Toxicity Of ZnO Nanoparticles Research Articles

Role of soluble zinc in ZnO nanoparticle cytotoxicity in Daphnia magna: A morphological approach

The role of soluble zinc has been determined in Daphnia magna by a morphological approach, integrating a previous paper in which the ultrastructural damages to gut epithelial cells have been studied after ZnO nanoparticles exposure. In the present paper, the toxicity and morphological effects of soluble zinc from ZnSO4 have been determined in a 48-h acute exposure test. Daphnids have been exposed to six nominal zinc concentrations (0.075, 0.15, 0.3, 0.6, 1.2, and 2.4mg Zn/L) and then fixed for microscopic analyses. Data from the acute toxicity tests gave an EC50 value of 0.99mg/L and showed that no immobilization appeared up to 0.3mg Zn/L. Ultrastructural analyses of samples from the two highest concentrations showed large vacuolar structures, swelling of mitochondria, multilamellar bodies, and a great number of autophagy vacuoles. These findings have been compared to those from our previous study, and similarities and/or differences discussed. Based on the overall results it can be concluded that dissolved zinc ions played a key role in ZnO nanoparticle toxicity and that the morphological approach is an extremely useful tool for comparing toxicological effects as well. A possible common toxic mechanism of soluble zinc and ZnO nanoparticles was also proposed.

Evaluating the Combined Toxicity of Cu and ZnO Nanoparticles: Utility of the Concept of Additivity and a Nested Experimental Design.

Little is understood regarding the effects of mixtures of different metal-based nanoparticles (NPs). Using concentration-addition (CA) and independent-action (IA) models, we evaluated the combined toxicity of Cu and ZnO NPs based on five nested combinations, i.e., Cu(NO3)2-CuNPs, Zn(NO3)2-ZnONPs, Cu(NO3)2-ZnONPs, Zn(NO3)2-CuNPs, and CuNPs-ZnONPs on root elongation of Lactuca sativa L. The CA and IA models performed equally well in estimating the toxicity of mixtures of Cu(NO3)2-CuNPs, Zn(NO3)2-ZnONPs, and Zn(NO3)2-CuNPs, whereas the IA model was significantly better for fitting the data of Cu(NO3)2-ZnONPs and CuNPs-ZnONPs mixtures. Dissolved Cu proved to be the most toxic metal species to lettuce roots in the tests, followed by Cu NPs, dissolved Zn, and ZnO NPs, respectively. An antagonistic effect was observed for ZnO NPs on the toxicity of Cu NPs. This antagonistic effect is expected to be the result of interactions between dissolved Cu and dissolved Zn, particulate Zn and dissolved Zn, particulate Cu and dissolved Zn, and between particulate Zn and dissolved Cu. In general terms, assuming additivity gives a first indication of the combined toxicity with soluble and insoluble metal particles, both being important in driving the toxicity of metal-based NPs to higher plants.

Effect of flexing and massage on in vivo human skin penetration and toxicity of zinc oxide nanoparticles.

We assessed the effects of flexing and massage on human skin penetration and toxicity of topically applied coated and uncoated zinc oxide nanoparticles (˜75 nm) in vivo. Noninvasive multiphoton tomography with fluorescence lifetime imaging was used to evaluate the penetration of nanoparticles through the skin barrier and cellular apoptosis in the viable epidermis. All nanoparticles applied to skin with flexing and massage were retained in the stratum corneum or skin furrows. No significant penetration into the viable epidermis was seen and no cellular toxicity was detected. Exposure of normal in vivo human skin to these nanoparticles under common in-use conditions of flexing or massage is not associated with significant adverse events.

The neglected nano-specific toxicity of ZnO nanoparticles in the yeast Saccharomyces cerevisiae.

