JPS6045640B2 - Houteimycin KO↓1 and its manufacturing method - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to horteimycin and its non-toxic acid addition salts and processes for their production.

Horteimycin 01 is a compound that has strong antibacterial activity against a wide range of Gram-positive and Gram-negative bacteria, and is a new compound that is expected to be used as an antibiotic.

For horteimycin compounds, horteimycin A (JP-A-50-29789), horteimycin B
(Japanese Unexamined Patent Publication No. 49-126892) and Horteimycin C (Japanese Unexamined Patent Publication No. 52-18888) are known to be produced by microorganisms belonging to the genus Micromonospora.

In addition, Horteimycin D (Japanese Patent Publication No. 58-34119), Horteimycin KE (Japanese Patent Publication No. 53-9024)
Horteimycin KF, Horteimycin KG (Japanese Patent Publication No. 54-6667), and Horteimycin KO (Japanese Patent Publication No. 58-5263) are known.

The present inventors have completed the present invention by discovering that microorganisms belonging to the genus Micromonos and Pora produce a new compound having antibacterial activity, which has been named Horteimycin-0. The present invention will be explained in detail below. Horteimycin KO (free base) of the present invention has the following physical and chemical properties.

1) Properties: Basic white powder (2) High resolution mass spectrum 349.2455 (M+1)+ (Cl5H33N
4O,)348.2352(M (Cl5H
3.

N4O5) 331.2115 (Cl5H29N3O5)
286.1791 (Cl3H24N3O4) 235
．． 1295 (C9Hl4N2O5) 207.1342
(C8Hl~204) 189.1229 (C8Hl7
N2O3) 143.1190 (C, Hl, N2O) As a result, the molecular weight of Horteimycin KOl free base is 34
F! 8 The molecular formula is Cl5H3.

N4O5 is given. (3) Melting point: 77-79CC (
4) Ultraviolet absorption spectrum: 220-360 for aqueous solution
It does not show a characteristic absorption maximum at r1m, but only shows terminal absorption.

(5) Specific rotation [α] w = +79 (C = 0.3, H2
0) (6) Infrared absorption Svector 2 The results measured by the KBr tablet method are shown in Figure 1.

The wave number (d-1) showing absorption maximum is 3370, 3300,
2940, 1590, 1455, 1375, 1088,
1020,930,910,660,565. (
7) Elemental analysis value C: 51.77% H: 9
．． 24% N: 16.10% (8) Carbon nuclear magnetic resonance svectometry was performed using JEOL
, JNM-FXlOO was used in a heavy water solution (PD=-10.
I went with 4).

The results are shown in FIG.

Signal assignments are as follows: 102.0 (C-'), 83.0 (C-6), 82.6
(C-3), 74.0 (C-5″), 73.6 (C-5″)
), 72.6 (C-.2), 67.4 (dioxane, internal standard substance), 59.9 (C-4), 57.8 (0CH
3), 55.9 (C-1), 51.0 (C-6''), 5
0.2 (C-7), 38.4 (NCH3), 27.3 (
C-4''), 26.6 (C-3''), 17.7 (6''-
CH3) (9) Color reaction: ninhydrin reaction
Positive. Potassium permanganate reaction positive
Elson Morgan reaction negative
Biuret reaction Negative QQ From the above physical and chemical properties, the steric structural formula of Horteimycin KOL is estimated as follows.

(11) Solubility in solvents: extremely soluble in water,
It is soluble in methanol and slightly soluble in ethanol and acetone, but is insoluble in organic solvents such as chloroform, benzene, ethyl acetate, butyl acetate, ether, butanol, petroleum ether, and n-hexane. (12) Add a developing agent to chloroform, methanol, 28% ammonia water (
2:1:1) (volume ratio; all solvents hereinafter indicate volume ratios) ascending vapor chromatography using the lower layer [Whatman NO. l(w&RBaIstOnLt
d) Use, room temperature, development time 4 hours] Rf values are shown in Table 1. Rf values of antibiotics considered to be close to Horteimycin KOL are also listed for comparison. Table 1 Antibiotic name Rf value Fortimjcin A (FOrtimjcin A) 0.41
FORTIMICIN B
0.72 Horteimycin C (FOrt
imicnC) 0.22 Fortimicin D
0.22 Follmic
inKE) 0.63 antibiotic name
Rf value Horteimycin Xiang (F
OrtimicinKF) 0.4
9Fortimicin KG
0.67 Horteimycin KO (
FORTIMICIN KO) 0.
53 FORTIMICIN KOL
Table 2 shows the Rf values of silica gel thin layer chromatography [Merc K&CO, hanging (the same applies hereinafter) room temperature, development time 3 hours].

