Surface Charge-dependent Bioaccumulation Dynamics And Toxicity Of Silver Nanoparticles In Chlorella Vulgaris

Silver nanoparticles will inevitably enter the water bodies if is widely used without suitable control measures,and will accumulate in aquatic organisms and cause toxic effects.Surface charge is an important factor affecting the accumulation of silver nanoparticles and their toxic effects on aquatic organisms.However,little is known about how the surface charge affects the accumulation of silver nanoparticles in organisms and their toxic effects.In this research,I used the bioaccumulation kinetics model,metabolomics and proteomics methods to reveal the bioaccumulation,toxic effects and toxic mechanisms of surface charge on silver nanoparticles in Chlorella vulgaris.The main results are as follows:(1)The bioaccumulation kinetics model combined with thermodynamic theory calculations are used to reveal the effect of surface charge on the accumulation kinetics of silver nanoparticles in Chlorella vulgaris.The 50%(EC50)and 10%(EC10)growth inhibition concentrations of negatively charged silver nanoparticles to Chlorella vulgaris were 70 ?g/L and 12?gg/L,respectively.The EC50 and EC10 of postively charged silver nanoparticles to algaes were 50 ?g/L and 5 ?g/L,respectively.The uptake phase of negatively charged silver nanoparticles in Chlorella vulgaris is consistent with the first-order kinetics model.Under the exposure concentration of EC50 and EC10,the uptake rate constant of negatively charged silver nanoparticles in Chlorella vulgaris was that(EC50)=0.36±0.13(L/g/h)and ku(EC10)=0.55±0.27(L/g/h),respectively.The rapidly uptake positively charged silver nanoparticles by algae and this accumulation behavior does not support the first-order kineties model.Under the exposure concentration of EC50 and EC10,the estimated uptake rate constant of positively charged silver nanoparticles in Chlorella vulgaris was that ku(EC50)=9.56(L/g/h)and ku(EC10)=13.28(L/g/h).By contrast,under the same inhibition concentration,the uptake rate constant(ku)of positively charged silver nanoparticles was nearly 20 times higher than that of negatively charged silver nanoparticles.The bioconcentration factors of negatively and positively charged silver nanoparticles in algae showed relatively low with BCF(EC50)=0.83,BCF(EC10)=1.41 for negatively charged silver nanoparticles,and that with BCF(EC50)=1.00,BCF(EC10)?0.80 for positively charged silver nanoparticles,respectively.The results of DLVO calculation shows that the interaction energy between silver nanoparticles and Chlorella vulgaris for negatively and positively charged silver nanoparticles group is 221.92 kaT and-792.73 kBT,respectively.This phenomenon indicates that the interaction energy between the two silver nanoparticles and algae is totally opposite,suggesting that the interaction mechanism may be quite different.The above results indicated that the surface charge of silver nanoparticles changes the bioaccumulation of silver nanoparticles in algae.(2)Combined with metabolomic analysis,fatty acid profiles and common ecotoxicological indicators,the effects of the surface charged influence of silver nanoparticles on the metabolism,fatty acid composition and the toxic effects of Chlorella vulgaris were explained.The maximum relative electron transport rate,the initial slope of the rapid light curves and minimum saturating irradiance of the positively charged group were lower than those of the negatively charged group when under the exposure concentrations of EC10 and EC50.From the transmission electron microscopy images of the Chlorella vulgaris cell structure,it was evident that the chloroplast of algae was more severely damaged by the positively charged silver nanoparticles than by those that were negatively charged.Following exposure to the positively and negatively charged silver nanoparticles(EC50)for 48 h,scanning electron microscopy imaging indicated that Chlorella vulgaris produced a large number of metabolites.Further analysis with infrared spectroscopy illustrated that the lipid-related functional groups,P=O(asymmetric stretching)and C=O(symmetric stretching)in the extracellular secretion of the algae changed.In addition,the ratio of saturated fatty acids to unsaturated fatty acids in the algae decreased when exposed to silver nanoparticles.These results indicated that both positively and negatively charged silver nanoparticles have a significant effect on the lipid metabolism of Chlorella vulgaris.To investigate the metabolic changes in algae exposed to both positively charged and negatively charged silver nanoparticles,LC-MS was used to examine the metabolic profiles of the control samples(algae cells only),as well as the algae samples exposed for 72 h to both the control solutions and the 30 ?g/L concentrations of positively charged and negatively charged silver nanoparticles.The exposure groups of the negatively charged silver nanoparticles(Cit-AgNPs),positively charged silver nanoparticles(PEI-AgNPs),as well as the control groups of the negatively charged(Cit-Ctrl)and positively charged silver nanoparticles(PEI-Ctrl)produced 32,31,34 and 36 differential metabolites(p<0.05),respectively,of which 70%(23 species)of the metabolites were shared by all exposure groups.The PEI-AgNPs has two specific metabolites namely PE(15:0/20:5(5Z,8Z,112,14Z,17Z))and PG(16:0/0:0)[U].The PEI-AgNPs' metabolites involve eight biological metabolic pathways,which are mainly related to lipid metabolism.The Cit-AgNPs has two specific metabolites namely Nystatin and Sucrose.The differential metabolites of Cit-AgNPs involve 14 metabolic pathways,which mainly include lipid metabolism and energy metabolism.The above results indicate that the positively charged silver nanoparticles exerted a greater influence on the photosynthetic system(PS?)of Chlorella vulgaris than those that were negatively charged.However,the effect of the negatively charged silver nanoparticles on the energy metabolism of algae exceeded that of the positively charged silver nanoparticles.(3)Proteomics analysis was used to elucidate the effect of surface charged silver nanoparticles on the protein expression and response of Chlorella vulgaris Following the exposure of Chlorella vulgaris to negatively and positively charged silver nanoparticles and their control solution for 24 h,Cit-AgNPs,PEI-AgNPs,Cit-Ctrl and PEI-Ctrl produced 45,110,343 and 126 differential proteins,respectively.After removal of the common proteins in the corresponding control solution groups,the Cit-AgNPs and PEI-AgNPs had 20 and 57 differential proteins,respectively.The KEGG analysis indicated that the Cit-AgNPs did not have any proteins involved in Cellular Processes pathway.Simultaneously,the Cit-AgNPs involved in Genetic Information Processing and Metabolism's pathways,expressed 1 and 6 pathways,respectively,which were significantly less than that of the PEI-AgNPs at 4 and 10,respectively.Gene Ontology analysis revealed that the proteins of the positively charged silver nanoparticles group were involved in 13 biological function expression related to DNA replication,RNA transcription and ribosome translation.The proteins of Cit-AgNPs were involved in 17 biological function expression related to mitochondria and oxidative phosphorylation processes.The above results indicate that positively charged silver nanoparticles mainly affect the transmission of genetic information for Chlorella vulgaris,while negatively charged nanoparticles mainly affect the energy metabolism pathway of Chlorella vulgaris.