AU573391B2

AU573391B2 - Xenocoumacins

Info

Publication number: AU573391B2
Application number: AU48014/85A
Authority: AU
Inventors: Richard Peter Gregson; Bernard Vincent Mcinerney
Original assignee: Biotech Australia Pty Ltd; Biotechnology Australia Pty Ltd
Current assignee: Inhibin Pty Ltd
Priority date: 1984-09-05
Filing date: 1985-09-05
Publication date: 1988-06-09
Anticipated expiration: 2005-09-05
Also published as: AU4801485A

Description

"XENOCOUMACINS" TECHNICAL FIELD The present invention relates to a new class of compounds, known as xenocoumacins, which compounds may be isolated from the culture of strains of bacteria of the genus Xenorhabdus.

BACKGROUND ART

Insect pathogenic nematodes of families Heterorhabitidae and Steinernematidae are known to be symbiotically associated with bacteria of the genus Xenorhabdus. It has been observed that these bacteria have the ability to inhibit the activity of other bacterial growth.

International Application No. PCT/AU83/00156 (W084/01775) discloses certain lipid soluble antibiotics isolated and characterised from cultures of the genus Xenorhabdus. The disclosures of that application are incorporated herein by reference.

DISCLOSURE OF INVENTION The compounds of the present invention include those isolated from the water soluble component of the culture supernatant of Xenorhabdus nematophilus and Xenorhabdus luminescens and their derivatives and have been found to have the structure of formula I.

wherein R₁ is hydrogen, straight or branched chain. alkyl or acyl

R₂ is straight or branched chain alkyl of at least 2 carbon atoms , unsubstituted or substituted by one or more substituents selected from hydroxy, acyl , acyloxy. halogen; R₃ and R₄ are hydrogen, hydroxy, alkoxy or acyloxy R₅ is hydrogen hydroxy. alkoxy or acyloxy; A is -(CH₂)_m-B. wherein m is 0, 1, 2 or 3 and B is an amino-containing radical. unsubstituted or substituted by acyloxy; or

A-CH-NHR₅ taken together represent a 5 to 7 membered heterocyclic ring which may contain a further nitrogen, oxygen or sulfur heteroatom, and the pharmaceutically acceptable salts and other derivatives thereof.

Particularly preferred compounds of the present invention have the structure of formulae II and III below, and are designated xenocoumacin 1 and xenocoumacin 2 respectively.

II

III The invention also provides a process for preparing xenocoumacin 1 and xenocoumacin 2 which comprises culturing an antibiotic producing strain of Xenorhabdus nematophilus. or Xenorhabdus luminescens. in a suitable culture medium and separating the compounds of the invention and their precursors from the culture broth.

Preferably, the process is a continuous process wherein culture medium is continously added to the fermenter and culture medium is continually removed from the fermenter at a rate to maintain the volume of the culture within predetermined limits. The compounds of the invention and their precursors are separated from the collected culture.

The invention also provides a process for preparing compounds of formula 1 as defined above, which process comprises

(a) to produce compounds of formula 1 wherein

(i) R₁ is hydrogen, R₂ is 2-methylpropyl. R₃ and R₄ are hydroxy. R₅ is hydrogen, n is 0 and A is -(CH₂)_m-B wnerein m is 3 and B is guanidino; and (ii) R₁ is hydrogen. R₂ is 2-methylpropyl, R₃ and R ₄ are hydroxy, R₅ is hydrogen, n is 0 and A-CH-NHR₅ is a pyrrolidinyl ring. culturing a xenocoumacin producing strain of Xenorhabdus nematophilus or Xenorhabdus luminescens in a suitable culture medium and separating the desired compounds from the culture broth;

(b) to produce compounds of formula 1 wherein R₁, R₂, R₃. R₄, R₅, R₆, n, A. m and B are other than as defined under (a)

(i) oxidising a compound of formula 1 as defined above, wherein R₁, R₂, R3, R₄, R₅, R ₆ , n, A, m and B are as defined under (a) to produce an aldehyde of the formula 2

(ii) reacting the compound of formula 2 with a phosphorane of formula 3:

wherein Ar is aryl and R₄, R₅, and A are as defined above to produce a compound of formula 4:

(iii)oxidising the compound of formula 4 to produce a compound of formula 1 in which R₃ is hydroxy;

(c) to produce compounds of formula 1 wherein R₁, R₂,

R₃, R₄, R₅, R₆, n, A, m and B are as defined under (b), alkylating compound of formula 1 as defined above wherein R₁ is hydrogen. R₃ and/or