Other hortimycins are also listed for comparison.
Table 2 Rf value Antibiotic name Developer 1* Developer ■※※Houteimycin AO. 36O. O9 Hortimycin BO. 7
5O. l3 Horteimycin CO. 23O. 2O Horteimycin DO. 2lO. O6 Hotei Maishi＞'KEO. 5OO. l2 Horteimycin KF' 0.350.12 Horteimycin KGO
．． 55O. l2 hortimycin KOO. 55O. 24 Horteimycin KOIO. 74O. l5*Developing agent 1
:Chloroform, methanol, 28% ammonia water (2
:1:1) Lower layer※※Developing agent H: 10% aqueous ammonium acetate solution, methanol, 28% aqueous ammonia (50:1)
50:1) Next, Table 3 shows the antibacterial spectrum of Horteimycin KOL against various microorganisms.

In Table 3, MIC is PH8. It was measured by the agar dilution method using Hanten medium. Table 3 Test microorganism MIC (Pg/ML) Bacillus●
Subtilis No. lO7O7 (KY4273) 0.40
Staphylococcus Aureus ATCC6538PO
．． O9 Klebsiella Newmoya ATCClOO3lO. 4O Test microorganism ■C (μg/ml) E. coli ATCC26O. 4O Etsushierihia●KoriKY83O2O. 4O (chloramphenicol, streptomycanicin, paromomycin, tetracycline, spectinomycin resistant) E. coli KY8327O. 35 (kanamycin dientamycin, tobramycin resistant) E. coli KY8348O. l8 (streptomycin, dieticin resistant) Proteus vulgaris ATCC6897O. 35 Shigella zonei ATCC929OO. 4O Salmonella●
Taihosa ATCC9992O. 2O Pseudomonas Aeruginosa BMH #15.3 As described above, Horteimycin KOL has strong antibacterial activity against a wide range of Gram-positive and Gram-negative bacteria.

Among its wide-ranging and powerful antibacterial properties, a particularly distinctive feature is that it is effective against certain water coli bacteria that are resistant to kanamycin, dientamicin, tobramycin, and the like. In addition, this hortimycin KOl can be used to treat various infectious diseases (in humans and animals) caused by the various bacteria mentioned above.
It has also been found that it has an extremely excellent therapeutic effect on. The extremely excellent antibacterial properties of Horteimycin KO1 indicate that this substance can be used for medical purposes, especially as an antibacterial antibiotic. Next, let's compare this substance with known antibiotics.

As a water-soluble basic antibacterial substance with a broad antibacterial spectrum produced by Micromonospora, dientamicin Complex (M●-J●Weinste
in et al.: ArltimicrObialAgentsA
ndChemOtherapy, 1963, 1 and D.
J. COOper et al., J. Infect. Dis. Hit,
342, 1969, J. A. WaitZ et al., AJlti
microOb. Ag. ChemOth. E1464,1
972), Antibiotic No. 46O (Tokuko Sho 4)
6-16153), Sisomicin (M.J.We
Instein et al., J. AntiblOtics? 55
1,555,559,1970), Antibiotics include Mycin KG and Horteimycin KO. Among these, Ho, Teimycin A,
Directly comparing B, C, D, KE, KF', KG and KO with KOl, as shown in Tables 1 and 2, K
Ol is vapor 0-chromatography [developing agent, chloroform, methanol, 28% aqueous ammonia (2:1:
1)] and silica gel thin layer chromatography [lower layer of developer 11: chloroform, methanol, 28% aqueous ammonia (2:1:1)] showed an Rf value close to that of Horteimycin B. In silica gel thin layer chromatography [developing solvent 1, 10% ammonium acetate aqueous solution, methanol, 28% aqueous ammonia (50:50:1)], it shows an Rf value close to that of Horteimycin B (Rf=0.13). However, in silica gel thin layer chromatography [lower layer of developing solvent: chloroform, methanol, 28% aqueous ammonia (7:2:2]), horteimycin KOL
is RfO. 4l, Horteimycin B (Rf=0
．． 49). Horteimycin KOL is also clearly distinguished from Horteimycin B in that it exhibits much stronger antibacterial activity. Horteimycin KOl has the exact same molecular formula as Horteimycin B, Cl5H.
3. It shows N4O5 and is represented by the same planar structural formula, but is thought to be a substance with a different three-dimensional structure of the fotamine moiety. In addition, dientamicins, sisomicin, XK
It differs from -62-2 in its physicochemical properties. Furthermore, water-soluble basic antibiotics with a broad antibacterial spectrum produced by actinomycetes other than Micromonospora include streptomycin, ribostamycin, lividomycin, spectinomycin, kasugamycin, neomycin, kanamycin, nebramycin, and paromomycin. However, it is clear that horteimycin KOI is different from any of these antibiotics in terms of physicochemical properties and vapor chromatography Rf values shown in Table 1. From the above, Horteimycin KOL is considered to be a new antibiotic.

