Abstract
Litophyton savignyi of the Red Sea is one of the underexplored soft corals that is a prolific producer of bioactive metabolites. The anti-inflammatory activity of the hexane fraction of L. savignyi was superior to that of the methanol fraction, as revealed by reducing superoxide anion generation (32.05% ± 8.06%) and elastase release in fMLF/CB-induced human neutrophils (96.89% ± 3.69%). The bioactive non-polar fraction was subjected to systematic chemical investigation using UPLC-qToF-MS/MS analysis, and 650 metabolites were found to be exclusively present in the hexane part, of which steroids were the main class representing 26% of all features. Further fractionation of the hexane extract revealed that the polar subfractions ND90-1 and ND90-3 displayed the best inhibition of superoxide generation (96.79% ± 0.16% and 94.27% ± 4.12%, respectively) and elastase release (92.35% ± 2.75% and 91.28% ± 3.41%, respectively). Bioactive molecular networking and multivariate analysis showed that bioactive nodes (r > 0.65 and p < 0.05) and metabolites with top VIP scores (>2) belonged to steroids, sesquiterpenes, fatty amides, and sphingolipids. Network pharmacological studies on these metabolites showed that their hub targets were SRC, PTGS2 (COX-2), HSP90AA1, PPARG, and HIF1A, and they were significantly enriched in inflammatory responses and nuclear receptor-mediated steroid hormone signaling pathways. Molecular docking on COX-2 showed steroids to display scores comparable to those of celecoxib and indomethacin (COX-2 inhibitors as controls), which highlighted the promising anti-inflammatory potential of the hexane extract of Litophyton savignyi. Quantitative real-time PCR (qPCR) analysis revealed that the bioactive subfractions reduced the expression of the proinflammatory cytokines IL-6 and IL-1β, hence supporting the involvement of COX-2 mediated anti-inflammatory effect. A detailed analysis on its anti-inflammatory potential requires future in vivo investigations.