Bacterial neuraminidase inhibitory chalcones from flowers of Coreopsis lanceolata, their kinetic characterization and antibiofilm effect.

BACKGROUND: Bacteria within biofilms are thousand times more resistant to antibiotics. Neuraminidase is a crucial enzyme for bacterial adhesion and biofilm formation, it hydrolyzes glycosidic residue of glycoproteins, glycolipids, and oligosaccharides. Coreopsis lanceolata L. flowers may have a significant potential of bacterial neuraminidase (BNA) inhibition because of high natural abundance of chalcones. PURPOSE: The investigation of bacterial biofilm inhibitors has emerged as a novel therapeutic strategy against antibiotic resistance. Therefore, individual chalcones were isolated from C. lanceolata and their capacity to inhibit BNA and formation of Escherichia coli biofilm were evaluated. METHODS: Different chromatographic techniques were used to isolate the compounds (1-12). Enzyme inhibition and detailed kinetic behavior of compounds was determined by estimation of kinetic parameters (Michaelis-Menten constants (Km), maximum velocity (Vmax), dissociation constant for binding with the free enzyme (KI) and enzyme-substate complex (KIS)). Binding affinities (KSV) and binding modes of inhibitors were elucidated by fluorescence quenching and molecular docking, respectively. The natural abundance of chalcones was established through UPLC-Q-TOF/MS. The most potent inhibitor (1) was tested for its ability to inhibit the formation of E. coli biofilm, which was examined by crystal violet assay, scanning electron microscope (SEM) and confocal laser scanning microscope (CLSM). RESULTS: A series of eight chalcones (1-8) and four chalcone glucosides (9-12), inhibited BNA in a dose-dependent manner with IC50 of 8.3 ∼ 77.0 µM. The most potent chalcones were butein (1, IC50 = 8.3 µM) and its glucoside 9 (IC50 = 13.8 µM). The aglycones (1-8) showed non-competitive inhibition, while chalcone glucosides (9-12) displayed a mixed type I (KI < KIS). Inhibitory behaviors were doubly confirmed by KSV and matched with tendency of IC50. The functional group responsible for BNA inhibition were disclosed as 4'-hydroxyl group on B-ring by structure activity relationship (SAR) and molecular docking experiments. Butein (1) suppressed E. coli biofilm formation by > 50 % at 100 µM according to crystal violet assay, which was confirmed by SEM and CLSM imaging. CONCLUSION: The results showed that chalcones (1-8) and chalcone glucosides (9-12), metabolites isolated from the flowers of C. lanceolata, had BNA inhibitory and antibiofilm formation effect on E. coli.

New Class of Tyrosinase Inhibitors, Rotenoids, from Amorpha fruticosa.

A series of rotenoids including a new one from the seeds of Amorpha fruticosa were found to have significant potential as tyrosinase inhibitors. All of the isolated rotenoids (1-6) displayed inhibitory activity against tyrosinase, both as a monophenolase for the oxidation of l-tyrosine and as a diphenolase for the oxidation of l-DOPA. The three most active compounds (1, 5, and 6) showed significant monophenolase inhibition with IC50 values of 2.1, 1.7, and 1.2 µM, respectively. They also inhibited diphenolase function with IC50 values in the range of 9.5-21.5 µM. The inhibition kinetics established all compounds to be competitive inhibitors of both oxidation processes. All rotenoids formed the Emet·I complex effectively around their IC50 values with long lag times. Tyrosinase inhibition of the new rotenoid 6 was additionally demonstrated using high-performance liquid chromatography (HPLC) analysis with N-acetyl-l-tyrosine. Molecular docking disclosed that the sugar moiety of 5 interacted with the bottom of the catalytic gorge.