Chemical Signals of Tropical Frogs

Die Dissertation befasst sich mit der Identifizierung und Totalsynthese von flüchtigen Signalstoffen aus den Madagaskarfröschen. Die Männchen der Unterfamilie Mantellinae bilden an der Unterseite ihrer Hinterschenkel Femoraldrüsen aus. Diese wurden entfernt, mit Dichlormethan extrahiert und anschließend mit GC/MS- und NMR-Methoden analysiert. Die identifizierten Strukturen wurden anschließend durch Totalsynthese überprüft und teilweise auf ihre Wirkung als Pheromon getestet. So wurden im Femoraldrüsenextrakt der Spezies Mantidactylus multiplicatus zwei unbekannte flüchtige Verbindungen durch GC/MS Analyse nachgewiesen. Für beide Verbindungen wurden Strukturvorschläge erarbeitet und durch den Vergleich mit synthetischem Material bestätigt. Nach der Synthese aller Enantiomere wurde die absolute Konfiguration durch chirale Gaschromatographie als (R)-8-Methylnonan-2-ol und (S)- Phoracantholid J bestimmt. Abschließend konnte in einem auf Madagaskar durchgeführten Biotest eine Bioaktivität der beiden Substanzen nachgewiesen werden. Eine Analyse der Drüsenextrakte der Spezies Mantidactylus femoralis zeigte einen unbekannten Naturstoff mit der Masse 184. Durch Synthese konnte die Struktur als (5R,10S)-5,10-Dimethyloxecan-2-on bestimmt werden. Der Femoraldrüsenextrakt von Gephyromantis boulengeri bestand hauptsächlich aus einer unbekannten Verbindung mit der Molekülmasse 224. Die Struktur konnte durch 2D-NMR Experimente aufgeklärt werden. Einer Synthese aller Enantiomere des (E)-3,7,11-Trimethyloxacyclododec-7-en-2-ons folgte die Bestimmung der absoluten Konfiguration, welche zeigte, dass der als Gephyromantolid A benannte neue Naturstoff in der (2S,6E,10R)-Konfiguration vorliegt. Neben diesen durch Totalsynthese identifizierten Naturstoffen wurden in über 150 Drüsenextrakten von über 35 verschiedenen Arten insgesamt über 250 Verbindungen nachgewiesen. Ungefähr die Hälfte dieser Verbindungen ist bis dato unbekannt.The thesis deals with the identification and total synthesis of volatile signal compounds isolated from the Mantellid frogs from Madagaskar. Males of the subfamily Mantellinae possess femoral glands on the ventral sides of their shanks. To reveal the composition of the gland constituents, the glands were excised, extracted with Dichloromethane and analyzed with GC/MS- and NMR-Methods. The identified structures were subsequently synthesized and tested for bioactivity. In the femoral gland extracts of Mantidactylus multiplicatus two unknown volatile compounds were detected. Both structures were elucidated using mass-spectrometric data and confirmed by synthesis. After the successful synthesis of all enantiomers, the absolute configuration was established as (R)-8-Methylnonan-2-ol und (S)-Phoracantholid J using chiral gas chromatography. Finally both compounds were tested for bioactivity in a bioassay carried out in Madagascar. Both compounds showed a significant bioactivity and are therefore the first volatile amphibian pheromones reported so far. The analysis of the femoral gland extracts of Mantidactylus femoralis revealed a new natural compound with the mass of 184. The absolute configuration of this new natural compound was determined as (5R,10S)-5,10-Dimethyloxecan-2-one. A single unknown compound with the mass of 224 could be detected in the femoral gland extract of Gephyromantis boulengeri, which was directly analyzed via 2D-NMR experiments. The synthesis of all enantiomers of (E)-3,7,11-Trimethyloxacyclododec-7-en-2-one was followed by the determination of the absolute configuration. It was shown, that the new natural compound, named Gephyromantolide A, has the (2S,6E,10R)-configuration. In addition to those identified compounds, more than 250 different compounds were detected in about 150 femoral gland extracts of 35 different species. The structures of about half of them are still unknown

