Development Of Fluorescent Method For The Detection Of Methamidophos And Methomyl Based On Enzyme Inhibition

Organophosphorus pesticides and carbamate pesticides play an active role in the production of agricultural and sideline products,which effectively solve the problem of crop diseases and insect pests.However,the long-term use and the misuse of pesticides conduced the fact that many agricultural products have drug residue exceeding the national standard,which becoming a serious threat to environmental safety and human health.The quickly circulating,large consumption and strong dispersion of agricultural products make it difficult to timely monitoring of pesticide residues by the common laboratory testing methods.Therefore,by the characteristics that organophosphorus pesticides and carbamate pesticides have inhibitive activity to acetylcholinesterase(AChE)and the good optical properties of CdTe quantum dots(CdTe QDs),we establish two fluorescence detection technologies which were used for detected organic phosphorus and Carbamate pesticides respectively.After a series of experimental verification and condition optimization,we achieve the high sensitivity and rapid detection to two pesticides.Specific research content is divided into the following two aspects:1.To establish the detection method of organophosphorus pesticides by optical analysis based on enzyme inhibition and the fluorescence effect of CdTe quantum dot.In the experiment,methamidophos(a kind of organophosphorus pesticide)was analyzed as an example.The interaction between AChE and its substrate thio acetylcholine(ATI)can affect the fluorescence emission of QDs,while organophosphorus pesticides can inhibit the activity of AChE and indirectly affect the fluorescence emission intensity of QDs.Therefore,the correlation between the concentration of organophosphorus pesticide and the fluorescence intensity of QDs is the core of the detecton method of the target pesticide.The surface of the synthesized CdTe QDs has a negative charge,while the ATI surface has a positive charge,so that the QDs fluorescence emission can be significantly enhanced by the approach of ATI.AChE can catalyze the hydrolysis of ATI and generate a positively charged thio choline with a mercapto group.Then,the electrostatic adsorption allows the CdTe QDs to bind to thiocholine,and the sulfhydrides are cover at the surface of the QDs causing a decrease in fluorescence intensity(AChE-ATI-QDs system);if the organophosphorus pesticide methamidophos with enzyme inhibition is added to the system,ATI will not be hydrolyzed and the fluorescence of the QDs will be illuminated by ATI and appear the red shift of the peaks(organophosphorus Pesticide-AChE-ATI-QDs system).The change of the fluorescence intensity of CdTe QDs can reflect the presence of organophosphorus pesticides in the system.The linear range between the fluorescence of AChE-ATI-QDs system with the different concentration of methamidophos was established.The linear range was 0.25-1.5 ?g/mL and the detection limit was 1.50 ng/mL.This method meets the requirements of rapid,simple and low-cost detection and analysis of organophosphorus pesticides,and has high detection sensitivity and strong anti-interference ability.2.A fluorescence method for the detection of carbamate pesticides was established based on the fluorescence inhalation effect(IFE)between the gold nanoparticles and CdTe QDs and the carbamate inhibition(take the methomyl as example).Gold nanoparticles(AuNPs)have strong absorption at ~ 520 nm,while the obtained CdTe QDs display a maximum fluorescence emission at 549 nm,which is just near the absorption maximum of AuNPs.Therefore,the IFE effect occurs after their mixture,and the fluorescence of QDs is quenched.ATI hydrolyses to produce thiocholine under the catalysis of AChE,and thiocholine with positive charge move towards negatively charged AuNPs,causing agglomeration and reunion cross of AuNPs,which further leads to a decrease in the absorbance of AuNPs.Based on the above knowledge background,carbamate pesticides were detected by fluorescence analysis based on enzyme inhibition and QDs.Firstly,the citrate-derived AuNPs(homogeneously dispersed and showing red)and thioglycolic acid-coated CdTe QDs were prepared,and both of them were negatively charged and were subjected to fluorescence quenching due to IFE after mixing;using ATI as the reaction matrix,adding AChE could cause its hydrolysis reaction,then ATI decomposed and formed thiocholine with positive charge.Thio cholines with mercapto were combined with negatively charged AuNPs causing the fluorescence emission of QDs restored to a high level due to the change of the absorbance value of AuNPs.When the carbamate pesticide tested was added,ATI hydrolysis is blocked due to the irreversible enzyme inhibition,the disintegration and absorbance of AuNPs were the same,and the fluorescence emission intensity of QDs was decreased again due to the effect of IFE.Therefore,the determination and detection of carbamate pesticides can be achieved by measuring the fluorescence changes of AChE-ATI-AuNPs-QDs system before and after the addition of the test substance.The fluorescence intensity of QDs was changed by adjusting the concentration of carbamate pesticides,the fluorescence quenching rate(F0-F)/F0 changes with the increase concentration of the measured pesticide(F0,F respectively indicate the fluorescence intensity when the system is not added and added to the carbamate pesticide).The linear range of the fluorescence quenching rate was 0.017-0.5 mg/mL and the detection limit was 0.011 mg/mL(S/N = 3).Compared with high performance liquid chromatography,the fluorescence analysis method has the advantages of simple detection steps,fast signal response and low detection limit.