JPS62174099A - Novel glycopeptide antibiotic substance pa-42867-a and pa-42867-b and production thereof - Google Patents

Abstract

NEW MATERIAL:The compound of formula (X is CH3 and Y is NH2 or X is H and Y is OH) having the following physical and chemical properties (as a compound wherein X is CH3 and Y is NH2). Appearance, white amorphous powder of ampholytic substance; [alpha]D<25> deg., -88.8+ or -3.1 deg. (C=0.41, water); mass spectrum analysis, MS(mz) 1557 (M+Z)<+>; elemental analysis (%), C 54.08, H 5.97, N 8.64, Cl 2.19; color reaction, positive to ninhydrin reaction; solubility, soluble in water and dimethyl sulfoxide, hardly soluble in alcohol, insoluble in ether, benzene, chloroform and ethyl acetate, etc. USE:An antibacterial agent exhibiting strong antibacterial activity against Gram- positive bacteria, especially methicillin-resistant bacteria and separable in high purity. In can be administered in the form of oral drug, injection, etc. PREPARATION:A microbial strain capable of producing PA-42867-A or -B [e.g. Nocardia orientalis PA-42867 (FERM P-8601)] is cultured at 28-32 deg.C for 60-100 hr preferably by deep-tank cultivation.

Description

DETAILED DESCRIPTION OF THE INVENTION A. Field of Industrial Application The present invention relates to a compound represented by formula 1; (wherein, X represents CH and Y represents NHt, or X represents H and Y represents OH. Antibiotics PA-42867-A and PA
-42867-B and their pharmaceutically acceptable salts, their production methods, and PA-42867-A and PA-
42867-B producing microorganism.

BACKGROUND OF THE INVENTION With the recent widespread use of antibiotics, the emergence of multidrug-resistant bacteria, particularly methicillin-resistant bacteria, has become a problem. Methicillin-resistant bacteria are not only resistant to methicillin, but also to many other antibiotics such as aminoglycoside antibiotics, tetracycline antibiotics, cephem antibiotics, penicillin antibiotics, carbapenam antibiotics, and macrolide antibiotics. It is resistant to many antibiotics.

It has been noticed that glycopeptide antibiotics, especially vancomycin antibiotics, are active against methicillin-resistant bacteria (ANTIMICROBIAL AGENT AND CHEMOTHERAPY).
AGENr AND CHEMOT) IERAPY)
28.660-662 (1985)). Vancomycin is an antibiotic that has been known for a long time.
450), and new vancomycin analogs have also been discovered (Antimicrobial Agents and Chemotherapy (ANrIMICROBIA)).
LAGEN'I AND CHEMOTHERAPY
) 28.660-662 (1985), The Journal Antibiotics fl) [EJOUR
NAL OF ANTIBIOTIC5) 37.44
6-453 (1984), Japan, 1-8 (1985
), 51-57 (1985), JP-A-60-1
39623.60-199397.60-231698
．． 60-237099, etc.). However, the antibiotics PA-42867-A and PA-42867-B of the present invention
is a vancomycin antibiotic with a completely new structure that does not match any of the above compounds.

C1 Problems to be solved by the invention Vancomycin currently on the market is for oral use and has low purity.
It is difficult to use it directly as an injection. Also,
There has been a long-awaited expectation for an antibiotic that has stronger activity against methicillin-resistant bacteria than conventional vancomycin antibiotics.

Means for Solving the 20 Problems The present inventors have determined that a bacterial strain belonging to the genus Nocardia has the formula: It was discovered that a compound represented by (Y represents OH), which has strong activity against methicillin-resistant bacteria, was produced.
and a compound in which Y is NHt PA-42867-A
, a compound in which X is H and Y is OH is referred to as PA-4286
It was named 7-B. The present invention encompasses not only these compounds, but also their pharmaceutically acceptable salts.

The physicochemical properties of the compound of the present invention are shown below.

