The Joint Toxicity Of Acetamiprid And Halosulfuron-methyl On The Zebra Fish,with Using Metabonomics Technology

Halosulfuron-methyl,a representative of the sulfonylurea herbicides,is used to control the broadleaf weeds in field.While acetamiprid,a representative variety of the chloronicotinyl pesticides,is widely used to control control pests on paddy fields,fruit trees and vegetable.The mixture use of pesticides and herbicides is very common in the process of the pesticide application.The solubility of halosulfuron-methyl and acetamiprid in water are both high,so the toxicological evaluation of these two kinds of pesticides on aquatic organisms is extremely urgent.1.Measured by the toxicological test,the LC50 after 96h of acetamiprid,Halosulfuron-methyl and mixture of them in zebrafish were 19.8 mg/L,24.3mg/L and 21.1mg/L respectively.The mixture use of these two pesticide showed a synergistic effect.Hydrological Static Method to investigate the accumulation of pesticides.8 days after exposed to the two pesticides,zebrafish's organs and whole body were tested.The concentrations of acetamiprid and Halosulfuron-methyl in liver,gill,muscle and whole body were lower significantly than that of in solution,and this means the enrichment of pesticides was not happened.2.The single and combined of acetamiprid-exposed and halosulfuron-methyl-exposed could led the content of MDA increase,and with the increase of pesticide dose,MDA content also increased,it is speculated the degree of oxidative damage of zebrafish was exacerbated.The CAT and GPx's main function is to decompose the peroxide into water and non-toxic compounds.They had the same trend in the single and combined of acetamiprid-exposed and halosulfuron-methyl-exposed,indicating that the synergy of CAT and GPx,together with SOD formed a complete anti-oxidation chain when the poison got into the fish's body.The high dose exposure under pesticides led the activity of SOD,GPx,CAT,Na+/K+-ATPase,Ca2+/Mg2+-ATPase and AchE in zebrafish to decrease.The reason may be that the lipid peroxide increased and the free radicals accumulated in body which caused damage to cells.3.Metabolites in the liver and serum of zebrafish exposed under acetamiprid and Halosulfuron-methyl were detected by GC-MS and analyzed by using the factorial analysis of variance(ANOVA).It was found that the combined exposure under the two pesticides could interact with the following metabolites(P?0.001):alanine,valine,leucine,serine,glycine,threonine,aspartic acid,methionine,glutamic acid,pyroglutamic acid,phenylalanine,tyrosine,ornithine,proline,lysine,lactic acid,phosphoric acid,gluconic acid,pyruvic acid,fumaric acid,malic acid,citric acid,and taurine,octadecanoic acid,erucylacetic acid,docosahexaenoic acid,mannose,glycerol,glycerol 3-phosphate,galactopyranoside,inositol,creatinine,adenosine,uridine,cholesterol,glucose,creatinine.It is speculated that the two pesticides can interact with the zebrafish,which is consistent with the results of the synergistic effect of the combined exposure under the two pesticides.4.A total of 51 kinds of metabolites in the liver and serum of zebrafish were identified by using GC-MS and LC-MS,including 17 kinds of amino acids,14 kinds of organic acids,3 kinds of carbohydrates and 17 kinds of other compounds.The contents of various metabolites in the single exposure groups under acetamiprid and halosulfuron-methyl were different significantly from those in the control and combined exposure groups.The metabolic pathways involved amino acid metabolism,lipid metabolism,urea circulation,polyamine metabolism and glycolysis.5.The content of amino acid as changed,the reason may be the self-protection regulation and the disturbance of metabolic pathway of the organism under adversity stress.Glucose was showed a downward trend in the exposure under Halosulfuron-methyl,it may be due to the sulfonylurea herbicides which have a similar chemical structure like sulfonylurea hypoglycemic agents,caused glucose content decrease.In addition,the levels of malic acid,citric acid and fumaric acid in the zebrafish were both changed under the single or combined exposure under the two pesticides,resulting in the destruction of the tricarboxylic acid cycle.Changes in organic acid content also led to changes in the pH value of zebrafish.Putrescine and spermidine as resistance substances,which balance body pH,then also changed significantly.Thymine,hypoxanthine and adenosine as the main metabolites of DNA and RNA,changed significantly,indicating that the genetic material in zebrafish had had a certain damage.6.The metabolites in liver were analyzed by PC A,PLS-DA and S-Plot:taurine can be used as potential markers of the sub-lethal and lethal dose of halosulfuron-methyl exposed.choline may be used as potential markers of the sub-lethal and lethal dose of acetamiprid-exposed.Dopamine and creatinine may be used as potential markers of the sub-lethal and lethal dose of mixed-exposed.The metabolites in serum were analyzed by PCA,PLS-DA and S-Plot:creatinine can be used as potential markers of the sub-lethal dose of halosulfuron-methyl exposed.taurine can be used as potential markers of the lethal dose of halosulfuron-methyl exposed.Glutamic acid,choline and spermidine can be used as potential markers of the sub-lethal and lethal dose of acetamiprid-exposed.5b-cyprinol sulfatecan be used as potential markers of the sub-lethal and lethal dose of mixed-exposed.The detection range and sensitivity of GC-MS and LC-MS were both overlap and were complemented each other,so the two-technical means become complementary in the field of metabolic research.In the result analysis,their overlapping parts of metabolites was be used as a means of verification.In the complementary part,metabolites be predicted by metabolic pathways.The combined using of LC-MS and GC-MS provides a new idea for the study of pesticide toxicology.