Nanoparticles (NPs) with unique physicochemical properties induce nano-specific (excess) toxicity in organisms compared with their bulk counterparts. Evaluation and consideration of nano-specific toxicity are meaningful for the safe design and environmental risk assessment of NPs. However, ZnO NPs have been reported to lack excess toxicity for diverse organisms. In the present study, the nano-specific toxicity of ZnO NPs was evaluated in the yeast Saccharomyces cerevisiae. Nano-specific toxicity of ZnO NPs was not observed in the wild type yeast. However, the ZnO NPs induced very similar nano-specific toxicities in the three mutants with comparable log Te (particle) values (0.64 vs 0.65 vs 0.62), suggesting that the mutants were more sensitive and specific for the NPs’ nano-specific toxicity. The toxic effects in the yeast were slightly attributable to dissolved zinc ions from the ZnO (nano or bulk) particles. Oxidative damage and mechanical damage contributed to the toxic effect of the ZnO particles. The mechanism of mechanical damage is proposed to be an inherent characteristic underlying the nano-specific toxicity in the mutants. The log Te (particle) was a useful parameter for evaluation of NPs nano-specific toxicity, whereas log Te (ion) efficiently determined the NPs toxicity associated with released ions.

Toxicity of ZnO nanoparticles on germinating Sesamum indicum (Co-1) and their antibacterial activity

A comparative study of chemically (ZnO) and biologically synthesized (nano-ZnO) nanoparticles were carried out to determine the effect on seed germination of Sesamum indicum (Co-1) by soaking method. Nano-ZnO is synthesized using Lantana aculeata aqueous extract. Chemical synthesis of ZnO nanoparticles by precipitate method and was characterized by ultraviolet–Visible spectroscopy (UV–Vis), Fourier transform infrared spectrometer (FT-IR), energy dispersive X-ray spectrometer (EDX), X-ray diffractometer (XRD), field emission scanning electron microscopy (FESEM) and high resolution transmission electron microscopy (HRTEM). Antibacterial activity against pathogens was determined using well diffusion method. All the characterization analysis revealed that ZnO and nano-ZnO nanoparticles were spherical in shape with an average particle size of 18±2 and 12±3 nm, respectively. Antibacterial studies conclude that nano-ZnO NPs have maximum zone of inhibition which was observed in Pseudomonas aeruginosa (15.60±1.0 mm) at 100 μg ml−1 concentration when compared to other ZnO NPs. Phytomediate ZnO have no adverse effects on seed germination, root elongation on S. indicum. But chemically synthesized ZnO nanoparticles significantly decreased in germination of S. indicum-treated samples and no changes were observed in bulk ZnO. These results clearly indicate the benefits of using bio-fabricate ZnO nanoparticles, i.e., more efficient in germination of S. indicum and can also act as antibacterial agent. It can be used as nanofertilizer in environmental aspect of agricultural development.

The toxicity of zinc oxide nanoparticles to Lemna minor (L.) is predominantly caused by dissolved Zn

Nano-ZnO particles have been reported to be toxic to many aquatic organisms, although it is debated whether this is caused by nanoparticles per sé, or rather dissolved Zn. This study investigated the role of dissolved Zn in nano-ZnO toxicity to Lemna minor. The technical approach was based on modulating nano-ZnO dissolution by either modifying the pH of the growth medium and/or surface coating of nano-ZnO, and measuring resulting impacts on L. minor growth and physiology. Results show rapid and total dissolution of nano-ZnO in the medium (pH 4.5). Quantitatively similar toxic effects were found when L. minor was exposed to nano-ZnO or the “dissolved Zn equivalent of dissolved nano-ZnO”. The conclusion that nano-ZnO toxicity is primarily caused by dissolved Zn was further supported by the observation that phytotoxicity was absent on medium with higher pH-values (>7), where dissolution of nano-ZnO almost ceased. Similarly, the reduced toxicity of coated nano-ZnO, which displays a slower Zn dissolution, is also consistent with a major role for dissolved Zn in nano-ZnO toxicity.

Continuous ultraviolet irradiation increases the adverse effects of photoreactive nanoparticles on the early development of Oryzias latipes.

The present study investigated the toxicity of photoreactive nanoparticles (NPs) on the development of Oryzias latipes. Buoyant fish embryos are potentially vulnerable to sunlight-derived ultraviolet (UV) irradiation. Zinc oxide (ZnO) NPs in surface water easily absorb UV irradiation from transmitted solar light. In the present study, O. latipes were exposed to ZnO NPs under irradiation with UV or visible light. The ZnO NPs exhibited considerable toxicity toward embryos and sac fry following UV irradiation, and these toxic effects resulted in increased mortality and abnormalities. The UV irradiation induced more serious effects on embryos than did visible light irradiation, and embryonic exposure resulted in irreversible developmental impairment or death of sac fry. The adverse effects of ZnO NPs may result from Zn ions released from photoreactive ZnO NPs. The present study demonstrates photo-dependent developmental impairment of O. latipes embryos as a result of exposure to ZnO NPs. The results demonstrate that the toxicity of photoreactive ZnO NPs could vary under environmentally relevant UV irradiation. These data could serve as a guide for evaluation of the toxicity of photo-activated NPs in natural surface waters and could be useful for the ecological risk assessment of photoreactive NPs.