Physicochemical properties■PA-42867-A Ultraviolet absorption spectrum included 0°01NHc1nm (E+:In)' 281
．． 8 (41,15)max input "lNN80H"qnm(E'): 302.6
(43,14>max 1cm Specific optical rotation [α 25°: -88,8±3.1° (c=0.
41, water) Infrared absorption spectrum (see Figure 1) IR (KBr): 3396.1654.1588 (s
h), 1505.1420.1396.1223.1
131.1064.101010(sh)” Mass spectrometry (
(See Figure 2) M S m/z: 1557 (M+H)"Elemental analysis Cy srs sow sN+.Theoretical value as Hlo for C1 and 3)'C; 54.08. H: 5.97.
Ni8.64. C112,19 Experimental value (%)'C;5
4.03. Ni6.14. N; 8.60. C11
2,19 nuclear magnetic resonance spectrum 'H-NMR: 400MHz, ds-DMSO+1
drop D, O, internal standard TMS, 100°C (3rd
(See figure) δppm: 0.888 (d, 6.5.31), 0.9
17 (d, 6° 0.3) 1), 1.093 (d, 6.0
．． 31(>, 1.156(s, 3H), 1.167(d
, 6.2.3H), 1.202(s, 3H), 1.42
Near 5 (m, IH), near 1525 (m, IH), 1.
630 (dd, 13.6.4.3, IH), 1.6
40 (dd, 136.4.3, IH), 1.766 (m
, IH), 1.866 (d-sama, 13.6, IH), 1.
915 (Mr. d, 13.6, IH), 2, 175 (dd
, 16.0.7.5.1) 1), 2.315(s , 3
H), 2.560 (dd, 16.0.5.0, IH),
2.892 (d, 9.5, IH), 2.947 (d, 9
．． 6.11 (), around 3,03 (overlapping with water in the solvent, IH), around 3.31 (m, 2H), 3.428 (like, around 8.7, IH), 3.540 ( dd, 11
．． 6.4.8, LH>, 3.609 (qd, 6.2
．． 9.5, IH), 3.688 (dd, 7.2.8.7
．． IH), 3.730 (dd, 11.6.2.1, IH
), 4.141(qd, 6.0.9.6.11(), 4
．． 233 (broad s, IH), 4.328 (d
d, 5.0.7,5, LM), 4.485 (Mr. s, I
H), 4.516 (s, IH), 4.674 (d-sama, 4
．． Near 3, IH), 4.699 (d, 40, IH), 5
．． 152 (d-sama, around 4.0, IH), 5.188 (b
road s-sama, IH), 5.190 (s-sama, IH),
5.305 (Mr. d, near 4.3, IH), 5.603 (
d, 7.2, LH>, 5.638 (br.

ad s, IH), 5.762 (broad s,
IH), 6, 375 (d, 23, IH), 6.388 (
d, 2.3.1) 1), 6, 712 (d, 8.3.1)
ri, 6.793 (dd, 8, 3.2.2, LH), 7.
087 (dd, 8.5.2.2, IH), 7.120 (
dd, 8.3.2.2, [), 7.148(d, 2.2
．． 11 (>, 7°215 (d, 8.4, IH), 7.2
86 (broad dd, around 8°3, around 2.1,
IH), 7.327 (dd, 8.4.2°0.1) 1)
, 7.586 (broad dd, near 8.5, near 2, IH), 7.863 (d, 2.0, IH)"C-N
MR' 50MHz, in DtO, external standard C) lsc
Nl, 7ppm, 60℃ (see Figure 4) δppm: 178.9.177.8.174.8.1
72.1°171.9.171.7.170.3.16
7.8.158.0°156.3. 155.9. 1
55.85. 155.85゜153.85,151,
138.8, 137, 136.85°136.7, 13
4.9. 133.9. 131. 130°128.
8. 128.4. 127.8. 127.1°12
6.2. 124.6. 124.2. 123.9.
122.8119.4. 118.6. 107.4
．． 107.3. 104.9104, 102.3.
99.2. 94. 80.7. 77.577.5
．． 76.9. 76.8. 75.2. 72.5°70.4. 67.4. 67.2. 63.4. 6
3.4°61.7. 60.5. 59.9. 56.
1. 55.5°55.5. 54.9. 52.7.
42. 41.4. 39.85°37.6. 34
．． 9. 25.2. 23.3. 22.5°20.6
．． 19.9. 18.4. 18.1 Thin Layer Chromatography Merck Precoated ILC Plate Silica Gel 60
F254 Developing solvent chloroform: methanol: concentrated aqueous ammonia: secondary butanol: water (5:10:5:5:2) Rf=0.28 High performance liquid chromatography (Takashi LC-6A) column: Chemcovac Nucleosil 5C18φ4. 6 x
150 mm (Chemco) Detection: UV 220 n
m Flow rate: 1 ml/min.

