JP2000316592A - New antibiotic tkr 2993 and its production - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a new antibiotic TKR 2993 which is useful as an antifungal agent effective for preventing and treating the fungal diseases of animals and plants, and so on, by culturing a strain which belongs to the genus Cylindrocarpon and has an ability to produce the antibiotic TKR 2993. SOLUTION: This new antibiotic TKR2,993 (salt) has the following physicochemical properties, mass spectrometry: FAB-MS m/z 1,410 [M+H]+; the number of carbon atoms: 72; the number of nitrogen atoms: 14; infrared absorption spectrum (KBr method): main absorption wavelengths (cm-1): 3,440, 2,960, 1,640, 1,470, 1,210, 840, 800, 720; solubility: soluble in methanol, slightly soluble in hexane, chloroform, water; amino acid analysis (detection by ninhydrin reaction): threonine, alanine, valine, leucine and proline are detected. The antibiotic is obtained by culturing a strain [Cylindrocarpon sp. TKR2993 (FERM P-17365)] belonging to the genus Cylindrocarpon and then isolating the product from the culture product.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a novel antibiotic TKR2993 useful as an antifungal agent and a method for producing the same.

[0002]

2. Description of the Related Art Fungi are known to infect humans, animals, plants and the like and cause various diseases. For example,
It causes superficial mycosis on human skin, oral cavity, etc., causes systemic mycosis on internal organs, brain, etc., and causes similar infections on animals such as pets and livestock. Furthermore, various diseases are caused on plants such as fruit trees and vegetables. Among these, the main fungi that cause human infection and cause systemic mycosis are Candida and Cryptococcus.
ococcus) and Aspergillus, etc., and it is considered that superficial mycosis is mainly caused by Candida which infects the skin, oral cavity, vagina, etc., Trichophyton which infects the skin of limbs and the like. Various other fungi exist in the living environment and are considered to be the cause of contamination of animals and plants.

[0003]

At present, only a very small number of antifungal agents which can be used for the treatment and protection against such fungal infections and contaminations are known. Absent. Among them, as a therapeutic agent for systemic infections of animals including humans in particular, amphotericin B, flucytosine, miconazole, fluconazole and the like can be mentioned. But these things
There were problems in efficacy, toxicity, antibacterial spectrum, etc., and they were not sufficient as therapeutic agents. The purpose of the present invention is
In view of such a situation, it is an object of the present invention to provide a novel antibiotic useful as an antifungal agent.

[0004]

Means for Solving the Problems In order to search for novel antibiotics, the present inventors isolated a large number of microorganisms from nature, isolated the antibiotics produced therefrom, and examined their biological properties. , Cylindrocarpon
It has been found that antibiotics exhibiting antibacterial activity against pathogenic fungi such as Candida albicans are present in cultures of microorganisms belonging to the genus. Thereafter, the present inventors
As a result of isolating this antibiotic and examining its physicochemical properties, it was confirmed that it was a novel substance having a unique physicochemical property and not listed in the literature, and this antibiotic was named TKR2993. Thus, the present invention provides the above antibiotic TKR2993 and a method for producing the same. Also, the present invention
Provided is a novel strain of the genus Cylindrocarpon having an ability to produce an antibiotic TKR2993 useful for this production method, and a processed product of the microorganism. The antibiotic TKR2993 of the present invention, or a microorganism capable of producing the antibiotic or a processed product thereof,
It is useful as a prophylactic or therapeutic agent for fungal diseases of humans, animals or plants, and the present invention also provides an antifungal agent containing the antibiotic as an active ingredient. In the present specification, the “antifungal agent” includes a medicine, an animal medicine, a cosmetic, an agricultural and horticultural medicine, and the like.

