A Physiologically Based Toxicokinetic/Toxicodynamic(PBTK/TD) Model Of Immature Rats Following The Oral Exposure To Chlorpyrifos

Objectives(1) To build Physiologically Based Toxicokinetic/Toxicodynamic (PBTK/TD) model of the immature rat following the oral exposure to chlorpyrifos(CPF), of which parameters could reflect the age-dependent difference.(2) To make a study on toxicokinetics and toxicodynamics in immature rats following the single or repeated oral exposure to chlorpyifos by using PBTK/TD model.(3) To explore the methods for applications of PBTK/TD model and the further research plan.To provide the xperience for establishing the PBTK/TD model of children exposure to CPF as well as a new method for risk assessment of dietary exposure to organophosphorus pesticides in children.Methods(1) A weight growth curve from neonate to adult rats was constructed based on the collected data.The organ coefficient of liver and brain tissues were plotted by animal age. The metabolism parameters of CYP450, PON1 and aliesterase were fitted as a function of body weight by using allometric scaling equation. We specified the model structure and physiological parameters and wrote the model equations. We also made a sensitivity analysis and optimization of model parameters. The verification and the estimation of the model were made after parameter adjustment.(2) 21-day-old female SD rats were randomly divided into 1 control group and 3 treated groups (12.5,25.0 and 50.0 mg/kg) according to the weight. Serum, cerebral cortex and liver were collected after single gavage at 1h,3 h,6 h, 12 h and 24 h, respectively. The concentration of liver CPF, serum CPF and serum TCP were determined, meanwhile the activity of acetylcholinesterase (AChE) in cerebral cortex and serum. Urine was collected continuously, and samples were taken for the period of 0-12 h and 12-24 h after administration to determine the accumulation of TCP in unne.(3) 7221-day-old female SD rats were given daily doses of CPF of 0,1.0,2.5,5.0,10.0 and 15.0 mg/kg/d via oral gavage for 10 days. Serum and cerebral cortex were collected on day 3,6,10 and 11. The concentration of CPF and 3,5,6-trichloropyridinol(TCP) in serum were detemined, meanwhile the activity of acetylcholinesterase (AChE) in cerebral cortex and serum. Urine samples were collected on day 3,6 and 11 to determine the accumulated amount of TCP in urine. To estimate the quantitative relationship between external, internal exposure and adverse effects at last.(4) To use the PBTK/TD model to analyze the dose-response relationship of chlorpyrifos concentration in immature rats and the corresponding dose of benchmark response, after incorporating the exposure background of animal experiment which is designed by benchmark dose methods the into the model. By adjusting the parameters and inputting daily dietary maximum exposure among the children aged 7-12 year in Jiangsu province, to predict the toxicokinetics of CPF and TCP as well as the extent of blood and brain AChE inhibition in children aged 7-12 year exposed daily to CPF over 2 months.Results(1) The equation of the body weight growth curve function is applied by the model as following:BW= 0.347 x (1-0.782 x e-0.024×Age)3.The age-dependent functional equation of liver and brain weight percentage is better fitting the data collected from animal experiments. The results of model fitting and validation were comparatively good, but not good for other literature data which would be overvalued or undervalued.(2) Experimental data of acute toxicity indicated that after 1-2h oral exposure, the concentration of CPF in the immature rat livers reaches the maximum. With the dose increasing, the peak concentration became higher. Under the same dose, the concentration of CPF in liver decreased with time. At one certain time point after administration, the concentration of CPF and TCP in serum increased with the increase of the dose. The concentration of CPF and TCP in serum increased at first and gradually decline. The peak concentration appeared at 3 h after exposure. The amount of TCP excreted in 24 h were significantly greater than that in 12 h. As the dose increases, the amount of TCP excreted increased at each time point. Within 1～6 h after exposure, the AChE activity in serum quickly declined to the lowest level, and then rose with time slowly. At one certain time point after administration, the AChE activity in serum declined as the dose increased. The minimum percentage of the AChE activity in serum in groups of low dose, middle dose and high dose were 17%,11%and 7%, respectively. The AChE activity in cerebral cortex declined to the lowest level within 1～12 h after exposure. The minimum level of activity in these three groups were 18%,8% and 2%, respectively and slowly recovered with time. At one certain time point, AChE activities in cerebral cortex decreased with the increase of doses.(3) The result of repeated dose test for 10 d:24 h was set for a cycle. The concentration of CPF and TCP in serum increased at first and gradually declined after each exposure. At one certain time point, the concentration of CPF and TCP in serum increased with the exposure dose. AChE activities in serum and in cerebral cortex cyclical changed over time and decreased with the increasing exposure dose. They recovered significantly in 11 d (1 d after stoppoing exposure). There was significant dose-effect relationship between repeated exposures and change of the AChE activity in cerebral cortex. With the increase of repeated exposure doses, the activity minimum values in cerebral cortex and serum of the immature rat declined.(4) The result of simulating BMD animal experiment:The corresponding dose of BMD was 1.92 mg/kg/d. The corresponding dose utilized by the lowest dose was 0.22 mg/kg/d. When the AChE activity in cerebral cortex decreased by 10%, the corresponding dose was 0.74 mg/kg/d, and the corresponding dose utilized by the lowest dose was 0.34 mg/kg/d. The predicting result of 7 year-old urban and rural children in Jiangsu province:internal exposure level forecast by the model was very low. The concentration of CPF was less than 10-3 μmol/L, and TCP was less than 0.5μmol/L. The internal exposure level in rural children was higher than that in urban children. Rural children had more AChE inhibition, although the degree of inhibition was under 10-3% which is far lower than the level of benchmark response.Conlusions(1) The hypothesis of structure and parameter for the PBTK/TD model is reasonable. The difference of age could be observed. The model could be used for prediction at the individual level with certain extrapolative ability, which still need further improvement.(2) The CPF exposed via oral gavage could be reabsorbed and metabolized in immature rats promptly. Immature rats are more sensitive to CPF for the weak metabolism. Recovery rate of AChE activity in cerebral cortex is slower than that in serum.(3)In the experiment of repeated infections, the index of PBTK/TD changes periodically. Metabolic ability of CPF and AChE activity level of immature rats could increase along with the age. AChE level in serum and cerebral cortex might be inhibited by exposed CPF continuously unless the exposure was removed. Dose-response relationship was investigated after repeated infections of CPF. PBTK/TD model combines exposure dose, internal dose and toxic effect organically together quantificationally.(4) This PBTK/TD model has great potential in determination of dose-response relationship, calculation of critical effect corresponding dose with BMD and risk assessment. It can also be used for evaluating toxic effect in children, analyzing the contribution of exposure by oral to the total exposure level, and providing a new method for children organophosphorus pesticide risk assessment, and a scientific basis for decision-making in public health.