Operation: 8 acetonitrile-0.05M phosphate buffer (pH 3.5) Retention time: 8.8 min.

:9z acetonitrile 0.05M phosphate buffer (
pH 3,5> Retention time: 5.6 min.

Color reaction: ninhydrin positive Solubility; Soluble in water, dimethyl sulfoxide, slightly soluble in alcohol, insoluble in ether, benzene, dichloroform, ethyl acetate Biampholyte, white amorphous powder PA-42867-B Ultraviolet Absorption spectrum λ0. OINHC1nm (Ej:ITl), 281.
6 (44,22)max λ”IN”OH”qnm(E'): 301 (45
,94) max ICm specific optical rotation [α]2": -97,3±3.3" (c=0.4
1. Water) Infrared absorption spectrum (see Figure 5) IR (KBr): 3360.1656.1587 (s
h), 1505.1421.1393.1230.1
129.1062.11013(sh)C” mass spectrometry (
(See Figure 6) M S m/z: 1544 (M+)I)"Elemental analysis Ct J* 102 ?Theoretical value as H and O in NIC1.1 (%): C; 55.01. H; 5.71 .N:
8.02. C1; 2.26 Experimental value C%>: C: 54.
83. H;5.81. N; 8.46. C1;2.
13 Nuclear magnetic resonance spectrum 'H-NMR: 400M) lz, ds-DMSO+
Internal standard IMS in 1 drop DtO, 100°C (see Figure 7) Sppm: 0.891 (d, 66.3H), 0.91
9 (d, 6°-1l = 5.3H), 1.080 (d, 6.2.3H),
1.159 (s, 3H), 1.173 (d, 6.0.
3H), around 1428 (m, IH), L 469 (m,
LH), around 1.525 (m, IH), 1,647 (d
d, 13.7.4.4, LH), 1.767 (m, LH
), 1.920 (d-sama, 13.7.1) 1), 2.0
42 (Mr. dd, around 130, around 5.3, IH), 2.
182 (dd, 16.0.7.5, IH), 2.314
(s, 3H), 2.565 (dd, 16.0.4.8.
1) 1), 2.817 (Sama, near 9.1, IH), 2
．． 895 (d, 9.7, LH), around 3.03 (overlapping with water in the solvent, IH), around 3.33 (m, 2H),
3.497 (like, around 8.5, IH), 3.547 (
dd, 11.6.4, 9, IH), 3.636 (q
d, 6.0.9.7, LH), 3.668 (dd, 7.
2.85, IH), around 3.685 (m, 1) 1), 3
．． 733 (dd, 11.6.2.3.LH), 4.09
2 (qd, 6°2.9.3, IH), 4.234 (br
d, around 1.5, IH), 4.330 (dd, 4.8.
7.5. LH), 4.489 (Mr. s, IH), 4.51
6 (s, IH), 4°686 (d-sama, around 4.4, IH
), 4.692(d, 3.9.IH〉, 5.156(d
, around 3.9, IH), 5.192Cbroad s
Mr., IH〉, 5.200 (Mr. s, IH), 5334 (d
, near 35, IH), 5.604 (d, 72, IH)
, 5.641 (broad s, IH), 5.771
(broadS, IH), 6.388 (s, 2H)
, 6.719 (d, 8.5.1) 1), 6.800 (d
d, 8.5.2.3.1) 1), 7.107 (dd, 8
．． 4.24, IH), 7.147(d, 23, LH>,
7, 170 (dd, 8.4.2.4.1) 1), 7.2
37 (d, 8.4, LH), 7.296 (broad
dd, around 84, around 2.0, IH), 7°331 (d
d, 8.4.2.0.18), 7,590 (broad
dd, around 8.4.2.2, IH〉, 7.857 (d
, 2.0.11 (>C-NMR' 50 MHz, in DtO, external standard cl
(scNl, 60°C as 7pp+n (see Figure 8) δppm: 177, 9.177, 8.174.8.1
72.2°172, L 171.7.170.3.16
7.8.157.6°156.1.156.155.8
．． 155.5.153.9゜151.2.139.13
7.136.7.136.7゜135, 134.2.1
31.4.130.9.128.8゜128.4.12
7.9.127.1.126.4.124.5°124
．． 3.124.1122.6.119.3.118.6
゜109.3.107.2.104.6.104.10
2.9°99.3. 93.7. 79.4. 78.
1. 77.9°76.7. 76.1. 75.2.
72.5. 70.2°69.6. 68.7. 6
7.3. 63.1. 63.1°61.7. 60.
4. 59.9. 57.1. 55.5°55.5.
53. 41.5. 40.2. 39.3. 37
．． 7°34, L 25.1. 23.3. 22.5
．． 19.3°18.3. 17.8 Thin Layer Chromatography Merck Bricoated TLC Plate Silica Gel 6
0F254 Developing solvent chloroform: methanol; concentrated aqueous ammonia: secondary butanol: water (5:10:5:5:2) Rf=0.22 High performance liquid chromatography (Takashi LC-6A) column: Nucleosil 5C18φ4.6 x 150 mm detection: UV 220 nm Flow rate: 1 ml/m
in.