[0005]

BEST MODE FOR CARRYING OUT THE INVENTION The antibiotic TKR2993 of the present invention has the following physicochemical properties (1), (2), (3), (4), (5) and (6). (1) Mass spectrometry: FAB-MS m / z 1410 [M + H] ⁺ . (2) Carbon number: 72, nitrogen number: 14. (3) Ultraviolet absorption spectrum (in methanol): shows the terminal absorption as shown in FIG. (4) Infrared absorption spectrum (KBr method): The main absorption wave numbers (cm ^-1 ) are 3440, 2960, 1640, 147 as shown in FIG.
0, 1210, 840, 800, 720. (5) Solubility: soluble in methanol. Poorly soluble in hexane, chloroform and water. (6) Amino acid analysis (detected by ninhydrin reaction):
Threonine, alanine, valine, leucine and proline are detected. The TKR2993 also has a ¹ H-NMR spectrum shown in FIG. 3,
It has the ¹³ C-NMR spectrum shown in FIG. 4, and has the property of being eluted at the position shown in FIG. 5 in reverse phase partition high performance liquid chromatography.

[0006] The antibiotic TKR2993 of the present invention is produced by culturing a strain belonging to the genus Cylindrocarpon and capable of producing TKR2993, and then isolating TKR2993 produced from the culture of the above strain. can do. The above strain used in the present invention includes TKR2
The strain is not particularly limited as long as it is a strain capable of producing 993. Such strains include, for example, Cylindrocarpon sp. TKR2993 (hereinafter simply referred to as TKR2993). TK
The R2993 strain is a novel strain that has not been described in the literature, and has been isolated and identified for the first time by the present inventors and has the property of producing TKR2993 in an advantageous manner. Below, TKR299
The mycological properties of the three strains will be described in detail. The TKR2993 strain has the color tone of colonies (hereinafter, also referred to as “communities”) in various media as shown in Table 1. In addition, the color tone in the table is based on the color name according to Japanese Industrial Standard JIS Z8102 (1985),
This is shown by the results of observation at 7 days after inoculation into the medium and culturing at 25 ° C.

[0007]

[Table 1] __________________________________________________________________________ Diameter of the village Color of the village Color of the back of the village State of the village (mm) Color Peach Fluffy Strawberry 2.5Y7 / 6 5YR8 / 3 Agar medium 3) Sabouraud 81 flesh-yellow Fluffy agar medium 5YR8 / 6 2.5Y8 / 6 4) Ypss 55 Naples yellow Bright gray yellow red Fluffy agar medium 5Y8 / 8 10R8 / 4 ________________________________________________________________________

[0008] The TKR2993 strain grows quickly on a potato dextrose agar medium, a Sabouraud agar medium, or the like, and the surface of the colony is fluffy, the center becomes moist, and it is slightly raised. The phialides of the TKR2993 strain are spindle-shaped or deformed cylindrical, and grow laterally from hyphae. The large conidium is cylindrical to kidney-shaped and has 1-3 partition walls, and its size is 10-14 × 2.6-4.0 μm. Small conidia are 1
~ 2 cells, from smooth elliptical to cylindrical, with a size of 3-8 x 1.2-2.2 m. In the above medium, ascospores and chlamydospores are not formed.

The physiological properties of the mycological properties of the TKR2993 strain are as follows. Growth temperature range: The growth temperature range is 10-37 ° C,
The optimum growth temperature is around 30 ° C. Growth pH range: The pH range at which growth is possible is pH3 to pH9,
The optimum pH for growth is pH 4-7. A bacterial species having the above mycological properties was identified by Joseph C. Gilman, A manual of soil fungi,
Constable and Company Limited (Cons
By searching for strains of the genus Cylindrocarpon described in the literature such as table and company ltd) (1959), the TKR2993 strain can be identified as a strain belonging to the genus Cylindrocarpon. it can. However, a strain belonging to the genus Cylindrocarpon, TKR2
There is no report on the ability to produce 993. Therefore, the present inventors designated this strain as a new strain, and used it as a strain of Cylindrocarpon sp.
drocarpon sp. TKR2993) and Cylindrocarpon s
p. TKR2993, and deposited on April 13, 1999 with the Research Institute of Biotechnology and Industrial Technology under the deposit number FERM P-17365. In the present invention, in addition to the above-mentioned TKR2993 strain, natural or artificial mutant strains of TKR2993 strain, other strains belonging to the genus Cylindrocarpone, and the like,
A microorganism capable of producing 3 can be used.

