Polyaniline nanofibers: acute toxicity and teratogenic effect on Rhinella arenarum embryos

The fate and effect of nanomaterials in the environment is of paramount importance towards the technological application of the materials. This work shows the ecotoxicological potential of polyaniline (PANI) nanofibers in the larvae Rhinella arenarum by means of AMPHITOX test. Acute toxicity of PANI nanofibers towards embryos of the common South American toad R. arenarum (Anura: bufonidae) was evaluated in the premetamorphosis (stage 25) larvae. The exposure of R. arenarum larvae to at dose of 150, 250 and 400 mg L⁻¹ resulted in 100% viability within 96 h exposure. The embryos at 2-4 blastomers stage (early life stage teratogenic test) revealed that embryos were not killed and no teratogenic effects were observed when embryos were incubated with PANI nanofibers (150 and 250 mg L⁻¹), while only a growth retardation of embryos was induced at levels of 250 mg PANI nanofibers L⁻¹. On the other hand, at 400 mg L⁻¹ concentration, a reduction in the body length of larvae and tail malformation was observed. This results suggest that a concentration-dependent toxicity is operative, typified by phenotypes that had abnormal body axes. The presence of PANI nanofibers in gut contents and its excretion by larval stages of R. arenarum was confirmed by UV-visible spectroscopy.     

Tributyltin chloride results in dorsal curvature in embryo development of Sebastiscus marmoratus via apoptosis pathway

Tributyltin (TBT) is a ubiquitous marine environmental contaminant characterized primarily by its reproductive toxicity. However, the embryotoxicity of TBT has not been extensively described, especially in fishes. The aim of this study was to investigate the developmental toxicity of waterborne TBT at environmental levels (0, 0.1, 1, and 10 ng L⁻¹ as Sn) on Sebastiscus marmoratus embryos. Our study showed that TBT reduced the hatchability and caused apparent morphological abnormalities including dorsal curvature, severely twisted tails and pericardial edema. In addition, localized apoptosis was found in the tail regions of embryos after TBT exposure. The study provided a possible mechanistic link between apoptosis and TBT-induced twisted tails abnormality. TBT exposure induced retinoid X receptor α expression in S. marmoratus embryos at the 0.1 and 1 ng L⁻¹ group, which would be responsible for the increasing apoptotic cells induced by TBT. The results of the present study have widespread implications for environmental ecological assessment, management and the etiology of developmental defects.     

Histopathological effects of copper and lithium in the ramshorn snail, Marisa cornuarietis (Gastropoda, Prosobranchia)

The aim of this study was to determine and quantify effects of copper and lithium in tissues of early juveniles of the ramshorn snail, Marisa cornuarietis. For this purpose, hatchlings of M. cornuarietis were exposed for 7 days to a range of five different sublethal concentrations of copper (5, 10, 25, 50, and 75 μg Cu²⁺ L⁻¹) and lithium (50, 100, 200, 1000, and 5000 μg Li⁺ L⁻¹). Both metals changed the tissue structure of epidermis, hepatopancreas, and gills, varying between slight and strong reactions, depending on the copper and lithium concentration. The histopathological changes included alterations in epithelial and mucous cells of the epidermis, swelling of hepatopancreatic digestive cells, alterations in the number of basophilic cells, abnormal apices of digestive cells, irregularly shaped cilia and changes in the amount of mucus in the gills. The most sensible organ in M. cornuarietis indicating Cu or Li pollution is the hepatopancreas (LOECs were 10 μg Cu²⁺ L⁻¹, or 200 μg Li⁺ L⁻¹). In epidermis, mantle and gills relevant effects occurred with higher LOECs (50 μg Cu²⁺ L⁻¹, or 1000 μg Li⁺ L⁻¹). Base on LOECs, our results indicated that histopathological endpoints are high sensitivity to copper and lithium compared to endpoints for embryonic developmental toxicity.     

Short-term exposure of the European sea bass Dicentrarchus labrax to copper-based antifouling treated nets: Copper bioavailability and biomarkers responses

We studied if the levels of copper released from antifouling treated nets used in finfish mariculture could affect the immune defense mechanism and/or induce oxidative stress in Dicentrarchus labrax, after short term exposure in laboratory experiments. Dissolved copper concentration released from the treated nets, copper bioavailability and a set of biomarkers responses were measured. Biomarkers included hemoglobin concentration, activities of lysozyme, total complement, respiratory burst, glutathione S-transferase and acetycholinesterase and concentration of thiobarbituric acid reactive substances. Results indicated elevated copper concentration in seawater (184 μg L⁻¹) but low concentration in muscle (1.5 μg g⁻¹) and liver (117 μg g⁻¹). Copper bioavailability was independent of copper complexes with dissolved organic carbon. However, formation of copper complexes with other matrices could neither be excluded nor justified. The released copper from the treated nets did not induce oxidative stress but affected the immediate immune defense mechanism of the exposed fish making them more easily vulnerable to diseases. Consequently, copper-based antifouling treated nets could be a risk factor for D. labrax health.     

