JPS6110595A - Novel antibiotic guanidylfungin b and preparation thereof - Google Patents

Abstract

NEW MATERIAL:An antibiotic guanidylfungin B expressed by the formula (either one of the carbon atoms at the 23- and 25-positions is malonic acid ester). Apperance; White powder. Melting point; 138-142 deg.C decomposition. Molecular weight; 1,115. Elementary analysis (%); C 60.87; H 9.30; N 3.55. Molecular formula; C57H101N3O18. Rf value; 0.17 (silica gel plate/CHCl3-CH3OH-H2O 65:25:4). Soluble in dimethyl sulfoxide and pyridine, sparingly soluble and insoluble in chloroform, ethyl acetate and acetone, etc. Positive to potassium permanganate, Dragendorff reactions, etc., and negative to the ferric chloride, 2,4-dinitrophenylhydrazine reactions, etc. USE:An antimicrobial agent. PREPARATION:A microorganism, belonging to the genus Streptomyces, and capable of producing guanidylfungin B is cultivated at 25-35 deg.C for 2-5 days preferably by the liquid culture medium cultivation method under stirring and aeration, and the aimed antibiotic is extracted from the resultant microbial cells.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a novel antibiotic guanidylphungin B and a method for producing the same.

[Conventional technology]

Antibiotics similar to guanidylfungin B are 66
2-mu (JP-A-58-170482, hereinafter referred to as guanidyl 7-in-jinmu), 662-A' (JP-A-58-170482)
No. 170482, hereinafter referred to as methyl-guanidyl7indi/A), azaromycin'*e'4e'i (
The Journal of Antibiotic Fever (J, Mun.
tibiotican volume 25.

107 pages (1970)], Copiamycin (Special Publications 1970)
1-15519), neocopiamycin A (The Journal of Antibiotics, Vol. 37, p. 105 (I984)), nifificin 2 (antibiotyphoid),
AntibiO-tikl) Volume 19, Page 579 (I9
1974)] have been reported. Their structures are shown below.

Guanidylfungin A (R=H), methyl-guanidylfungin A (R=OHs) Azapmycin? , (R1=R-value=H), F4(i'j
l=mouth H,, R,==H), F'S (R1=
Rx=OHm) copiamycin (R: 0H
j), Neocopiamycin (R=H) Niphimycin I [Problems to be Solved by the Invention] The purpose of the present invention is to develop a structurally novel antibiotic guanidyl 7
The present invention aims to provide B and a method for producing the same.

[Friendly means of solving problems]

To summarize the present invention, the first invention of the present invention relates to guanidylphungin B, and is represented by the following formula (I): (wherein, the 0-25th position and the C-25th position are malonic acid is a compound represented by An invention relating to a method for Streptomyces (8)
The method is characterized in that guanidyl funsif B-producing l@ belonging to the genus Treptomycea) is cultured, and guanidyl funsif B is collected from the culture.

The present inventors conducted a search for new antibiotics using bacterial strains belonging to the genus Streptomyces. As a result, they discovered that a new antibiotic, which had not been described in any literature, was produced in a culture of a certain strain of the genus, and succeeded in isolating and purifying this active ingredient. It was named.

The present invention will be explained in detail below.

The physicochemical properties and biological properties of the antibiotic guanidylphungin B according to the present invention are as follows.

(I) Color of substance: White powder (2) Melting point: 215B-142℃ (decomposition) (3) Molecular weight: 1115 (by secondary ion mass spectrum) (4) Elemental analysis (weight%) 2 Analysis value Carbon 6187
゜Hydrogen 9.50.

Nitrogen

(6) Infrared absorption spectrum: Figure 1 of the attached drawing [wave number (
6n''''1) (graph of the 7'CKBr method shown by the relationship between the horizontal axis and the transmittance (%) (vertical axis)), it shows the following maximum value.

3350.2950.2900.1710.1660.
1580.1450.1370.1080.1060'
(tM-'') (7) Thin layer chromatography: The following Rf value is shown in thin layer chromatography using a silica gel plate (No. 5554, manufactured by Merck & Co.).

a) 0HO2s-methanol-'H,065225
:4 (capacity/capacity) α17b)
5ec-butanol-H,04: 1 (volume/volume)
n1O(8) Solubility Soluble in dimethyl sulfoxide and pyridine, but difficult to bathe or insoluble in chloroform, ethyl acetate, acetone, methanol, and water.

(9) Positive for color reaction potassium dipermanganate, Dragendorff, Sakaguchi reaction, ferric chloride,
Negative for 2,4-dinitrophenylhydrazine reaction.

(I0 Antibacterial Spectrum: Table 1 shows the results obtained by determining the minimum growth inhibitory concentration (M-C) by the Kite dilution method.

From the above, it is clear that the antibiotic guanidyl 7injin B of the present invention is a new substance that has not been described in any literature, and its chemical structure is as shown in formula (I) above. Met.

Next, a method for producing the antirealistic substance guanidylphungin B of the present invention will be explained.

Guanidylfungin BH of the present invention A guanidylfungin B-producing bacterium belonging to the genus Streptomyces is cultured in a nutrient medium, and the antibiotic guanidylfungin B is produced and accumulated in the culture. It can be produced by isolation.

Examples of guanidylphungin B-producing strains belonging to the genus Streptomyces include, for example, Streptomyces-bygroscopicus (8treptomycθθhygroscope).
icus) No. 662 strain (Feikoken Bacteria No. 6434).

Streptomyces vigroscopicus no.

The mycological properties of the 662 strain are disclosed in JP-A-58-170482.
It is clearly stated in the issue.

