Studies of highly N-methylated cyclodepsipeptides, the aureobasidins

Submission note: A thesis submitted in total fulfilment of the requirements for the degree of Doctor of Philosophy to the School of Molecular Sciences, Faculty of Science, Technology and Engineering, La Trobe University, Bundoora.

Studies on the synthesis of a class of highly N-methylated cyclodepsipeptides, the aureobasidins, have been undertaken. Aureobasidins are important cyclic depsipeptides because of their antifungal properties and potential medicinal application. However, these compounds are difficult to obtain either by isolation or total synthesis. This work aimed to develop a new efficient method to synthesise aureobasidin derivatives through a combination of solution- and solid-phase peptide synthesis, where [2S,3S-Hmp]- aureobasidin L was selected as the model compound to develop synthetic techniques due to the ready availability of residues. [2S,3S-Hmp]-Aureobasidin L has been successfully synthesised through a combination of solution- and solid-phase peptide synthesis. All of the Fmoc-protected residues including a depsidipeptide, Fmoc-MeVal-Hmp-OH, were prepared in solution phase. These residues were chain elongated on chlorotrityl resin using selected coupling reagents including HBTU/HOBt, HATU/HOAt and BTC/collidine. The latter coupling reagent was found to be the best agent for the attachment of N-methylated residues and also the depsidipeptide. Cyclisation was carried out in solution using HATU in dichloromethane. The desired cyclic product was obtained after the purification using normal-phase flash chromatography. ESI-MS and NMR were used to confirm the compound. The structure of [2S,3S-Hmp]-aureobasidin L was assigned using 1D- and 2D-NMR (DEPT 135, COSY, HSQC, HMBC and NOESY) and comparison with the spectra of aureobasidin A (conformer A and B) and aureobasidin E. The two conformers which were found to be present in deuterated chloroform were successfully determined. The conformation of the amide bonds was confirmed by further 2D-NMR analysis using NOESY. The successful method developed for the synthesis of [2S,3S-Hmp]-aureobasidin L was then applied to aureobasidin L. Almost all steps were applicable to the new compound, except the cleavage step of the linear nonadepsipeptide which resulted in several problematic by-products which were unable to be completely removed by HPLC purification. The cyclisation of the impure crude linear peptide gave material which also proved very difficult to purify. The small amount of impure aureobasidin L obtained was subjected to ESI-MS analysis, giving the correct molecular ion. A1 H-NMR spectra of the impure cyclic product showed a similar profile to the [2S,3S-Hmp]-AbL spectra. In the last part of this work, simpler aureobasidin analogues were designed in order to make their synthesis more straightforward but to retain activity. The synthesis of these compounds was then developed. These compounds included both N-methylated cyclononapeptides and smaller N-methylated cycloheptapeptides. This project has successfully developed methods which can be applied to the synthesis and purification of aureobasidins but also other N-methylated cyclodepsipeptides and Nmethylated cyclopeptides.