engleski

Ultrastructural changes in tobacco cells induced by differently coated silver nanoparticles

Nanoparticles (NPs) research is currently an area of intense scientific surveys due to a variety of their potential applications. However, the characteristics making NPs attractive for their exploitation in new products have led to concerns that they may pose a risk for environment and human health. Silver NPs (AgNPs) are of particular interest because of their well-known antibacterial and antifungal properties. Therefore, they can be found in various consumer products. Since plants play a significant role in accumulation and biodistribution of many environmentally released substances, they could be influenced by AgNPs, serving as a potential pathway for AgNP-transport and bioaccumulation into food chains. The aim of the study is to reveal effects of differently coated laboratory-synthesized AgNPs: slightly negatively charged AgNP-PVP, strong negatively charged AgNP-citrate and strong positively charged AgNP-CTAB as well as of silver nitrate (AgNO3) on tobacco cell ultrastructure. Two months-old plants grown in vitro on a solid MS medium were exposed to 100 μM AgNPs of each type and to 100 μM AgNO3 in ½ strength MS medium for 7 days. For localization of AgNPs in plant cells and ultrastructural analyses, small pieces of tobacco leaf and root were fixed with 1% glutaraldehyde in 50 mM cacodylate buffer (pH 7.2) for 1 h at +4 °C, washed twice and post-fixed with 1% osmium tetroxide in the same buffer for 1 h at +4 °C followed by 10 min wash in ice-cold water. After dehydration in a graded series of ethanol, the tissue was embedded in Spurr´s resin. Semi-thin sections of fixed material were stained with mixture of 2% toluidine blue and 2% borax and examined using light microscope. Ultrathin sections were stained with 2% uranyl acetate and 2% lead citrate and examined using the FEI Morgagni 268D electron microscope for ultrastructural study and the monochromated TF20 (FEI Tecnai G2) TEM for confirmation of AgNPs localization in the tobacco cells. Ultrastructural studies revealed changes in root meristematic cells and leaf chloroplasts in both AgNPs- and AgNO3-treated plants. The root meristematic cells became highly vacuolated and due to large vacuoles only nuclei were observed. AgNPs were localized in the root cells and in the intermembrane space. In the leaf cells, the chloroplasts of AgNP-citrate treated plants were swollen and ruptured with no difference in thylakoid system formation compared to control. Leaves of AgNP-PVP treated plants were characterized with thinner and longer chloroplasts with stacked thylakoids compared to control. In AgNP-CTAB treated plants, chloroplasts were well developed with higher content of plastoglobules compared to control. Big chloroplasts with well-developed thylakoid system were characteristic for leaves of AgNO3 treated plants. The obtained results show the impact of AgNPs and silver ions on the ultrastructure of root meristematic cells and leaf chloroplasts, which is also dependent on the applied coating.

silver nanoparticles ; tobacco ; transmission electron microscopy ; ultrastructure ; localization

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