No abstract

Rhizocticin A, the main component of the antifungal, hydrophilic phosphono-oligopeptides of Bacillus subtilis ATCC 6633, was used for sensitivity testing and experiments into the molecular mechanism of the antibiotic action. Budding and filamentous fungi as well as the cultivated nematode Caenorhabditis elegans were found to be sensitive, whereas bacteria and the protozoon Paramecium caudatum were insensitive. Rhizoctonia solani was inhibited in agar dilution tests but not in diffusion tests. The antifungal effect of rhizocticin A was neutralized by a variety of amino acids and oligopeptides. Oligopeptide influence was mainly understood as transport antagonism, and it was concluded that the antibiotic enters the recipient cell via the peptide transport system. L- and D-cystine were also identified as potent, general antagonists of the oligopeptide transport. The rhizocticin-antagonism of four other amino acids was taken as a clue to the site of action. Provided that rhizocticin A is split by peptidases of the target cell into inactive L-arginine and toxic L-2-amino-5-phosphono-3-cis-pentenoic acid (L-APPA), the latter may interfere with the threonine or threonine-related metabolism.

Of the two antifungal antibiotics produced by Badllus subtilis F-29-3, the dipeptide compound bacilysin inhibits yeasts (and bacteria), whereas the formerly unknown fengymycin, a complex of closely related lipopeptide components, shows antibiotic activity against filamentous fungi.Bacilysin production, formerly known for a few strains only, could be demonstrated for all 12 wild-type cultures of Badllus subtilis tested during this study. The antibiotic also occurs in some strains of three other Badllus species considered as closely realted to B. subtilis.Members of the lipopeptide class of antifungal Badllus metabolites were formed by 8 of 12 Badllus subtilis-isolztes and several other Badllus strains.The antibiotics of F-29-3 were compared with antifungal metabolites of other Badllus isolates using TLC, agar-diffusion techniques and tests demonstrating the capacity of six lipopeptide and peptide preparations to protect rice seedlings from phytomycosis due to Rbizoctonia solani.Fengymycin proved to be different from the other compounds tested. It was less toxic to the test plants and protected them better from Rhizoctonia disease than the other antibiotics of the study did.

ZUSAMMENFASSUNG 1235 4,5,6-Trimethoxy-N, N-dimethyl-tryptamin (XI) und zwei dem Mescalin nahestehende Verbindungen, fi-(3,4,5-Trimethoxyphenoxy)-athyl-amin (VI) und ,4,5-Trimethoxyphenoxy)-athyl-dimethylamin (III), wurden synthetisiert. Keine der drei Substanzen zeigte in vorlaufigen Humanversuchen psychotrope Aktivitat. Vor etwa 28 Jahren haben OXFORD et ~1 .~) aus dem Kulturfiltrat von Penicillium brefeldianum DODGE die Fulvinsaure (I) z, 3, isoliert. Der gleiche Pilz produziert auch Grise~fulvin~). Bei der Untersuchung eines Stammes (S 464) dieses Pilzes haben wir als Hauptprodukt Palitantin (11) 4, gefunden. In kleinerer Menge konnten wir das mit Palitantin eng verwandte Frequentin (111) 5, 6 , sowie zwei neue Stoffwechselprodukte isolieren, die wir Brefeldin A und Brefeldin B nennen. Fulvinsaure und Griseofulvin liessen sich bisher in keiner unserer Ziichtungen, die unter den verschiedensten Bedingungen durchgefiihrt wurden, nachweisen. Dagegen konnten wir Brefeldin A auch aus dem Kulturfiltrat eines weiteren Penicillium-Stammes (S 498) als Hauptmetabolit gewinnen. Als Nebenprodukt erhielten wir aus diesem Stamme noch 1-0-Stearylglycerin (cc-Monostearin)

