Thallium Toxicity and its Interference with Potassium Pathways Tested on Various Cell Lines.

Thallium (Tl) is a highly toxic heavy metal whose mechanism of toxicity is still not completely understood. The aim of this study was to test Tl cytotoxicity on several cell lines of different tissue origin in order to clarify specific Tl toxicity to a particular organ. In addition, possible interference of Tl with cell potassium (K) transport was examined. Human keratinocytes (HaCaT), human hepatocellular carcinoma (HepG2), porcine kidney epithelial cells (PK15), human neuroblastoma (SH-SY5Y) and Chinese hamster lung fibroblast cells (V79) were treated with thallium (I) acetate in a wide concentration range (3.9-500 µg/mL) for 24 h, 48 and 72 h. To assess competitive interaction between Tl and K, the cells were treated with four Tl concentrations close to IC50 (15.63, 31.25, 62.50, 125 µg/mL) in combination with/or without potassium (I) acetate (500 µg/mL). The cells' morphology was monitored, and cytotoxic effect was assessed by 3-(4, 5-dimethylthiazole-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) test. The most sensitive to Tl exposure were SH-SY5Y cells, while HepG2 were the most resistant. The combined exposure to thallium (I) acetate and potassium (I) acetate for every cell line, except V79 cells, resulted in higher cell viability compared to thallium (I) acetate alone. The results of our study indicate that cell sensitivity to Tl treatment is largely affected by tissue culture origin, its function, and Na+/K+-ATPase activity.

The Occurrence of Oxidative Stress Induced by Silver Nanoparticles in Chlorella vulgaris Depends on the Surface-Stabilizing Agent.

Silver nanoparticles (AgNPs) are of great interest due to their antimicrobial properties, but their reactivity and toxicity pose a significant risk to aquatic ecosystems. In biological systems, AgNPs tend to aggregate and dissolve, so they are often stabilized by agents that affect their physicochemical properties. In this study, microalga Chlorella vulgaris was used as a model organism to evaluate the effects of AgNPs in aquatic habitats. Algae were exposed to AgNPs stabilized with citrate and cetyltrimethylammonium bromide (CTAB) agents and to AgNO3 at concentrations that allowed 75% cell survival after 72 h. To investigate algal response, silver accumulation, ROS content, damage to biomolecules (lipids, proteins, and DNA), activity of antioxidant enzymes (APX, PPX, CAT, SOD), content of non-enzymatic antioxidants (proline and GSH), and changes in ultrastructure were analyzed. The results showed that all treatments induced oxidative stress and adversely affected algal cells. AgNO3 resulted in the fastest death of algae compared to both AgNPs, but the extent of oxidative damage and antioxidant enzymatic defense was similar to AgNP-citrate. Furthermore, AgNP-CTAB showed the least toxic effect and caused the least oxidative damage. These results highlight the importance of surface-stabilizing agents in determining the phytotoxicity of AgNPs and the underlying mechanisms affecting aquatic organisms.

Phytotoxic Effects of Polystyrene and Polymethyl Methacrylate Microplastics on Allium cepa Roots.

Plastic contamination has become one of the most pressing environmental issues due to rapidly increasing production of disposable plastic products, their fragmentation into smaller pieces, and long persistence in the environment, which affects all living organisms, including plants. In this study, Allium cepa roots were exposed to 0.01, 0.1, and 1 g L-1 of commercial polystyrene (PS-MPs) and polymethyl methacrylate microparticles (PMMA-MPs) for 72 h. Dynamic light scattering (DLS) analyses showed high stability of both types of MPs in ultrapure water used for A. cepa treatment. Morphometric analysis revealed no significant change in root length compared to control. Pyrolysis hyphenated to gas chromatography and mass spectrometry (Py-GC-MS) has proven PS-MPs uptake by onion roots in all treatments, while PMMA-MPs were recorded only upon exposure to the highest concentration. Neither MPs induced any (cyto)toxic effect on root growth and PMMA-MPs even had a stimulating effect on root growth. ROS production as well as lipid and protein oxidation were somewhat higher in PS-MP treatments compared to the corresponding concentrations of PMMA-MP, while neither of the applied MPs induced significant damage to the DNA molecule assayed with a Comet test. Significantly elevated activity of H2O2 scavenging enzymes, catalase, and peroxidases was measured after exposure to both types of MPs. Obtained results suggest that onion roots take up PS-MPs more readily in comparison to PMMA-MPs, while both types of MPs induce a successful activation of antioxidant machinery in root cells that prevented the occurrence of toxic effects.

