LC-Q-Orbitrap HRMS profiling and functional characterization of Ocimum basilicum L. var. basilicum extract as an ampicillin-modulating agent against Escherichia coli

Plant-derived polyphenol-rich extracts are of interest as resistance-modifying agents because they can influence bacterial redox balance, membrane-associated bioenergetics, and antibiotic susceptibility. In this study, we evaluated the antibiotic-modulatory potential of an ethanol extract of purple-leaved Ocimum basilicum L. var. basilicum (OBEE) against Escherichia coli NM111, a katG::lacZ reporter strain derived from E. coli K-12. LC-Q-Orbitrap HRMS tentatively annotated 102 constituents in OBEE. Hydroxycinnamic acids and their derivatives were the largest group (32 compounds), followed by flavonoids (29 compounds); prominent constituents included rosmarinic acid, chicoric acid, sagerinic acid, salvianolic acid K, caftaric acid, and quercetin, luteolin, and cyanidin derivatives. OBEE has high total phenolic content (317.75 ± 4.11 µg gallic acid equivalents mg⁻¹ dry extract) and showed strong DPPH radical-scavenging activity with an IC 50 of 19.37 ± 0.38 µg mL⁻¹, close to that of catechin, with rosmarinic acid identified as the dominant antioxidant compound. Although OBEE did not exhibit direct antibacterial activity against E. coli NM111 up to 2 mg mL⁻¹, it potentiated ampicillin at non-inhibitory concentrations, reducing the ampicillin MIC two-fold (modulation factor ≥ 2) and showing synergistic interaction (FICI < 0.5). No synergy was observed with kanamycin. The OBEE–ampicillin combination reduced colony-forming ability by ~ 65%, decreased the specific growth rate ~ 2-fold, prolonged the generation time, and markedly suppressed proton flux (~ 2.1-fold) and total ATPase activity (~ 4.2-fold). OBEE also increased SOD (~ 12%) and catalase activities and induced katG::lacZ expression. These findings identify OBEE as an antibiotic-modulating extract and support impaired bacterial bioenergetics as a mechanistic hypothesis; confirmation will require active-compound isolation and direct target validation.