GC–MS profiling and antivirulence potential of limonene-rich Citrus medica cv. ‘Rugosa’ essential oil: in vitro and in silico evaluations

Abstract

Antimicrobial therapy is a primary approach for treating microbial infections, yet increasing antibiotic resistance continues to hinder effective disease management, highlighting natural products as promising alternative sources of novel anti-infective agents. In this context, Citrus medica cv. ‘rugosa’ EO was chemically profiled by GC-MS, and its antimicrobial, antibiofilm, and anti-quorum sensing inhibitory action was firstly tested using both in vitro and computational studies. C. medica cv ‘‘rugosa ‘’ EO revealed a predominance of monoterpene hydrocarbons (79.12%) with limonene (76.15%) as the major component, along with minor compounds such as limonene oxide (3.32%), trans-carvyl acetate (1.93%), nerol (1.75%), cis-carveol (1.37%), myrcene (1.31%), carvacrol (1.14%), and β-bisabolene (1.10%). The EO exhibited potent broad-spectrum antimicrobial activity, with inhibition zones up to 30.00 ± 1.73 mm, surpassing limonene (6.00–25.00 mm) and ampicillin (6.00–8.67 mm), and antifungal effects (12.33–14.33 mm) comparable to amphotericin B (11.67–15.33 mm). MICs values ranged from 0.097–1.562 mg/mL with MBC/MFC ratios mostly 2–8, indicating strong bactericidal and fungicidal activity. The EO also demonstrated dose-dependent antibiofilm inhibition (B. subtilis 84.57%; P. aeruginosa 43.19%), quorum-sensing suppression in C. violaceum (100% at MIC; 62.70% at MIC/2; 34.78% at MIC/4) and reduced swarming motility (77.17–46.74%), highlighting its anti-virulence potential. Computational analyses of C. medica cv. ‘rugosa’ EO phytocompounds, particularly valencene (− 9.3 kcal/mol) and aromadendrene (− 8.3 kcal/mol), showed strong binding affinities and stable 200 ns MD interactions, along with favorable ADMET profiles, confirming their drug-like potential. These results indicate that phytochemicals from these medicinal plants possess promising anti-infective and anti-virulence properties and merit further investigation for therapeutic applications.