Effects of Silver Nanoparticles on Physiological and Proteomic Responses of Tobacco (Nicotiana tabacum) Seedlings Are Coating-Dependent.

The harmful effects of silver nanoparticles (AgNPs) have been confirmed in many organisms, but the mechanism of their toxicity is not yet fully understood. In biological systems, AgNPs tend to aggregate and dissolve, so they are often stabilized by coatings that influence their physico-chemical properties. In this study, the effects of AgNPs with different coatings [polyvinylpyrrolidone (PVP) and cetyltrimethylammonium bromide (CTAB)] on oxidative stress appearance and proteome changes in tobacco (Nicotiana tabacum) seedlings have been examined. To discriminate between the nanoparticulate Ag form from the ionic one, the treatments with AgNO3, a source of Ag+ ions, were also included. Ag uptake and accumulation were found to be similarly effective upon exposure to all treatment types, although positively charged AgNP-CTAB showed less stability and a generally stronger impact on the investigated parameters in comparison with more stable and negatively charged AgNP-PVP and ionic silver (AgNO3). Both AgNP treatments induced reactive oxygen species (ROS) formation and increased the expression of proteins involved in antioxidant defense, confirming oxidative stress as an important mechanism of AgNP phytotoxicity. However, the mechanism of seedling responses differed depending on the type of AgNP used. The highest AgNP-CTAB concentration and CTAB coating resulted in increased H2O2 content and significant damage to lipids, proteins and DNA molecules, as well as a strong activation of antioxidant enzymes, especially CAT and APX. On the other hand, AgNP-PVP and AgNO3 treatments induced the nonenzymatic antioxidants by significantly increasing the proline and GSH content. Exposure to AgNP-CTAB also resulted in more noticeable changes in the expression of proteins belonging to the defense and stress response, carbohydrate and energy metabolism and storage protein categories in comparison to AgNP-PVP and AgNO3. Cysteine addition significantly reduced the effects of AgNP-PVP and AgNO3 for the majority of investigated parameters, indicating that AgNP-PVP toxicity mostly derives from released Ag+ ions. AgNP-CTAB effects, however, were not alleviated by cysteine addition, suggesting that their toxicity derives from the intrinsic properties of the nanoparticles and the coating itself.

Silver Nanoparticle Effects on Antioxidant Response in Tobacco Are Modulated by Surface Coating.

The antimicrobial properties of silver and enhanced reactivity when applied in a nanoparticle form (AgNPs) led to their growing utilization in industry and various consumer products, which raises concerns about their environmental impact. Since AgNPs are prone to transformation, surface coatings are added to enhance their stability. AgNP phytotoxicity has been mainly attributed to the excess generation of reactive oxygen species (ROS), leading to the induction of oxidative stress. Herein, in vitro-grown tobacco (Nicotiana tabacum) plants were exposed to AgNPs stabilized with either polyvinylpyrrolidone (PVP) or cetyltrimethylammonium bromide (CTAB) as well as to ionic silver (AgNO3), applied in the same concentrations, either alone or in combination with cysteine, a strong silver ligand. The results show a higher accumulation of Ag in roots and leaves after exposure to AgNPs compared to AgNO3. This was correlated with a predominantly higher impact of nanoparticle than ionic silver form on parameters of oxidative stress, although no severe damage to important biomolecules was observed. Nevertheless, all types of treatments caused mobilization of antioxidant machinery, especially in leaves, although surface coatings modulated the activation of its specific components. Most effects induced by AgNPs or AgNO3 were alleviated with addition of cysteine.

Surface Coating-Modulated Phytotoxic Responses of Silver Nanoparticles in Plants and Freshwater Green Algae.

Silver nanoparticles (AgNPs) have been implemented in a wide range of commercial products, resulting in their unregulated release into aquatic as well as terrestrial systems. This raises concerns over their impending environmental effects. Once released into the environment, they are prone to various transformation processes that modify their reactivity. In order to increase AgNP stability, different stabilizing coatings are applied during their synthesis. However, coating agents determine particle size and shape and influence their solubility, reactivity, and overall stability as well as their behavior and transformations in the biological medium. In this review, we attempt to give an overview on how the employment of different stabilizing coatings can modulate AgNP-induced phytotoxicity with respect to growth, physiology, and gene and protein expression in terrestrial and aquatic plants and freshwater algae.

