Toxicity Mechanism Of Copper Oxide Engineered Nanoparticles In Chlorella Pyrenoidosa Effected By Dissolved Organic Matter

Engineered nanoparticles（ENPs） with the special nature of the small particle size,large surface area, quantum size effect and macroscopic quantum tunneling effect,have been used extensively in recent years and have gradually become a hot research.ENPs are released into the environment inevitablely during production andapplication, which bring the potential environmental risks. Copper oxide engineerednanoparticles （CuO ENPs） has been widely used in commercial areas, such as gassensors, photovoltaic cells, a catalyst, and a semiconductor material. Algae are theprimary producers of the ecosystem. The algal cells exposed to ENPs will ingestENPs and ENPs will be delivered to other organisms by the food chain. This study isintended to analysis systematically of toxicity mechanism to Chlorella pyrenoidosainfluenced by fulvic acid （SRFA）, and observe the process of algal cell uptaking CuOENPs, in order to provide the basis for the environmental safety assessment for CuOENPs ultimately.（1） In this paper, the Chlorella pyrenoidosa grwth-inhibition test showed that theCuO ENPs can inhibit the growth of algal cells with a72h a median effectiveconcentration (EC50,45.6mg L^-1). Analyzing effects of SRFA on the toxicity of40mgL^-1CuO ENPs, we choose20mg L^-1SRFA which has a significant differencescompared with the control in subsequent experiments.（2） SRFA significantly enhanced the toxicity of CuO ENPs Chlorellapyrenoidosa with a lower72h EC50(10.26mg L^-1). One reason is the SRFA promotingthe release of Cu ion whose toxicity can not be ignored; the othe is enhancing thesuspension stability of CuO ENPs, thus reducing their aggregation and increasing theopportunity that ENPs interacted with algal cell and internalized into cells.（3） SRFA can promote the Cu content in algal cells. Increased Cu is mainly come from the internalization CuO ENPs.（4） It was the first time to observe the algal cell without cutting into slices tosaving extracellular polymer of algae makes extracellular polymer visible clearly inTransmission Electron Microscope （TEM） observation. Compared with the control,the cell treated by CuO ENPs have a thicker extracellular polymer layer. CuO ENPshave no changed at≥100nm from the cell wall. However,（200） CuO ENPs in thesize of1-3nm was closed to the cell wall. It concluded that CuO ENPs’ structuralmodification at <100nm from cell wall.（5） By adding the swallow inhibitors, we found intracellar Cu content decreasedby26%. It concluded that endocytosis is one of the ways of ENPs transmembrane. Wealso observed the algal cell uptake ENPs by endocytosis in TEM observation.（6） Treated by CuO ENPs, algal cell plasmolysis, photosynthesis chloroplastlamella rupture, nuclear membrane damage were observed. CuO ENPs less than3nmdispersed in cytoplasm, nuclei, chloroplast lamella in the form of a single or a fewparticles gathered. ENPs existed in membrane structure in aggregate form.（7） The accumulation of additional reactive oxygen radicals （ROS） causedoxidative stress, which resulted in membrane lipid peroxidation and mitochondrialdepolarization, at last algal cells apoptosis. Therefore, intracellular ROS accumulationcaused by CuO ENPs leading to oxidative stress in Chlorella pyrenoidosa is theimportant toxic mechanism.