# The Influence of Silver Nanoparticle Form on the Toxicity in Freshwater Mussels
## 4 Discussion
The reasons for these non-linear changes are unclear but it could be related to SA-induced crowding effects. In marine clams, AChE activity was also decreased in the gills and digestive glands of clams exposed to nTiO\({}_{2}\) and Au-coated nTiO\({}_{2}\)[32]. This decrease was associated with decreased feeding activity and filtration rates. While catalase was induced by either nTiO\({}_{2}\) or Au-coated nTiO\({}_{2}\), oxidative damage (LPO) was more strongly expressed with uncoated nTiO\({}_{2}\). Dissolved but not nAu-inhibited AChE in the hemolymph of _Mytilus galloprovicincallis_ suggests that ionic forms of the metal acted on the enzyme [33]. Inhibition of AChE in fish exposed to nAgwas observed in muscles and brains [34]. However, the contribution of dissolved Ag was not examined in this study. Nanoparticles are recognized to produce effects not only from the release of dissolved elements but from other properties, such as form, size and coatings [35]. The capacity of nanoparticles to absorb existing chemicals, such as PAHs and other chemicals, e.g., pharmaceuticals or pesticides, is another property of nanomaterials referred to collectively as \"Trojan horse effects\" [36; 37]. Surface properties could also be changed by surface interactions, as shown above, involving the adsorbtion of proteins, lipids or other macromolecules (natural organic matter) at the surface of nanoparticles [33]. The formation of protein or lipid coronae at the surface of nAg was shown to preserve the Ag core, thereby limiting rapid diffusion of ionic Ag\({}^{+}\) into the medium and toxicity [38]. Serum bovine albumin binds at the surface of nAg, forming a corona which, in turn, could interact with lipid vesicles [39]. The fluidity of lipid vesicles was increased in the presence of albumin-coated nAg compared to uncoated nAg. This is in agreement with the observation of decreased viscosity in found nAg but not with ionic/dissolved Ag in the present study. Moreover, decreased viscosity was stronger with lower SA prismatic nAg compared to spheric and cubic nAg (Table 1). This suggests that weakened protein coronae increase interactions with lipid vesicles, perhaps by hydrophilic interactions of the PVP-coating of nAg. Based on correlation analysis, viscosity was significantly correlated with lipids, protein aggregation and labile Zn (Table 2). It is noteworthy that labile Zn levels and viscosity changes followed a non-linear U-shaped curve, suggesting that the observed changes occur in a biphasic manner. For example, the drop in viscosity at low concentrations of cubic and prismatic nAg could be associated with changes in protein-lipid interactions at the surface, where viscosity is returned to control values as the concentration of nAg increases in the cytoplasm. These biophysical changes and labile Zn changes were shown to induce protein denaturation and turnover in cells exposed to nAg [40], which could influence viscosity. This suggests that the geometry and corona surfaces could influence the availability, the biophysical characteristics of the cytoplasm and the toxicity of nAg. Indeed, various forms of nAg in the same size range were shown to induce partially organized nematic liquid crystals in the cytoplasm, leading to increased protein denaturation and turnover in cells [10]. Another consequence with respect to these interactions caused by the fine nanoparticles is the reduction of free space in the cytoplasm, in which complex macromolecules interact during metabolism. As shown in this study, the _fD_ of AChE was reduced, thereby slowing down reaction rates at high substrate concentrations, perhaps by crowding (traffic) by nanoparticles or changes in liquid crystals, as evidenced by viscosity and lipid mobilisation. Decreased enzyme activity by steric interactions was also shown with the pyruvate kinase-lactate dehydrogenase complex, where various forms of nAg in mussels reduced the (fractal) dimension of the enzyme complex activities [10].