Coating-Dependent Effects of Silver Nanoparticles on Tobacco Seed Germination and Early Growth.

Silver nanoparticles (AgNPs) are used in a wide range of consumer products because of their excellent antimicrobial properties. AgNPs released into the environment are prone to transformations such as aggregation, oxidation, or dissolution so they are often stabilised by coatings that affect their physico-chemical properties and change their effect on living organisms. In this study we investigated the stability of polyvinylpyrrolidone (PVP) and cetyltrimethylammonium bromide (CTAB) coated AgNPs in an exposure medium, as well as their effect on tobacco germination and early growth. AgNP-CTAB was found to be more stable in the solid Murashige and Skoog (MS) medium compared to AgNP-PVP. The uptake and accumulation of silver in seedlings was equally efficient after exposure to both types of AgNPs. However, AgNP-PVP induced only mild toxicity on seedlings growth, while AgNP-CTAB caused severe negative effects on all parameters, even compared to AgNO3. Moreover, CTAB coating itself exerted negative effects on growth. Cysteine addition generally alleviated AgNP-PVP-induced negative effects, while it failed to improve germination and growth parameters after exposure to AgNP-CTAB. These results suggest that the toxic effects of AgNP-PVP are mainly a consequence of release of Ag+ ions, while phytotoxicity of AgNP-CTAB can rather be ascribed to surface coating itself.

Effect of coal mining activities and related industry on composition, cytotoxicity and genotoxicity of surrounding soils.

Coal mining and related industries each leave their characteristic "metal fingerprint" in the surrounding soils. Although geochemical investigations of such soils most often indicate heavy contamination with certain metals and bioassays point to their cytotoxic and genotoxic effects, the majority of studies are based on only one of the mentioned approaches. Here, the presented study investigated the effect of coal mining activities and related industry on surrounding soils by means of both geochemical and biological tools. The multielement composition of soils and associated eluates were used for the assessment of soil contamination level and the element bioavailable fractions, respectively. For cytotoxicity and genotoxicity evaluation, shallot (Allium ascalonicum L.) roots were exposed to selected soil eluates. Root growth, frequency of mitosis, mitotic and chromosomal abnormalities in root meristem cells, level of lipid peroxidation, and DNA damage evaluated by a comet assay were scored as toxicity endpoints. The results point to significant differences in the composition of collected soils and a variety of factors that contribute not only to their total metal load but also to the observed cytotoxic and genotoxic effects; all of which emphasize the necessity of a multidisciplinary approach in assessing the impact of anthropogenic activities on the environment, especially in historical mining areas.

Comparative proteomic study of phytotoxic effects of silver nanoparticles and silver ions on tobacco plants.

Widespread application of silver nanoparticles (AgNPs), due to their antibacterial and antifungal properties, increases their release into the environment and potential detrimental impact on living organisms. Plants may serve as a potential pathway for AgNPs bioaccumulation and a route into the food chain, hence investigation of AgNP phytotoxic effects are of particular importance. Since proteins are directly involved in stress response, studies of their abundance changes can help elucidate the mechanism of the AgNP-mediated phytotoxicity. In this study, we investigated proteomic changes in tobacco (Nicotiana tabacum) exposed to AgNPs and ionic silver (AgNO3). A high overlap of differently abundant proteins was found in root after exposure to both treatments, while in leaf, almost a half of the proteins exhibited different abundance level between treatments, indicating tissue-specific responses. Majority of the identified proteins were down-regulated in both tissues after exposure to either AgNPs or AgNO3; in roots, the most affected proteins were those involved in response to abiotic and biotic stimuli and oxidative stress, while in leaf, both treatments had the most prominent effect on photosynthesis-related proteins. However, since AgNPs induced higher suppression of protein abundance than AgNO3, we conclude that AgNP effects can, at least partially, be attributed to nanoparticle form.

Physiological, ultrastructural and proteomic responses of tobacco seedlings exposed to silver nanoparticles and silver nitrate.