Next, an example of a method for producing the compound hortimycin KOL of the present invention will be shown.

A microorganism belonging to the genus Micromonospora and having the ability to produce horteimycin KOl is cultured in a medium, horteimycin KOl is accumulated in the culture, and the horteimycin KOl is accumulated in the culture. Horteimycin KOL is collected from the feed.

As the microorganism used, any strain can be used as long as it belongs to the genus Micromonospora and can produce and accumulate Horteimycin KOL in the medium.
Specific preferred examples include Micromonospora Olivoasterosbora MK-70 (Feikoken Bacterial Report No. 15 (1))
(ATCC2l8l9), Micromonosvora Olivo Asterospora MK8O (Feikoken Bacteria Collection No. 2194) (AT
CC3lOlO), Micromonospora ●Olibo Asterospora Mm744 (Feikoken Bibori No. 2193) (ATC
C3lOO9), etc. The mycological properties of these strains are specified in JP-A-50-297. As the medium, a medium commonly used for culturing actinomycetes is used.

As the carbon source, glucose, starch, mannose, fructose, sucrose, molasses, etc. are used alone or in combination.

Furthermore, depending on the ability of bacteria to assimilate hydrocarbons, alcohols, organic acids, etc., may be used. Examples of inorganic and organic nitrogen sources include ammonium chloride, ammonium sulfate, urea, ammonium nitrate, and sodium nitrate. As such, peptone, meat factory scratches, yeast factory scratches, dry yeast, corn stew liquor, soybean flour, casamino acids, solvable vegetable double protein, etc. are used alone or in combination.In addition, salt, In addition to adding inorganic salts such as potassium chloride, calcium carbonate, and phosphates, organic or inorganic substances that promote the growth of the strain used and the production of Horteimycin KOL can be appropriately added.As a culture method, liquid culture method is used. The deep agitation culture method is particularly suitable.

It is desirable that the culture be carried out at a culture temperature of 25 to 40°C and a pH near neutral. When culture is carried out in liquid culture for usually 2 to 15 days, horteimycin KOL is produced and accumulated in the culture solution. When the production amount of the target substance in the culture solution reaches the maximum, the culture is stopped, and the target substance is purified and isolated from the culture solution. Purification and isolation of Horteimycin KOL is carried out as follows.

For purification and isolation of Horteimycin KOL from culture furnace solution,
Conventional separation and purification methods are utilized to isolate microbial metabolic products from their culture fluids.

Since Horteimycin KOL is a basic substance that is well soluble in water but not well soluble in general organic solvents, it is purified by a method that is often used for the purification of so-called water-soluble basic antibiotics.

Namely, adsorption/desorption method using cation exchange resin, cellulose column chromatography, Sephadex LH
An appropriate combination of methods such as adsorption/desorption using 1-20 columns and silica gel column chromatography can be used. Next, one example of a specific purification method for hortimycin KOL will be shown.

That is, after adjusting the culture pH to 7.5, the cation exchange resin Amperlite 1RC-50 (ammonia type) (R
Ohm&HaasCO. (manufactured by), and after washing with water, 0.
Elute with 5N aqueous ammonia. After collecting the active fractions and concentrating them under reduced pressure, the concentrated liquid was transferred to the cation exchange resin Amperlite CG-
50 (ammonia type) (manufactured by ROhm & Haas CO.)
After adsorption and washing with water, 0.1MNF [0.1 containing 4CI
Elute with N ammonia water. At this time, hortimycin KO and hortimycin C are eluted first, and then hortimycin KO1 is eluted together with hortimycin B.

