New macrodiolide antibiotics, 11-O-monomethyl- and 11, 11'-O-dimethylelaiophylins, from Streptomyces sp. HKI-0113 and HKI-0114.

Elaiophylin (1) and two new methyl derivatives, 11-O-monomethylelaiophylin (2) and 11,11'-O-dimethylelaiophylin (3), were isolated from the mycelium cake of Streptomyces strains HKI-0113 and HKI-0114. The structures of 2 and 3 were determined by mass spectrometric and NMR investigations. Compounds 2 and 3 display antimicrobial and moderate cytotoxic activities.

Lipohexin, a new inhibitor of prolyl endopeptidase from Moeszia lindtneri (HKI-0054) and Paecilomyces sp. (HKI-0055; HKI-0096). I. Screening, isolation and structure elucidation.

Lipohexin was isolated as a novel lipohexapeptide (I) (C39H68N6O9) from three fungal strains, Moeszia lindtneri HKI-0054, Paecilomyces sp. HKI-0055 and Paecilomyces sp. HKI-0096. The structure was elucidated by detailed mass spectrometric and NMR experiments. The proline-containing peptide displays moderate antibacterial activity against Bacillus subtilis ATCC 6633 and inhibits competitively the prolyl endopeptidase from human placenta.

Chrysospermins, new peptaibol antibiotics from Apiocrea chrysosperma Ap101.

Four new members of peptaibol antibiotics, designated as chrysospermins A, B, C, and D, were isolated from the mycelium of Apiocrea chrysosperma Ap101 by solvent extraction, silica gel chromatography and preparative recycling HPLC. Their structures as new nonadecapeptides were settled by detailed spectroscopic analysis and chemical degradation experiments. The chrysospermins display antibacterial and antifungal activity, and induce pigment formation by the fungus Phoma destructiva.

Aurantimycins, new depsipeptide antibiotics from Streptomyces aurantiacus IMET 43917. Production, isolation, structure elucidation, and biological activity.

Aurantimycins A (1), B (2) and C (3) were isolated from the mycelium of Streptomyces aurantiacus JA 4570 as new representatives of the azinothricin group of hexadepsipeptide antibiotics. Their structures were settled by X-ray diffraction analysis of crystalline aurantimycin A (1), high field homo- and heteronuclear 2D NMR experiments, high-resolution mass spectrometry and amino acid analysis. Aurantimycins are characterized by a new side chain containing fourteen carbon atoms. They display strong activity against Gram-positive bacteria and cytotoxic effects against L-929 mouse fibroblast cells.

Helioferins; novel antifungal lipopeptides from Mycogone rosea: screening, isolation, structures and biological properties.

Helioferins A and B were detected as novel aminolipopeptides in cultures of Mycogone rosea DSM 8822 in the course of a screening for mediators of helianthate anion transfer from aqueous to toluene phases. Their structures as novel antibiotics and cytotoxic agents were elucidated by mass spectrometry and NMR spectroscopic methods. Antimicrobial activity was estimated against Candida albicans and Gram-positive bacteria including Mycobacterium spp.

[Biosynthesis of anthracycline: a new interpretation of the results for daunomycin biosynthesis]. / Biosynthese der Anthracycline: eine Neuinterpretation der Ergebnisse zur Daunomycin-Biosynthese.

On the basis of literature data and our own experiments the "late" biosynthetic pathway to daunomycin has been interpreted from a new point of view considering both the in vivo biosynthesis and formation of shunt products. In contrast to existing hypotheses proposed by other authors we discuss a modified sequence leading to C-11 oxidation and, as a consequence, understand epsilon-rhodomycinone as a shunt product instead of a biosynthetic intermediate. In addition, a new hypothesis about the "early" steps of the ring formation from polyketides by a sequence of enzyme reactions has been proposed.

Physiochemical characterization of substituted chromeno[4,3-b][1,5]benzodiazepine stereoisomers designed as cell membrane active antitumor agents.