R₄ are hydroxy, to produce compounds of formula 1 as defined above wherein one or more of R₁, R₃ and/or R₄ are alkyl; or (d) to produce compounds of formula 1 wherein R₁, R₂, R₃, R₄, R₅, R₆, n, A, m and B are as defined under (c). acylating a compound of formula 1 as defined above wherein R₁ is hydrogen. R₂ and/or R₄ is hydroxy, or R₅ is hydrogen to produce compounds of formula 1 as defined above wherein R₁ is acyl. one or more of R₃, R₄ and R₅ is acyloxy and/or B is substituted by acyloxy. In a further aspect the invention provides pharmaceutical formulations characterised in that the active ingredient comprises a compound of the invention.

Xenocoumacins have been found to possess antibacterial, antifungal. acaricidal. anti-inflammatory and antiulcerogenic properties. The invention therefore also provides a method for the prevention or control of such conditions in a mammal requiring said prevention or control, which method comprises administering to said mammal a therapeutically effective amount of at least one compound of the invention.

Preferably, the compounds of the invention are isolated from cultures of Xenorhabdus using typical extraction techniques such as liquid chromotography and subsequent extraction by an organic or aqueous solvent.

Suitable culture media for the antibiotic strains of Xenorhabdus include materials containing suitable carbon and energy sources such as glucose or other carbohydrates, glycerol, or lipids, suitable nitrogen sources such as ammonia, urea, amino acids, peptides or proteins, appropriate quantities of inorganic nutrients such as phosphate, potassium, magnesium, calcium and trace elements, and preferably a source of vitamins and growth factors , e . g . yeast extract .

The continuous culture processes of the invention are preferably carried out between 23°C and 37°C, most preferably at about 28°C. They are preferably carried out at a pH of between 6.3 and 7.5, most preferably about 6.8. In continuous culture, fresh culture medium is preferably added to give a dilution rate of between 0.01 hr^-1 and 0.5 hr^-1, most preferably between 0.04 hr^-1 and 0.1 hr^-1. MODES FOR CARRYING OUT THE INVENTION Notwithstanding other forms which may fall within the scope of the present invention, preferred forms will now be described with references to the following examples: Example 1. Culture of Xenorhabdus

Xenorhabdus nematophilus strain All/1 (ATCC 53200) can exist in two morphologically different forms known as primary (1°) and secondary (2°) as reported by R.J.Akhurst in J. Gen. Micro. 121. 303-309. (1980). These forms can be distinguished by their gross cell morphology, the fact that only the primary forms elicit xenorhabdins and xenocoumacins, and by their colour when grown on 0.004% triphenyl tetrazolium chloride and 0.0025% bromothymol blue, as secondary colonies appear red since they do not absorb bromothymol blue whereas colonies of the primary form have a red core overlaid by blue.

Culture 1 X. nematophilus strain All/1 (ATCC 53200) (symbiont of Steinernema feltiae All) was cultured in the following medium

(5L) for 48hr. glycerol 5gL ^-1; yeast extract 15gL-1;

MgSO₄ 5mL^-1(lM); (NH₄)₂SO₄ 2gL^-1;

KH₂PO₄ 5mlL^-1(lM); K₂HPO₄ 5mlL^-1(lM);

Na₂SO₄ 10mlL^-1(lM).

The cells were harvested by centrifugation (9000 RPM, 0.25h) and the supernatant was decanted. A sintered Pyrex funnel was uniformly packed with dry octadecyl silica (50-70um, 6cmx12.5cm) and covered with filter paper. The solvent flow through the silica was induced by a vacuum (10 kPa) and the eluate collected in a filter flask. The silica was washed with methanol (1L). water (4L) and then the supernatant (5L) was applied followed by water (2L) and acetonitrile: ammonium acetate (1:1) (0.2M). (pH4.5.2L). This latter fraction was evaporated in vacuo to yield a crude mixture of xenocoumacins (3%. 21g).

A solution of this mixture (5g) in water (20ml) was chromatographed on Sephadex G10 (84x5cm. 1650 ml) in aqueous acetic acid (0.5%) at a flow rate of 3.2ml min^-1. The eluate was monitored continuously at 254 nm and absorbances corresponding to xenocoumacin I and II occurred at 1150-1400ml and 1300-1600ml respectively. A total of 3.7g (20%) of xenocoumacins was recovered as a brown solid.

Culture 2

The following medium is found to be suitable for the culture of Xenorhabdus nematophilus CC 39497.