Neben Xanthomegnin (16b), 3,4-Dehydroxanthomegnin (29 a), Viomellein (23) und Vioxanthin (3) wurde aus Penicillium citreo-oiride unter veranderten Kulturbedingungen semi-Vioxanthin (7) isoliert. Konstitution 7 folgl aus den Speklren und dem Befund, da8 mit Fremy-Salz 3,4,6,9-Tetrahydro-1O-hydroxy-7-methoxy-3-melhyl-1,6,9-trioxo-lH-naphtho[2.3-c]pyran (9a = semiXanthomegnin) entsteht. -Das durch Entmethylierung von 9a zugangliche 9c lieB sich mit KaliumpersulfatjNatriumhydroxid zu 16a dimerisieren, das zu 16b methyliert wurde. Synthetisches 16b war mit nativem Xanthomegnin identisch, was die von Hi$e und Rijser') revidierte Konstitution bestatigt und beweist, daB die Farbstoffe der Xanthomegnin-Reihe biogenetisch einheitlich sind. -Die Stoffwechselprodukte des Stammes hemmen das Wachstum von Bakterien, 16b und 23 wirken ferner gegen Insekten. Metabolic Products of Microorganism, 172 ') Isolation of the Antibiotic semi-Vioxantbin from Penicillium citueo-uivide and Synthesis of XantbomegninBesides xanthomegnin (16 b), 3,4-dehydroxanthomegnin (29a), viomellein (23) and vioxanthin (3) from Penicillium citreo-oiride under different culture conditions semi-vioxanthin (7) was isolated. Structure 7 has been established by its spectra and the fact that oxidation with Fremy's salt gives 3,4,6,9-tetrahydro-1O-hydroxy-7-methoxy-3-methyl-1,6,9-trioxo-1H-naphtho~2,3-c]pyran (9a = semi-xanthomegnin). -9c, prepared by demethylation of 9a, was dimerized by potassium peroxydisulfate,'sodium hydroxide and methylated, giving 16b. Synthetic 16b and authentic xanthomegnin were identical, which verifies the structure revised by Hofle and Riiser 9' and shows, that the dyes of the xanthomegnin series are biogenetically homogenous. -The metabolic products of the strain inhibit the growth of bacteria. 16b and 26 furthermore have an effect against insects.Der Pilzstamm Tii 553 (Penicillium citreo-viride) produziert ein Gemisch gelber und gelbroter Farbstoffe, die antibiotische Aktivitat zeigen und auf Insekten fraljhemmend wirken. Das fettfreie Farbstoff-Rohprodukt, das durch Aceton-Extraktion des Mycels und Petrolether-Fallung des oligen Eindampfriickstandes gewonnen wurde, trennte sich beim wiederholten Chromatographieren an Oxalsaure-Kieselgel in sechs Komponenten

An unknown antibiotic, ophiocordin, C21H22N2O8, MW: 430, was isolated from submerged cultures of Cordyceps ophioglossoides, strain TU 276, grown in a glycerol soybean meal medium at 27 degrees C. The antibiotic was extracted from acidified culture fluids with n-butanol and purified by subsequent column chromatography on DEAE-Sephadex and cellulose. Studies including nuclear magnetic resonance and mass spectrometry resulted in proposals of partial structures of the molecule. Inhibition by ophiocordin could be demonstrated for a small number of fungi belonging to different taxonomic groups. Bacteria were not inhibited. The antifungal effect was antagonized by ammonia and nitrate ions and by certain amino acids.

