Phytochemical characterization and evaluation of antimicrobial activity of Eugenia sp.: antibacterial, antibiofilm and ampicillin-potentiating activities against resistant Staphylococcus aureus and Staphylococcus epidermidis

This study investigated the phytochemical composition, antibacterial, antibiofilm, synergistic, and cytotoxic effects of hydroethanolic extracts and fractions from Eugenia astringens and Eugenia mattosii . HPLC–ESI-MS/MS analysis revealed the presence of nine compounds were identified in E. astringens , which was characterized by a high content of gallic acid (542.4 ± 8.2 μg g -1 ), whereas E. mattosii contained eleven identified compounds, with epicatechin (533.2 ± 90.4 μg g -1 ) and catechin (359.8 ± 10.8 μg g -1 ) as major compounds. Gallic acid, protocatechuic acid, syringic acid, p -coumaric acid, scopoletin, p -anisic acid, abscisic acid, quercetin, and galangin were identified for the first time in E. astringens , while epicatechin, vanillic acid, and ferulic acid were reported for the first time in E. mattosii . The extracts exhibited activity against Gram-positive bacteria, with E. astringens showing the highest potency (MIC = 15.62 μg mL -1 against Staphylococcus aureus and MRSA; 7.81 μg mL -1 against Staphylococcus epidermidis ), whereas E. mattosii showed MIC values of 125 μg mL -1 against all strains. No relevant activity was observed against the Gram-negative bacteria tested. Active extracts and fractions inhibited biofilm formation at 7.81–15.62 μg mL -1 , while mature biofilm eradication was observed mainly against MRSA, with EBH ≥125 μg mL -1 and DCM ≥62.5 μg mL -1 . Growth kinetics confirmed sustained inhibition by selected fractions. Checkerboard assays revealed synergistic interactions between ampicillin and E. astringens against MRSA, (fractional inhibitory concentration index (FICI) = 0.24–0.28), and against S. epidermidis (FICI = 0.35–0.36). These findings are relevant because potentiation of ampicillin activity may enhance β-lactam efficacy against resistant Staphylococcus . SYTOX green assays demonstrated that the active samples, especially DCM from E. astringens and FAE from E. mattosii increased bacterial membrane permeability, suggesting membrane disruption as one of the mechanisms underlying their antibacterial activity. Cytotoxicity assays in RAW 264.7 macrophages demonstrated low toxicity at MIC concentrations. Collectively, the potent anti-staphylococcal activity, inhibition of biofilm formation, synergistic interactions with ampicillin, membrane permeabilization, and low cytotoxicity at MIC concentrations highlight E. astringens as a promising source of antibacterial compounds and antibiotic adjuvants. These findings support further investigation of E. astringens against resistant Gram-positive pathogens.