Cultivation in the production of guanidylphinsif B of the present invention is carried out as follows. In culturing guanidylphungin B-producing bacteria belonging to the genus Streptomyces, conventional culture methods for producing antibiotics are generally used.Nutritional media useful for this purpose are nutrient media that can be used by actinomycetes. The medium may contain sugars, starches, assimilable carbon sources such as glycerin, and organic or inorganic nitrogen sources such as corn steep liquor, yeast extract, meat extract, peptone, and ammonium salts. It may also contain phosphates, metal salts such as calcium, iron, etc. If necessary, silicone, vegetable oil, or synthetic antifoaming agent may be added to the medium as an antifoaming agent.

As a culture method, a liquid culture method is suitable, and the culture is carried out at a culture temperature of 25 to 55° C. with aeration for usually 2 to 5 days.

Guanidylfungin Bi can be obtained from the culture thus obtained by appropriately using any means commonly used to collect metabolites. for example,
Any of the following methods may be used alone: one that utilizes the difference in adsorption affinity between guanidylphungin B and impurities, one that utilizes the difference in molecular weight, one that utilizes the difference in ionic bonding force, and one that utilizes the difference in solubility. used in combination or repeatedly.

Specifically, guanidylphungin B accumulates in particularly large amounts in cultured bacterial cells, and is extracted from V bacterial cells with an organic solvent such as aqueous acetone or methanol. It can be adsorbed onto a solvent and eluted with an organic solvent such as methanol or a mixture of chloroform, methanol and water, or a mixture of methanol and water. Furthermore, it can be collected as a white powder by recrystallization with water-containing acetone, methanol, etc.

〔Example〕

The present invention will be explained in detail below with reference to Examples, but the present invention is not limited thereto.

Example 1 Glucose 1 f/cLt, polypeptone (L 2 t
/61% Meat
2) was dispensed into a Sakalo flask, and after sterilization, Streptomyces vigroscopicus No. 662 strain was bridged with one platinum loop, cultured with shaking at 27° C. for 48 hours, and this was used as an incubation culture solution.

Soluble starch 1 t/At, glucose 1 f, 4
L corn steep force -1.5f/dt, yeast: j-
+7゜α1 t/eLL% N-acetylgelcosagun α025t/dL -KxHPO4 α05 f/eL
I 5NH4C2α05t/61. %FeSO4 [
L O1t/eLL, α6t/dt of 0aOOs (
After sterilization, 20 t of a medium consisting of PH 7.2) was placed in a 50 t stainless steel fermentor, and after sterilization, 100 t of the above seed culture solution was inoculated thereto, and the mixture was aerated and stirred at 27°C for 6 days (rotation = 500 r, p, m, ventilation: 10t
/min).

The bacterial cells are removed by filtration from the culture solution prepared as above.
2 tons of acetone-water (7:5, volume/volume) mixed solvent was added to this, the bacterial cells were thoroughly immersed, and after stirring,
The extract was collected. This operation was repeated three times, and the extracts were combined and concentrated under reduced pressure to obtain 50f'i of an oily substance. After adsorbing this oily substance on about 2Of Celite, silica gel (N
o, 7734) was packed at the top of the IL column. Mixed solvent of chloroform-methanol (5:1) 2
After washing with t, chloroform-methanol-water (5:4
It was bathed with 4 t of the mixed solvent of 21). The eluted active fractions were collected and the solvent was removed under reduced pressure to form a crude powder 101F.
I got it. This coarse powder was mixed with 2 mixtures of acetone and water (7:5).
The mixture was dissolved by heating at 00 dVC and left at room temperature for 3 days, and the resulting precipitate was collected to obtain 5 ft of pale yellow powder.

This powder 17f was dissolved in pyridine 2- and methanol-α01 molar ammonium acetate aqueous solution (7
6:24) and adjusted to saturation with a mixture of
Pak)■500/0ss (manufactured by Waters), developed and eluted with the same solvent to obtain an active fraction. After collecting these fractions and distilling off the bath medium under reduced pressure, a mixture of acetone and water (7:3) was dissolved at 100 dlC, and the solution was heated at room temperature for 2 hours.
The mixture was left to stand for several days, and the resulting precipitate was collected to obtain a white powder of α4f. This powder α4f was mixed with steam-butanol-water (9:1).
After dissolving in a mixed solution of 10-10, it was applied to a silica gel column (IOOG) which had been saturated with 5ec-butanol in advance. After washing with 5ec-butanol 100-, 5ec
-Butanol-water (8:1 mixture 400-1) followed by development with a mixture of the same solvent (4:1) 400-1, followed by bathing.
Guanidylfungin A, a known substance, was obtained from the ec-butanol-water (8:1) elution fraction, and the same solvent (4:1) was obtained.
:1) Guanidylfungin B was obtained from the elution fraction.

The fractions containing this guanidylphungin B were collected, the solvent was distilled off under reduced pressure, and a mixture of acetone and water (7:5) was
The mixture was reconsolidated to obtain a white powder.

〔Effect of the invention〕

As explained above, the present invention provides a novel antibiotic guanidylphungin B and a method for producing the same.

[Brief explanation of drawings]

FIG. 1 is an infrared absorption spectrum (KBr) graph of guanidylphungin B of the present invention.

Claims (1)

[Claims] 1. Guanidylfungin B represented by the following structural formula (I). ▲There are mathematical formulas, chemical formulas, tables, etc.▼...(I) (In the formula, the C-23 and C-25 positions indicate that malonic acid is ester bonded to either of them.) 2. Streptomyces Guanidyl fungin B belonging to the layer
Cultivate the producing bacteria and extract guanidylphungin B from the culture.
A method for producing guanidylphungin B, which comprises collecting guanidylphungin B.