Impact of polyethylene microbeads on the floating freshwater plant duckweed Lemna minor.

Microplastics (MP), small plastic particles below 5 mm, have become one of the central concerns of environmental risk assessment. Microplastics are continuously being released into the aquatic environment either directly through consumer products or indirectly through fragmentation of larger plastic materials. The aim of our study was to investigate the effect of polyethylene microbeads from cosmetic products on duckweed (Lemna minor), a freshwater floating plant. The effects of microbeads from two exfoliating products on the specific leaf growth rate, the chlorophyll a and b content in the leaves, root number, root length and root cell viability were assessed. At the same time, water leachates from microbeads were also prepared to exclude the contribution of cosmetic ingredients on the measured impacts. Specific leaf growth rate and content of photosynthetic pigments in duckweed leaves were not affected by polyethylene microbeads, but these microbeads significantly affected the root growth by mechanical blocking. Sharp particles also reduced the viability of root cells, while the impact of microbeads with a smooth surface was neglected. It was concluded that microbeads from cosmetic products can also have negative impacts on floating plants in freshwater ecosystems.

Comparative study of acetylcholinesterase and glutathione S-transferase activities of closely related cave and surface Asellus aquaticus (Isopoda: Crustacea).

The freshwater isopod crustacean Asellus aquaticus has recently been developed as an emerging invertebrate cave model for studying evolutionary and developmental biology. Mostly morphological and genetic differences between cave and surface A. aquaticus populations have been described up to now, while scarce data are available on other aspects, including physiology. The purpose of this study was to advance our understanding of the physiological differences between cave A. aquaticus and its surface-dwelling counterparts. We sampled two surface populations from the surface section of the sinking Pivka River (central Slovenia, Europe), i.e. locality Pivka Polje, and locality Planina Polje, and one cave population from the subterranean section of the sinking Pivka River, i.e. locality Planina Cave. Animals were sampled in spring, summer and autumn. We measured the activities of acetylcholinesterase (AChE) and glutathione S-transferase (GST) in individuals snap-frozen in the field immediately after collection. Acetylcholinesterase is likely related to animals' locomotor activity, while GST activity is related to the metabolic activity of an organism. Our study shows significantly lower AChE and GST activities in the cave population in comparison to both surface A. aquaticus populations. This confirms the assumption that cave A. aquaticus have lower locomotor and metabolic activity than surface A. aquaticus in their respective natural environments. In surface A. aquaticus populations, seasonal fluctuations in GST activity were observed, while these were less pronounced in individuals from the more stable cave environment. On the other hand, AChE activity was generally season-independent in all populations. To our knowledge, this is the first study of its kind conducted in A. aquaticus. Our results show that among closely related cave and surface A. aquaticus populations also physiological differences are present besides the morphological and genetic. These findings contribute to a better understanding of the biology of A. aquaticus and cave crustaceans in general.

Uptake and effects of microplastic textile fibers on freshwater crustacean Daphnia magna.

Microplastic fibers (MP) from textile weathering and washing are increasingly being recognized as environmental pollutants. The majority of studies on the bioavailability and effects of microplastic focused on small polystyrene spherical plastic particles, while less data are available for fibers and for other materials besides polystyrene. We investigated the ingestion and effects of ground polyethylene terephthalate (PET) textile microfibers (length range: 62-1400 µm, width 31-528 µm, thickness 1-21.5 µm) on the freshwater zooplankton crustacean Daphnia magna after a 48 h exposure and subsequent 24 h of recovery in MP free medium and algae. The majority of ingested fibers by D. magna were around 300 µm, but also some very large twisted MP fibers around 1400 µm were found inside the gut. Exposure to these fibers results in increased mortality of daphnids after 48 h only in the case where daphnids were not pre-fed with algae prior to experiment, but no effect was found when daphnids were fed before the experiments. Regardless of the feeding regime, daphnids were not able to recover from MP exposure after additional 24 h incubation period in a MP free medium with algae. The uptake and effects of PET textile MP on D. magna are presented here for the first time.

In field conditions, commercial pigment grade TiO2 was not harmful to terrestrial isopods but reduced leaf litter fragmentation.