Since silver nanoparticles (AgNPs) are a dominant nanomaterial in consumer products, there is growing concern about their impact on the environment. Although numerous studies on the effects of AgNPs on living organisms have been conducted, the interaction of AgNPs with plants has not been fully clarified. To reveal the plant mechanisms activated after exposure to AgNPs and to differentiate between effects specific to nanoparticles and ionic silver, we investigated the physiological, ultrastructural and proteomic changes in seedlings of tobacco (Nicotiana tabacum) exposed to commercial AgNPs and ionic silver (AgNO3) from the seed stage. A higher Ag content was measured in seedlings exposed to AgNPs than in those exposed to the same concentration of AgNO3. However, the results on oxidative stress parameters obtained revealed that, in general, higher toxicity was recorded in AgNO3-treated seedlings than in those exposed to nanosilver. Ultrastructural analysis of root cells confirmed the presence of silver in the form of nanoparticles, which may explain the lower toxicity of AgNPs. However, the ultrastructural changes of chloroplasts as well as proteomic study showed that both AgNPs and AgNO3 can affect photosynthesis. Moreover, the majority of the proteins involved in the primary metabolism were up-regulated after both types of treatments, indicating that enhanced energy production, which can be used to reinforce defensive mechanisms, enables plants to cope with silver-induced toxicity.

Phytotoxic effects of silver nanoparticles in tobacco plants.

The small size of nanoparticles (NPs), with dimensions between 1 and 100 nm, results in unique chemical and physical characteristics, which is why they are implemented in various consumer products. Therefore, an important concern is the potential detrimental impact of NPs on the environment. As plants are a vital part of ecosystem, investigation of the phytotoxic effects of NPs is particularly interesting. This study investigated the potential phytotoxicity of silver nanoparticles (AgNPs) on tobacco (Nicotiana tabacum) plants and compared it with the effects of the same AgNO3 concentrations. Accumulation of silver in roots and leaves was equally efficient after both AgNP and AgNO3 treatment, with predominant Ag levels found in the roots. Exposure to AgNPs did not result in elevated values of oxidative stress parameters either in roots or in leaves, while AgNO3 induced oxidative stress in both plant tissues. In the presence of both AgNPs and AgNO3, root meristem cells became highly vacuolated, which indicates that vacuoles might be the primary storage target for accumulated Ag. Direct AgNP uptake by root cells was confirmed. Leaf ultrastructural studies revealed changes mainly in the size of chloroplasts of AgNP-treated and AgNO3-treated plants. All of these findings indicate that nano form of silver is less toxic to tobacco plants than silver ions.

Toxicity of silver ions and differently coated silver nanoparticles in Allium cepa roots.

Silver nanoparticles (AgNPs) are the dominating nanomaterial in consumer products due to their well-known antibacterial and antifungal properties. To enhance their properties, different surface coatings may be used, which affect physico-chemical properties of AgNPs. Due to their wide application, there has been concern about possible environmental and health consequences. Since plants play a significant role in accumulation and biodistribution of many environmentally released substances, they are also very likely to be influenced by AgNPs. In this study we investigated the toxicity of AgNO3 and three types of laboratory-synthesized AgNPs with different surface coatings [citrate, polyvinylpyrrolidone (PVP) and cetyltrimethylammonium bromide (CTAB)] on Allium cepa roots. Ionic form of Ag was confirmed to be more toxic than any of the AgNPs applied. All tested AgNPs caused oxidative stress and exhibited toxicity only when applied in higher concentrations. The highest toxicity was recorded for AgNPs-CTAB, which resulted with increased Ag uptake in the roots, consequently leading to strong reduction of the root growth and oxidative damage. The weakest impact was found for AgNPs-citrate, much bigger, negatively charged NPs, which also aggregated to larger particles. Therefore, we can conclude that the toxicity of AgNPs is directly correlated with their size, overall surface charge and/or surface coating.

Modulation of urokinase plasminogen activator system by poly(ADP-ribose)polymerase-1 inhibition.

The urokinase plasminogen activator (uPA) system is a complex regulator of extracellular proteolysis which is involved in various physiological and pathological processes. The major components of this system are the serine protease uPA, two inhibitors PAI-1 and PAI-2, and the receptor uPAR. It has been previously shown by several groups that the uPA system has an important role in cancer progression and therefore its possible prognostic and therapeutic value has been evaluated. The aim of this study is to tackle the role of poly(ADP-ribosyl)ation in the induction of uPA activity in a glioblastoma cell line, A1235. This cell line is sensitive to alkylation damage and is a model for drug treatment. The components of the uPA system and the level of DNA damage were analyzed after alkylation agent treatment in combination with poly(ADP-ribose)polymerase-1 (PARP-1) inhibition. Here we show that the increase in uPA activity results from the net balance change between uPA and its inhibitor at mRNA level. Further, PARP-1 inhibition exerts its influence on uPA activity through DNA damage increase. Involvement of several signaling pathways, as well as cell specific regulation influencing the uPA system are discussed.

