Study On Combined Effects Of Pesticide Residue Mixtures

Pesticide residues found in agricultural products, do not appear singly and usually occur as mixtures, causing combined toxicities to environment and human health. Pesticides are commonly present in agricultural products and have attracted much attention recently. Current food safety regulations in China only focus on single pesticide, and the maximum limits concerning multi-components still lacking. This phenomenon is of increasing public concern in the past few years in the country.The present study selected organophosphate (OP) and N-methyl carbamate (CB) insecticides, which are two classes of pesticides inhibiting the enzyme acetylcholinesterase (AChE) by interfering with cholinergic neurotransmission in both humans and wildlife. In this study we exposed freshwater carps to binary combinations of OP insecticides, such as chlorpyrifos, triazophos, and malathion, and CB insecticides, such as fenobucarb and carbosulfan. In addition, OP, CB insecticides and other commonly used pesticides such as imidacloprid, phoxim, butachlor, X-cyhalothrin were included in the earthworm bioassays in artificial soil and filter paper tests to evaluate their individual and combined toxicities, including binary, ternary and complex mixtures. As the acute toxicity endpoint, the mortality rate of the earthworms was used. On the basis of the measured LC50(50%mortality rate) values of the individual chemicals, a mixture ratio where50%of the effect came from each individual chemical was chosen. This thesis also used zebrafish embryonic testing method to investigate the acute toxic effects of above mentioned pesticides on the survival rate of zebrafish embryo and larvae. The equi-toxic mixture ratio design was used in the embryonic testing experiment where50%of the effect came from each individual chemical based on the measured LC50values of the individual chemicals. We determined the type of combined action that occurred in these mixtures and whether the use of the concentration addition (CA) or independent addition (IA) models of mixture toxicity effectively estimated the risk posed by the mixtures. Results were also analyzed by the method of the Combination Index (CI)-isobologram equation method, which has been widely used in evaluating drug interactions in pharmacology. The thesis studied the influence of atrazine, imidacloprid and chromium, as well as their mixtures, on a series of anti-oxidant enzymes (CAT, POD, MDA and SOD) in earthworms. Finally, the changes of the earthworms’ physiological related genes’ expression were detected using real-time PCR technique under low-dose soil fortified with imidacloprid and chromium as well as their mixtures.The results indicate that the majority of the binary combinations were found to have deviation factors from CA model between0.5and1. Although the components of the tested chemical mixtures may have different modes of action, CA model exhibited well performance to predict the mixture toxicity with the assumption that all the components have the same mode of action, we could draw the conclusion that the majority of the binary mixtures conform to the CA model. While for mixtures consisting of more than three components, a strong synergism would be expected, such as the complex mixtures containing imidacloprid, phoxim, butachlor, λ-cyhalothrin and cadmium, even if the individual components do not exert their effects. The results indicate that the evaluation of mixture toxicity from single component data using CA approach may underestimate combined toxicity and we propose that the combination-index isobologram equation method could serve as a useful tool in risk assessment. The activity of the anti-oxidant enzyme SOD reflected the stress well when the earthworms were exposed in atrazine-chromium mixture, which could be obviously induced along with the experimental process. We propose that SOD could be used as sensitive biomarkers for earthworms exposed in atrazine-chromium mixture. The HSP70and ANN expressions increased significantly in earthworms’ treatments of artificial soil treated with imidacloprid-chromium mixture and chromium, which could be considered as sensitive biomarkers for the specific chemicals in ecological risk assessment.The results of the thesis suggest that the nature of combined effects of chemicals found in agro-products could be targeted specifically through measures of exposure reduction. The use of risk assessment outputs is also essential to develop practical risk management strategies in pesticide residue limit regulations.