Transmission Electron Microscopy and Scanning Transmission X-Ray Microscopy Studies on the Bioaccumulation and Tissue Level Absorption of TiO<SUB>2</SUB> Nanoparticles in <I>Daphnia magna</I>

Abstract

In this study, bioaccumulation and tissue-level absorption of TiO2 nanoparticles (NPs) in freshwater invertebrates were investigated using transmission electron microscopy (TEM) and scanning transmission X-ray microscopy (STXM). The TiO2 NPs were used to test impacts of core sizes (i.e., 5 ± 2 nm and 23 ± 7 nm for TiO2(SYN) and TiO2(P25), respectively) and agglomerations (i.e., well dispersed vs. highly agglomerated) on the uptake of TiO2 NPs in Daphnia magna (D. magna). Highly agglomerated TiO2 NPs, regardless of their core sizes, were heavily taken up into the digestive tract of D. magna and no detectable penetration of both TiO2 NPs into the gut epithelial cells of D. magna was observed in TEM and STXM images. However, significant damages involving morphological changes in the microvilli and gut epithelial cells (e.g., irregular shaped microvilli, epithelial cell protrusion, and dilatation of cytoplasmic inclusion) were observed only with the commercial TiO2 NPs (TiO2(P25)) with larger core size and mixed crystalline phase, while the laboratory synthesized TiO2 NPs (TiO2(Syn)) with smaller core size and single crystalline phase showed slight morphological changes in the gut microvilli and epithelial cells. In the case of D. magna exposed to the well dispersed synthetic TiO2 NP ((Cit)TiO2(Syn)), only a negligible amount of TiO2 NPs were found within the digestive tract of the D. magna without any significant damages in the gut microvilli and epithelial cells and any detectable penetrations of TiO2 NPs into epithelial cells of D. magna gut. These TEM and STXM observations confirmed us that uptake of NP into D. magna are strongly dependent on their agglomeration (i.e., hydrodynamic sizes), rather than their core sizes, while direct penetration of NPs into tissues of digestive tract seems unlikely without significant morphological changes (e.g., collapse of the epithelial tissue) caused by high toxicity of NPs or released metal ions.

Colloidal properties and stability of aqueous suspensions of few-layer graphene: Importance of graphene concentration

“…The yolks were taken out of the embryos, rinsed with PBS buffer, and fixed with 3 % glutaraldehyde for 24 h, which was followed by a post-fixation step with 1.0 % osmium tetroxide for 1 h. After dehydration in a series of (50, 70, 80, 95, and 100) % ethanol, the samples were embedded with propylene oxide and epoxy mixture. Ultra-thin sections (the thickness of each section was about 50 nm to 60 nm) were placed on formvar-coated copper grids (300 mesh)(Kwon et al, 2015). The sections of the yolks were stained with uranyl acetate and lead citrate and observed using a FEI Tecnai TF20 high resolution transmission electron microscope (HRTEM) to verify the presence of S-FLG in the yolk.…”

Aquatic Ecotoxicity Testing of Nanoparticles—The Quest To Disclose Nanoparticle Effects

“…113, 114, TiO 2 ENPs: Ref. 115). In contrast to the above observations, some studies did observe uptake through the microvilli (ZnO ENPs: Ref.…”

“…Protrusion of gut epithelia in fish was observed after exposure to TiO 2 ENPs, without evidence of internalization 115. Reduction in food intake and energy inputs, changes in gut mobility, or effects on nerves or smooth muscle fibers have also been proposed as effects not necessarily associated with the internalization of ENPs 123.…”

Aquatische Ökotoxizität von Nanopartikeln – Versuche zur Aufklärung von Nanopartikeleffekten

“…, TiO 2 ‐ENPs: Lit. ). In anderen Studien wurde dagegen eine Passage durch die Mikrovilli beobachtet (ZnO‐ENPs: Lit.…”

“…Es sollte auch erwähnt werden, dass Effekte möglicherweise auch ohne eine Internalisierung oder Translokation der ENPs im Organismus auftreten könnten, und zwar allein aufgrund der Gegenwart der ENPs im Darm. Bei Fischen ist bei einer Exposition gegenüber TiO 2 ‐ENPs eine Hervorwölbung von Darmhautschichten beobachtet worden, ohne Zeichen einer Internalisierung . Eine Reduktion der Nahrungsaufnahme und Energieeinträge, eine Veränderung der Darmmobilität sowie Auswirkungen auf Nerven oder glatte Muskelfasern sind ebenfalls als Effekte vorgeschlagen worden, die nicht notwendigerweise mit einer Internalisierung der ENPs verbunden sind .…”