Removal of landfill leachate toxicity and genotoxicity by two treatment methods.

Leachates from active and closed municipal solid waste landfills can be a major source of contamination to groundwater and surface waters. In the present study the toxic and genotoxic potential of leachate from an old sanitary landfill prior to and following chemical and electrochemical treatments were assessed using Lemna, Allium, and comet tests. Photosynthetic pigments, malondialdehyde (indicator of lipid peroxidation) and antioxidant enzyme activities were evaluated as additional indicators of toxicity in duckweed. Following duckweed exposure to 25 % dilution of landfill leachate, growth rate and photosynthetic pigments content significantly decreased while lipid peroxidation increased despite stimulation of antioxidative defence mechanisms. Diluted leachate induced DNA strand breaks in duckweed cells as evidenced by the comet assay. Regarding the Allium test, untreated leachate caused inhibition of Allium cepa cell division and induction of mitotic and chromosomal aberrations. Although both water treatments completely reduced genotoxicity of leachate, the electrochemical method was found to be more efficient in removing toxic substances present in landfill leachate and thus more suitable for treating such leachates prior to their discharge into the environment. As landfill leachates pose a risk to human health and environment in general due to their (geno)toxicity, the present study demonstrates that the ecotoxicity/genotoxicity assays should be used in leachate risk assessment together with physicochemical analysis.

Proteomics of heavy metal toxicity in plants.

Plants endure a variety of abiotic and biotic stresses, all of which cause major limitations to production. Among abiotic stressors, heavy metal contamination represents a global environmental problem endangering humans, animals, and plants. Exposure to heavy metals has been documented to induce changes in the expression of plant proteins. Proteins are macromolecules directly responsible for most biological processes in a living cell, while protein function is directly influenced by posttranslational modifications, which cannot be identified through genome studies. Therefore, it is necessary to conduct proteomic studies, which enable the elucidation of the presence and role of proteins under specific environmental conditions. This review attempts to present current knowledge on proteomic techniques developed with an aim to detect the response of plant to heavy metal stress. Significant contributions to a better understanding of the complex mechanisms of plant acclimation to metal stress are also discussed.

The effects of cadmium-zinc interactions on biochemical responses in tobacco seedlings and adult plants.

The objective of the present study was to investigate the effects of cadmium-zinc (Cd-Zn) interactions on their uptake, oxidative damage of cell macromolecules (lipids, proteins, DNA) and activities of antioxidative enzymes in tobacco seedlings as well as roots and leaves of adult plants. Seedlings and plants were exposed to Cd (10 µM and 15 µM) and Zn (25 µM and 50 µM) as well as their combinations (10 µM or 15 µM Cd with either 25 µM or 50 µM Zn). Measurement of metal accumulation exhibited that Zn had mostly positive effect on Cd uptake in roots and seedlings, while Cd had antagonistic effect on Zn uptake in leaves and roots. According to examined oxidative stress parameters, in seedlings and roots individual Cd treatments induced oxidative damage, which was less prominent in combined treatments, indicating that the presence of Zn alleviates oxidative stress. However, DNA damage found in seedlings, and lower glutathione reductase (GR) and superoxide dismutase (SOD) activity recorded in both seedlings and roots, after individual Zn treatments, indicate that Zn accumulation could impose toxic effects. In leaves, oxidative stress was found after exposure to Cd either alone or in combination with Zn, thus implying that in this tissue Zn did not have alleviating effects. In conclusion, results obtained in different tobacco tissues suggest tissue-dependent Cd-Zn interactions, which resulted in activation of different mechanisms involved in the protection against metal stress.

The efficiency of combined CaO/electrochemical treatment in removal of acid mine drainage induced toxicity and genotoxicity.