Working: 9z acetonitrile 0.05M phosphate buffer (pH 3,5) Retention time: 9.4 m1n1. 10@A acetonitrile-0.05M phosphoric acid 7-heffer (pH 3.5) Retention time: 6.9 min.

PA-42867-B, which is insoluble in loloform and ethyl acetate, has a steric structure represented by the following formula: It is estimated that

As above, PA-42867-A and PA-428
67-B has properties different from those of known glycopeptide antibiotics, and is considered to be a novel glycopeptide antibiotic.

The producing bacteria of PA-42867-A and PA-42867-B include: - PA-428 isolated from a soil sample;
There are 67 stocks. Based on the results of taxonomic examination, this strain is classified as Nocardia orientalis (Nocardia orientalis).
Nocardia orientalis PA-4 was identified as the same species as Nocardia orientalis PA-4.
2867 (No-cardia orientalis
PA-42867), which has been deposited as Microtechnical Research Institute No. 8601.

The mycological properties of this strain are as shown below.

(1) Morphological properties Aerial hyphae adhere well on yeast malt agar, tyrosine agar, Pennet agar medium, etc., and are spore-forming abundantly. Branches are simple, with no whorled branches. Aerial hyphae are relatively long and have straight or wavy tips. As a result of observation using an electron microscope, the spores are long cylindrical, with a width of 0.3 to 0.5 μm and a length of 1.2 to 1.7 μm.
Its surface structure is smooth. No sporangia, flagellated spores, or sclerotia are observed.

(Margins below) (2) Various culturing properties (cultivation at 28°C for 14 days) The color name display is based on the one-color standard (Japan Color Research New Edition).

Growth temperature (Pennet agar medium, cultured for 14 days at each temperature) 10°C: Quite good growth, but no aerial mycelium was formed. 28°C: Growth, aerial mycelium formation, and spore formation were all good.

37°C: no growth 45°C: no growth (3) Physiological properties (culture at 28°C for 114 days) Melanoid pigment production ability Negative tyrosinase reaction Coagulation of negative milk Peptonization of negative milk Positive gelatin liquefaction ability Positive starch Ice-melting ability Negative (blank below) (4) Carbon source utilization ability (5) Cell wall composition The type of diaminopimelic acid is meso-type.

Based on the above characteristics, it can be concluded that this strain belongs to the genus Nocardia. Waxman, The Aetinomycetes, No. 2
Volume (1961), Shearing and Boratribe I, S, P, (International
al StreptomycesProject)
Report [International Journal of Systematic Bacteriology]
nal Journal of Systematic
13acteriology) Volume 18 (1968), Volume 19 (1969), Volume 22 (1972)
) and Bargey's Manual of Determinative Tapir, Theriology (Bargey's
Manual of Determina-tive
Bacteriology) 8th edition (1974) and other literature announcing new species of actinomycetes for related strains of this strain, Nocardia orientalis (
Nocardia oriantalis, Streptomycas orientalis in the following literature
is) [International
nal Journal of Systematic
Bacteriology Vol. 18, pp. 154-157 (1968), The Actinomycetes
, Vol. 2, pp. 254-255 (1961)] are listed as the most closely related species. As a result of comparing the properties of Nocardia orientalis and strain PA-42867, a difference was observed in the ability to utilize arabinose and rhamnose, but other major properties were in good agreement. Therefore, PA-42
Strain 867 was identified as homologous to Nocardia orientalis, and Nocardia orientalis PA-42.
867 (Nocardia orientalis PA
-42867).

