Isolation and characterization of antibacterial compounds from marine organisms

Abstract

Marine bioprospecting is a promising field of research that has gained interest worldwide for the past few decades, and can be characterised as a systematic and purpose oriented search for bioactive compounds or genetic material from marine sources. Several compounds have been isolated and some have found their way into the commercial market. The main aim of this master thesis was to investigate the antibacterial activity of bioactive compounds in the extracts of the marine ascidian Halocynthia pyriformis. Two other marine organisms Metridium senile and Parastichopus tremulus were also screened for antibacterial activity, but they were purified in two other parallel projects. Aqueous and organic extracts from H. pyriformis were purified using preparative HPLC and screened for antibacterial activity. The antibacterial activity was mostly found in the organic fractions against the Gram-positive bacteria E.faecalis, S. aureus and MRSA. The active fractions from the primary screening were subjected to a secondary screening where they did not display any clear activity. Three of the fractions showed some activity and was pooled and refractionated and later subjected to a third screening. The active fractions were analysed by ESI-TOF and ESI-Q without obtaining any results. A dose-response test was carried out to investigate the potency of the raw-extract. Due to weak results it was decided to fractionate the organic extract of H. pyriformis once again before it was subjected to a new primary screening. This time the fractions were concentrated and three of them displayed antibacterial activity and were subjected to MS-analyses in positive and negative mode. The results from the MS-analyses indicated that one of the fractions contained two active compounds with the m/z values 269.2094 and 505.4248. The calculated elemental composition proposed the empirical formula C16H28O3 for the molecular weight 268.2021. A search in databases containing known bioactive compounds proposed four different compounds with the same empirical formula. The elemental composition of the m/z value 505.4248, proposed two empirical formulas C32H56O4 and C31H54N4. None of these two formulas gave any hits in the database over known bioactive compounds. This could indicate that this is a novel compound. It was impossible to know the identity of these compounds without analysing them in NMR. NMR analyses require large amounts of pure compounds which was difficult to obtain in this thesis.