Study On The Concentration-time-structure Effects Of 17?-Estradiol,Bisphenol A And Nonylphenol On Zebrafish Vtg1 Gene Expression

This study determined and evaluated the estrogenic potentials on hepatic vtg1 gene of adult male zebrafish which were exposed to low level concentrations?ng/L-?g/L levels?of individual and binary mixtures of E₂,BPA and NP through the use of real-time qPCR,reduced life expectancy?RLE?model and molecular docking analysis,and explored the RLE model for the reduced life expectancy in teleost fish caused by the estrogen effects.It is important to study the ecological safety assessment of water environment under low-dose and long-term exposure of typical EDCs.The results are as follow:?1?Binary mixtures of E₂,BPA and NP had weak synergistic effects under the exposure concentration ranges?ng/L-?g/L?.The results of qPCR analysis show that the order of in vivo estrogenic potentials of individual and binary exposure of E₂,BPA and NP was as follows:E₂+BPA>E₂>E₂+NP>BPA>BPA+NP>>>NP.?2?The RLE model analyses can be useful to predict the variation tendency of reduced life expentency of zebrafish using EC₅₀0 as toxicity endpoint.All plots of the linear RLE model had negative slopes indicating that EC₅₀0 was negatively correlated with the natural logarithm of exposure time(lnET₅₀).The slope of E₂,BPA,E₂+BPA,E₂+NP and BPA+NP is-4.43E-9,-4.18E-5,-0.2300,-0.2217 and-0.2104,respectively.The correlation coefficient was greater than0.96.The reported NLT ranging from 930 to 1980 days and the calculated NLT ranging from1123 to 1240 days.The calculated NLT was within the reported NLT of the literature.?3?Molecular docking analysis can be useful to understand interactive effects of E₂,BPA and NP.Molecular docking analysis revealed that the interaction potential of E₂?Binding energy:-10.1 kcal/mol?for the zebrafish ER?-LBD was the most potent?stable?,followed by BPA?-8.0kcal/mol?and NP?-6.8 kcal/mol?.The phenolic hydroxyl of E₂ on one side formed hydrogen-bonds to stabilize the combination with the side chain atoms of the key amino acids of Arg89 and Glu48?polar residue?.The docking simulation revealed that BPA adopted a binding mode with one phenol ring to the key amino acids of Arg89 on one side,which has a Pi-sigma interaction to stabilized combination.Also,the phenolic hydroxyl of BPA on one side was hydrogen-bonded to the N atom of Ala45?non-polar residue?skeleton and the O atom of Leu41?non-polar residue?skeleton,in order to stabilize the combination.However,NP shows no stronger hydrogen-bonded and Pi-sigma interactions and its binding capacity in three ligands was the weakest.?4?The RLE model can be used to analyze the reduced life expectancy in the teleost fishes caused by the estrogen effect.The results on the regression analysis over different exposure times has demonstrated that the EC₅₀0 falls with increasing exposure times in a predictable manner.It is indicates that slopes of E₂,EE₂,BPA and NP between-9.18E-05 and-3.91E-08 and R² value between 0.5530 and 0.9264.Reported NLT and calculated NLT of teleost fish are in the range of 540-1400 d and 13-84 d respectively.The ratio of averages is 1006/35 and the ratio of the standard deviation is 311/33.The correlation coefficient obtained from the plots of reported NLT against calculated NLT was greater than 0.91 in a general accord.The reduction of toxicity over time can be interpreted based on the bioconcentration process,which can be used to understand transfer routes of the compounds from water to teleost fish body.