Isolation and characterization of vitilevuamide from the ascidians Didemnum cuculliferum and Polysyncraton lithostrotum

This dissertation was oriented towards the isolation and structure elucidation of novel biologically active metabolites from marine organisms collected in the South Pacific. In this investigation I focused on the characterization and biological evaluation of vitilevuamide, a novel bicyclic depsipeptide isolated from two Fijian ascidians, Polysyncraton lithostrotum and Didemnun cuculliferum. The first chapter reviews cyclic peptides of marine origin along with their biological activity. This provides an overview of the types of peptides isolated from marine sources along with their structural uniqueness and similarities. The subsequent chapters deal with different aspects of the chemical and biological characterization of vitilevuamide. The alcohol extract of the ascidians was fractionated to yield vitilevuamide which displayed potent cytotoxic activity toward the human colon tumor cell line HCT 116. The structure of this peptide was elucidated primarily using nuclear magnetic resonance (NMR) spectroscopic techniques and tandem mass spectrometry (MS/MS). The peptide was partially linearized using alcoholic ammonia in order to facilitate analysis by tandem mass spectrometry. Chapter 3 deals with the synthesis of two not commercially available amino acids. The first is homoisoleucine (Hil) and the second N-methyl methoxinine (Nmm), a novel amino acid. The synthesis of Hil was successfully completed according to reported procedure. In the case of Nmm, several synthetic routes were attempted prior to successful synthesis of the molecule. Chapter 4 deals with stereospecific assignments of the amino acids. Their absolute configurations were determined by HPLC analysis of derivatized constituent amino acids obtained from acid hydrolysis of the peptide. New conditions were determined for the unambiguous assignment of isoleucine (He), Nmm and Hil. Liquid chromatography mass spectrometry (LCMS) was utilized in order to verify the stability of lanthionine (Lan) prior to assignment of stereochemistry. The biological profile of vitilevuamide is discussed in Chapter 5. The cytotoxicity of the peptide was evaluated in a panel of six human tumor cell lines. Vitilevuamide also scored positive in an in-house mechanism based antitumor screen geared towards the detection of tubulin interactive compounds.