[0010] In the present invention, TKR2993 is
A strain having a 993 producing ability is inoculated into a nutrient source-containing medium,
It is produced by culturing. Among the nutrient sources, examples of the carbon source include glucose, fructose, saccharose, starch, dextrin, glycerin, molasses, starch syrup, oils and fats, and organic acids. Examples of the nitrogen source include soybean flour, cottonseed flour, corn steep liquor, casein, peptone, yeast extract, meat extract, germ, urea, amino acids, organic nitrogen compounds such as ammonium salts, inorganic nitrogen compounds, and the like. it can. Examples of the salts include inorganic salts such as sodium salt, potassium salt, calcium salt, magnesium salt, and phosphate. Each of these may be used alone or in combination as appropriate. The aforementioned nutrient source-containing medium, further, if necessary, iron salts, copper salts, zinc salts, heavy metals such as cobalt salts; biotin, vitamins such as vitamin B _1; other, helping the growth of bacteria, the TKR2993 Organic substances, inorganic substances, and the like that promote production can be appropriately added. In addition to the above-mentioned nutrient sources, if necessary, an antifoaming agent such as silicone oil and polyalkylene glycol ether, a surfactant and the like can be added.

When culturing a strain capable of producing TKR2993 in the above-described nutrient-containing medium, a method generally used for producing an antibiotic by culturing a microorganism can be used. A culture method, in particular, a shaking or deep aeration stirring culture method can be suitably used. The above culture is preferably carried out at 15 to 25 ° C., and the pH of the medium is usually pH 3 to 8, but preferably around pH 5. In general, a sufficient production amount can be obtained in a culture period of 6 to 11 days. By the above culture method, TKR2
993 is contained in the culture solution and cells and accumulates in the culture. In the present invention, TKR2 accumulated in the culture
993 can be isolated from a culture using the physicochemical properties of these antifungal substances, and further purified and obtained as necessary.

Separation from the culture can be carried out by extracting the whole culture with a non-hydrophilic organic solvent such as ethyl acetate, butyl acetate, chloroform, butanol, methyl isobutyl ketone and the like. Alternatively, the culture may be separated into a culture solution and cells by filtration or centrifugation, and then separated from the culture solution and cells. In order to separate TKR2993 from the culture solution, the above extraction method with a non-hydrophilic organic solvent can also be adopted.Also, the culture solution is brought into contact with an adsorptive carrier, and the TKR2993 in the culture solution is adsorbed on the carrier. After that, a method of eluting with a solvent may be employed. As the carrier, for example, activated carbon, powdered cellulose,
Adsorbent resins and the like can be mentioned. The solvent may be used singly or in combination of two or more depending on the type, properties and the like of the carrier. For example, aqueous solvents such as water-containing acetone and water-containing alcohols may be appropriately combined. And the like.
In order to separate TKR2993 from the above cells, a method of extracting with a hydrophilic organic solvent such as acetone can be adopted.

In the present invention, the crude extract of TKR2993 thus separated from the culture can be subjected to a step of further purification, if necessary. Purification can be performed by a method commonly used for separation and purification of fat-soluble antibiotics. Examples of such a method include column chromatography using a carrier such as silica gel, activated alumina, activated carbon, and an adsorptive resin. And high performance liquid chromatography. When the column chromatography method using silica gel as a carrier is employed, examples of the elution solvent include chloroform, ethyl acetate, methanol, acetone, water, and the like, and two or more of these can be used in combination. . When the high-performance liquid chromatography method is employed, examples of the carrier include chemically bonded silica gel in which an octadecyl group, an octyl group, and a phenyl group are bonded; a polystyrene-based porous polymer gel, and the like. For example, an aqueous solution of a water-soluble organic solvent such as aqueous methanol and aqueous acetonitrile can be used.

The microorganism capable of producing TKR2993 of the present invention produces TKR2993 in a cell or outside a cell. Thus, the microorganisms of the present invention can be used as needed to provide TKR2
When used together with a nutrient medium suitable for the production of 993, it can be used as an active ingredient of an agricultural and horticultural drug. Furthermore, when a microorganism having a TKR2993 producing ability of the present invention is subjected to a known sterilization method, for example, heat sterilization, ethylene oxide gas sterilization, or ultraviolet sterilization, the treated product has lost the growth ability as a microorganism, TKR2993 is retained in the treated cells. Therefore, the microorganism capable of producing TKR2993 and / or its processed product can be used as an active ingredient of an agricultural and horticultural drug for preventing or treating a fungal disease of a plant.