Volatile amphibian pheromones: Macrolides from mantellid frogs from madagascar

Amphibians like water, but do they also notice volatile compounds in the air? Yes, they do. Macrolides, such as phoracantholide-J (see picture; upper right structure) or the newly discovered natural product gephyromantolide-A (left structure), are used for communication by mantelline frogs from Madagascar. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Macrolides and Alcohols as Scent Gland Constituents of the Madagascan Frog <i>Mantidactylus femoralis</i> and Their Intraspecific Diversity

Acoustic and, to a lesser degree, visual signals are the predominant means of signaling in frogs. Nevertheless, certain lineages such as the mantelline frogs from Madagascar use the chemical communication channel as well. Males possess femoral glands on the hind legs, which recently have been shown to contain volatile compounds used in communication as pheromones. Many mantelline species occur in sympatry, and so far species recognition is regarded to occur mainly by acoustic signals. The analysis of the gland constituents of <i>Mantidactylus femoralis</i> by GC/MS revealed the presence of volatile macrolides and secondary alcohols. The new natural products mantidactolides A (<b>4</b>) and B (<b>6</b>), as well as several methyl carbinols, were identified, and their structures were confirmed by synthesis. The analysis of individuals from different locations of Madagascar revealed the presence of two groups characterized by specific patterns of compounds. While one group contained the alcohols and mantidactolide B, the other showed specific presence of the macrolides phoracantholide I (<b>1</b>) and mantidactolide A (<b>4</b>). Genetic analysis of some individuals showed no congruence between genetic relatedness and gland constituents. Several other individuals from related species had different gland compositions. This suggests that a basic set of biosynthetic machinery might be available to a broader group of related species

Discovery of skin alkaloids in a miniaturized eleutherodactylid frog from Cuba

Four phylogenetically independent lineages of frogs are currently known to sequester lipid-soluble skin alkaloids for which a dietary source has been demonstrated. We report here a remarkable fifth such instance, in Eleutherodactylus iberia and Eleutherodactylus orientalis, two species of miniaturized frogs of the family Eleutherodactylidae from Cuba. Six pumiliotoxins and two indolizidines were found in E. iberia, one of the smallest frogs in the world and characterized by a contrasting colour pattern for which we hypothesize an aposematic function. Analyses of stomach content indicated a numerical prevalence of mites with an important proportion of oribatids—a group of arthropods known to contain one of the pumiliotoxins detected in E. iberia. This suggests that miniaturization and specialization to small prey may have favoured the acquisition of dietary skin alkaloids in these amphibians

Take time to smell the frogs: vocal sac glands of reed frogs (Anura: Hyperoliidae) contain species-specific chemical cocktails

Males of all reed frog species (Anura: Hyperoliidae) have a prominent, often colourful, gular patch on their vocal sac, which is particularly conspicuous once the vocal sac is inflated. Although the presence, shape, and form of the gular patch are well-known diagnostic characters for these frogs, its function remains unknown. By integrating biochemical and histological methods, we found strong evidence that the gular patch is a gland producing volatile compounds, which might be emitted while calling. Volatile compounds were confirmed by gas chromatography-mass spectrometry in the gular glands in 11 species of the hyperoliid genera Afrixalus, Heterixalus, Hyperolius, and Phlyctimantis. Comparing the gular gland contents of 17 specimens of four sympatric Hyperolius species yielded a large variety of 65 compounds in species-specific combinations. We suggest that reed frogs might use a complex combination of at least acoustic and chemical signals in species recognition and mate choice.(c) 2013 The Authors. Biological Journal of the Linnean Society published by John Wiley & Sons Ltd on behalf of The Linnean Society of London

Mass Spectrometry of Aliphatic Macrolides, Important Semiochemicals or Pheromones

Macrolides are a relatively common structural motif prevalent in Nature. However, the structures of these large ring lactones have been relatively difficult to elucidate via NMR spectroscopy due to the minute amounts of compounds that are sometimes obtainable from natural sources. Thus, GC-MS analysis of individual macrolactones has become the method of choice for the structural identification of these compounds. Here we discuss the mass spectrometric behavior of aliphatic macrolides, evaluating spectra from numerous compounds of various ring size, including derivatives containing methyl branches as well as double bonds. The specific fragmentation of these macrolactones under electron impact conditions allows for the development of a general rule set aimed at the identification of similar compounds by mass spectrometry. In addition, the mass spectra of dimethyl disulfide adducts of unsaturated macrolides are discussed. The mass spectra of almost 50 macrolides are presented