Acid mine drainage (AMD) is a by-product of the mining industry that has a detrimental effect on aquatic plant and animal life due to high load of heavy metals and sulfates. In the present study, the toxic and genotoxic potential of AMD prior to and following combination of neutralization/electrocoagulation processes was evaluated using several bioassays and selected parameters. Regardless of pH correction of AMD prior to Daphnia bioassay, high acute toxicity was observed in Daphnia magna. The mine leachate also induced strong phyto-, cyto- and genotoxicity to Allium cepa roots. Short term exposure to AMD inhibited duckweed growth and chlorophyll a content and simultaneously promoted lipid peroxidation and DNA damage despite duckweed capability to upregulate antioxidative defense mechanisms. The results show that observed (geno)toxicity could be related to oxidative stress most probably induced by toxic metal action. However, influence of low pH as a contributing factor in the phytotoxicity of AMD cannot be excluded. The application of combined treatment eliminated genotoxicity and was highly efficient in reducing toxicity of AMD. Thus, the method seems to be suitable for treatment of AMD waters enabling their safe discharge to an aquatic environment.

Assessment of surface water in the vicinity of fertilizer factory using fish and plants.

The genotoxic and toxic potential of polluted surface water exposed to a fertilizer factory effluent was evaluated using assays with fish (Cyprinus carpio) and plant (Lemna minor) model organisms. Beside classical physicochemical parameters, the contents of fluorides, some heavy metals and polycyclic aromatic hydrocarbons were analyzed as well. Surface water caused inhibition of plant growth and decrease of photosynthetic pigment content. Regarding DNA damage and oxidative stress parameters, both fish and plants showed similar response to the surface water. In confirmation to biochemical markers, histopathological analysis of gill and liver tissues revealed a higher incidence of lesions in fish exposed to polluted surface water. Generally, results obtained by biological monitoring were mostly in agreement with chemical analysis of the surface water, although several discrepancies were observed which might be due to difference in sensitivity of model organisms or in experimental conditions (laboratory and field exposure). The results imply that conventional chemical analysis should be extended to genotoxicity/toxicity assays as measured biological effects and the potential health hazard cannot be predicted based on the physicochemical characteristics of water samples alone.

Toxicological characterization of the landfill leachate prior/after chemical and electrochemical treatment: a study on human and plant cells.

In this research, toxicological safety of two newly developed methods for the treatment of landfill leachate from the Piskornica (Croatia) sanitary landfill was investigated. Chemical treatment procedure combined chemical precipitation with CaO followed by coagulation with ferric chloride and final adsorption by clinoptilolite. Electrochemical treatment approach included pretreatment with ozone followed by electrooxidation/electrocoagulation and final polishing by microwave irradiation. Cell viability of untreated/treated landfill leachate was examined using fluorescence microscopy. Cytotoxic effect of the original leachate was obtained for both exposure periods (4 and 24 h) while treated samples showed no cytotoxic effect even after prolonged exposure time. The potential DNA damage of the untreated/treated landfill leachate was evaluated by the comet assay and cytokinesis-block micronucleus (CBMN) assay using either human or plant cells. The original leachate exhibited significantly higher comet assay parameters compared to negative control after 24 h exposure. On the contrary, there was no significant difference between negative control and chemically/electrochemically treated leachate for any of the parameters tested. There was also no significant increase in either CBMN assay parameter compared to the negative control following the exposure of the lymphocytes to the chemically or electrochemically treated landfill leachate for both exposure periods while the original sample showed significantly higher number of micronuclei, nucleoplasmic bridges and nuclear buds for both exposure times. Results suggest that both methods are suitable for the treatment of such complex waste effluent due to high removal efficiency of all measured parameters and toxicological safety of the treated effluent.

Cadmium and zinc induced similar changes in protein and glycoprotein patterns in tobacco (Nicotiana tabacum l.) seedlings and plants.

The effects of 10 µmol L-1 and 15 µmol L-1 cadmium (Cd), a nonessential toxic element and 25 µmol L-1 and 50 µmol L-1 zinc (Zn), an essential micronutrient, on proteins and glycoproteins of Nicotiana tabacum L. seedlings and plants were investigated after exposure to each metal alone or to their combinations. Changes in only few polypeptides related to heavy metal treatments were observed in tobacco seedlings and leaves of adult plants, while the greatest change in total soluble protein pattern was observed in plant roots. Differences between control and treated tobacco tissues were more pronounced in the glycoprotein pattern, which was analysed by application of different lectins. The majority of the detected glycoproteins in leaves and roots of adult plants can be considered as a result of enhanced glycosylation due to heavy metal stress. The difference in glycoproteins between Cd and Zn application on tobacco seedlings and adult plants could not be determined since enhanced glycosylation was noticed after treatment with either metal alone or in combination. Therefore, it can be concluded that both metals induced N- and Oglycosylation as a result of changed environmental conditions.