Discovery of potent anti-inflammatory flavonoid-sesquiterpenoid hybrids from Syzygium simile: Structure–activity relationships and nanomolar inhibition of human neutrophil activation

Current anti-inflammatory drugs often cause significant adverse effects, underscoring the need for safer alternatives. Natural products with rare and distinctive scaffolds represent a promising source of leads for discovering new anti-inflammatory agents. This study aimed to find anti-inflammatory flavonoid-sesquiterpenoid hybrids from Syzygium simile (Myrtaceae), and to clarify the structural-activity relationship underlying their inhibition of biomarkers in activated human neutrophils. A range of chromatographic techniques was sequentially applied to isolate pure compounds, the structures of which were elucidated by spectroscopic and spectrometric analyses. Anti-inflammatory activities were evaluated in fMLP/CB-induced human neutrophils by measuring inhibitory effects on superoxide anion generation and elastase release. Twenty-two flavonoid-sesquiterpenoid hybrids, including thirteen previously undescribed structures, were isolated from S. simile leaves, comprising five flavonoid scaffolds (types I–V) and five sesquiterpenoid moieties (germacrene-, cubebene-, oplopene-, cadinene-, and copaene-types) generated via distinctive electron transfer-mediated biosynthesis. Most compounds potently inhibited superoxide anion generation and elastase release, with IC50 values up to 32-fold lower than that of the positive control, without exhibiting cytotoxicity. Structure-activity relationship analysis revealed that the combination of a type I flavonoid and a germacrene-type sesquiterpenoid skeleton, integrated through an rA-type ring fusion, represents the optimal pharmacophore. The lead compound, simisyzygin C (3), achieved nanomolar potency with IC50 values of 0.04 μM (superoxide anion) and 0.16 μM (elastase release). These findings identify flavonoid-sesquiterpenoid hybrids as a structurally distinctive class of anti-inflammatory natural products, providing a structural basis for lead optimization.