Development Of Peptide Mimics And Quantum Dots-based Sensing System For Application In Detecting Of Organophosphorus Pesticides And Organochlorine Pesticides

Pesticides have been widely used in agricultural production since their birth and have played a great role in increasing agricultural production,but they have also led to serious environmental pollution and food safety problems.Pesticides can persist in the environment and accumulate in organisms.Commonly used pesticides,organophosphates（OPs）and organochlorine pesticides（OCPs）,OPs are neurotoxic and inhibit the activity of acetylcholinesterase（ACh E）,causing acetylcholine to accumulate in the central nervous system and organ failure and even death;OCPs inhibit inositol metabolism in the body,strongly stimulate the central nervous system and damage organs such as the liver and kidneys.Therefore,pesticide detection is very important for food safety,but traditional methods require complex pretreatment and expensive instrumentation,which is not conducive to real-time pesticide detection.Therefore,it is of great significance and value to develop simple,rapid and sensitive pesticide detection methods.In this paper,sensor systems for OPs and OCPs are constructed based on peptide mimics and quantum dots.The research in this paper is as follows:（1）Based on the binuclear metal active sites of phosphotriesterase（PTE）and the characteristics of peptide fibrosis,Zn²⁺bonding peptide was designed to mimic the activity of PTE.According to the degradation products of OPs,the peptide mimics with the highest activity were selected for the construction of OPs sensors.Firstly,the influence of binuclear metal center active sites and fiber formation on the catalytic activity of peptide mimics were investigated by ultraviolet spectrum,circular dichroism,fluorescence and transmission electron microscopy.Next,the role of Zn²⁺in peptide fibrosis and catalysis was systematically explored.It was found that both active sites ofαmetal andβmetal were necessary for metal bonding peptide,and that Zn²⁺played an important role in promoting peptide fiber formation and participated in the degradation of OPs as a catalytic cofactor.Furthermore,the optimal conditions for peptide hydrolysis were selected by exploring the effects of p H,temperature and metal ions on P-nitrophenol（PNP）production.Under optimal conditions,the hydrolysis rate of the peptide mimics to methyl parathion was 1.729μM/min,3.25 times higher than that of the enzyme mimics based on the catalytic triad Ser-His-Asp（0.532μM/min）.（2）Based on the electrochemical activity of the degradation product of PNP,a non-toxic and efficient peptide mimics was selected and the electrochemical sensor of metalloenzyme mimics was constructed for detection of OPs.First,to improve the bonding efficiency of Ni Co₂O₄,Ni Co₂O₄ was modified with a dendritic macromolecule of PAMAM which contained a large number of carboxyl groups,and the peptide mimics were connected to Ni Co₂O₄ to form the Ni Co₂O₄-PAMAM-peptide compsites,modifying the glassy carbon electrode（GCE）to form an electrochemical sensor.Secondly,the signal of PNP which was the degradation product of OPs,can be monitored by square wave voltammetry（SWV）to determine the amounts of OPs in actual samples within 15 min,with good stability,selectivity,and anti-jamming capability of the sensor.Compared with traditional methods,the electrochemical sensor of the peptide is sensitive,reliable,non-toxic and simple,with a detection limit of 0.08μM for methyl parathion.（3）Based on the peptide active sites in Chapter 1（active sites ofαmetal andβmetal）,metal bonding peptide were designed to significantly increase catalytic activity and were combined with carbon quantum dots（CQDs）and cadmium telluride quantum dots（Cd Te QDs）to development ratio fluorescence sensors for detection of OPs.Firstly,adding amino acids that promote proton transfer and altering the position of Asp at theαmetal active site caused the hydrolysis activity of metal bonding peptide increased about 50%and 27%,respectively.We also investigated divalent cations including Zn²⁺and found that Zn²⁺was the optimum ion to promote polypeptide fibrosis and catalytic cofactor.Secondly,the catalytic activities of peptides to methyl paraoxion and ethyl paraoxion were compared.Through molecular dynamics simulation,the non-covalent interactions and binding free energy among methyl paraoxion,ethyl paraoxion with peptide mimics were calculated,revealing the catalytic mechanism of peptide mimics on OPs.Finally,in the sensor system,metal bonding peptide was employed to recognize and hydrolyze OPs,and produce PNP.Cd Te QDs with red fluorescence were used as a response unit,and CQDs with blue fluorescence were used as an internal reference and enhancer in response.After adding OPs into the sensing system,OPs were hydrolyzed by metal bonding peptide to produce PNP,which quenched the fluorescence of Cd Te QDs.The fluorescence of the peptide composite sensor changed from red to blue,and the detection limit of methyl paraoxion was 0.035μM.In this paper,a ratio sensor based on Zn²⁺bonding peptide and quantum dots had high sensitivity and anti-jamming ability for visual detection of actual samples.The results provided new ideas for peptide immobilization and real-time detection of OPs.（4）MAPB-QDs were first applied to the detection of polar OCPs based on phenomenon that the fluorescence spectra of MAPB-perovskite quantum dots（MAPB-QDs）exhibited the blue shift in the presence of OCPs.The sensing mechanism was revealed by ¹H NMR,FTIR,XPS,and XRD characterization.In the presence of polar OCPs,MAPB-QDs ligands oleic acid and oleamine were substituted by OCPs and the chlorine element was adequately doped into QDs,resulting in the increase of the MAPB-QDs’bandgap and blue shifts in fluorescence wavelengths of MAPB-QDs.Mixing MAPB-QDs with polydimethylsiloxane（PDMS）solved the problem of poor stability of MAPB-QDs in the presence of water,and PDMS fluorescence colorimetric cards were prepared to detect OCPs and realized the visual detection of polar OCPs in real samples.