Glycerol 21gL^-1; yeast extract 10gL^-1; (NH₄)₂SO₄. 20gL^_1; KH₂PO₄. 10gL^-1; MgSO₄.7H₂O, 2.5gL^-1; CaCl₂.2H₂O.0.29gL^-1 FeSO₄.7H₂O. 27.8mgL^_1; MnSO₄.H₂O, 8.45mgL^-1; ZnSO₄.7H₂O, 14.4mgL^-1; CoCl₂.6H₂O, 0.10mgL^-1; and CuSO₄.5H₂O, 0.19mgL^-1.

The medium was innoculated and growth proceeded for 6 days.

The supernatant (136L) was applied to a column (150mm by lm) of Amberlite XAD-2 resin (17.1L) at 500ml.min^-1. After washing the column with 20L of water, methanol was pumped onto the column at a rate of 500ml/min and the eluent collected in 20L aliquots.

500ml of the first aqueous methanolic fraction was lyophilised. extracted with ethylacetate (3 x 200ml) and then chromatographed on Sephadex G25 in water.

The column (2.5cm x 76cm) was washed thoroughly with water, and then eluted with aqueous acetic acid (10% V/V). The eluent was lyophilised.

Substantially pure xenocoumacin 1 was obtained by subjecting the acetic acid fraction to HPLC on a Whatman Partisil M9 10/50 ODS column with acetonitrile: ammonium acetate (0.2M.pH4.5) 30:70 as eluant. At 4ml/min on a 254mm column the retention time was 19 minutes for xenocoumacin I.

Culture 3

High yeilds of xenocoumacins 1 and 2 can be extracted from the supernatant of cultures of Xenorhabdus nematophilus strain All/1 grown on tryptone soya broth as shown in Table 1.

Xenocoumacin 1 and 2 ¹³C NMR 100.62 MHz (D₂O.) Referenced to dioxan 67.8 ppm

Xenocoumacin 1 Hvdrochloride

400 MHz H spectrum (D₂O) ref. Dioxan 3.70 wrt TMS.

8.22, 0.90, 2 x d, J6.6 Hz. 1'-2'-Me; 1.41. q of d. J4a'. 4'b 13.2 Hz J4'a.3' 7.2 Hz. J4'a.5' 4.0 Hz. H4'a; 1.58. m. H3'; 1.64. m. H12'a; 1.70. m. H4'b; 1.74. m. H11'a; 1.79. m. H12'b; 1.83. m. H11'b; 2.96. m. (H4)₂; 3.19, t. J6.4Hz, (H13)₂; 3.47. sextet. J10',9'. 4 Hz. J10'11'a 3.5 Hz. J10'11'b 8.4Hz. H10'; 4.11. dd. J9',10' 4HZ. J9'.8'. 6.1 Hz H9', 4.20. dt, J5'.4'a 4.0 Hz. J5'.4'b 9.8 Hz. J5'.3 4.0 Hz. H5'; 4.59. dt, J3.4a 8.1 Hz. J3.4b 4.5Hz. J3.5' 4.0Hz. H3 ; 6.8, 6.82. 2 x d. J8 Hz. H5. H6; 7.45. t. J8 Hz, H6

Xenocoumacin 1 Hexaacetate

400 MHz ¹H NMR CDCl₃:C₆D₆, 3:2 referenced to TMS

0.89. 0.91, 2 x d. 1'-.2'-Me: 1.20. dq. H4'a; 1.37. m. H11'a; 1.5. m. H11b. (H12)₂; 1.66. m. H3'; 1.81. m. H4'b; 1.80. 1.85, 1.89.1.92. 2.10. 2.20. 6 X S. OAc; 2.53. dd. J4a.4b 14.1 Hz, J4a,3 2.8 Hz, H4a; 2.97, dd, J4a4b 14.1 Hz, J4b, 3 12.6 Hz. H4b; 3.39, q, J13.14 6Hz, J13'.12' 6 Hz, (H13')2; 4.09, dq, J3,4b 12.6 Hz, J3,4a 2.8 Hz. J3,5' 3.5 Hz, H3; 4.24. dt, J5',4'a 9.1 Hz, J5',4'b. 9.1 Hz, J5',3 3.5 Hz, H5'; 4.43, qd. J10',9' 7.2 Hz. J10'.11'a 7.2 Hz. J10'11'b 2Hz. H10'; 5.09. dd. J9',10' 7.2 Hz. J9',8' 1.7 Hz, H9'; 5.32, d, J8',9' 1.7 Hz H8'; 6.59. d, J8 HZ. 10' NH; 6.84, 6.86, 2 X d, J 8 Hz, H5, H7; 7.06, d. J 9 Hz, H6'; 7.2. t. J 8 Hz, H7; 9.0. t, J 6 Hz, H14'.

