A Study Of The Mechanisms Of Inhibitory Effect Of Phenolic Acids Derived From Rice Straw On Staphylococcus Aureus Biofilm

Phenolic acids(PAs)are abundant in rice straw.Exploiting the high value of PAs could solve the problem of straw resource utilization.For this,PAs were extracted from rice straw to investigate their antimicrobial effect in this study.Chemical composition of PAs was analyzed and 16 compounds were identified.The yield of PAs was 11.4±0.4 g from 1 kg of rice straw,and the major compounds were p-coumaric acid(46.0%w/w),ferulic acid(23.3%w/w),vanillic acid(5.7%w/w)and vanillin(1.2%w/w).We found that PAs have a strong inhibitory effect on Staphylococcus aureus.The antimicrobial activity of PAs was mainly attributed to p-coumaric acid and ferulic acid.The MIC values of p-coumaric acid and ferulic acid were 0.6 g/L and 1.2 g/L,respectively.More importantly,the combination of the phenolic acids could provide a synergistic effect.Therefore,the MIC of PAs derived from rice straw decreased by 1.5-to 3-fold relative to the MICs of p-coumaric acid and ferulic acid used individually.From assays of metabolites,ATP,ADP,NAD~+/NADH,NADP~+/NADPH,and the hydroxyl radical,we found that the stimulation of the Fenton reaction,which consumed ATP and NADH,increased hydroxyl radicals,resulting in oxidative damage to cells,which was similar to the common mechanism of cell death induced by bactericidal antibiotics.Confocal laser scanning microscopic and atomic force microscopic images revealed that PAs destroyed the biofilm structure,changed the morphology of cells in the biofilm,and reduced their viability,suggesting potential medical applications for PAs.This study could provide theoretical guidance for the development of a plant-based antimicrobial agent,thus promoting straw utilization in a biorefinery.p-Coumaric acid(CA)is a phenolic acid that is abundant in rice straw,which is a crop waste.The aim of this study was to determine the inhibitory effect of CA on growth and biofilm formation of Staphylococcus aureus.It was found that CA caused morphological changes of S.aureus biofilm.The CLSM and FTIR assays demonstrate that the morphological changes resulted from an increase in cellulose content of the biofilm.The data of resazurin assay and CFU showed that CA decreased the viability of the cells in biofilm.The observation of AFM images revealed that at high CA concentration,cell surface became rough and uneven.The determination of intracellular GSH,ATP,NAD~+/NADH,NADP~+/NADPH and peroxides in cells suggested that CA regulated intracellular redox status of cells in biofilm and led to intracellular hydrogen peroxide formation,resulting in the death of cells in biofilm.Moreover,in light of the change of component of biofilm,cellulase and amylase were chosen to amplify the antibacterial effect of CA.As a result,biofilm structure was destroyed completely with little cell viability.These finds suggest the feasibility of CA as an antimicrobial agent and provides a theoretical basis for straw applications.