After that, Horteimycin A, D, KE, KF, and KG are eluted. After collecting the fractions containing Horteimycin KOL and neutralizing them with sulfuric acid, they were added to Amperlite IRC-50 (
After washing with water, the active fraction was eluted with 0.5N ammonia water and concentrated to dryness to obtain a crude powder containing hortimycin KOL. This crude powder was placed on a silica gel column and developed and eluted with a lower layer of chloroform, methanol, and 28% aqueous ammonia (5:1:1). In this case, the mixed hortimycin B is eluted first, and then the hortimycin KOI is eluted. Fractions containing hortimycin KOL are collected, concentrated under reduced pressure, and freeze-dried to obtain white free base of hortimycin KO1. In this way, silica gel column chromatography can yield fairly clean Horteimycin KO, but since some impurities are mixed in, Biorex-70 (ammonia type) can be obtained.
(BiORadLalx) manufactured by RtOries), and after washing with water, 0.0% containing 0.04M ammonium acetate
Elute with 4M aqueous ammonia. Horteimycin KO,
The fractions containing Amperlite CG-50 (
After washing with water, elution with 0.5N ammonia water, collecting and concentrating the active fraction, and freeze-drying the pure sample (free base) of Horteimycin KOL can be obtained. can. Column chromatography using carboxymethyl cellulose is also effective for removing mixed ninhydrin-positive substances. That is, in this case, a solution of coarse powder is passed through a column filled with carboxymethyl cellulose (ammonia type) to adsorb the active substance. After that, it was thoroughly washed with water to elute most of the pigments and inorganic salts, and then 0.211!4-citric acid.
Phosphate buffer PH3. If it is eluted with O), the active substance will be eluted with b. Purified hortimycin KOl can be obtained by collecting and lyophilizing fractions containing hortimycin KOl. The trend of the active fraction of hortimycin KOL during the above purification process is checked by silica gel thin layer chromatography 1-.

As a developing solvent, chloroform, methanol, 28%
The lower layer of aqueous ammonia (3.5:1:1), 10% aqueous ammonium acetate, methanol, and 28% aqueous ammonia (50:50:1) was used for detection, and the detection was performed using ninhydrin color method and Pulse subtilis NO. The bacteria were isolated using bioautography using lOO7 as a test bacterium. The non-toxic acid addition salt of Horteimycin KOll is Horteimycin KOll.
It refers to mono-, di-, tri-, or tetra-salts produced by the reaction of the molecule with 1 to 4 equivalents of a pharmacologically acceptable non-toxic acid. Acids include hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, carbonic acid, inorganic acids such as nitric acid, acetic acid, fumaric acid, malic acid, citric acid, mandelic acid, ascorbic acid, tartaric acid, succinic acid, etc. of organic acids. Hereinafter, the production of the compounds of the present invention will be shown in Examples, but these are merely illustrative and do not limit the present invention in any way.

Example 1) A. Culture of MK Ichijo strain Micromonospora oliboasterospora (MiCrOmOrK) SPOraOIivOaste as the inoculum
rOspOra)K. K-70 (ATCC2l8l9)
(Feikoken Bibori No. 1560), glucose 2g/d1, peptone 0.5g/DIl, yeast scratch 0.5g/d1, and calcium carbonate 0.1y/d1 as the first type medium.
1 (pH 7.5 before sterilization) was used.

Place one platinum loop of inoculum in a 50mL thick test tube at 10T! The above medium of Ll is inoculated and cultured at 30°C for 5 days. 10 ml of this seed culture solution is inoculated into a 30 mt second seed medium placed in a 250 ml Erlenmeyer flask. The composition of the second type medium is the same as that of the first type medium. Type 2 culture is 3
Culture with shaking at 0°C for 2 days. 2 30ml of this seed culture
30 in an Erlenmeyer flask with 1 baffle
Inoculate 0ml of third type medium. The composition of the third type medium is the same as that of the first type medium. Type 3 culture is 2 at 30℃.
Incubate with shaking for 1 day. This third type medium 1.51 (for 5 flasks) is inoculated into the fourth type medium 151 in a 301 capacity stainless steel jar fermenter. The composition of the fourth type medium is the same as the composition of the first type medium.