,

Xenocoumacin 2 Tetraacetate

¹H NMR 400 MHz (CDCl₃) referenced to TMS

0.93, 0.97, 2 x d, J 6.5 Hz, 1'-,2'-Me; 1.48, qd, J4'a,4'b 13.5

Hz, J4'a,3' 8.5 Hz, J4'a,5' 5.5 Hz, H4'a; 1.63. dd (b), J 6 Hz,

13.5 Ha, H11'a; 1.7, m, 3H. H3', H12'a, H11'b; 1.88, qd,

J4'b,4'a 13.5 Hz, J4'b,5' 10 Hz, J4'b,3' 5.5 Hz, H4'b; 1.95. s.

OAc; 2.01. m. H12'a; 2.10. 2.11, 2.40. 3 x s, OAc; 2.89. dd, J

16.7 Hz, 2.5 Hz, H4a; 3.30, dd, J 16.7, 13 Hz, H4b; 3.45, q(b),

J_gem 10 Hz, J13'a,12a, J13'a,12b 10 Hz, H13'a; 3.54, td,

J_gem 10 Hz. J13'b,l2a 3 Hz, J13'b,12b 10 Hz, H13'b; 4.34, m,

H5'; 4.51. dt. J 12.5, 1.5 Hz, H3; 4.56. m, H10'; 5.14, dd, J9',10' 9.9 Hz, J9',8' 1.5 Hz, H9'; 5.21, d, J8',9' 1.5 Hz. H8'; 7.03, 7.12, 2 d, J 8 Hz, H5 , H7; 7.52, t, J 8 Hz, H6; 8.69, d, J 8.5 HZ. H6'

Xenocoumacin 2 1H NMR 400 MHZ (D₂O) referenced to Dioxan 3.70

0.82, d, J 6.5 Hz, 1'-Me; 0.89, d, J 6.5 Hz, 2'-Me; 1.39, qd, H4'a; 1.55. m, H3'; 1.70, m, H4'b; 1.92, m, 2H, H11'a, H12'a; 2.06. m, H12'b; 2.1, m, H11'b; 2.96, m, (H4)₂; 3.27, t, J 8 Hz, (H13')₂; 3.61, m, H10'; 4.10, dd, H9'; 4.16, m, H8', H5'; 4.58, m, H3; 6.78, 6.81, 2 x d, J 8 Hz, H5. H7; 7.44, t, J 8 Hz, H6.

Xenocoumacin 2 Monoacetate

¹H NMR 400 MHz (CDCl₃/D₂O) referenced to TMS

0.93, 0.96, 1'-,2'-Me; 1.50, qd, J 13.5. 8.6, 5 Hz, H4'a; 1.68, m. H3'; 1.79, m. H4'b; 1.88, m, H12'a; 1.91, m. H11'a; 2.04, m, H12'b 2.12, S, OAc; 2.27, m, H11'b; 2.83. dd, J 16.7, 2.5 Hz. H4a; 3.11, dd, J 16.7, 13 Hz, H4b; 3.53, m, H13'a, H13'b; 3.70, dd. J 7.8. 4.4 Hz, H9'; 3.90, d, J 7.8 Hz, H8'; 4.21, m, H10'; 4.35, m, H5'; 4.60, dt, J 12.7, 2.4 Hz, H3 ; 6.71, 6.90, 2 x d, H5, H7; 7.42, t, H6 ; 7.9, d, 9Hz, H6'; 10.8, x, 8-OH. MASS SPECTRA

Xenoucoumacin 1 Hexaacetate

Negative Ion Chemical lonisation (isobutane, gold probe, 200°) m/z: 717M- (25), 675 (52) (M-CH₂CO), 657 (42), 615 (52),

403 (5), 361 (20), 254 (10), 163 (38), 150 (100),

Electron lonisation, 70 EV, 200°, gold probe m/z: 717 M⁺ (2),

702 (4), 675 (15), 658 (10), 632 (10), 616 (10), 512 (20) 470

(15), 341 (5), 267 (28), 184 (90), 170 (28), 112 (30), 86 (50),

70 (100), Observed Mass 184.1076. Calculated for

C₈H₁₄N₃O₂ 184.1086

Positive Ion Chemical lonisation (ammonia, gold probe, 200°) m/z: 718 (M + H, 75), 676 (MH-CH₂CO, 100), 658 (15) 634 (10), 617 (12), 593 (10), 557 (7), 513 (15), 471 (12), 267 (22), 184 (40), 170 (15), 112 (15).

