Isolation Characterization, Synthesis, and Biological Applications of Novel 2-methoxylated Insaturated Fatty Acids of Marine Origin
Montano Morales, Nashbly
Carballeira, Nestor M. (Consejero)
The discovery of new bioactive compounds in nature is an active area of research. Marine-‐derived biologically active natural products continue to be a source of potential therapeutic compounds as well as a motivation to develop new synthetic strategies to construct them. We report the identification and characterization of a group of more than 40 phospholipids and free fatty acids from the Caribbean sponge Asteropus niger using mass spectrometry, circular dichroism (CD), magnetic resonance studies, and chain length correlations to determine the structures. Some of these fatty acids were unprecedented in nature, such as the (2R,5Z,9Z,13Z)-‐2-‐methoxyoctadeca-‐5,9,13-‐trienoic acid, (2R,5Z,9Z)-‐2-‐ methoxy-‐25-‐methylhexacosa-‐5,9-‐dienoic acid, (2R,5Z,9Z)-‐2-‐methoxy-‐24-‐methylhexacosa-‐5,9-‐ dienoic acid, (2R,5Z,9Z)-‐2-‐methoxyoctacosa-‐5,9-‐dienoic acid, (2R,5Z,9Z)-‐2-‐methoxy-‐26-‐ methylheptacosa-‐5,9-‐dienoic acid, and the (2R,5Z,9Z)-‐2-‐methoxy-‐27-‐methyloctacosa-‐5,9-‐dienoic acid. An improved synthetic methodology for the preparation of the 2-‐methoxylated unsaturated fatty acids was used to prepare two 2-‐methoxylated Δ5,9-‐diunsatruated fatty acids, namely the (5Z,9Z)-‐(±)-‐2-‐methoxy-‐5,9-‐eicosadienoic acid and the (±)-‐2-‐methoxy-‐5,9-‐ eicosadiynoic acid, in seven and eight steps with overall yields of 6% and 11%, respectively. Five 2-‐methoxylated monounsaturated fatty acids were prepared, including the (5Z)-‐(±)-‐2-‐methoxy-‐ 5-‐eicosenoic acid, (±)-‐2-‐methoxy-‐5-‐eicosynoic acid, (±)-‐2-‐methoxy-‐9-‐eicosynoic acid, (6Z)-‐(±)-‐2-‐ methoxy-‐6-‐octadecenoic acid, and the (±)-‐2-‐methoxy-‐6-‐octadecynoic acid in seven to eight steps with overall yields between 6-‐20%. With the synthesis of the (5Z,9Z)-‐(±)-‐2-‐methoxy-‐5,9-‐ eicosadienoic acid we were able to corroborate the structure of the natural compound by gas chromatographic co-‐injection of the methyl ester with the whole fatty acid methyl ester (FAME) mixture from the sponge A. niger. The inhibition of the relaxation of Topoisomerase IB (TopIB) by a mixture of 2-‐ methoxylated fatty acids mainly composed of the (2R,5Z,9Z)-‐2-‐methoxy-‐25-‐methylhexacosa-‐5,9-‐ dienoic acid and (2R,5Z,9Z)-‐2-‐methoxy-‐24-‐methylhexacosa-‐5,9-‐dienoic acid, isolated from A. niger, was studied displaying inhibitory concentrations at an EC50 of ~4.3 μM for the human TopIB and an EC50 of ~11.7 μM for the Leishmania TopIB (LdTopIB). These results are compared to short-‐chain fatty acids, such as the (±)-‐2-‐methoxy-‐5,9-‐eicosadienoic acid, and the (±)-‐2-‐ methoxy-‐6-‐octadecenoic acid, that only inhibited the LdTopIB at concentrations of 22-‐31 μM and 89-‐90 μM, respectively, while neither of the latter acids significantly inhibited hTopIB (EC50 > 100 μM). These short-‐chain acids were toxic against L. infantum promastigotes (EC50 = 237-‐250 μM), as compared to the very long-‐chain 2-‐methoxylated unsaturated fatty acids (EC50 ~ 776 μM). The antibacterial activity of the 2-‐methoxylated unsaturated fatty acids against multi-‐ drug resistant bacteria was studied and the (±)-‐2-‐methoxy-‐6-‐octadecynoic acid inhibited the methicillin-‐resistant S. aureus (MRSA) with an IC50 of 17 μg/ml. The long-‐chain C20 2-‐ methoxylated fatty acids did not inhibit the studied bacterial strains. Our results indicated that the formation of micelles was responsible for their antibacterial activity. It was found that 2-‐ methoxylation reduces the concentration where micelles are formed as compared to the corresponding non-‐methoxylated fatty acids.