The widely uscd and wcll-known bacterial strain Bucillrcs suh/i/is ATCC 6633 was found to produce two novel. antifungal hydrophilic pcptidc anlibiotics, 1.-arginyl-~-2-amino-5-phosphono-3-cis-pcnlenoic acid (I.-A~~-L-APPA, rhizocticin A) and r-valyl-L~irgit~yI-~-2-amino-5-phosphono-3-ci.~-pentenoic acid (L-Val-rArp-L-APPA. rhizocticin B). Besides rhizocticin A and B. the main components. small amounts of related tripeptidcs were detcctcd. Instead of the t-Val of rhizocticin B thcy contain r-Ile or I.-LCU and are rcfcrred to as rhizocticins C and D, respectively. The C-tcrminal rcsiduc was idcntificd by NMR spectroscopy as the unsaturated phosphono amino acid L-APPA. known till now only as D cnantiomcr. Enzymatic cleavages of rhizocticin B yielded both I.-APPA and rhizocticin A.In cultures of Bacillus subtilis ATCC 6633, known to produce the antibacterial antibiotic subtilin'), Michener and Snell" demonstrated two antifungal activities referred to as "aspergillus factor" and "rhizoctonia factor". No chemical structures were then implicated, but during a preliminary study on hydrophilic antibiotics from strains of Bacillus subtilis it became obvious that s u b~p o r i n s~~ and aspergillus factor are based on the same activity and that the bacterial strain used in both investigation^^,^) was the same, although the latter authors3) had designated it as "PCI 219".In the investigation described here, the structures of the antibiotics representing the rhizoctonia factor have been elucidated, and the name "rhizocticins" applied to them. Chro- Rbizocticine -ncue Phosphono-Oligopeptide mit antifungi.wher matographic and spectroscopic analyses of the highly hydrophilic metabolites revealed them to be formerly unknown antifungal di-and tripeptides, possessing the unusual Cterminal amino acid ~-2-amino-5-phosphono-3-cis-pentenoic acid (L-APPA, Scheme 1). APPA has been described as a part of the molecule of the plumbemycins, threonine antagonists from Streptomyces plumbeus4). However, D configuration of APPA was assumed, whereas rhizocticins contain L-APPA. Plumbemycins are active against bacteria, whereas rhizocticins show antifungal effects. Fermentation and Isolation of RhizocticinsRhizocticins can be detected in complex culture media (yeast extract, soy bean flour) of Bacillus subtilis ATCC 6633. During optimization experiments the complex medium was replaced by a chemically defined production medium. The change of the nutrient medium resulted in advantages for the detection and purification of the antibiotics. The fermentation broth was harvested after 72 h of incubation in 5-1 and 25-1 fermenter vessels at 27 "C. Preliminary examination had indicated that two naturally occurring components of the antibiotic, rhizocticins A and B, exist. They are of low molecular mass, hydrophilic, amphoteric, highly charged, and could not be extracted from the culture broth with organic solvents.After acidification, the culture broth was centrifuged to remove the bacterial growth together with precipitated components (Scheme 2). Other ...

Volumen XLV, Fasciculus 111 (1962) -No. 98-99 839 Ausgangsmenge betrugen. Die auf diese Art gewonnene Essigsaure enthielt kein Acetanhydrid und ihr Wassergehalt, der durch Bestimmen des Schmelzpunktes auf & 0,Ol Gew.-Proz. genau ermittelt w~r d e *~) , war durchwegs kleiner als 0,l Gew.-Proz.Zur Durchfuhrung der vorliegenden Arbeit standen Mittel aus der ROBERT GNEHM-StiftUng und dem Schulmittelfonds der ETH zur Verfiigung, wofiir wir aufrichtig danken. SUMMARYThe glass electrode is shown to respond to the concentration of protonated acetic acid within the ranges 0 to 0.50% (w/w) water and about to 1 0 -l~ perchloric acid. There exists a linear relationship between the alkaline error of glass electrodes and the logarithm of their potential drift in acetic acid. Some electrode glasses that are specially well suited for measurements in acetic acid are mentioned. Die GattungMyrothecium TODE ex FRIES gehort zu den Fungi imfierfecti, Ordnung Moniliales, Familie Tuberculariaceae. Mehrere Arten auf pflanzlichen Substraten sind beschrieben worden. Myrothecium roriduum TODE ex FR., Gattungstypus bei FRIES 2, 3), und, in unseren Versuchen sogar haufiger, Myrotkecium verrzlcaria (AL-BERTINI et SCHWEINITZ) DITMAR ex FRIES wurden auch aus dem Erdboden isoliert4).Schon seit langerer Zeit ist bekannt, dass Kulturen der beiden erwahnten Arten cellulolytische Eigenschaften besitzen, die fur die Vorbehandlung der Cellulose bei der Herstellung von Textilien nutzbar gemacht werden konnen5). Die fur diesen enzymatischen Abbau der Cellulose verantwortliche Cellulase ist von zahlreichen Autoren eingehend untersucht wordenb).Die Unterscheidung von Species innerhalb der Gattung scheint oftmals Schwierigkeiten bereitet zu haben, und auch die Gattungszugehorigkeit untersuchter Stamme wurde nicht immer erkannt. So isolierten BRIAN & MCGOWAN~) erstmals eine l) Jetzige Adresse :