Xenocoumacin 2 Hexaacetate

Negative Ion Chemical lonisation (isobutane. gold probe. 200°) m/z: 573 (M- H), 531 (M - CH₃CO, 100), 514 (8), 454 (18), 412

(12), 394 (12), 265 (10), 222 (10), 163 (8), 130 (10).

Electron lonisation (70 EV, gold probe. 200°) m/z: 574 (M⁺,

10), 369 (10), 242 (11), 196 (10), 154 (12), 135 (5). 112 (35),

70 (100).

Positive Ion Chemical lonisation (ammonia, gold probe, 200°) m/z: 575 (M + H, 100) 533 (MH-CH₂O, 12). 370 (10), 242 (8),

196 (12), 154 (8), 112 (15), 70 (17).

Xenocoumacin 1

Fast Atom Bombardment: Observed Mass 464.2574

Theoretical Mass for C₂₂H₃₄N₅O₆ (M - H) 464.2509. Example 3. Production of Derivatives

The following synthetic pathways serve to illustrate how derivatives of xenocoumacins may be produced.

Ar is an aryl and A.R ₄ and R₅ are as defined above.

As will be understood by those skilled in the art. the stereospecificity may be altered or controlled by altering the oxidant used. e.g. H₂O₂/SeO₂ will confer a different stereochemistry on the vicinal diol to OsO₄.

R₁, R₃ and R₄ of formula I may be varied by known processes of alkylation and acylation as per the following reactions:

Example 4. Antiulceroqenic activity

The antiulcerogenic activity of the xenocoumacins of the present invention was demonstrated by the following tests: Test 1

Male Wistar rats were dosed perorally (p.o.) dosed with varying amounts of xenocoumacin 1. The animals were then exposed to ulcer inducing stress conuitions. killed and examined for ulcer induction. The results were compared to a control using the Wilcoxon rank sum test. Significance is taken as p< 0.05. Xenocoumacin 1 was shown to be 74% effective when a dosage of 25 mg/kg.po was administered, but only 8% effective for a dosage of 10mg/kg.po. Test 2

Test 1 was repeated using xenocoumacin 2 , which was found to be 70% effective for a dosage of 25 mg/kg.po., 61% effective for a dosage of 10mg/kg.po and 26% effective for a dosage of 5mg/kg.po.

The xenocoumacins of the present invention may be used in the treatment of bacterial or fungal infections or in the treatment or prophylaxis of inflammatory diseases or ulcers.

The compounds or mixtures of them may be administered in standard pharmaceutical formulations, in admixture with known pharmaceutically acceptable excipients. adjuvants and diluents. Formulations of the invention may also contain other active substances. Example A

Xenocoumacins 1 and 2 can be used independently or in admixture as the active ingredient in the manufacture of the following pharmaceutical formulations:

The xenocoumacins of the invention may also be administered topically or intravenously.

Preferred dosage rates are within the range of 0.1mg/kg to 50mg/kg when administered periorally, and 0.01 to 20 mg/kg when administered intravenously. Topical formulations can contain up to approximately 10% active substance.

Claims

CLAIMS Compounds of the formula I

wherein R₁ is hydrogen, straight or branched chain. alkyl or acyl

R₂ is straight or branched chain alkyl of at least 2 carbon atoms, unsubstituted or substituted by one or more substituents selected from hydroxy. acyl. acyloxy, halogen;

R₃ and R₄ are hydrogen, hydroxy. alkoxy or acyloxy

R₅ is hydrogen hydroxy. alkoxy or acyloxy; and A is -(CH₂)_m-B, wherein m is 0, 1, 2 or 3 and B is an amino-containing radical. unsubstituted or substituted by acyloxy; or