We investigated the effects of a commercial pigment grade rutile TiO2 on the terrestrial isopod Porcellio scaber in three locations that differed in terms of abiotic and biotic conditions: the laboratory, open air, and the closed barn. Mortality and isopod energy reserves (digestive gland total proteins, lipids and carbohydrates) were not affected following 14days exposure to up to 1000mg TiO2 per kg dry leaves (mg/kg) under any experimental scenario. However, in the field tests, isopods consumption of TiO2-coated leaves was reduced compared to that of uncoated leaves and the decrease was not dose-dependent. The highest reduction was in the closed barn (45-56%) rather than in the open-air (38-40%). In laboratory-based food choice tests, isopods neither preferred nor avoided leaves coated with TiO2, suggesting that rather than sensing the TiO2 on the leaves directly, the isopods under open-air and barn exposure responded to altered attractiveness and/or palatability of the TiO2 amended leaves. We propose that this could be due to altered microbial population on the leaves, a hypothesis that requires further investigation. Although short-term exposure to atmospheric deposition of up to 1000mg/kg commercial TiO2 is unlikely to pose an immediate threat to isopod mortality and energy balance, reduced leaf feeding may have implications for the decomposition of plant material.

The role of PVP in the bioavailability of Ag from the PVP-stabilized Ag nanoparticle suspension.

We assessed the bioavailability of Ag from Ag nanoparticles (NPs), stabilized with polyvinylpyrrolidone (PVP), to terrestrial isopods which were exposed to 10, 100 and 1000 µg Ag NPs/g of dry food. Different Ag species were determined in the NP suspension that was fed to isopods: (i) total Ag by atomic absorption spectroscopy, (ii) the sum of Ag-PVP complexes and free Ag+ by anodic stripping voltammetry at the bismuth-film electrode, and (iii) free Ag+ by ion-selective potentiometry. The amounts of Ag species in the consumed food were compared to the masses of Ag accumulated in the isopod digestive glands. Our results show that all three Ag species (Ag NPs, Ag-PVP complexes and free Ag+) could be the source of bioaccumulated Ag, but to various degrees depending on the exposure concentration and transformations in the digestive system. We provide a proof that (i) Ag NPs dissolve and Ag-PVP complexes dissociate in the isopod digestive tract; (ii) the concentration of free Ag+ in the suspension offered to the test organisms is not the only measure of bioavailable Ag. The type of NP stabilizer along with the NP transformations in the digestive system needs to be considered in the creation of new computational models of the nanomaterial fate.

Multilaboratory evaluation of 15 bioassays for (eco)toxicity screening and hazard ranking of engineered nanomaterials: FP7 project NANOVALID.

Within EU FP7 project NANOVALID, the (eco)toxicity of 7 well-characterized engineered nanomaterials (NMs) was evaluated by 15 bioassays in 4 laboratories. The highest tested nominal concentration of NMs was 100 mg/l. The panel of the bioassays yielded the following toxicity order: Ag > ZnO > CuO > TiO2 > MWCNTs > SiO2 > Au. Ag, ZnO and CuO proved very toxic in the majority of assays, assumingly due to dissolution. The latter was supported by the parallel analysis of the toxicity of respective soluble metal salts. The most sensitive tests/species were Daphnia magna (towards Ag NMs, 24-h EC50 = 0.003 mg Ag/l), algae Raphidocelis subcapitata (ZnO and CuO, 72-h EC50 = 0.14 mg Zn/l and 0.7 mg Cu/l, respectively) and murine fibroblasts BALB/3T3 (CuO, 48-h EC50 = 0.7 mg Cu/l). MWCNTs showed toxicity only towards rat alveolar macrophages (EC50 = 15.3 mg/l) assumingly due to high aspect ratio and TiO2 towards R. subcapitata (EC50 = 6.8 mg Ti/l) due to agglomeration of TiO2 and entrapment of algal cells. Finally, we constructed a decision tree to select the bioassays for hazard ranking of NMs. For NM testing, we recommend a multitrophic suite of 4 in vitro (eco)toxicity assays: 48-h D. magna immobilization (OECD202), 72-h R. subcapitata growth inhibition (OECD201), 30-min Vibrio fischeri bioluminescence inhibition (ISO2010) and 48-h murine fibroblast BALB/3T3 neutral red uptake in vitro (OECD129) representing crustaceans, algae, bacteria and mammalian cells, respectively. Notably, our results showed that these assays, standardized for toxicity evaluation of "regular" chemicals, proved efficient also for shortlisting of hazardous NMs. Additional assays are recommended for immunotoxicity evaluation of high aspect ratio NMs (such as MWCNTs).

A case study to optimise and validate the brine shrimp Artemia franciscana immobilisation assay with silver nanoparticles: The role of harmonisation.