The TKR2993 of the present invention can be used as it is or
It can be used in medicine as its pharmacologically acceptable salt. The salt is not particularly limited as long as it is pharmacologically acceptable. For example, salts of mineral acids such as hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrofluoric acid and hydrobromic acid;
Acetic acid, tartaric acid, lactic acid, citric acid, fumaric acid, maleic acid, succinic acid, methanesulfonic acid, ethanesulfonic acid,
Salts of organic acids such as benzenesulfonic acid, toluenesulfonic acid, naphthalenesulfonic acid and camphorsulfonic acid;
Examples thereof include salts of alkali metals such as sodium, potassium and calcium and salts of alkaline earth metals and the like.
When TKR2993 of the present invention or a pharmacologically acceptable salt thereof is administered as a medicament, the TKR2993 of the present invention or a pharmacologically acceptable salt thereof is used as it is or a pharmaceutically acceptable non-toxic and non-toxic. In an active carrier, for example, 0.1
It is administered to animals including humans as a pharmaceutical composition containing 999.5%, preferably 0.5-90%. Examples of the carrier include, for example, solid, semi-solid or liquid diluents, fillers and other formulation aids, and the like.
One or more of these can be used. The pharmaceutical composition is preferably administered in the form of a dosage unit, and can be made into a dosage form suitable for the administration method by a method known per se, for example, oral administration, intra-tissue administration, topical administration (transdermal administration) Etc.) or rectally. Oral administration can be carried out in solid, powder or liquid dosage units, for example, powders, powders, tablets, sugar coatings, capsules, drops, sublinguals, other dosage forms, and the like. Parenteral administration can be accomplished, for example, by using liquid dosage unit forms for subcutaneous, intramuscular or intravenous injection, such as solutions and suspensions. These include suspending or dissolving a fixed amount of TKR2993 of the present invention, or a pharmacologically acceptable salt thereof, in a nontoxic liquid carrier suitable for injection, such as an aqueous or oily vehicle. And
It is then produced by sterilizing the suspension or solution. Topical administration (such as transdermal administration) can be carried out, for example, by using external preparations such as liquids, creams, powders, pastes, gels, and ointments. These can be used to apply a certain amount of TKR2993 of the present invention, or a pharmaceutically acceptable salt thereof, to flavors, coloring agents, fillers, surfactants, moisturizers, emollients that are suitable for the purpose of external preparations. It is produced by combining with one or more of agents, gelling agents, carriers, preservatives, stabilizers and the like. Rectal administration according to the invention
TKR2993, or a certain amount of its pharmacologically acceptable salts, for example, suppositories mixed with low-melting solids such as higher esters such as myristyl palmitate, polyethylene glycol, cocoa butter, and mixtures thereof And the like.

When the TKR2993 of the present invention or a pharmacologically acceptable salt thereof is administered as a medicament, the dose as an antifungal agent depends on the condition of the patient such as age and body weight, the administration route, the nature and extent of the disease, etc. It is desirable to adjust in consideration of, but usually, for humans, the amount of the active ingredient of the present invention for adults is in the range of 10 to 2000 mg per day, and severe side effects in this range Without the desired medicinal effect. It can also be administered in several divided doses per day.

The TKR2993 of the present invention can be used as it is or as a salt thereof as an active ingredient of agricultural and horticultural medicine. Further, as described above, the TK of the present invention
A microorganism capable of producing R2993 and / or a processed product thereof can also be used as an active ingredient of an agricultural or horticultural drug. The agricultural and horticultural drug of the present invention only needs to contain at least one component selected from the above-mentioned active ingredients, and may also contain other active ingredients. Other active ingredients include, for example, JP-A-4-45792,
JP-A-9-25286, JP-A-9-249680, WO95
/ 18147, WO96 / 28456, WO97 / 26367, WO97 / 34012, WO98 / 2
1196, WO98 / 23767, WO98 / 56755 and PCT / JP98 / 05797, each of which includes an antibiotic, a microorganism capable of producing an antibiotic, a processed product thereof, or a compound described in WO99 / 1289. In addition, these antibiotics, microorganisms capable of producing antibiotics or processed products thereof, or the above compounds can be used alone as an active ingredient of agricultural and horticultural drugs.