As an alternative to naturally occurring pyrrolo[2,1-c][1,4]benzodiazepines (e.g., antramycin) which possess properties of DNA alkylation, we have designed several antileukemic chromeno[4,3-b][1,5]benzodiazepine derivatives with potential activity toward leukemia cell membranes and the cyclic nucleotide system. The cis and trans diastereoisomers were characterized by NMR. The absolute configurations of the enantiomers were established by X-ray diffraction and circular dichroism (CD) measurements. By means of absorption spectroscopy and determinations of fluorescence and fluorescence decay, it was found that the cancerostatically active compound (+)(6aR, 13aS)-3,4-dimethoxy-10,11-dimethyl-6,6a,7,8,13, 13a-hexahydrochromeno[4,3-b][1,5]benzodiazepine (ZIMET 54/79) and its biologically inactive (-) enantiomer (ZIMET 55/79) interact with liposomal membranes. At pH values of 6.0 and 7.3 the long-wave absorption bands of these agents showed weak bathochromic and hypochromic effects upon addition of neutral, and positively and negatively charged phosphatidylcholine and phosphatidylcholine/cholesterol liposomes. Such spectral changes are interpreted as resulting from the binding of both agents to phospholipid bilayers. Steady-state determinations using the membrane probe 1-anilino-8-naphthalenesulfonic acid (1,8-ANS) led to the observation of a small decrease in fluorescence intensity in the presence of either agent. Time-resolved measurements demonstrate that the mechanism of action of the agents occurs mainly through the partial displacement of probe molecules from regions of hydrophobic binding to areas of greater solvent accessibility. No significant differences in binding between the cancerostatically active and inactive enantiomers with liposomes (archiral systems) were detectable on the basis of spectrophotometric and fluorescence determinations. Cell membrane bound adenylate cyclase is stimulated by ZIMET 54/79, resulting in an increase of 103% in the level of cAMP in mouse L1210 leukemia cells. On examination of structure-activity relationships, it was found that the biological activity (leukemia L1210, P388, Lewis lung carcinoma, melanoma B16, increase in cAMP) is correlated with the particular configuration (6aR,13aS) and type of substituent at positions 3 and 4 of the benzo ring in the case of alkoxy groups and positions 10 and 11 for methyl groups. No activity was detected toward DNA/RNA using microbial test systems.

Kinetics of secondary metabolite formation by wild type and mutant strains of Streptomyces griseus.

In a particle containing medium, the kinetics of formation of secondary metabolites of a Streptomyces wild type and a mutant strain were determined by spectrophotometric measurements of the anthracyclinone pigments and by the analysis of antibiotic activity of the substances produced. The results were related to the physiological state of the culture derived from the oxygen supply. The total cultivation time was about 400 hours. The production of secondary metabolites started at the end of the logarithmic growth phase (after about 30 hours). During the following cultivation remarkable differences between the two strains regarding the formation of pigments and antibiotics occurred. In the wild type strain producing daunomycinone glycosides and related compounds with antibiotic activity, these antibiotics are active as inhibitors (negative feedback). Therefore, at a certain concentration of antibiotics, the de-novo formation of anthracycline molecules was impaired. In the mutant strain, as the result of a genetic change, the secondary metabolite production was shifted to nonglycosylated compounds. The production of these substances is not influenced by feedback inhibition as in the case of the wild type strain. From that reason the amount of pigment increased slightly throughout the course of the experiment. Under our experimental conditions the measurement of oxygen supply was a suitable indicator of the physiological state of culture.

[Relationships between structure and activity of anthracycline antibiotics of the rhodomycin group: inhibition kinetics of DNA and RNA viruses of the double- and single-strand types]. / Beziehungen zwischen Struktur und Wirkung von Anthracyclin-Antibiotika der Rhodomycingruppe: Hemmungskinetik der DNS- und RNS-Viren vom Doppelstrang- und Einstrang-Typ.

Antiviral activity from anthracycline antibiotics of rhodomycin-type was investigated by two independent methods, which determined the infectious units and antigenity of incomplete virions. The action of rhodomycin was dependent on structure, number and position of amino sugar, which is in aglycon. The viruses of double-stranded DNA and RNA viruses was stronger inhibited as single-stranded RNA viruses. The antiviral activity were found to increase in the serial order iremycin less than adriamycin less than daunomycin less than alpha-rubicin I less than beta-rhodomycin I less than violamycin BI-complex by inhibition kinetic determined from Adeno virus.

Interaction of anthracycline antibiotics with biopolymers: comparative studies of DNA binding and antimicrobial activity of rhodomycin-type anthracycline antibiotics.

The binding of the anthracyclines beta-rhodomycin-I and beta-rhodomycin-II to calf thymus DNA was investigated by both equilibrium and kinetic methods taking into account ligand dimerization (ionic strength I = 0.2 M, pH 6.0). The analysis was based upon a cooperative single-step binding mechanism with overlapping of potential binding sites on a linear homogeneous lattice. Equilibrium binding parameters were estimated from spectrophotometric titration experiments by means of a nonlinear fitting program. The results were compared with those obtained previously for the related antibiotic iremycin and were complemented by kinetic parameters determined from temperature-jump experiments at high binding ratio. The binding constants and the mean attachment times of the drugs were found to increase in the serial order iremycin, beta-rhodomycin-I and beta-rhodomycin-II, which is in line with their increasing antimicrobial activity on Bacillus subtilis ATCC 6633.