Brine shrimp Artemia sp. has been recognised as an important ecotoxicity and nanotoxicity test model organism for salt-rich aquatic environments, but currently there is still no harmonised testing protocol which would ensure the comparable results for hazard identification. In this paper we aimed to design the harmonised protocol for nanomaterial toxicity testing using Artemia franciscana and present a case study to validate the protocol with silver nanoparticles (AgNPs). We (i) revised the existing nanotoxicity test protocols with Artemia sp. (ii) optimised certain methodological steps based on the experiments with AgNPs and potassium dichromate (K2Cr2O7) as a soluble reference chemical and (iii) tested the optimised protocol in an international inter-laboratory exercise conducted within the EU FP7 NanoValid project. The intra- and inter-laboratory reproducibility of the proposed protocol with a soluble reference chemical K2Cr2O7 was good, which confirms the suitability of this assay for conventional chemicals. However, the variability of AgNPs toxicity results was very high showing again that nanomaterials are inherently challenging for toxicity studies, especially those which toxic effect is linked to shed metal ions. Among the identified sources for this variability were: the hatching conditions, the type of test plate incubation and the illumination regime. The latter induced variations assumingly due to the changes in bioavailable silver species concentrations. Up to our knowledge this is the first inter-laboratory comparison of the Artemia sp. toxicity study involving nanomaterials. Although the inter-laboratory exercise revealed poor repeatability of AgNPs toxicity results, this study provides valuable information regarding the importance of harmonisation of all steps in the test procedure. Also, the presented AgNPs toxicity case study may serve as a platform for further validation steps with other types of NMs.

The impact of humic acid on chromium phytoextraction by aquatic macrophyte Lemna minor.

Studies assessing chromium phytoextration from natural waters rarely consider potential implications of chromium speciation in the presence of ubiquitous humic substances. Therefore, the present study investigated the influence of environmentally relevant concentration of humic acid (TOC = 10 mg L(-1)) on chromium speciation (Cr = 0.15 mg L(-1)) and consequently on phytoextraction by aquatic macrophyte duckweed Lemna minor. In absence of humic acid, only hexavalent chromium was present in water samples and easily taken up by L. minor. Chromium uptake resulted in a significant reduction of growth rate by 22% and decrease of chlorophyll a and chlorophyll b contents by 48% and 43%, respectively. On the other hand, presence of humic acid significantly reduced chromium bioavailability (57% Cr uptake decrease) and consequently it did not cause any measurable effect to duckweed. Such effect was related to abiotic reduction of hexavalent chromium species to trivalent. Hence, findings of our study suggest that presence of humic acid and chromium speciation cannot be neglected during phytoextraction studies.

An interlaboratory comparison of nanosilver characterisation and hazard identification: Harmonising techniques for high quality data.

Within the FP7 EU project NanoValid a consortium of six partners jointly investigated the hazard of silver nanoparticles (AgNPs) paying special attention to methodical aspects that are important for providing high-quality ecotoxicity data. Laboratories were supplied with the same original stock dispersion of AgNPs. All partners applied a harmonised procedure for storage and preparation of toxicity test suspensions. Altogether ten different toxicity assays with a range of environmentally relevant test species from different trophic levels were conducted in parallel to AgNP characterisation in the respective test media. The paper presents a comprehensive dataset of toxicity values and AgNP characteristics like hydrodynamic sizes of AgNP agglomerates and the share (%) of Ag(+)-species (the concentration of Ag(+)-species in relation to the total measured concentration of Ag). The studied AgNP preparation (20.4±6.8 nm primary size, mean total Ag concentration 41.14 mg/L, 46-68% of soluble Ag(+)-species in stock, 123.8±12.2 nm mean z-average value in dH2O) showed extreme toxicity to crustaceans Daphnia magna, algae Pseudokirchneriella subcapitata and zebrafish Danio rerio embryos (EC50<0.01 mg total Ag/L), was very toxic in the in vitro assay with rainbow trout Oncorhynchus mykiss gut cells (EC50: 0.01-1 mg total Ag/L); toxic to bacteria Vibrio fischeri, protozoa Tetrahymena thermophila (EC50: 1-10 mg total Ag/L) and harmful to marine crustaceans Artemia franciscana (EC50: 10-100 mg total Ag/L). Along with AgNPs, also the toxicity of AgNO3 was analyzed. The toxicity data revealed the same hazard ranking for AgNPs and AgNO3 (i.e. the EC50 values were in the same order of magnitude) proving the importance of soluble Ag(+)-species analysis for predicting the hazard of AgNPs. The study clearly points to the need for harmonised procedures for the characterisation of NMs. Harmonised procedures should consider: (i) measuring the AgNP properties like hydrodynamic size and metal ions species in each toxicity test medium at a range of concentrations, and (ii) including soluble metal salt control both in toxicity testing as well as in Ag(+)-species measurements. The present study is among the first nanomaterial interlaboratory comparison studies with the aim to improve the hazard identification testing protocols.

