Inhibitory effects of biochanin A on the efflux pump of methicillin-resistant Staphylococcus aureus (MRSA)

Abstract:

Abstract: ［Objective］ To study the inhibitory effect of biochanin A on efflux system of Methicillin-resistant Staphylococcus aureus (MRSA)． ［Methods］ Inhibitory effects of biochanin A on efflux system of Strain MＲSA41577 were evaluated using double dilution method，two plate method and fluorescence spectrophotometry．Real time PCＲ and SDS-PAGE were applied to detect the expression of MRSA41577 norA and to analyze the changes of MRSA41577 efflux protein before and after dosing biochanin A in association with liquid chromatography mass spectrometry to determinate protein variation．［Results］Biochanin A alone had no inhibitory effect on MRSA41577，but it showed synergy effect with ciprofloxacin in inhibition MRSA41577 in which 40μg/mL biochanin A decreased the minimum inhibitory concentration (MIC) value of ciprofloxacin from 64 μg/mL to 8 μg/mL．Biochanin A significantly increased the accumulation of ciprofloxacin in MRSA41577 in a time-dependent manner．At 15 min，biochanin A increased ciprofloxacin in MRSA41577 by 83%，which is similar to that of reserpine (positive control)．Further mechanism studies indicated that biochanin A could reduce the expression of norA in ciprofloxacin-treated MＲSA41577．After incubated with biochanin A and ciprofloxacin for 16 h，the relative expression of norA of MＲSA41577 was reduced by 65%．SDS-PAGE analysis showed that the total protein profiles of MRSA41577 were significantly changed after treatment with biochanin A for 16h，in which both norA protein and efflux system ABC transporter ATP-binding protein were significantly decreased．［Conclusion］Biochanin A could inhibit Methicillin- resistant Staphylococcus aureus efflux system through reducing pathogen’s expression of norA and norA protein.

Key words:Keywords: Biochanin A，MRSA (Methicillin-resistant Staphylococcus aureus)，norA gene，efflux protein