[Leukaemomycin-blocked mutants of Streptomyces griseus and their pigments. III. 11-Desoxydaunomycinone derivatives from the mutant ZIMET 43699/G44]. / Leukaemomycin-geblockte Mutanten des Streptomyces griseus und ihre Pigmente. III. 11-Desoxydaunomycinonderivate aus der Mutante ZIMET 43699/G44.

The isolation and identification of several anthracyclinones (designated as G44-K4/5, G44-G1, G44-G2) and anthracyclines (G44-A, B, C, D, E, F, G) produced by the mutant strain IMET JA 3933/G44 of the daunomycin-producing Streptomyces griseus strain IMET JA 3933 are described. G44-K4/5 was found to be identical with 7,11-dideoxy-13-dihydrodaunomycinone previously isolated from a mutant strain of Streptomyces coeruleorubidus. Compound G44-G1 was identified as 11-deoxydaunomycinone, the aglycone of the antibiotic 11-deoxydaunomycin. G44-G2 was found to be a stereoisomer of G44-G1. The NMR and CD spectral data suggest strongly that the compound is 7-epi-11-deoxydaunomycinone. Of the 7 isolated G44-glycosides only the major component G44-B could be identified. Comparison with an authentic sample revealed that this compound is 11-deoxydaunomycin which had previously been isolated from cultures of Streptomyces peucetius var. aureus and Micromonospora peucetica. As reported for S. coeruleorubidus, S. peucetius var. aureus, and Micromonospora peucetica the 11-deoxydaunomycinone derivatives described in this paper were isolated from the fermentation broth of a mutant of a daunomycin-producing wild type strain. This suggest that in general the accumulation of 11-deoxydaunomycinone derivatives may be the result of a block of C11-hydroxylation in the normal biosynthetic pathway of daunomycin and its analogues.

[Anthracycline antibiotics from genetically modified streptomycetes. The isolation, spectroscopic structural elucidation and biologic effects of beta-rhodomycin I]. / Anthracyclinantibiotica genetisch modifizierter Streptomyceten. Zur Isolierung, spektroskopischen Strukturaufklärung und biologischen Wirkung von beta-Rhodomycin-1.

By mutagenic treatment and selection procedures the mutant ZIMET 43678 was obtained from a population of the interspecific recombinant Streptomyces violaceus subsp. iremyceticus ZIMET 43615, which showed a changed spectrum of secondary metabolites. The main component isolated from the fermentation broth was a pure anthracycline evidenced by TLC. By means of acid hydrolysis, identification of the degradation products and also by spectroscopic UV/VIS-, IR-, MS-, 1H/13C-NMR- and CD-investigations with intact anthracycline the structure 7-(alpha-L- rhodosaminyl )-beta- rhodomycinon with the absolute configuration 7S, 9R , 10R was found. The anthracycline called beta- rhodomycin -1 (1) exhibits antimicrobial and cytostatic activity in vitro and is also effective on tumour cells in tumour bearing animals.

Inducers of both cytodifferentiation and anthracycline biosynthesis of Streptomyces griseus and their occurrence in actinomycetes and other microorganisms.

Many taxonomically different strains of actinomycetes as well as several other procaryotic and eucaryotic microorganisms were shown to produce inducers of cytodifferentiation and anthracycline biosynthesis being active towards blocked mutant ZIMET 43682 of Streptomyces griseus. 37 out of 40 strains of the species of S. griseus (92.5%) yielding different kinds of antibiotics were able to induce both the formation of aerial mycelium in the indicator strain ZIMET 43682 concomitant with the biosynthesis of anthracycline antibiotics. 80 (26.3%) out of 304 strains belonging to 94 different Streptomyces species were also producers of inducing agents. Among 77 strains of actinomycetes belonging to 28 different genera being apart from Streptomyces, 12 strains displayed inducing activity (15.6%). The results obtained so far support the conclusion that the formation of inducing agents was neither associated with a particular species of Streptomyces nor its capacity to produce a given antibiotic. The occurrence of inducers has been observed even with members of other genera. Few strains were capable of inducing only one of the differentiation-associated functions. Some strains of Streptomyces species produced at least two different kinds of inducers discernible by their Rf values on TLC.