In the present invention, not only the above-mentioned strain PA-42867 and its natural and artificial mutant strains, but also all strains that belong to the genus Nocardia and produce PA-42867-A and/or -B can be used. within the range of

The production of PA-42867-A and/or -B is
-42867-A and/or -B producing strains were cultured in a nutrient medium under aerobic conditions, and after the culture was completed, PA-
This is done by separating and collecting 42867-A and/or -B. A general method for producing PA-42867-A and/or -B is described below.

As for the culture medium composition, culture conditions, etc., those generally used in the production of antibiotics may be used. In principle, the medium contains a carbon source, a nitrogen source, inorganic salts, etc. Vitamins, precursors, etc. may be added as necessary. As the carbon source, for example, glucose, starch, dextrin, glycerin, molasses, organic acids, etc. are used alone or in a mixture. As the nitrogen source, for example, soybean flour, corn stew liquor, meat extract, yeast extract, cottonseed flour, peptone, wheat germ, ammonium sulfate, ammonium nitrate, etc. are used singly or as a mixture. Examples of inorganic salts include calcium carbonate, sodium chloride, potassium chloride, magnesium sulfate, steel sulfate, manganese chloride, zinc sulfate, cobalt chloride, and various phosphates.
Add to the medium as needed.

Cultivation may be carried out according to methods generally used for the production of antibiotics, preferably liquid culture, but for mass production, deep aeration culture is preferred.

If the pH of the medium is variable, a buffer such as calcium carbonate may be added to the medium. Culture at approximately 20-40℃
The temperature is preferably 28 to 32°C. The culture time depends largely on the scale of fermentation, but is approximately 60 to 100
This is the culture time required for mass production.

If severe foaming occurs during culturing, an antifoaming agent such as vegetable oil, lard, or polypropylene glycol may be appropriately added before or during culturing.

After completion of the culture, PA-42867-A and/or
In order to separate and collect -B, a conventional method for separating and collecting fermentation products is appropriately used. For example, filtration, centrifugation,
Adsorption/desorption using various ion exchange resins or other active adsorbents, chromatography, extraction using various organic solvents, etc. may be appropriately combined. It may be desirable to form PA-42867-A and -B into salts for convenience of separation operations and use as medicines and veterinary drugs. P.A.
Bases that can form salts with -42867-A and -B include alkali metals such as potassium and sodium, alkaline earth metals such as aluminum and magnesium, and acids include inorganic acids such as hydrochloric acid, sulfuric acid, and nitric acid, and acetic acid; Examples include organic acids such as fumaric acid.

PA-42867-A and -B and their pharmaceutically acceptable salts can be administered orally or parenterally to humans or animals as active ingredients of antibacterial agents. Using commonly used excipients, stabilizers, preservatives, wetting agents, surfactants, etc., it can be administered orally in the form of tablets, capsules, and powders, and parenterally as injections, liniments, and suppositories. It can also be administered directly. The dosage varies depending on the therapeutic purpose, age of the patient, symptoms, etc., but when administered intravenously, it is approximately 0.1 to 10 g per day for adults.

20 Effects The compounds PA-42867-A and -B of the present invention exhibit strong antibacterial activity against Durham-positive bacteria, particularly against methicillin-resistant bacteria, and are useful as pharmaceuticals for humans or animals.

Furthermore, since it can be collected with high purity, it is suitable for use not only as an oral preparation but also as an injection.

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

(a) Fermentation process: soluble starch 0.5%, glucose 0.5%, polypeptone 0.5%, beef extract 0.5%, yeast extract 0.25
%, salt 0.25%, deionized water (pH 7.0, before sterilization) containing 800 ml of medium 2! Inoculate a seed culture slant of Nocardia sp.
Shaking culture is carried out at 28°C for 548 hours at 0 rotations. 800ml of this culture solution was mixed into a medium 2 consisting of the same components as above.
Inoculate a 3012-volume J-Fermenku containing 012, aeration rate 2 fl/min, internal pressure 0, 5 kg/cm"G,
Culture at 28° C. for 24 hours at a stirring speed of 20 rpm.