The bioavailability and toxicity of ZnO and Ni nanoparticles and their bulk counterparts in different sediments

The intensive development of nanotechnology raises a question of the potential consequences of the presence of nanoparticles (NPs) in the different components of the environment, including sediments. The aim of this study was to evaluate the toxicity of nanoparticles of ZnO and Ni and their bulk counterparts in bottom sediments (SD1, SD2) with different properties collected from the Vistula River in Poland. Sediment samples with NPs at a concentration of 100 mg kg−1 were incubated for 17 months in the dark or under a photoperiod of 12 h light/12 h dark. The Microtox® (bacteria, Vibrio fischeri) and OSTRACODTOXKIT F™ (ostracods, Heterocypris incongruens) tests were used to evaluate toxicity. In addition, the contents of Zn and Ni were determined in extracts (H2O and CaCl2) of the bottom sediments. The Zn concentration was much lower in the SD1 sediment with the addition of NPs/bulk particles (30–230 μg kg−1) compared to the SD2 sediment (280–1140 μg kg−1). The toxicity of ZnO and Ni was determined by the type of bottom sediment and the parameter studied. Both nano- and bulk-ZnO and Ni caused the mortality of H. incongruens at a level of 13.3–53.3 %. The influence of ZnO and Ni on the growth of H. incongruens was observed to be the opposite. ZnO resulted in growth stimulation, while Ni resulted in growth inhibition of H. incongruens. Both ZnO and Ni stimulated V. fisheri luminescence. In most cases, the incubation of ZnO and Ni under the photoperiod increased the toxicity or decreased the stimulation of V. fisheri bioluminescence and H. ingongruens growth compared to the dark-incubated sediments. The study provides new and important information on the ecotoxicological effects of ZnO and Ni nanoparticles in different sediments and under various environmental conditions that may be useful for the risk assessment of this new group of contaminants.

Sequestration of nanoparticles by an EPS matrix reduces the particle-specific bactericidal activity.

Most artificial nanomaterials are known to exhibit broad-spectrum bactericidal activity; however, the defence mechanisms that bacteria use based on extracellular polymeric substances (EPS) to detoxify nanoparticles (NPs) are not well known. We ruled out the possibility of ion-specific bactericidal activity by showing the lack of equivalent dissolved zinc and silicon toxicity and determined the particle-specific toxicity of ZnO and SiO2 nanoparticles (ZnONPs/SiO2NPs) through dialysis isolation experiments. Surprisingly, the manipulation of the E. coli EPS (i.e., no EPS manipulation or EPS removal by sonication/centrifugation) showed that their particle-specific bactericidal activity could be antagonized by NP-EPS sequestration. The survival rates of pristine E. coli (no EPS manipulation) reached 65% (ZnONPs, 500 mg L−1) and 79% (SiO2NPs, 500 mg L−1), whereas survival rates following EPS removal by sonication/centrifugation were 11% and 63%, respectively. Transmission electron microscopy (TEM) combined with fluorescence micro-titration analysis and Fourier-transform infrared spectroscopy (FTIR) showed that protein-like substances (N-H and C-N in amide II) and secondary carbonyl groups (C=O) in the carboxylic acids of EPS acted as important binding sites that were involved in NP sequestration. Accordingly, the amount and composition of EPS produced by bacteria have important implications for the bactericidal efficacy and potential environmental effects of NPs.

Uptake and toxicity of nano-ZnO in the plant-feeding nematode, Xiphinema vuittenezi: the role of dissolved zinc and nanoparticle-specific effects.