The agricultural or horticultural drug according to the present invention can use the above-mentioned active ingredient as it is, but usually, it contains various preparations such as powders, wettable powders, granules and emulsions together with auxiliary preparations. Use as At this time, one or more active ingredients are contained in the preparation in an amount of 0.1 to 95% by weight, preferably 0.5 to 90% by weight, more preferably 2 to 70% by weight.
% By weight. Examples of carriers, diluents and surfactants used as formulation auxiliaries include, as solid carriers, talc, kaolin, bentonite, diatomaceous earth, white carbon, clay, etc., and liquid diluents such as water, xylene, toluene, chlorobenzene, Cyclohexane, cyclohexanone, dimethylsulfoxide, dimethylformamide, N-methylpyrrolidone alcohol, etc. may be mentioned, and the surfactant may be properly used depending on its effect, and as an emulsifier, polyoxyethylene alkylaryl ether, polyoxyethylene sorbitan monolaurate Examples of the dispersant include lignin sulfonate and dibutylnaphthalene sulfonate, and examples of the lubricant include alkyl sulfonate and alkylphenyl sulfonate.

The above-mentioned preparations may be used as they are,
Some are used after being diluted to a predetermined concentration with a diluent such as water. When diluted and used, the concentration of the compound of the present invention is 0.00
A range of 1 to 1.0% is preferred. In addition, the amount of the compound of the present invention is determined in agricultural and horticultural land 1 such as fields, fields, orchards, greenhouses,
5 to 5000 g, preferably 5 to 500 g, per ha
More preferably, it is 50 to 500 g. The concentration and amount of use vary depending on the dosage form, period of use, method of use, place of use, target crop, and the like, and are not limited to the above ranges. Incidentally, the agricultural and horticultural drug in the present invention, other agricultural and horticultural agents, for example, fungicides, insecticides,
It can be used in combination with acaricides, herbicides and the like.

[0020]

The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to these examples. Example 1 One loop of a loop of a TKR2993 strain (FERM P-17365) was placed in a 100 ml liquid medium (Difco yeast nitrogen base 0.67% (w / v), glucose 2.0% (w / v).
v)) and inoculate a 500 ml Erlenmeyer flask
After shaking for 3 days, a seed culture was obtained. 1.0 ml of this seed culture was inoculated into five 500 ml Erlenmeyer flasks containing 140 ml of the above liquid medium, and subjected to shaking culture (shaking 220 rpm) at 25 ° C. for 10 days. The culture solution thus obtained was centrifuged to separate a supernatant and cells. 500 m of methanol in the obtained cells
After adding l, mixing well and performing an extraction operation, concentration under reduced pressure was performed. Water and butanol (300 ml each) were added to the obtained residue, mixed well, and a butanol extraction operation was performed.
Methanol was added to the residue obtained by concentrating this extract under reduced pressure and dissolved. Water and ethyl acetate (300 ml each) were added to the residue obtained by concentrating the methanol solution under reduced pressure, mixed well, and washed with ethyl acetate.
The aqueous layer was concentrated under reduced pressure to obtain 489 mg of a residue. This was dissolved in 6 ml of 50% methanol, applied to a YMC-GEL ODS-A 60-230 / 70 (YMC) column (100 ml) equilibrated with water, and washed with 50% methanol. %
The active fraction was obtained by eluting with methanol. This fraction was concentrated under reduced pressure to obtain 110 mg of a residue.
This was dissolved in methanol (0.8 ml) and subjected to high performance liquid chromatography to obtain an active fraction. The fraction was concentrated under reduced pressure to obtain 77 mg of a residue. The conditions for the high performance liquid chromatography were as follows. Apparatus: LC8A (manufactured by Shimadzu Corporation) Column: PEGASIL C8 (2.0 cm x 25 cm) (manufactured by Senshu Kagaku) Mobile phase: 45% (v / v) acetonitrile / water containing 0.05% trifluoroacetic acid 2.8 ml of methanol And subjected to high performance liquid chromatography to obtain an active fraction. This fraction was concentrated under reduced pressure to obtain 36.2 mg of a crudely purified TKR2993.
The conditions for the high performance liquid chromatography were as follows. Apparatus: LC8A (manufactured by Shimadzu Corporation) Column: PEGASIL C8 (2.0 cm x 25 cm) (manufactured by Senshu Kagaku) Mobile phase: 40% (v / v) acetonitrile / water containing 0.05% trifluoroacetic acid 1.2 ml of methanol And subjected to high performance liquid chromatography again to obtain an active fraction. The fraction was concentrated under reduced pressure to obtain 15.7 mg of a purified product of TKR2993 as a white powder. The conditions for the second high-performance liquid chromatography were as follows. Apparatus: LC8A (manufactured by Shimadzu Corporation) Column: PEGASIL C8 (2.0 cm x 25 cm) (manufactured by Senshu Kagaku) Mobile phase: 40% (v / v) acetonitrile / water containing 0.05% trifluoroacetic acid

