pearl to 3cb, Sand] are grown on the plant, and the soluble pigment is only yellowish brown. (2) Glucose-asparagine agar medium (cultured at 27℃) Growth is colorless to pale yellow [2gc, Bam-
boo] ~ Dull Rust [4pe, Orange Rust], does not grow aerial mycelia, and the soluble pigment is only reddish brown. (3) Glycerin-asparagine agar medium (ISP-
Medium 5.27℃ culture) Light brown [3ic, Lt Amber] ~ Light brown [3le, Maple] ~ Tea ash [4lg, Lt Spice Brown]
On the growing surface, thin white aerial mycelium grows, and the soluble pigment is only reddish-brown. (4) Starch/inorganic salt agar medium (ISP-medium 4.27
℃ culture) Light yellow brown [3lg, Adobe Brown] ~ Light brown [4ie, Cork Tan] ~ White to brownish white [3dc, Natural] aerial mycelia are grown on the growing brown ash, and the soluble pigment is red. It has a brownish tinge. (5) Tyrosine agar medium (ISP-medium 7.27℃ culture) White to brownish aerial mycelium grows on the growth of light brown to grayish reddish brown [5ng, Brick Red] and produces reddish soluble pigment. do. (6) Nutrient agar medium (cultured at 27℃) Yellow Maple [3ng, Yellow Maple] ~ Tea [4ni,
From around 20 days after culturing, a slight white aerial mycelium grows on the growing [Chestnut Brown], and the soluble pigment is only brownish. (7) Yeast/malt agar medium (ISP-Medium 2, 27
°C culture) Yellow tea to light tea [3le, Cinnamon to 3lg, Adobe
Brown], white aerial mycelium grows on it, and the soluble pigment is only reddish-brown. (8) Oatmil agar medium (ISP-Medium 3, cultured at 27℃) Light Daidai [3gc, Lt Tan] ~ Dull Daidai [3ic, Lt Amber ~ 4ic, Pastel Oran-ge] ~ Light Red Daidai [5ic, Lt Persi- On the growth of [mmon], white to brownish white [5 cb] aerial mycelium grows, and the soluble pigment is only yellowish brown. (9) Glycerin/nitrate agar medium (cultured at 27℃) The growth is colorless to pale yellow [2gc, Bamboo] to light yellow brown [3ic, Cinnamon], with no aerial mycelia attached.
The soluble pigment has a brownish tinge. (10) Starch agar medium (cultured at 27℃) Light yellowish brown to dull yellowish brown [4ic, Pastel Oran−
ge], a thin white aerial mycelium grows on it, and the soluble pigment is only slightly brownish. (11) Malate lime agar medium (cultured at 27°C) White to brownish aerial mycelium grows on the colorless to light yellowish brown growth, and the soluble pigment is only slightly yellowish. (12) Cellulose (synthetic solution added with paper strips, cultured at 27°C) White to brownish aerial mycelium grows on the growth of light yellow to light yellowish brown, and no soluble pigments are observed. (13) Gelatin puncture culture Simple gelatin medium (cultured at 20℃), glucose/
In both peptone and gelatin media (cultured at 27°C), growth is colorless, no aerial mycelia are attached, and no soluble pigments are observed. (14) Skimmed milk (cultured at 37℃) Growth is light yellow to light brown, no aerial mycelia attached,
The soluble pigment has a brownish tinge. 3 Physiological properties (1) Growth temperature range Yeast starch agar medium [Soluble starch (manufactured by Koso Kagaku Co., Ltd.) 1.0%, yeast extract (manufactured by Daigo Nutrient Chemical Co., Ltd.) 0.2%, string agar 3.0%, PH7.0] using
As a result of tests at 20℃, 24℃, 27℃, 30℃, 37℃, and 50℃, it grew at all temperatures except 50℃, but the optimal temperature was around 27℃ to 37℃. Presumed. (2) Liquefaction of gelatin (15% simple gelatin, cultured at 20°C; glucose peptone gelatin, cultured at 27°C) In simple gelatin medium, liquefaction begins around 14 days after culture, and its effect is weak. In the glucose-peptone-gelatin medium, no liquefaction was observed during observation for 30 days after culturing. (3) Hydrolysis of starch (starch. Inorganic salt agar medium and starch agar medium, both cultured at 27°C) In the case of starch/inorganic salt agar medium, 5 days after cultivation.
On starch agar medium, water decomposition was observed from around 10 days after cultivation, and the effect was moderate. (4) Coagulation and peptonization of skimmed milk (skimmed milk, 37
(℃ culture) Coagulation begins around 14 days after culture, and immediately after completion, peptonization begins. Peptonization occurs 25 days after incubation.
It will be completed around the day. The effects are both moderate to strong. (5) Production of melanin-like pigments (tryptone yeast prosthesis, ISP-medium 1: peptone yeast, iron agar, ISP-medium 6: tyrosine agar,
(ISP-Medium 7, all cultured at 27°C) No production of melanin-like pigment was observed in any of the media. (6) Utilization of carbon sources (Pridham-Godelive agar medium, ISP-medium 9, 27°C culture) D-glucose, L-arabinose, D-xylose, D-fructose. It grows using L-rhamnose, raffinose, and D-mannitol, but does not use inositol. It is presumed that sucrose is probably used. (7) Dissolution of malate lime (malate lime agar, 27
(℃ culture) From around 14 days after culturing, malate lime around the growth is dissolved, and its effect is moderate. (8) Nitrate reduction reaction (peptone water containing 0.1% potassium nitrate, ISP-medium 8, cultured at 27°C) Positive. To summarize the above properties, the MG802-AF1 strain is
No sporangia are observed, aerial hyphae are hook-shaped to spiral-shaped, and whorled branches are not observed. The surface of the spore has short thorn-like projections and sclerotium-like structures are observed. In various media, white to brownish white aerial mycelium grows on the growth of light yellow to light brown to brown ash, and the soluble pigment is
Yellow to reddish-brown in color. The production of melanin-like pigments was negative, and the hydrolyzability of starch was moderate. Regarding protein decomposition power, gelatin has weak liquefaction ability, and milk coagulation and peptonization ability is moderate to strong. The cell wall composition of the MG802-AF1 strain is that 2,6-diaminopimelic acid is LL
- type, does not contain arabinose as a sugar component. From these properties, it is estimated that the MG802-AF1 strain is an actinomycete belonging to the genus Cainia that has a sclerotia-like structure. Furthermore, we searched for known bacterial species similar to strain MG802-AF1 from the genera Cainia and Streptomyces, and found that Chainia rubra [International Journal of Systematic Bacteriology]
Bacteriology, Vol. 22, p. 347 (1972)] was listed as a closely related species. Table 1 shows Cainia rubra and MG802 in the literature.
-Comparison of AF1 strains is shown.

