Objective To establish a method for determining the content of luteolin in Humulus scandens (Lour.) Merr. from Anhui province, and explore its mechanism against Escherichia coli in vitro.

Methods The content of luteolin was determined by reversed-phase high-performance liquid chromatography (RP-HPLC) using a ZORBAX SB-C₁₈ column (250 mm×4.6 mm, 5 μm), with a mobile phase of acetonitrile (A)-0.2% phosphoric acid solution (B) in gradient elution mode. The flow rate was 1.0 mL/min, the column temperature was maintained at 30 ℃, and the detection wavelength was set at 350 nm. The antimicrobial activity of luteolin from Humulus scandens (Lour.) Merr. extract was assessed using the Oxford cup method and the microdilution method. Changes of morphology in cell surface and intracellular ultrastructure were observed by transmission electron microscopy. The antimicrobial mechanism of luteolin was investigated through prokaryotic transcriptome analysis.

Results When the concentration of luteolin was in the range of 5.65-45.20 μg/mL, it exhibited a linear relationship with the peak area. The average recovery rate was 97.63% (RSD=1.08%, n=6). Luteolin demonstrated inhibitory effects against Escherichia coli, induced the lysis of bacterial capsules and pili, while caused organelle dissolution with vacuole-like distribution. Furthermore, luteolin was found to affect multiple metabolic pathways in Escherichia coli, including amide biosynthesis, ribosomal structure composition, organelle components, oxidative phosphorylation, fatty acid biosynthesis, and nucleotide metabolism.

Conclusions RP-HPLC is simple, feasible, accurate, and sensitive, and can be used for the detection of luteolin content in Humulus scandens (Lour.) Merr. Additionally, luteolin exhibites significantly inhibitory effects on the growth of Escherichia coli, a Gram-negative bacterium. These findings provide a theoretical foundation for the potential application of luteolin derived from Humulus scandens (Lour.) Merr. extract.