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Effect of silver nanoparticles and ions on oxidative stress formation and antioxidative machinery of Chlorella vulgaris

Silver nanoparticles (AgNPs) are used in a variety of industries and due to their tendency to agglomerate in various media, they are commonly stabilised with various surface coatings such as carboxylic acids (citrate) or surfactant cetyltrimethylammonium bromide (CTAB) among others. As the use of AgNPs increases, so does the potential for their release into the aquatic environment. Since Chlorella vulgaris is one of the most ubiquitous algae inhabiting aquatic ecosystems, it is widely used as a model organism for assessing the impact of materials of anthropogenic origin, e.g. AgNPs, on aquatic habitats. To evaluate the impact of AgNP on C. vulgaris, algal cultures were grown in a liquid BBM nutrient medium for four days after which they were exposed to AgNPs coated with citrate or CTAB, and to AgNO3 which was used as ionic Ag control. Concentration endpoints were obtained by growth inhibition test (72 h) and the estimated 25% inhibition of growth rate (EC25) values for the AgNP-citrate, AgNP-CTAB and AgNO3 were 0.188 mg L- 1, 0.895 mg L-1 and 0.130 mg L-1, respectively. After 72 h treatment, the quantity of newly synthesised oxygen, damage to membrane lipids as measured through malondialdehyde (MDA) content, levels of reactive oxygen species (ROS) and activity of antioxidant enzymes (superoxide dismutase-SOD, catalase-CAT, ascorbate and pyrogallol peroxidase-APX and PPX) were analysed. To understand the ROS formation, algal suspensions were incubated with fluorogenic dyes through which hydroxyl and peroxyl activity within the cells (H2DCFDA) or superoxide (O2.−) generation (DHE) were determined, with both dyes providing a measure of other ROS activity through non-specific oxidation. Moreover, to further examine the interaction between AgNPs and algal cells, the treated organisms were observed directly by transmission electron microscopy (TEM). Obtained results showed detrimental effect of all treatments on the photosystem of C. vulgaris since significant decrease in oxygen synthesis was reported. Similarly, all treatments have increased ROS formation and MDA content, which is indicative of membrane lipids damage. Interestingly, only increases in PPX and CAT activity were observed after treatment with AgNP-citrate, while for APX and for other treatments a trend of decreasing activity of antioxidant enzymes were observed. TEM analysis of alge cells showed multiple NPs in extracellular polymeric substances (EPS) of algae cells which could aid in decreasing AgNPs toxic effect on algal cells. In conclusion, all silver nanoparticles (AgNP-citrate and AgNP-CTAB) and ionic silver showed damaging effects on the photosynthetic apparatus of C. vulgaris algae. The parallel increase in ROS formation and decrease in activity of the antioxidant machinery after 72 h treatment can be explained by an overproduction of ROS which consequently results in a change of enzyme structure and activity.

Chlorella vulgaris ; silver nanoparticles ; oxidative stress

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