FTIR microscopy reveals distinct biomolecular profile of crustacean digestive glands upon subtoxic exposure to ZnO nanoparticles.

Biomolecular profiling with Fourier-Transform InfraRed Microscopy was performed to distinguish the Zn(2+)-mediated effects on the crustacean (Porcellio scaber) digestive glands from the ones elicited by the ZnO nanoparticles (NPs). The exposure to ZnO NPs or ZnCl2 (1500 and 4000 µg Zn/g of dry food) activated different types of metabolic pathways: some were found in the case of both substances, some only in the case of ZnCl2, and some only upon exposure to ZnO NPs. Both the ZnO NPs and the ZnCl2 increased the protein (∼1312 cm(-1); 1720-1485 cm(-1)/3000-2830 cm(-1)) and RNA concentration (∼1115 cm(-1)). At the highest exposure concentration of ZnCl2, where the effects occurred also at the organismal level, some additional changes were found that were not detected upon the ZnO NP exposure. These included changed carbohydrate (most likely glycogen) concentrations (∼1043 cm(-1)) and the desaturation of cell membrane lipids (∼3014 cm(-1)). The activation of novel metabolic pathways, as evidenced by changed proteins' structure (at 1274 cm(-1)), was found only in the case of ZnO NPs. This proves that Zn(2+) are not the only inducers of the response to ZnO NPs. Low bioavailable fraction of Zn(2+) in the digestive glands exposed to ZnO NPs further supports the role of particles in the ZnO NP-generated effects. This study provides the evidence that ZnO NPs induce their own metabolic responses in the subtoxic range.

High surface adsorption properties of carbon-based nanomaterials are responsible for mortality, swimming inhibition, and biochemical responses in Artemia salina larvae.

We investigated the effects of three different carbon-based nanomaterials on brine shrimp (Artemia salina) larvae. The larvae were exposed to different concentrations of carbon black, graphene oxide, and multiwall carbon nanotubes for 48 h, and observed using phase contrast and scanning electron microscopy. Acute (mortality) and behavioural (swimming speed alteration) responses and cholinesterase, glutathione-S-transferase and catalase enzyme activities were evaluated. These nanomaterials were ingested and concentrated in the gut, and attached onto the body surface of the A. salina larvae. This attachment was responsible for concentration-dependent inhibition of larval swimming, and partly for alterations in the enzyme activities, that differed according to the type of tested nanomaterials. No lethal effects were observed up to 0.5mg/mL carbon black and 0.1mg/mL multiwall carbon nanotubes, while graphene oxide showed a threshold whereby it had no effects at 0.6 mg/mL, and more than 90% mortality at 0.7 mg/mL. Risk quotients calculated on the basis of predicted environmental concentrations indicate that carbon black and multiwall carbon nanotubes currently do not pose a serious risk to the marine environment, however if uncontrolled release of nanomaterials continues, this scenario can rapidly change.

The hazard assessment of nanostructured CeO2-based mixed oxides on the zebrafish Danio rerio under environmentally relevant UV-A exposure.

The effect of nanomaterials on biota under realistic environmental conditions is an important question. However, there is still a lack of knowledge on how different illumination conditions alter the toxicity of some photocatalytic nanomaterials. We have investigated how environmentally relevant UV-A exposure (intensity 8.50 ± 0.61 W/m(2), exposure dose 9.0J/cm(2)) affected the toxicity of cerium oxide (CeO2)-based nanostructured materials to the early-life stages of zebrafish Danio rerio. Pure cerium oxide (CeO2), copper-cerium (CuO-CeO2) (with a nominal 10, 15 and 20 mol.% CuO content), cerium-zirconium (CeO2-ZrO2) and nickel and cobalt (Ni-Co) deposited over CeO2-ZrO2 were tested. It was found that under both illumination regimes, none of the tested materials affected the normal development or induced mortality of zebrafish early-life stages up to 100mg/L. Only in the case of CuO-CeO2, the growth of larvae was decreased (96 h LOEC values for CuCe10, CuCe15 and CuCe20 were 50, 50 and 10mg/L, respectively). To conclude, CeO2-based nanostructured materials are not severely toxic to zebrafish and environmentally relevant UV-A exposure does not enhance their toxicity.

