Optimize Of The Platform For Pesticide Residues On Tea Leaves And Preliminary Research On Degradation Of Imidaclothiz

In our country,neonicotinoid insecticides were frequently used in the tea garden to kill the pests the tea leaves,because of its effectiveness,selectiveness and security to mammals.However,these insecticides could be absorbed by the plant and involved in its metabolism.It was proved that the parent of neonicotinoid insecticides had low toxicity to mammals,but it was still needed to study the toxicity of its metabolites.Since the magnitude of metabolites in actual environment was in trace amount,these metabolites were difficult to be collected.Therefore midaclothiz,the agricultural neonicotinoids developed and commercialized by our country first,was selected in this experiment as a model,using the method of tissue culture to get the suspension of tea leaf's callus first;then directly added the imidaclothiz to the fluid culture medium of callus,made it take part in the metabolism and the metabolites would be accumulated in the culture solution;it was expected to separate the metabolites andout the analytical investigations.The main research results were as follows:1.Leaves of Longjing 43 sterile strain was selected as the experimental material and its primary induction conditions of callus cells were optimized.The result showed that the best induction condition of primary callus was: using MS(Murashige &Skoog,1962)medium;plant growth regulator 2,4-D(2.0 mg/L)+KT(0.5 mg/L);p H5.8;the induction time is about 30 days.2.The subculture conditions of callus of Longjing 43 leaves were optimized.The proportioning of plant growth regulator has been fine-tuned on the basis of the preliminary experiments.Determined the optimum plant growth regulator composition for subculture of callus is 2.4-D(0.1 mg/L)+ KT(0.1 mg/L);PH 5.8;According to growth status of callus of sterile plant leaves,25 to 30 days was selected as the best growth cycle of subculture.3.The Longjing 43 leaf callus liquid culture system was established: using B5 medium as the basic medium;the loaded liquid of the conical flask of 250 m L was100 m L,split the callus into small fritters with the diameter of about 3 mm,with about 150 fritters every flask;the combination of plant growth regulator is 2,4-D(0.1mg/L)+KT(0.1 mg/L);p H 5.8;the appropriate revolving speed was 120 r/min.4.The UPLC-MS/MS analysis and detection method of eight kinds of neonicotinoid was established: 1 g tea powder was weighed into a 150 m L flask.Then 2 m L deionized water was added in the tea surface,left to stand for 30 min at room temperature prior to extraction.Afterward,the mixture was extracted with 20 ml of acetonitrile whirled extract for 2 min,then filtered into 2 g anhydrous sodium added in 150 m L flask,whirled for 1 min,assimilate 5 m L supernatant into 35 m L stoppered centrifuge tube contained 0.4 g PVPP,shaked vigorously for 2min.The extracted liquid was centrifuged at speed of 8000 rpm/min for 8 min;2 m L supernatant was transferred into a 50 m L centrifuge tubes containing 25 mg PSA,50 mg C18,100 mg GCB,and 150 mg anhydrous magnesium sulfate.Then,the solution was shaken for 2min and then left to stand for 5 min,one milliliter of the cleaned-up extract was reduced to near-dryness using a nitrogen evaporator and then dissolved with 0.5ml mixture of acetonitrile and deionized water(15:85,v/v).The final extract was filtered through 0.22 ?m filter into an autosampler vial.The results showed that this method could quantify 8 kinds of neonicotinoid insecticides at the concentration range of 0.01 ~ 2 mg/kg in the tea.The recoveries of eight kinds of insecticides were between 77% to 95% and the relative standard deviations were all less than 9.0%.The LOD was about 0.001-0.002?g/L.5.The metabolism of imidaclothiz was studied in the liquid nutrient medium of tea leaves callus.It was determined the methods of obtaining and making samples.At the bases of the UPLC-MS/MS analysis and detection method of eight kinds of neonicotinoid,the UPLC-MS/MS analysis and detection method had been optimized for imidaclothiz,According to this method,we concluded the degradation tendency chart of imidaclothiz.