Next, 10ρ of this culture solution was mixed with tomato paste 2°4%,
Dextrin 2.4%, dry yeast (Beast, manufactured by Iwaki Pharmaceutical) 1.2%, cobalt chloride hexahydrate 0.0006%
, Antifoaming agent P-2000 (manufactured by Inu Nippon Ink>O, OS%,
250 containing 14CB of medium made of tap water (pH 7.0, before sterilization)! Inoculate the fermentation tank with bacteria, aeration rate 150ρ/min, internal pressure 5%1, stirring speed 325 rpm, 28℃,
Incubate for 64 hours.

(b) Separation step: The culture solution obtained in the above step was adjusted to pH 10.5 with 10% sodium hydroxide, and the supernatant liquid was separated by centrifugation.
5! get. After adjusting this supernatant to pH 4.0, 13ρ
of DOWEX 50 After washing with water, elution is carried out with 40 ρ of 30% acetone water containing 1% triethylamine. Fraction 22 showing activity was collected by pulp disk diffusion method using Bacillus subtilis and adjusted to pH 5. This is concentrated under reduced pressure to remove acetone. Then, it is passed through a 2 column HP-20 (manufactured by Mitsubishi Kasei Corporation). After washing with 20ml of water, 50ml
Elute with % acetone water. Collect active fraction 612. When this is concentrated under reduced pressure and freeze-dried, PA-428
35.6 g of coarse powder of No. 67 is obtained.

(c) Purification step: A solution of 150 ml of 0.01N hydrochloric acid is prepared using 12 g of the above crude powder, and chromatographed using 100 ml of M'ICGEL CHP-20P (manufactured by Mitsubishi Chemical Corporation). ) Elute with 0.0 IN hydrochloric acid while checking the content using IPLc, and PA-4
The fraction containing 2867=27--AXB was adjusted to pH 7.0, and then added to CHP-2 again.
Chromatograph at 0F. After applying the solution containing PA-42867-A and B, it was thoroughly washed with 15% methanol water and eluted with 15% methanol-0,005N hydrochloric acid.
-42867-A containing fraction and PA-42867-B
Separate into containing fractions. PA-42867-B containing fraction was p
After adjusting to H7.0, it was concentrated and freeze-dried to obtain 3 mg of residue 6B. The PA-42867-A-containing fraction was adjusted to pH 7.0, concentrated, filtered with 10 ml of CHP-20F for decolorization, eluted with dilute hydrochloric acid (pH 5.0) aqueous solution, and purified with PA-42867-A.
Fractions containing -42867-A are collected, concentrated and lyophilized to obtain 571 mg of residue (purity 70%). This 571
After dissolving mg in water, add dilute hydrochloric acid to adjust the pH to 5.0, apply it to a CUP-20P10n+1 column, elute with water, collect the fraction containing PA-42867-A, adjust the pH to 7.0, and concentrate. CHP-20P 10 again for desalination purposes
ml (0.05 M phosphate buffer (stabilized at pH 7.0)), washed with 0.05 M phosphate buffer (pH 7.0), and washed with water, then 50% methanol water. The fractions containing PA-42867-A were collected, concentrated and lyophilized to yield 256 mg of PA-42867-A (95% purity).
〉obtained. After dissolving 683 mg of the residue containing PA-42867-B in water, add gold-hydrochloric acid to adjust the pH to 4.0, apply it to a 5 ml column of C) IF-20P for decolorization, and elute with a dilute aqueous hydrochloric acid solution (pH 4.0). The fractions containing PA-42867-B were collected, adjusted to pH 7.0, concentrated, and then applied to backed column RQ-2 (Fujigel Sales Co., Ltd.). HP
7% acetonitrile-0 while checking purity by LC.
,05M phosphate buffer (pH 4,9>), then 8% acetonitrile-0,05M phosphate buffer (pH 4,9>).
), collect fractions with purity of 95% or higher, adjust to pH 7.0, concentrate and desalt with 10 ml of σF-20P (0,
05M phosphate buffer (pH 7.0)). After washing with water, elute with 50% methanol-water.
Fractions containing A-42867-B were collected, concentrated, and lyophilized to yield 102 mg of PA-42867-B (98% purity).
get.