This jar fermentor culture was carried out at 3 rC for 2 days using an aeration agitation method (rotation speed 350 r'Pm, aeration rate 151/M).
in). 300% of this fourth type culture solution 151
] Inoculate the fermentation medium 1501 in a fermenter with a capacity of 1. Fermentation medium composition: Soluble starch 4g/d1, soybean meal 2g/Dll cornstarch liquor 1y/D
l. ．． K2HPO4O. O5g/d1, MgSO47H
2OO. O5g/d1, KClO. O3g/Dl,. C
aCO3O. A medium of lg/d1 (pH 7.5 before sterilization) was used. This fermenter culture was carried out at 37℃ for 4 days using an aeration agitation method (rotation speed 150 r'Pm, aeration rate 801/Mi).
n). Isolation of B-Hoteimycin KOL ●After the purification culture, the pH of the culture solution was adjusted to P by adding concentrated sulfuric acid.
Adjust to H25 and stir 3 powders.

Thereafter, approximately 7 kg of Radiolite #600 (manufactured by Showa Kagaku Kogyo) was added as a purifying agent to separate the bacterial cells. To liquid? -Adjust the pH to 7.5 by adding sodium hydroxide. This Oki liquid is used as cation exchange resin Amperlite IRC.
Pass through a column packed with -50 (ammonia type) approx. 201, and discard the effluent. Since the active substance is adsorbed on the resin, after washing the resin with water, the adsorbed active substance is eluted with 0.5N aqueous ammonia. Bacillus Zuptilis NQ. The eluate was measured by the vapor disk method using a 1O7O7 agar plate. Measure activity. The active fractions are combined and concentrated under reduced pressure to ca. This concentrated solution was adjusted to pH 7 with concentrated hydrochloric acid, and then passed through a column of Amperlite CG-50 (ammonia type) 31.

The active substance is adsorbed, so after washing with water, 20.1MN11
Elute with 0.1N NH40H containing 4C1. The eluate was fractionated into 500 ml portions, and the components in each fraction were separated from Bacillus.
Subtilis No. Antibacterial activity against 7O7O7 (KY4273), silica gel thin layer chromatography [Developer: chloroform-methanol-ammonia water (2:1:
1)] and ninhydrin coloring. Horteimycin KOL is eluted at about 301 points. Fractions containing hortimycin KOL were collected, ammonia was removed by vacuum concentration, and then neutralized and passed through a column containing 500 nt of Amperlite IRC-50 (ammonia type), washed with water and eluted with 0.5N aqueous ammonia. After concentration under reduced pressure, the residue was lyophilized to obtain about 1 kg of white powder. Approximately 400J in this powder! It included 9 Horteimycin KOs. C. Preparation of pure hortimycin KOl The crude hortimycin KOl powder obtained in the above step B was placed on a 1.51 silica gel column (silica gel, Wakogel C-200 manufactured by Fuwako Pure Chemical Industries, Ltd., column diameter approximately 5C7). Chloroform, methanol, aqueous ammonia (5:1:
1) Develop and elute in the lower layer.

The eluate was fractionated into 20ml portions, and the components in each fraction were examined by silica gel thin layer chromatography. Horteimycin B was eluted first, followed by Horteimycin KOL.
is eluted. Fractions of Horteimycin KOL were collected, concentrated under reduced pressure, and lyophilized to form Horteimycin Kol.
36Om9 was obtained. The activity was 930 units. (Activity is defined as 1000 units per 1 mg of pure product) 360 mg of Horteimycin KO1 obtained was dissolved in water, the pH was adjusted to 7, and the mixture was adsorbed on 50 ml of Biorex 70 (ammonium type). After washing with water, 0.04 M acetic acid was added. 0. Contains ammonium.
Elute with 04N ammonia water.

The eluate was fractionated into 10 ml portions, and each fraction was subjected to silica gel thin layer chromatography.
Fractions containing only Ol were collected, ammonia was removed by vacuum concentration, and the pH was adjusted to 7.
Adsorb onto 100ml of C-50 (ammonia type), wash with water, and elute with 0.5N ammonia water. The active fractions were collected, concentrated under reduced pressure, and finally lyophilized to yield 330m9 of pure Horteimycin KOL free base as a white powder. Example 2 A culture solution obtained by culturing in the same manner as in Section A of Example 1 is treated in the same manner as in Section B of Example 1.

