Phytochemical composition and synergistic antimicrobial effects of Rosmarinus officinalis essential oils during flowering in an arid mediterranean region

Rosmarinus officinalis has attracted significant attention due to its broad-spectrum antimicrobial activity, largely attributed to its bioactive essential oils (EOs). Several studies indicate that the flowering period is crucial for harvesting rosemary’s aerial parts for optimal EO extraction. However, its prolonged flowering period complicates the determination of an optimal harvest time, potentially affecting yield, chemical composition, and efficacy. This study provides, for the first time, a systematic month-by-month evaluation of EO yield, chemical composition, and synergistic antimicrobial potential of rosemary cultivated under arid Mediterranean conditions during its flowering period (September to March). EO samples were analyzed by GC-MS and assessed for antimicrobial activity against clinically relevant pathogens, including S. aureus, E. coli, S. enterica, and four Candida species. The synergistic potential was further evaluated with two conventional antimicrobials (streptomycin and amphotericin B). The results showed that the EO yields ranged from 1.73% to 2.75%, with a clear peak in autumn. GC-MS analysis identified 31 compounds, dominated by 1,8-cineole (27.57 ± 0.76%–36.28 ± 0.26%), α-pinene (15.36 ± 0.23%–28.97 ± 0.10%), and camphor (7.12 ± 0.00%–15.37 ± 0.12%), confirming the prevalence of the 1,8-cineole/α-pinene/camphor chemotype. Antimicrobial assays demonstrated stronger activity against fungal strains, particularly C. krusei and C. albicans, with enhanced efficacy observed in EOs collected in October-November. Synergy assays showed significant potentiation of streptomycin activity, particularly against E. coli in autumn, with up to a 32-fold increase in efficacy. In contrast, only limited synergistic effects were observed with amphotericin B. Overall, our findings emphasize the clinical relevance of optimizing harvest timing, as autumn-harvested rosemary EOs exhibit the greatest potential as natural antibiotic adjuvants against multidrug-resistant pathogens. Nevertheless, the partial antagonism with amphotericin B highlights the need for strain-specific compatibility assessments to avoid compromising drug efficacy in combinatory therapies.