【table】

[Manufacturing method of MG802-AF1-B]

A culture containing MG802-AF1-B can be obtained by inoculating a MG802-AF1-B producing strain belonging to the genus Cainia into a nutrient-containing medium and growing it aerobically. As the nutrient source, those that can be used as a nutrient source for actinomycetes are used. For example, commercially available nitrogen sources such as peptone, meat extract, corn, stave liquor, cottonseed flour, peanut flour, soybean flour, yeast extract, NZ-amine, casein hydrolyzate, sodium nitrate, ammonium nitrate, ammonium sulfate, and Commercially available carbohydrates such as glycerin, sucrose, starch, glucose, galactose, mannose, and molasses, or carbon sources such as fat, and inorganic salts such as common salt, phosphate, calcium carbonate, and magnesium sulfate can be used. . In addition, trace amounts of metal salts, antifoaming agents such as animal, vegetable, and mineral oils may be added as necessary. These substances are used by production bacteria, and MG802−
Any culture material for known actinobacteria can be used as long as it is useful for producing AF1-B. Liquid culture is preferred for mass production of MG802-AF1-B, and the culture temperature is set at a temperature that allows the production bacteria to grow.
Applicable within the range of producing MG802-AF1-B,
The temperature is usually 20-40°C, preferably 27-37°C. MG802−AF1− was cultured using the conditions described above.
The method can be appropriately selected depending on the properties of the B-producing bacteria. MG802-AF1-B exists in both the culture solution and the bacterial cells. The culture solution can be extracted with a water-immiscible organic solvent such as butyl acetate, chloroform, or butanol at a pH of 10 or lower. Rather than bacterial cells,
After extraction with an organic solvent such as methanol or acetone, the extract can be concentrated under reduced pressure and further extracted in the same manner as the culture solution. In addition to the above-mentioned extraction methods, known methods used to collect fat-soluble substances, such as adsorption chromatography, gel permeation chromatography, scraping from thin layer chromatography,
Pure samples can be obtained by appropriately combining or repeating high-performance liquid chromatography and the like. [MG802-AF1-B1, MG802-AF1-B2,
Physical and chemical properties of MG802-AF1-B3] Shape, mass spectrometry, molecular formula, specific rotation, ultraviolet absorption maximum, infrared absorption maximum, solubility, acidity, neutrality, base of MG802-AF1-B1, B2, B3 gender distinction,
Table 2 shows the Rf values of silica gel thin layer chromatography.

[Table] Lum-methanol

Expanded in Le 40〓1)
Merck Art. 0.24
0.34
13724 (developed with methanol-water 95〓5)

Claims (1)

[Claims] 1. The following general formula: (In the formula, R ₁ represents hydrogen, R ₂ represents halogen, or
or R ₁ and R ₂ together indicate a conjugate, and R ₃
is a halogen). 2 The compound represented by the general formula has the following formula: The antibiotic MG802-AF1-B1 according to claim 1, which is a compound represented by: 3 The compound represented by the general formula has the following formula: The antibiotic MG802-AF1-B2 according to claim 1, which is a compound represented by: 4 The compound represented by the general formula has the following formula: The antibiotic MG802-AF1-B3 according to claim 1, which is a compound represented by: 5. Cultivate MG802-AF1-B producing bacteria belonging to the genus Cainia, and extract the antibiotic MG802-B from the culture.
AF1−B1, MG802−AF1−B2 and MG802−
Antibiotics MG802-AF1-B1, MG802-AF1-B2, characterized in that at least one antibiotic selected from the group consisting of AF1-B3 is isolated and collected;
Manufacturing method of MG802−AF1−B3.