500 mg of the obtained crude powder is dissolved in 5 ml of water and passed through a column packed with about 200 ml of carboxymethyl cellulose (ammonia type). When approximately 1000 ml of water is subsequently flushed, the active ingredient is adsorbed onto the carboxymethylcellulose, and most of the dyes and inorganic salts that are not adsorbed onto the carboxymethylcellulose are washed out. Then 0. ? )
A-Citrate/Phosphate Buffer Swamp H3. Elute the column with O) (flow rate approximately 50 mL/hr). The eluted solution is collected in 10 ml fractions, and the activity of each fraction is assayed by the vapor disk method. The active fractions are subjected to silica gel thin layer chromatography, and fractions containing components corresponding to hortimycin KOL are collected. This fraction is passed through 50 mL of Amperlite IRC-50 (ammonium type) to adsorb the active ingredient. After washing with water, elution is performed with 0.5N aqueous ammonia, and the active fraction is lyophilized to obtain about 100 mg of horteimycin KOL free base.
The activity was 98 positions/Mg. Example 3 Micromonospora as inoculum Olivo Asterospora M
m744, KYllO67 strain (Feikoken Bibori No. 2193, ATCC3lOO Crocodile) was used, and the first type medium was glucose 2g/Dll peptone 0.5g/Dtl yeast engineering scratch 0.3g/Dll calcium carbonate 0.1g/DL. (
A medium with a pH of 7.2) before sterilization was used.

Seed culture (first type culture ~
Type 4 culture) was performed. The compositions of the second type medium to the fourth type medium are all the same as the composition of the first type medium. The thus obtained type 4 culture solution 151 is inoculated into the main fermentation medium 1501 in a 3001 volume fermenter made of stainless steel. The main fermentation medium composition is soluble starch 2g/Dll, soybean meal 0.5g/Dll glucose 2g/Dll
Corn staple liquor 1g/DLl yeast factory scratches 1g/
DlK2HPO4O. O5g/DlMgSO,7H2O
O. O5g/DlKClO. O3g/DlCaCO3O
．． A medium of lg/DL (pH 7.0 before sterilization) is used.

This main fermentation culture was carried out at 30'C for 4 days using an aeration stirring method (rotation speed: 150 r''Pm, aeration rate: 801/min).The main fermentation liquid thus obtained was subjected to the method shown in Example 1, Section B. Therefore, when the crude hortimycin KOl powder was isolated, about 1 g of crude hortimycin KOl with an activity of about 22 positions/M9 was obtained.
When purification was carried out in the same manner as shown in section 1, horteimycin KO1 free base with an activity of about 98 molecules/Mg was purified.
60m9 was obtained. Example 4 Micromonospora oliboasterospora MK8OlKYllO55 as inoculum
strain (Feikoken Bibori No. 2192, ATCC 31O■), the first type medium was glucose 1g/Dll soluble starch 1g/DL, yeast scratch 0.5g/Dll peptone 0.5g/Dll calcium carbonate. A medium of 0.1 g/dl (pH 7.0 before sterilization) was used.

Seed culture (first type culture ~
Type 4 culture) was performed. The compositions of the second type medium to the fourth type medium were all the same as the composition of the first type medium of this example. The thus obtained type 4 culture solution was subjected to main fermentation culture in the same manner as shown in Example 3. The main fermentation medium composition was the same as the main fermentation medium shown in Example 3. The main fermentation culture solution thus obtained was used in Example 1 and B.
Horteimycin KOl was prepared by the same method as described in Section
Isolation of the crude powder yielded approximately 900 mg of crude Horteimycin KOL with an activity of approximately 28 bullets/M9. This crude horteimycin KOL powder was purified in the same manner as shown in Example 1, Section C, to obtain a purified sample of about 200 m9 of horteimycin KO1 free base. The activity of this purified hortimycin KOL was approximately 98 molecules/7n9. j Example 5 Horteimycin KO
Dissolve 1 g of free base in a small amount of water and ? After adjusting the pH to 4.5 using monosulfuric acid, about 1 volume of acetone is added to precipitate horteimycin KOL sulfate.

The precipitate was collected in a centrifuge and dried to obtain about 1.5 g of white powder of Horteimycin KOL sulfate. The activity was 60 bullets/M9.

[Brief explanation of the drawing]

FIG. 1 shows the infrared absorption 5 spectrum of Horteimycin KOl, and FIG. 2 shows the carbon nuclear nuclear magnetic resonance spectrum of Horteimycin KOl.

Claims (1)

[Claims] 1 Horteimycin KO_1 and its non-toxic acid addition salt represented by the formula ▲ Numerical formula, chemical formula, table, etc. ▼. 2 Belongs to the genus Micromonospora, Horteimycin KO
A method for producing horteimycin KO_1, which comprises culturing a microorganism capable of producing _1 in a medium, accumulating horteimycin KO_1 in the culture, and collecting horteimycin KO_1 from the culture.