Bioavailability of cobalt and iron from citric-acid-adsorbed CoFe2O4 nanoparticles in the terrestrial isopod Porcellio scaber.

The aim of this study was to determine whether citric acid adsorbed onto cobalt ferrite (CoFe2O4) nanoparticles (NPs) influences the bioavailability of their constituents Co and Fe. Dissolution of Co and Fe was assessed by two measures: (i) in aqueous suspension using chemical analysis, prior to application onto the food of test organisms; and (ii) in vivo, measuring the bioavailability in the model terrestrial invertebrate (Porcellio scaber, Isopoda, Crustacea). The isopods were exposed to citric-acid-adsorbed CoFe2O4 NPs for 2 weeks, and tissue accumulation of Co and Fe was assessed. This was compared to pristine CoFe2O4 NPs, and CoCl2 and Fe(III) salts as positive controls. The combined data shows that citric acid enhances free metal ion concentration from CoFe2O4 NPs in aqueous suspension, although in vivo, very similar amounts of assimilated Co were found in isopods exposed to both types of NPs. Therefore, evaluation of the dissolution in suspension by chemical means is not a good predictor of metal assimilation of this model organism; body assimilation of Co and Fe is rather governed by the physiological capacity of P. scaber for the uptake of these metals. Moreover, we propose that citric acid, due to its chelating properties, may hinder the uptake of Co that dissolves from citric-acid-adsorbed CoFe2O4 NPs, if citric acid is present in sufficient quantity.

Hindered disulfide bonds to regulate release rate of model drug from mesoporous silica.

With the advancement of drug delivery systems based on mesoporous silica nanoparticles (MSNs), a simple and efficient method regulating the drug release kinetics is needed. We developed redox-responsive release systems with three levels of hindrance around the disulfide bond. A model drug (rhodamine B dye) was loaded into MSNs' mesoporous voids. The pore opening was capped with ß-cyclodextrin in order to prevent leakage of drug. Indeed, in absence of a reducing agent the systems exhibited little leakage, while the addition of dithiothreitol cleaved the disulfide bonds and enabled the release of cargo. The release rate and the amount of released dye were tuned by the level of hindrance around disulfide bonds, with the increased hindrance causing a decrease in the release rate as well as in the amount of released drug. Thus, we demonstrated the ability of the present mesoporous systems to intrinsically control the release rate and the amount of the released cargo by only minor structural variations. Furthermore, an in vivo experiment on zebrafish confirmed that the present model delivery system is nonteratogenic.

Antioxidant responses and whole-organism changes in Daphnia magna acutely and chronically exposed to endocrine disruptor bisphenol A.

Bisphenol A (BPA) is a well known endocrine disruptor. Significantly less is known about its ability to induce antioxidant defense and oxidative stress in aquatic organisms. This is an interesting subject because BPA can act both as a prooxidant (induces the formation of reactive oxygen species) and an antioxidant. The aim of the present study was to elucidate if BPA induces changes in the activities of antioxidant enzymes catalase (CAT) and glutathione S-transferases (GSTs) and the level of lipid peroxidation in crustacean Daphnia magna. These biomarkers were followed after acute (48h) and chronic (21d) exposure period. Interestingly, CAT and GSTs were changed in the same manner after both exposure durations. The most observable change in daphnids was the increase of GSTs activities, measured with 1,2-chloro-4-dinitrobenzene as a substrate. No GSTs activities with ethacrynic acid and 1,2-dichloro-4-nitrobenzene as substrates were detected. The activity of CAT and the level of lipid peroxidation remained unchanged in daphnids exposed to sublethal concentrations of BPA. The reproduction rate was considerably affected already at 1.73mg/L while the growth was only affected at the highest concentration (13.8mg/L), where also significant mortality was observed.

Upon exposure to Cu nanoparticles, accumulation of copper in the isopod Porcellio scaber is due to the dissolved Cu ions inside the digestive tract.