Nanoparticulate ZnO is one of the most commonly applied nanomaterials. As ZnO is more soluble than many other oxide nanoparticles, its toxicity beyond the nanoparticle-specific effects can be attributed to the dissolved ionic zinc. The investigation of uptake and toxicity of nano-ZnO in the plant-feeding nematode, Xiphinema vuittenezi, which was used in previous studies as a biological model organism, was aimed. The establishment of the role of dissolved zinc and nanoparticle-specific effects in the toxicity was also the objective of our study. Zn uptake was found to be significantly higher for bulk and nano-ZnO than for ZnSO4 solution; however, treatments caused loss of potassium in the worms in a dissolved-zinc-dependent manner. The toxicity was the lowest for bulk ZnO, and it was very similar for nano-ZnO and ZnSO4 solution. Accordingly, the toxicity of ZnO nanoparticles is a combination of dissolved-zinc-caused toxicity and nanoparticle-specific effects.

Iron sulfide attenuates the methanogenic toxicity of elemental copper and zinc oxide nanoparticles and their soluble metal ion analogs

Elemental copper (Cu0) and zinc oxide (ZnO) nanoparticle (NP) toxicity to methanogens has been attributed to the release of soluble metal ions. Iron sulfide (FeS) partially controls the soluble concentration of heavy metals and their toxicity in aquatic environments. Heavy metals displace the Fe from FeS forming poorly soluble metal sulfides in the FeS matrix. Therefore, FeS may be expected to attenuate the NP toxicity. This work assessed FeS as an attenuator of the methanogenic toxicity of Cu0 and ZnO NPs and their soluble salt analogs. The toxicity attenuation capacity of fine (25–75μm) and coarse (500 to 1200μm) preparations of FeS (FeS-f and FeS-c respectively) was tested in the presence of highly inhibitory concentrations of CuCl2, ZnCl2 Cu0 and ZnO NPs. FeS-f attenuated methanogenic toxicity better than FeS-c. The results revealed that 2.5× less FeS-f than FeS-c was required to recover the methanogenic activity to 50% (activity normalized to uninhibited controls). The results also indicated that a molar FeS-f/Cu0 NP, FeS-f/ZnO NP, FeS-f/ZnCl2, and FeS-f/CuCl2 ratio of 2.14, 2.14, 4.28, and 8.56 respectively, was necessary to recover the methanogenic activity to >75%. Displacement experiments demonstrated that CuCl2 and ZnCl2 partially displaced Fe from FeS. As a whole, the results indicate that not all the sulfide in FeS was readily available to react with the soluble Cu and Zn ions which may explain the need for a large stoichiometric excess of FeS to highly attenuate Cu and Zn toxicity. Overall, this study provides evidence that FeS attenuates the toxicity caused by Cu0 and ZnO NPs and their soluble ion analogs to methanogens.

Zinc-Oxide Nanoparticles Exhibit Genotoxic, Clastogenic, Cytotoxic and Actin Depolymerization Effects by Inducing Oxidative Stress Responses in Macrophages and Adult Mice.

Zinc oxide nanoparticles (ZnO-NPs) have wide biological applications, which have raised serious concerns about their impact on the health and environment. Although, various studies have shown ZnO-NP toxicity on different cells underin vitroconditions, sufficient information is lacking regarding toxicity and underlying mechanisms underin vivoconditions. In this work, we investigated genotoxic, clastogenic, and cytotoxic effects of ZnO-NPs on macrophages and in adult mice. ZnO-NP-treated mice showed signs of toxicity such as loss in body weight, passive behavior and reduced survival. Further mechanistic studies revealed that administration of higher dose caused severe DNA damage in peripheral blood and bone marrow cells as evident by the formation of COMET tail, micronuclei, chromosomal fragmentation, and phosphorylation of H2A histone family member X. Moreover, ZnO-NPs inhibited DNA repair mechanism by downregulating the expression offen-1andpolBproteins. Histopathological examinations showed severe inflammation and damage to liver, lungs, and kidneys. Cell viability and wound healing assays revealed that ZnO-NPs killed macrophages in a dose-dependent manner, caused severe wounds and inhibited cellular migration by irreversible actin depolymerization and degradation. Reduction in the viability of macrophages was due to the arrest of the cell cycle at the G0/G1 phase, inhibition of superoxide dismutase and catalase and eventually reactive oxygen species. Furthermore, treatment with an antioxidant drug N-acetyl cysteine significantly reduced the ZnO-NP induced genotoxicity bothin vitroandin vivo Altogether, this study gives detailed pathological insights of ZnO-NP that impair cellular functions, thus will enable to arbitrate their biological applications.