A-CH-NHR₅ taken together represent a 5 to 7 membered heterocyclic ring which may contain a further nitrogen, oxygen or sulfur heteroatom. and the pharmaceutically acceptable salts and other derivatives thereof.
2. A compound of formula 1 as defined in claim 1, wherein R₁ is hydrogen. R₂ is 2-mechylpropyl. R₃ and R ₄ are hydroxy, R₅ is hydrogen, n is O and A-CH-NHR₅ is a pyrrolidinyl ring.
3. A compound of formula 1 as defined in claim 1, wherein R₁ is hydrogen, R₂ is 2- methylpropyl, R₃ and R₄ are hydroxy, R₅ is hydrogen, n is O, A is -(CH₂)_m-B wherein m is 3 and. B is. guanidino.
4. A compound of formula 1 as defined in claim 1, wherein R₁ is acetyl, R₂ is 2-methylpropyl. R₃, R₄ and

R₅ are acetyloxy. n is O, A is -(CH₂)_m-B wherein m is 3 and B is 1,3-diacetyloxyguanidino.
5. A compound of formula 1 as defined in claim 1, wherein R₁ is acetyl. R₂ is 2-methylpropyl. R₃, R₄ and R₅ are acetyloxy, n is O and A-CH-NHR₅ is a

N-acetyloxypyrrolodinyl ring.
6. A compound of formula 1 as defined in claim 1, wherein R₁ is hydrogen, R₂ is 2-methylpropyl. R₃ and R₄ are hydroxy, R₅ is acetyloxy, n is O and A-CII-NHR₅ is an

N-acetyloxypyrrolidinyl ring.
7. A process for preparing compounds of formula 1 as defined in claim 1. which process comprises

(a) to produce compounds of formula 1 wherein

(i) R- is hydrogen. R₂ is 2-methylproρyl. R₃ and R₄ are hydroxy, R₅ is hydrogen, n is O and A is -(CH₂)_m-B wherein m is 3 and B is guanidino; and (ii) R₁ is hydrogen, R₂ is 2-methylpropyl, R₃ and R₄ are hydroxy, R₅ is hydrogen, n is O and A-CH-NHR₅ is a pyrrolidinyl ring, culturing a xenocoumacin producing strain of Xenorhabdus nematophilus or Xenorhabdus luminescens in a suitable culture medium and separating the desired compounds from the culture broth:

(b) to produce compounds of formula 1 wherein R₁, R₂, R₃, R₄, R₅, R₆, n, A, K and B are other than as defined under (a) (i) oxidising a compound of formula 1 as defined in claim 1. wherein R₁, R₂, R₃, R₄, R₅,

R₆, n, A, m and B are as defined under (a) to produce an aldehyde of the formula 2

O 2

(ii) reacting the compound of formula 2 with a phosphorane of formula 3:

3

wherein Ar is aryl and R₄, R_5, and A are as defined in claim 1 to produce a compound of formula 4:

4

(iii)oxidising the compound of formula 4 to produce a compound of formula 1 in which R₃ is hydroxy: (c) to produce compounds of formula 1 wherein R₁, R₂, R₃, R₄, R₅, R₆, n, A, m and B are as defined under (b), alkylating a compound of formula 1 as defined in claim 1 wherein R₁ is hydrogen. R₃ and/or R₄ are hydroxy, to produce compounds of formula 1 as defined in claim 1 wherein one or more of R₁, R₃ and/or R₄ are alkyl; or

(d) to produce compounds of formula 1 wherein R₁, R₂, R₃, R₄, R₅, R₆, n. A, m and B are as defined under (c), acylating a compound of formula 1 as defined in claim 1 wherein R₁ is hydrogen. R₂ and/or R₄ is hydroxy, or R₅ is hydrogen to produce compounds of formula 1 as defined in claim 1 wherein R₁ is acyl, one or more of R₃, R₄ and

R₅ is acyloxy and/or B is substituted by acyloxy.
8. A process as defined in claim 7. wherein the xenocoumacin producing strain is Xenorhabdus nematophilus strain XQ1 (ATCC 39497).
9. A process as defined in claim 7. wherein the xenoconmacin producing strain is Xenorhabdus nematophilus strain All/1. (ATCC 53200)
10. A process as defined in claim 9. wherein the medium is tryptone soya broth.
11. A compound of formula 1 as defined in claim 1. whenever produced by the process claimed in claim 7.
12. A pharmaceutical composition, which comprises at least one compound of formula 1 as defined in claim 1 together with a pharmaceutically acceptable carrier or diluent therefor.
13. A method for the prevention or control of infectious disease in a mammal requiring said prevention or control, which method comprises administering to said mammal an effective amount of at least one compound of formula 1 as defined in claim 1.
14. A method of prophylaxis or treatment of ulcers in a mammal requiring such prophylaxis or treatment, which process comprises administering to said mammal an effective amount of at least one compound of formula 1 as defined in claim 1.