The fate of nanoparticles in organisms is of significant interest. In the current work, we used a test system with terrestrial isopods (Porcellio scaber) fed with food spiked with Cu NPs or soluble Cu salt for 14 days. Two different doses were used for spiking to yield final concentrations of 2000 and 5000 µg Cu/g dry food. After the exposure period, part of the exposed group of animals was transferred to clean food to depurate. Cu content was analyzed in the digestive glands, gut, and the 'rest' of the body. Similar patterns of (i) assimilated and depurated amounts of Cu, (ii) Cu body distribution, and (iii) effect on isopods feeding behavior were observed regardless of whether the animals were fed with Cu NPs or soluble Cu salt spiked food. Thus, Cu ions and not Cu NPs were assimilated by the digestive gland cells. Solubilization of the Cu NPs applied to the leaves was also analyzed with chemical methods and recombinant Cu-sensing bacteria. The comparison of the in vitro data on solubilization of Cu NPs and in vivo data on Cu accumulation in the animal tissues showed that about 99% of accumulated copper ions was dissolved from ingested Cu NPs in the digestive system of isopods.

Effects of four CeO2 nanocrystalline catalysts on early-life stages of zebrafish Danio rerio and crustacean Daphnia magna.

Effects of four different nanocrystalline CeO(2)-based catalysts on crustaceans Daphnia magna and early-life stages of zebrafish Danio rerio were studied. Pure CeO(2) and CuO-CeO(2) mixed oxides with a nominal 10, 15 and 20 mol.% CuO content were tested. Pure CeO(2) provoked no effects, but CuO-CeO(2) mixed oxides induced some sublethal effects on fish and affected daphnids' survival. The most pronounced effects were found on fish body growth, which was reduced at 10 mg/L in case of CuCe20 and 50 mg/L in cases of CuCe10 and CuCe15. Daphnids' survival was affected above 80 mg/L of CuCe20, while CuCe10 and CuCe15 did not affect daphnids. None of the materials was highly toxic to daphnids and fish in comparison to some other environmental pollutants. Differences in effects between the materials could not be explained by their specific physicochemical properties. This work indicates that more attention should be placed at potential toxicity of nanostructured materials, such as nanocrystalline mixed-oxides.

Assessment of landfill leachate toxicity reduction after biological treatment.

In the present article, the efficiency of biological treatment of landfill leachates was evaluated by implementation of physicochemical characterisation and a complex toxicity assessment. An array of toxicity tests using bacterium Vibrio fischeri, alga Desmodesmus subspicatus, crustacean Daphnia magna, and embryo of fish Danio rerio, as well as unconventional methods using biochemical biomarkers (protein content, enzymes cholinesterase, and glutathione-S-transferase), were employed. Toxicity of leachates varied depending on the season of collection in relation to their different physicochemical characteristics. Uncommon effects of leachates on organisms, such as hormetic-like increases of algal growth and reproduction of daphnids, were identified. New approaches using the activities of enzymes were found unsuitable for routine hazard assessment of leachates. Although physicochemical parameters and toxicity decreased significantly after biological treatment, the effluents did not meet the demands of the current Slovenian legislation; thus, the existing biological treatment was found inappropriate. The development of advanced treatment techniques for landfill leachates is thus encouraged.

The link between antioxidant enzymes catalase and glutathione S-transferase and physiological condition of a control population of terrestrial isopod (Porcellio scaber).

The aim of this work was to investigate if the activities of catalase and glutathione S-transferase in a control population of terrestrial isopods (Porcellio scaber) are correlated with the physiological condition of the isopods. For this purpose, the activities of these enzymes were analysed in isopods from a stock population and in parallel, the physiological condition of the same specimens was assessed using a histological approach based on epithelial thickness and lipid droplets. We found a correlation between antioxidant enzymes and the physiological condition of the isopods. This implies that these enzymes could be used as predictive indicators of the physiological condition in a stock population before comprehensive toxicological studies are conducted and also in control group after the experiment. When a control group is found to be very heterogeneous in terms of physiological condition, the experiment should be repeated with a larger number of experimental animals. The findings of this study will contribute to more accurate experimental design of toxicity tests when using biomarkers. This should encourage other researchers to increase their effort to know the physiological state of their test organisms.

Ecotoxicity of nanosized TiO2. Review of in vivo data.

This report presents an exhaustive literature review of data on the effect of nanoparticulate TiO(2) on algae, higher plants, aquatic and terrestrial invertebrates and freshwater fish. The aim, to identify the biologically important characteristics of the nanoparticles that have most biological significance, was unsuccessful, no discernable correlation between primary particle size and toxic effect being apparent. Secondary particle size and particle surface area may be relevant to biological potential of nanoparticles, but insufficient confirmatory data exist. The nanotoxicity data from thirteen studies fail to reveal the characteristics actually responsible for their biological reactivity because reported nanotoxicity studies rarely carry information on the physicochemical characteristics of the nanoparticles tested. A number of practical measures are suggested which should support the generation of reliable QSAR models and so overcome this data inadequacy.