Toxicity of zinc oxide nanoparticles to the annelid Enchytraeus crypticus in agar-based exposure media

AbstractToxicity of zinc oxide nanoparticle (ZnO-NPs) powder and water soluble salt of Zn (ZnCl

Metal homeostasis disruption and mitochondrial dysfunction in hepatocytes exposed to sub-toxic doses of zinc oxide nanoparticles

Increased production and use of zinc oxide nanoparticles (ZnO-NPs) in consumer products has prompted the scientific community to investigate their potential toxicity, and understand their impact on the environment and organisms. Molecular mechanisms involved in ZnO-NP toxicity are still under debate and focus essentially on high dose expositions. In our study, we chose to evaluate the effect of sub-toxic doses of ZnO-NPs on human hepatocytes (HepG2) with a focus on metal homeostasis and redox balance disruptions. We showed massive dissolution of ZnO-NPs outside the cell, transport and accumulation of zinc ions inside the cell but no evidence of nanoparticle entry, even when analysed by high resolution TEM microscopy coupled with EDX. Gene expression analysis highlighted zinc homeostasis disruptions as shown by metallothionein 1X and zinc transporter 1 and 2 (ZnT1, ZnT2) over-expression. Major oxidative stress response genes, such as superoxide dismutase 1, 2 and catalase were not induced. Phase 2 enzymes in term of antioxidant response, such as heme oxygenase 1 (HMOX1) and the regulating subunit of the glutamate-cysteine ligase (GCLM) were slightly upregulated, but these observations may be linked solely to metal homeostasis disruptions, as these actors are involved in both metal and ROS responses. Finally, we observed abnormal mitochondria morphologies and autophagy vesicles in response to ZnO-NPs, indicating a potential role of mitochondria in storing and protecting cells from zinc excess but ultimately causing cell death at higher doses.

Effects of Stabilized Nanoparticles of Copper, Zinc, Manganese, and Iron Oxides in Low Concentrations on Lettuce (Lactuca sativa) Seed Germination: Nanotoxicants or Nanonutrients?

Information on the phytotoxicity of nanoparticles (NPs) at low concentrations (e.g., ppb to low ppm) is scarce. Therefore, this study was conducted to assess the effects of laboratory-prepared Cu, Zn, Mn, and Fe oxide NPs in low concentrations (<50 ppm) on the germination of lettuce (Lactuca sativa) seeds in a water medium. The data showed that CuO NPs were slightly more toxic than Cu ions while the toxicity of ZnO NPs was similar to that of Zn ions, and MnOx NPs and FeOx NPs were not only less toxic than their ionic counterparts but also significantly stimulated the growth of lettuce seedlings by 12–54 %. This study showed that manufactured NPs were not always more toxic than other chemical species containing the same elements. Instead, Mn or Fe NPs can significantly enhance plant growth and have the potential to be effective nanofertilizers for increasing agronomic productivity.

Mechanism of oxidative stress involved in the toxicity of ZnO nanoparticles against eukaryotic cells

ZnO NPs (zinc oxide nanoparticles) has generated significant scientific interest as a novel antibacterial and anticancer agent. Since oxidative stress is a critical determinant of ZnO NPs-induced damage, it is necessary to characterize their underlying mode of action. Different structural and physicochemical properties of ZnO NPs such as particle surface, size, shape, crystal structure, chemical position, and presence of metals can lead to changes in biological activities including ROS (reactive oxygen species) production. However, there are some inconsistencies in the literature on the relation between the physicochemical features of ZnO NPs and their plausible oxidative stress mechanism. Herein, the possible oxidative stress mechanism of ZnO NPs was reviewed. This is worthy of further detailed evaluations in order to improve our understanding of vital NPs characteristics governing their toxicity. Therefore, this study focuses on the different reported oxidative stress paradigms induced by ZnO NPs including ROS generated by NPs, oxidative stress due to the NPs-cell interaction, and role of the particle dissolution in the oxidative damage. Also, this study tries to characterize and understand the multiple pathways involved in oxidative stress induced by ZnO NPs. Knowledge about different cellular signaling cascades stimulated by ZnO NPs lead to the better interpretation of the toxic influences induced by the cellular and acellular parameters. Regarding the potential benefits of toxic effects of ZnO NPs, in-depth evaluation of their toxicity mechanism and various effects of these nanoparticles would facilitate their implementation for biomedical applications.

