Chloride/Nitrite Promoted CuO-Fenton Chemistry For The Inhibition Or Removal Of Biofilm

Biofilm(BF)is aggregate of microorganisms in which cells that are frequently embedded within a self-produced matrix of extracellular polymeric substance(EPS)adhere to each other or to a surface.The microbial biofilm is a fixed system that can be adapted internally to environmental conditions by its inhabitants and be likely to form a biofilm in any part of food processing,which lead to the contamination of food to harm the health of people.With the development of the nanotechnology,the nanoparticles have provided many broad platforms,by generating reactive species to combat unwanted bacteria and biofilm.Copper oxide nanoparticles act as one of the high efficiency catalyst,catalyzing hydrogen peroxide to produce hydroxyl free radical,which has extremely strong electronic ability and the oxidability is second only to fluorine in nature.It is widely used because the oxidbillity can effectively for the inactivation of broad-spectrum microorganisms.Compared with reactive oxygen species(ROS),reactive nitrogen species(RNS)and reactive chlorine species(RCS)were less studied in the anti-bacterial treatment and disinfection fied.Pseudmonas aeruginosa,a gram-negative bacteria and widely distributed in the environment,is one of the main opportunistic pathogen in the hospital and food industry.In this paper,the influence of inorganic anions(Cl-、NO2-)on the inhibition and removal of biofilm was studied on the basis of simple Fenton-like reaction catalyzed by copper oxide nanoparticles using pseudomonas aeruginosa as the object with the 96-well microtiter plates method to form biofilm,the intermediates and the related mechanism was preliminary inquiry.The relative work was divided into the following aspects:1.The effect of the removal of biofilm through Chloride-Accelerated CuO-Fenton(CA-CuO-Fenton)Chemistry was studied.The reactive chlorine species induced by copper oxide nanoparticles can kill planktonic bacteria,which can decrease the levels of the beginning adhesive bacteria in the formation process of biofilm,and produce a chain reaction in a short time to oxidize the organic compounds such as sugar,protein and lipid into carbon dioxide and water.At the same time,a variety of methods were used to quantitatively and qualitatively detect the biofilm removal effect,we found that the CA-CuO-Fenton system can remove the biofilm of Pseudomonas aeruginosa growth in static and dynamic environment,and can also deform the single cell which the effect is stronger than that of CuO-H2O2 system.The reactive chlorine species generated in the system not only removal of biofilm,but also kill the bacteria protected by EPS,and hinder the formation of new biofilm.Under acidic conditions,copper oxide nanoparticles are dissolved to release Cu2+ to induce the production of reactive chlorine species.The cinnamyl alcohol was analyzed as a model molecule to prove the produce of chlorine free radicals,which halogenated products of cinnamyl alcohol was found Then,the reactive chlorine species were applied to the chromogenic reaction of 3,3’,5,5’-tetramethylbenzidine(TMB).It was found that the color effect was stronger than that of the CuO-H2O2 reaction,which further explained the influence of chloride ion on the Fenton reaction of copper.2.The effect of the inhibition of biofilm through nitrite-enhanced CuO-Fenton(NE-CuO-Fenton)Chemistry was studied.In this experiment,through the preparation of eight groups of different reagents to deal with Pseudomonas aeruginosa biofilm growth in static and dynamic environment respectively,and followed by crystal violet staining and then observed under inverted microscope,fluorescence microscopy images and field emission scanning electron microscope(FE-SEM)to found that the NE-CuO-Fenton reaction inhibited biofilm formation is better than that by the CuO-H2O2 system and there are obvious synergistic effect between the three kinds of reagent.The reactive nitrogen species produced in the system not only inhibited the growth of planktonic bacteria,but also reduced the growth activity of the biofilm bacteria.By using gel electrophoresis to degrade nucleic acid,the degradation rate of NE-CuO-Fenton system is slightly faster than that of CuO-H2O2 system,which sterilize more effectively.The formation process of biofilm is mainly regulated by quorum sensing system,and which N-acyl homoserine lactine(AHL)family is the quorum sensing signals of Gram-negative bacteria.So we studied the effect of NE-CuO-Fenton reaction response to AHLs,which to be found can be degraded effectively.Under acidic conditions,the existence of nitroxide radical in NE-CuO-Fenton reaction was proved by the determination of the product of cinnamyl alcohol and fluorescence quenching experiments.Finally,the NE-CuO-Fenton reation was applied to the chromogenic reaction of TMB,which further proved the synergy effect among the reagents.In addition,the addition of hydroxyl radical scavengers does not affect the color effect indicated that hydroxyl was not the main function substance.3.Based on the results of previous study,we simply coated copper oxide nanoparticles(CuO NPs)on the surface of copper foil and used for the test of antibacterial properties in seawater,which the antibacterial effect was found stronger than that of CuO NPs in distilled water.