The increasing prevalence of antibiotic-resistant bacteria highlights the urgent need for alternative antimicrobial agents derived from natural sources. In this study, we investigated the organ-specific phytochemical composition and antibacterial potential of hydroethanolic extracts obtained from flowers, leaves, and stems of Ebenus pinnata Aiton, an endemic Moroccan species. Comprehensive metabolite profiling was performed using UHPLC–ESI–MS, while total phenolic (TPC), flavonoid (TFC), and proanthocyanidin (TPrC) contents were quantified spectrophotometrically. Antibacterial activity was evaluated using agar diffusion, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) assays against representative Gram-positive and Gram-negative bacteria.

UHPLC–ESI–MS analysis revealed pronounced organ-specific chemical signatures. Flowers were enriched in flavanols, particularly epigallocatechin, alongside flavonoids and phenolic glycosides. Leaves were dominated by glycosylated flavonoids and hydroxycinnamic acids, whereas stems accumulated diverse phenolic acids, proanthocyanidins, and triterpenoids, including ursolic acid. Quantitative assays confirmed significant inter-organ variability, with stems exhibiting the highest TPC, TFC, and TPrC values, followed by leaves and flowers. Antibacterial assays demonstrated a clear selectivity toward Gram-positive bacteria. Flower and stem extracts showed the strongest activity, particularly against Micrococcus luteus and Staphylococcus aureus, with low MIC values (2.60–5.20 mg/mL) and bactericidal effects (MBC/MIC ≤ 2.50). In contrast, Gram-negative strains were markedly less susceptible.

Antibacterial efficacy correlated more strongly with the qualitative composition of phenolic constituents, especially flavanols and proanthocyanidins, than with total phenolic abundance. These findings highlight the importance of organ-specific phytochemical allocation in shaping antimicrobial activity and position E. pinnata, particularly its floral and stem tissues, as a promising source of bioactive phenolics with potential relevance for the development of plant-derived antibacterial agents.