[Biological effects of coordination compounds of transitional metals. The beta-lactamase inhibitory effect of cis-platin, 2-aminopyridine palladium chloride, clavulanic acid and penicillanic acid sulfonate CP 45,899 in the nitrocefin test and Titertek/microtiter automatic system]. / Biologische Wirkungen von Koordinationsverbindungen der Ubergangsmetalle. beta-Lactamasen-inhibitorische Wirksamkeit von Cis-Platin, 2-Aminopyridin-palladium-chlorid, Clavulansäure und Penicillansäure-Sulfon CP 45 899 im Nitrocefin-Test und Titertek/Microtiter-Automatensystem.

The beta-lactamase-inhibiting activity of 6m-ethyl-pyrid-2-yl-ammine palladium-dichloride (Pd 25681) and cis-dichloro-diammine-platinum(II) was studied and compared with the enzyme inhibitory action of potassium clavulanate and the penicillanic acid sulfone CP 45899. Using the nitrocefin test method and the Titertek/Microtiter equipment CP 45899 and potassium clavulanate were the strongest inhibitors of the Bacillus cereus beta-lactamase. Cis-dichloro-diammine-platinum(II) was fourfold less active than the palladium complex PD 25681 in äquimolar concentration. The following ID50 values were found: CP 45899: 0.0281 microgram; K-clavulanate: 0.1274 microgram; Pd 25681: 3.8603 microgram; cis-dichlorodiammine-platinum(II): 12.5120 microgram/100 microliter.

New anthracycline antibiotics produced by interspecific recombinants of streptomycetes. IV. Antimicrobial activity of iremycin.

The in vitro antimicrobial activity of iremycin (10-(alpha-L-rhodosaminyl)-gamma-rhodomycinone) was determined in comparison to that of doxorubicin, a 14-hydroxy-derivative of daunorubicin, which exhibited a strong antitumor activity and is useful in chemotherapy of human tumors. The MIC values determined by means of a standardized agar diffusion plate test indicated a lower antimicrobial activity of iremycin in vitro in comparison to that of doxorubicin. In contrast to doxorubicin, iremycin was highly active against Mycobacterium smegmatis, but five-fold less active than doxorubicin against Staphylococcus aureus, seven-fold less active against Bacillus subtilis, and twenty five-fold less active against Commamonas terrigena. Furthermore, iremycin was hundred-fold less active against a highly sensitive permeation mutant of Pseudomonas aeruginosa. No inducing activity on prophages in lysogenic E. coli cells was demonstrable for iremycin and no growth inhibition in the repair test was observable. In contrast, iremycin inhibited the multiplication of gamma-phages in the BIP test, but the MIC values of violamycin BI, doxorubicin and iremycin in this test system indicated that iremycin is two hundred fifty-fold less active than violamycin BI and ten-fold less active than doxorubicin. No serum binding was demonstrable for iremycin.

[Leukaemomycin-blocked mutants of Streptomyces griseus and their pigments]. / Leukaemomycin-geblockte Mutanten des Streptomyces griseus und ihre Pigmente.

In a continued search for leukaemomycin-blocked mutants of three leukaemomycin-producing strains IMET JA 3933, IMET JA 5142 and IMET JA 5570 of Streptomyces griseus, 32 mutants producing aerial mycelium and spores were detected. Furthermore, in all mutants cosynthetic capability has been observed. This report describes characterization of leukaemomycin-blocked mutants obtained by mutagenic treatment experiments using NTG and combined UV-/X-rays. According to the biosynthetic capability for anthracyclinones or other pigments the mutants could be divided into six classes. The first class contains 14 leukaemomycin-blocked mutants unable to synthesize anthracyclinones. Besides two classes of mutants (12)synthesizing well-known anthracyclinones as epsilon-rhodomycinone, 7-deoxy-epsilon-rhodomycinone, 11-deoxy-derivatives of daunomycinone, three new classes of mutants (6) synthesizing reddish-brown, brown and blue-violet pigments on solid media with structures not elucidated as yet, will be described.

[Antineoplastic activity of lambdamycin in different murine test models compared with cyclophosphamide, 6-mercaptopurine and 5-fluorouracil (author's transl)]. / Antineoplastische Wirksamkeit von Lambdamycin an verschiedenen murinen Testmodellen im Vergleich zur Wirkung von Cyclophosphamid, 6-Mercaptopurin and 5-Fluoruracil.

Lambdamycin, a chromoglycoside antibiotic like chartreusin was found to be very active against leukemias L 1210 and P 388 and moderate effective against melanoma B 16 and Lewis lung carcinoma of mice. It was nearly without any effect when tested on Walker 256 carcinoma of rats. The effectiveness of lambdamycin was compared with that of cyclophosphamide, 5-fluorouracil and 6-mercaptopurine.