Phenolic compounds from Origanum majorana with biofilm-inhibitory activity against methicillin-resistant Staphylococcus aureus and Escherichia coli strains.

Context: Antibiotic resistance in bacteria is a growing global problem, with biofilm formation and efflux pumps playing crucial roles in this issue.

Objective: This study explores the effects of phenolic compounds of Origanum majorana against Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA) strains by inhibiting biofilm formation and efflux pumps.

Materials and methods: The methanolic extract of O. majorana was fractionated guided by an antibiofilm assay, and the active fractions were analyzed by multistep chromatographic separation to yield five pure compounds. Their structures were then determined using 1D and 2D nuclear magnetic resonance spectroscopy. The minimum inhibitory concentrations of the extracts, fractions, and isolated compounds were determined via the microdilution method in a 96-well plate. Antibiofilm activity was assessed using the crystal violet method, and the effect on efflux pumps was tested by a real-time ethidium bromide accumulation assay.

Results: Arbutin (1), apigenin 7-O-glucoside (2), 6'-caffeoylarbutin (3), rosmarinic acid (4), and 2-deoxy-d-1,4-ribonolactone (5) were isolated from the aqueous methanolic extract. Compounds 1, 2, and 4 reduced E. coli biofilm formation by 24.82%-42.98% at 100 µM, whereas only arbutin (1) moderately suppressed biofilm formation of MRSA (23.15 ± 1.56% at 50 µM). Arbutin also demonstrated efflux pump inhibitory activity against MRSA (relative fluorescence index of 0.49 at 100 µM).

Discussion and conclusions: The newly discovered natural antibiofilm agents show promise as candidates for treating biofilm-associated infections and combating antibiotic-resistant bacteria.