Example 2 The physicochemical properties of TKR2993 obtained in Example 1 were measured as follows. For mass spectrometry, a JMS-DX302 type mass spectrometer (manufactured by JEOL Ltd.) was used. ¹ H-NMR spectrum (in deuterated methanol, standard substance: tetramethylsilane), and
^{A 13} C-NMR spectrum (in deuterated methanol, standard substance: deuterated methanol) was measured using a JNM-A500 nuclear magnetic resonance apparatus (manufactured by JEOL Ltd.). For UV absorption spectrum analysis (in methanol), a UV-250 type self-recording spectrophotometer (manufactured by Shimadzu Corporation) was used. Infrared absorption spectrum analysis (KBr method)
270-30 type infrared spectrophotometer (manufactured by Hitachi, Ltd.). (1) Mass spectrometry A purified white powder obtained by subjecting to a high performance liquid chromatography and concentrating the obtained active fraction under reduced pressure is as follows:
FAB-MS measurement by mass spectrometry revealed that the m / z was 1410 [M + H] ⁺ . (2) Carbon number and nitrogen number The purified white powder obtained by subjecting to high performance liquid chromatography and concentrating the obtained active fraction under reduced pressure is as follows:
^{According to 1} H-NMR spectrum measurement and ¹³ C-NMR spectrum measurement and analysis thereof, it was found that the compound had 72 carbon atoms and 14 nitrogen atoms.
It turned out to be. The ¹ H-NMR spectrum is shown in FIG.
FIG. 4 shows the ¹³ C-NMR spectrum.

(3) Ultraviolet absorption spectrum The purified white powder obtained by subjecting the active fraction obtained to high-performance liquid chromatography and concentrating under reduced pressure to
It was found that the ultraviolet absorption spectrum in methanol showed terminal absorption as shown in FIG. (4) Infrared absorption spectrum A white powder purified product obtained by subjecting to a high performance liquid chromatography and concentrating the obtained active fraction under reduced pressure,
The results of infrared spectrum measurement by the KBr method are as follows. IR (KBr) (cm ^-1 ): 3440, 2960, 1640, 1470, 1210, 8
40, 800, 720. FIG. 2 shows the infrared absorption spectrum. In addition, the solubility of this substance in various solvents was soluble in methanol and poorly soluble in hexane, chloroform and water. From the results of the above analysis, it was revealed that the purified product of white powder obtained by subjecting to a high performance liquid chromatography and concentrating the obtained active fraction under reduced pressure was TKR2993. TKR2993 above
Was subjected to analysis by reversed-phase partition high performance liquid chromatography (HPLC) using an LC-10A type high performance liquid chromatography apparatus (manufactured by Shimadzu Corporation). The conditions for the high performance liquid chromatography were as follows. Column: CAPCELL PAK _C18 (6 mm x 150 mm) (manufactured by Shiseido) Mobile phase: 50% (v / v) acetonitrile / water containing 0.05% trifluoroacetic acid Column temperature: 40 ° C Detection UV wavelength: 220 nm As a result, the above It was revealed that TKR2993 was eluted at the position shown in FIG.