Physicochemical transformation and algal toxicity of engineered nanoparticles in surface water samples

Most studies on the behavior and toxicity of engineered nanoparticles (NPs) have been conducted in artificial water with well-controlled conditions, which are dramatically different from natural waters with complex compositions. To better understand the fate and toxicity of NPs in the natural water environment, physicochemical transformations of four NPs (TiO2, ZnO, Ag, and carbon nanotubes (CNTs)) and their toxicities towards a unicellular green alga (Chlorella pyrenoidosa) in four fresh water and one seawater sample were investigated. Results indicated that water chemistry had profound effects on aggregation, dissolution, and algal toxicity of the NPs. The strongest homoaggregation of the NPs was associated with the highest ionic strength, but no obvious correlation was observed between the homoaggregation of NPs and pH or dissolved organic matter content of the water samples. The greatest dissolution of ZnO NPs also occurred in seawater with the highest ionic strength, while the dissolution of Ag NPs varied differently from ZnO NPs. The released Zn2+ and especially Ag+ mainly accounted for the algal toxicity of ZnO and Ag NPs, respectively. The NP-cell heteroagglomeration occurred generally for CNTs and Ag NPs, which contributed to the observed nanotoxicity. However, there was no significant correlation between the observed nanotoxicity and the type of NP or the water chemistry. It was thus concluded that the physicochemical transformations and algal toxicities of NPs in the natural water samples were caused by the combined effects of complex water quality parameters rather than any single influencing factor alone. These results will increase our knowledge on the fate and effects of NPs in the aquatic environment.

Nanoinformatics for environmental health and biomedicine.

Nanoinformatics for environmental health and biomedicine.

Evaluation of Different Oxidative Stress Parameters and Apoptosis in Human Cervical Cancer Cells Exposed to Rod and Spherical Shaped Zinc Oxide Nanoparticles

Zinc oxide nanoparticles are presently being used in cosmetics, paints, biosensors, and medical applications. However, zinc oxide nanoparticles are broadly used in the cosmetic industry for many years; recently, they have been explored for use in cancer therapy. This study was intended to comprehend the mechanism of the differential cytotoxicity of rod and spherical shaped zinc oxide nanoparticles to cervical cancer cells HeLa and SiHa. The role of ion dissolution in the toxicity of zinc oxide nanoparticles was also investigated. Zinc oxide nanorod induced significantly higher oxidative stress, reactive oxygen species compared to spherical shaped zinc oxide nanoparticles. Additionally, rod shaped nanoparticles activated the NF-κB signaling pathway. Besides, zinc oxide nanorod also induced mitochondrial membrane damage leading to apoptosis as observed by PARP cleavage and reduction of Phospho-Bad. In conclusion, zinc oxide nanorod, after proper tailoring, may have potential to use as chemotherapeutic agents against cervical cancer.

Testing nanoeffect onto model bacteria: Impact of speciation and genotypes

The gram-negative bacteria Escherichia coli (E. coli) is a very useful prokaryotic model for testing the toxicity of ZnO nanoparticles (nano-ZnO). This toxicity is often linked to Zn2+ released from nanoparticles in the culture medium, and nano-ZnO dissolution in different media is clearly established. Here, two model E. coli strains MG1655 and W3110 both descendant from the original K-12 showing slight differences in their genome were submitted to nano-ZnO or Zn2+ in order 1 > to refine the nano-ZnO toxicity mechanisms to E. coli, and 2 > to investigate whether toxicity resulted from a real “nanoparticle” effect or from the release of Zn2+ in solution. To do so, both strains were submitted to various concentrations (i.e., 0.1–1 mM) of nano-ZnO or Zn2+ in Luria Bertani (LB) medium. These toxicity studies take into account the nano-ZnO solubility in the culture medium by specifically monitoring the Zn2+ release in our experimental systems. In our experimental conditions, differences in tolerance to nano-ZnO or Zn2+ between both strains were clearly evidenced. W3110 is generally more tolerant to metal than MG1655, the latter showing no real difference in its sensitivity to the two zinc added forms unlike W3110. The differences in behavior between both strains could be attributed to differences in the two genomes as a mutation named “amber” in W3110. Moreover, by using these two closely E. coli strains, a real “nano” effect is here clearly demonstrated providing a model to study the toxicity of ZnO nanoparticles.