Enantioselective Toxicity Of Two Novel Chiral Pesticieds On HepG2 Cells

The study has demonstrated that the enantioselectivity of chiral pesticides in bioaccumulation,environmental fate,reproductive toxicity,and the metabolomics.Therefore,it is necessary to assess the environmental risks caused by chiral pesticides and explore the selective toxicity and mechanism of enantiomers of chiral pesticides and environmental risk assessment of chiral pesticides from the level of the enantiomers in-depth.However,the research on the mechanism of toxicity caused by enantiomers of chiral pesticides is still limited.The selective toxicity studies of chiral pesticides: penthiopyrad,prothioconazole and its metabolites prothioconazole-desthio have not been found yet.Therefore,in this study,the human hepatocellular carcinoma cell HepG2 was used to an in vitro model to evaluate the cell viability and oxidative stress between enantiomers of penthiopyrad,prothioconazole and its metabolites.Both the mechanism and enantioselectivity of toxicities were investigated using molecular biology methods,the results in the study are as follows:(1)Study on the selective toxicity and oxidative stress of penthiopyrad and its enantiomersThe penthiopyrad peak-1,penthiopyrad peak-2,and the racemate of penthiopyrad was formulated into 10,20,40,50,and 60 mg/L Venom with the cell culture medium.After 24 hours,cell viability(MTT)and lactate dehydrogenase(LDH)activity were measured.The results showed that with the increase of drug concentration,cell viability was significantly inhibited under penthiopyrad peak-1,penthiopyrad peak-2 and the racemate of penthiopyrad,the LDH activity wassignificantly increased.At the same time,the effect of enantiomers on cell viability and LDH activity was significantly different,showing enantioselective toxicity.The effects of different concentrations of penthiopyrad and their enantiomers on the viability of the cells: penthiopyrad peak-1> penthiopyrad peak-2> penthiopyrad racemate,The increase in extracellular LDH activity induced by penthiopyrad peak-1 was higher than that penthiopyrad peak-2 and penthiopyrad racemate.The indexes of Malondialdehyde(MDA),superoxide dismutase(SOD)and catalase(CAT)were used to detect the oxidative damage of penthiopyrad enantiomers to HepG2 cells.Under the induction of penthiopyrad peak-1,the content of malondialdehyde(MDA),the activity of SOD and CAT were significantly increased,and the effect of enantiomers on oxidative damage have significantly different.The results showed that the level of oxidative stress caused by penthiopyrad peak-1 was higher than that of its enantiomers and racemates,and the oxidative stress caused by both penthiopyrad and its enantiomers: penthiopyrad peak-1> penthiopyrad-2> penthiopyrad racemate.(2)Study of selective toxicity and oxidative stress of prothioconazole and its enantiomers on cellsProthioconazole peak-1,prothioconazole peak-2,prothiothioconazole racemate with cell culture solution with the concentrations of 20,40,60,80,100 mg/L,after 24 hours treatment.The results of measuring cell viability(MTT)and lactate dehydrogenase(LDH)activity was revealed that cell viability was significantly inhibited as compared with the control group when the concentration was increased,and with prothioconazole peak-2 inhibition of viability is significantly stronger than its enantiomers.At different concentrations,the inhibitory effect of prothioconazole and its enantiomers on cell viability was as follows: prothioconazole peak-2> prothioconazole racemate> prothioconazole peak-1.LDH activity showing with the prothioconazole peak-2treatment,the LDH activity increased the most,followed by prothioconazole racemate and prothioconazole peak-1.The index of MDA,CAT,and SOD as evaluation the oxidative stress,it was found that with the increase of concentration,the content of MDA increased,indicating that lipid peroxidation occurred.In response,the SOD and CAT activities were also significantly increased compared to the control group.At the same concentration,under prothiothiazole peak-2 the increase MDAcontent,SOD activity,and CAT activity was higher than prothioconazole peak-1,prothioconazole racemate,indicating that prothioconazole was responsible for cellular oxidation.The stress injury is enantioselective,and the extent of oxidative stress: prothioconazole peak-2> prothioconazole racemate> prothioconazole peak-1.(3)Study on the Selective Toxicity and Oxidative Damage of Dimeconazole and Its Enantiomers on the Metabolites of Prothioconazole(Prothioconazole-desthio)The stock solution containing Prothioconazole-desthio peak-1,Prothioconazole-desthio peak-2,and Prothioconazole-desthio racemate was formulated into 20,40,60,80,and 100 mg/L venoms.The HepG2 cells were treated for 24 h and the cells were assayed.The results of viability(MTT)and lactate dehydrogenase(LDH)activity showed that,with the increase concentration,the cell viability of Prothioconazole-desthio peak-1,Prothioconazole-desthio peak-2,Prothioconazoledesthio racemate on significant Inhibitory effect,similarly,there was a significant difference in the effect of enantiomers on cell viability.At different concentrations,the order of cell viability inhibition was Prothioconazole-desthio peak-2> Prothioconazole-desthio racemate>Prothioconazole-desthio peak-1.Determination the LDH activity,LDH activity induced by Prothioconazole-desthio peak-2 was increase highest,Prothioconazole-desthio peak-1 was the lowest,and Prothioconazole-desthio racemate was in center.MDA,SOD,and CAT were selected as evaluation indicators of oxidative stress.MDA content was increased under the induction of Prothioconazole-desthio and its enantiomers,and the activity of SOD and CAT also increased.Indicating that the cells were oxidatively stressed.as follows: Prothioconazole-desthio peak-2> Prothioconazole-desthio racemate>Prothioconazole-desthio peak-1.Above all,results of our study suggest that enantiomer-specific chiral pesticide-induced oxidative stress is possibly the mechanism of chiral pesticide-induced enantioselctive cytotoxicity.This study provides a scientific basis for evaluating the environmental safety of two new chiral pesticides and screening high enantiomerically safe and biologically low toxic enantiomeric monomers.