Example 3 The biological properties of TKR2993 obtained in Example 1 were measured as follows. The antibacterial spectrum against various microorganisms was examined using the obtained TKR2993. In the measurement, the concentration at which the growth of the bacteria was almost completely inhibited was determined as the minimum growth inhibitory concentration (μg / ml) by the liquid medium dilution method. The results are shown in Table 2. In the table, YNBG represents a medium containing 0.67% of yeast nitrogen base (manufactured by Difco) and 1.0% of glucose.

[0024]

[Table 2] ____________________________________________________________________________ Test bacteria culture medium Minimum growth inhibitory concentration (μg / ml) __________________________________________________

From the results in Table 2, it can be seen that the antifungal substance TK of the present invention
R2993 was found to have antibacterial activity against pathogenic fungi such as Candida albicans. Also obtained
TKR2993 was intraperitoneally administered to ICR mice at 50 mg / kg, but no toxicity was observed.

[0026]

Industrial Applicability According to the present invention, there can be provided an antifungal substance TKR2993 which is useful as a preventive or therapeutic agent for fungal diseases of humans, animals and plants, and a method for producing the same.

[Brief description of the drawings]

FIG. 1 is a view showing an ultraviolet absorption spectrum of an antifungal substance TKR2993. The horizontal axis indicates the wavelength (nm).

FIG. 2 is a view showing an infrared absorption spectrum of an antifungal substance TKR2993. The horizontal axis indicates the wave number (cm ^-1 ).

FIG. 3 is a diagram showing a ¹ H-NMR spectrum of an antifungal substance TKR2993. The horizontal axis indicates the chemical shift value (ppm).

FIG. 4 is a view showing a ¹³ C-NMR spectrum of an antifungal substance TKR2993. The horizontal axis indicates the chemical shift value (ppm).

FIG. 5 is a diagram showing elution positions of the antifungal substance TKR2993 by HPLC. The vertical axis indicates the relative ultraviolet absorption intensity, and the horizontal axis indicates the retention time (minute).

 ──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Yoshimi Onishi 3-4-1, Seta, Otsu City, Shiga Prefecture Inside Takara Shuzo Co., Ltd. Central Research Laboratory (72) Inventor Ikunoyuki Kato 3-4, Seta, Otsu City, Shiga Prefecture No.1 Takara Shuzo Co., Ltd. Central Research Laboratory F-term (reference) 4B064 AH19 BA10 BE09 BG01 BG09 BH02 BH04 BH05 BH06 BH10 BH20 CA05 DA03 DA04 DA12 4B065 AA58X AC14 BA22 CA02 CA44 4C087 AA01 AA02 AA04 ZA35 CA22 ZC61 4H055 AA01 AA02 AA03 AB29 AC62 AD20 AD32 BA50 CA40 DA10 DA79 DA88 DA89 DA91

Claims (6)

[Claims] 1. The following (1), (2), (3), (4),
An antibiotic TKR2993 or a salt thereof having the physicochemical properties of (5) and (6): mass spectrometry: FAB-MS m / z 1410 [M + H] ⁺ ; (2) carbon number: 72; Nitrogen number: 14; (3) Ultraviolet absorption spectrum (in methanol): shows terminal absorption; (4) Infrared absorption spectrum (KBr method): The main absorption wave numbers (cm ^-1 ) are as follows: 3440, 2960, 1640, 1470, 12
10, 840, 800, 720; (5) Solubility: soluble in methanol. Poorly soluble in hexane, chloroform and water; (6) Amino acid analysis (detected by ninhydrin reaction):
Threonine, alanine, valine, leucine and proline are detected. 2. Cylindrocarpon
A method for producing an antibiotic TKR2993, comprising culturing a strain belonging to the genus and having the ability to produce the antibiotic TKR2993, and isolating the antibiotic TKR2993 from the culture. 3. Cylindrocarpon
A microorganism capable of producing the antibiotic TKR2993 belonging to the genus. 4. Cylind carpone sp.
rocarpon sp.) TKR2993 (FERM P-17365). 5. A processed product of the microorganism according to claim 3 or 4. 6. An antibiotic TKR2293 according to claim 1 and a salt thereof, a microorganism according to claims 3 to 4, and at least one active ingredient selected from the group consisting of a treated product according to claim 5. An antifungal agent characterized by the following.