Effects of the invention PA-42867-A and PA"42867-B i
The antibacterial activity in vitro was measured under the following conditions according to the method prescribed by the Japanese Society of Chemotherapy.

(2) Preparation of bacterial cell suspension A platinum loop of test bacterial strain 1 on a slant medium is inoculated into tml of a growth medium (trypto soy broth, Eiken Chemical Co., Ltd.) and cultured at 37°C for 18 to 20 hours. A 100-fold dilution of the culture is used as a cell suspension for inoculation.

■ Accurately weigh 9-10 mg of the sample solution sample and add it to distilled water for 2 m.
Dissolve to give a concentration of g/ml.

■ Dilute the agar plate sample solution with sterile water (2000 ~
0.25 μg/ml). Pour 0.5 ml of each diluted sample solution into a sterile plastic Petri dish (9 cm diameter).
9.5 ml of agar medium (susceptibility test medium, Eiken Kagaku Co., Ltd.) was poured therein, stirred gently, and solidified.

■Measurement of MIC value One platinum loop (0,5-
1 μm) onto the surface of an agar plate containing various concentrations of samples prepared as described above. After culturing at 37°C for 18-20 hours, observe the growth of bacteria on the agar surface. The lowest concentration at which bacterial growth is observed to be completely inhibited is defined as MIC and expressed in μg/ml.

■ Addition of horse serum in special medium Agar medium for measuring MIC against Streptococcus is supplemented with 0.5% (V/V > horse serum) before injection.

The results are shown in Table 1.

(The following is a blank space) Next, Table 2 shows the antibacterial activity of PA-42867-A in vivo.

Test method: Test bacteria are intraperitoneally inoculated into 5lc-ICR female mice (group of 8 mice), and PA-42867-A (multiple dilution) is administered subcutaneously twice, 1 hour and 5 hours later.

Results = 50% effective dose (ED) based on survival rate of mice after 7 days
,. ) is calculated.

(Margin below)

[Brief explanation of drawings]

Figure 1 is the infrared absorption spectrum of PA-42867-A, Figure 2 is the mass spectrometry spectrum of PA-42867-A, and Figure 3 is the ')l-NMR of PA-42867-A.
1 Figure 4 shows the "C-NMR of PA-42867-A. Figure 5 shows the infrared absorption spectrum of PA-42867-B, Figure 6 shows the mass spectrometry spectrum of PA-42867-B, and Figure 7 shows the "C-NMR of PA-42867-A. In-NM of PA-42867-B
R, Figure 8 shows the "C-NMR of PA-42867-B.

Claims (9)

[Claims] (1) Formula I: ▲There are mathematical formulas, chemical formulas, tables, etc.▼ I (In the formula, does X represent CH_3 and Y represents NH_2?
Or X represents H and Y represents OH. ) Antibiotic PA-42867-A or PA
-42867-B and its pharmaceutically acceptable salts. (2) In formula I, X is CH_3 and Y is NH_2
PA-42867 according to claim 1, which is
-A. (3) PA-42867-B according to claim 1, wherein in formula I, X is H and Y is OH. (4) Cultivate PA-42867-A and/or PA-42867-B-producing bacteria belonging to the genus Nocardia in a medium, and extract PA-42867-A and/or PA-42867-B from the culture.
P characterized by separating and collecting A-42867-B
A-42867-A and/or PA-42867-
Manufacturing method of B. (5) The PA-42867-A and/or PA-4
5. The production method according to claim 4, wherein the 2867-B producing bacterium belongs to the species Nocardia orientalis. (6) The PA-42867-A and/or PA-4
The 2867-B producing bacterium is Nocardia orientalis P.
A-42867, the manufacturing method according to claim 4. (7) PA-42867-A and/or PA-42867-B producing bacteria belonging to the genus Nocardia. (8) PA-42867-A according to claim 7 belonging to Nocardia orientalis species and/or
Or PA-42867-B producing bacteria. (9) Nocardia orientalis PA-42867
PA-42867 as set forth in claim 8 which is
-A and/or PA-42867-B producing bacteria.