Effects of nitrogen deposition and soil fertility on cover and physiology of Cladonia foliacea (Huds.) Willd., a lichen of biological soil crusts from Mediterranean Spain

We are fertilizing a thicket with 0, 10, 20 and 50 kg nitrogen (N) ha⁻¹ yr⁻¹ in central Spain. Here we report changes in cover, pigments, pigment ratios and FvFm of the N-tolerant, terricolous, lichen Cladonia foliacea after 1-2 y adding N in order to study its potential as biomarker of atmospheric pollution. Cover tended to increase. Pigments increased with fertilization independently of the dose supplied but only significantly with soil nitrate as covariate. β-carotene/chlorophylls increased with 20-50 kg N ha⁻¹ yr⁻¹ (over the background) and neoxanthin/chlorophylls also increased with N. (Neoxanthin+lutein)/carotene decreased with N when nitrate and pH seasonalities were used as covariates. FvFm showed a critical load above 40 kg N ha⁻¹ yr⁻¹. Water-stress, iron and copper also explained variables of lichen physiology. We conclude that this tolerant lichen could be used as biomarker and that responses to N are complex in heterogeneous Mediterranean-type landscapes.     

Food utilization and growth of cutworm Spodoptera litura Fabricius larvae exposed to nickel,and its effect on reproductive potential

Food utilization and growth of the 5th and 6th instar Spodoptera litura Fabricius larvae, and its effect on reproduction potential was evaluated by feeding larvae diets with different doses of Ni for 3 generations. Dose-dependent relationships between Ni levels and food consumption and growth were variable with different larval developmental period and Ni exposure duration. RCR, AD and RGR of the 6th instar larvae were much more affected by Ni exposure than those of 5th instar larvae, and the effects were strongest in the 3rd generation. It was found that RCR was significantly stimulated after 1 and 20 mg kg⁻¹ Ni exposure, while AD was significantly inhibited after 1, 5, 10 and 40 mg kg⁻¹ Ni exposure. However, lower levels of Ni (?5 mg kg⁻¹) significantly increased and higher levels of Ni (?10 mg kg⁻¹) significantly decreased RGR. In 3 successive generations, 10 mg kg⁻¹ Ni significantly increased the ECI and ECD of the 5th instar larvae, and 5 mg kg⁻¹ Ni significantly increased the ECD of the 6th instar larvae. However, ECD were all significantly inhibited with 20 mg kg⁻¹ Ni exposure. Results also revealed that durations of larvae were shortened at low levels of Ni, but extended at high levels of Ni. Fecundity was inhibited by the highest Ni doses in each generation, while improved by low Ni doses in the 3rd generation. Hatching rates in all treatments were significantly decreased in a Ni dose-dependent manner. Study indicated that effects of Ni on these parameters were predominant with the increasing Ni exposure period.     

The toxicity of titanium dioxide nanopowder to early life stages of the Japanese medaka (Oryzias latipes)

In this study, fertilized Japanese medaka (Oryzias latipes) embryos were exposed from fertilization to 5 d post-hatch using static non-renewal assays to aqueous suspensions of titanium dioxide nanoparticles (nTiO₂) ranging in nominal concentrations between 0 and 14 μg mL⁻¹. The average size of the nTiO₂ in the stock solution before addition to the test treatments was 87 nm (±14 nm). TiO₂ materials accumulated in a concentration dependent manner on the chorionic filaments of developing medaka embryos with evidence of pericardial edema occurring during embryo development. However, no significant (p > 0.05) increases in mortality relative to control treatments were observed for the nTiO₂ exposed embryos. A concentration dependent increase in cumulative percent hatch was observed at 11 d, indicating that exposure to increasing concentrations of nTiO₂ resulted in the premature hatch of medaka embryos. Post-hatch, a significant proportion of sac fry from the nTiO₂ exposure groups exhibited moribund swimming behavior and these individuals also experienced greater mortality at 15 d post-hatch. Combined, these results demonstrate that exposure to nTiO₂ can impact the development of early life stages of fish.     

Effects of endosulfan exposure and Taura Syndrome Virus infection on the survival and molting of the marine penaeid shrimp, Litopenaeus vannamei

Molting in crustaceans is an important endocrine-controlled biological process that plays a critical role in growth and reproduction. Many factors can affect this physiological cycle in crustaceans including environmental stressors and disease agents. For example the pathology of Taura Syndrome Virus (TSV) of shrimp is closely related to molting cycle. Similarly, endosulfan, a commonly used pesticide is a potential endocrine disruptor. This study explores interrelationships between pesticide exposure, virus infection and their interactions with physiology and susceptibility of the shrimp. Litopenaeus vannamei (Pacific white shrimp) were challenged with increasing doses of endosulfan and TSV (TSV-C, a Belize reference strain) to determine the respective median lethal concentrations (LC₅₀s). The 96-h endosulfan LC₅₀ was 5.32 μg L⁻¹, while the 7-d TSV LC₅₀ was 54.74 mg L⁻¹. Subsequently, based on their respective LC₅₀ values, a 20-d interaction experiment with sublethal concentrations of endosulfan (2 μg L⁻¹) and TSV (30 mg L⁻¹) confirmed a significant interaction (p < 0.05, χ² = 5.29), and thereby the susceptibility of the shrimp. Concurrently, molt-stage of animals, both at the time of exposure and death, was compared with mortality. For animals challenged with TSV, no strong correlation between molt-stage and mortality was observed (p > 0.05). For animals exposed to endosulfan, animals in the postmolt stage were shown to be more susceptible to acute toxicity (p < 0.05). For animals exposed to both TSV and endosulfan, interference of endosulfan-associated stress lead to increasingly higher susceptibility at postmolt (p < 0.05) during the acute phase of the TSV disease cycle.     

Copper kinetics and internal distribution in the marbled crayfish (Procambarus sp.)

Metal pollution e.g. copper, in water bodies occurs worldwide. Although copper is an essential trace metal, at certain levels it is still considered as pollutant. The aim of this study was to investigate the effect of exposure concentration on copper bioaccumulation in marbled crayfish (Procambarus sp.) by determining uptake and elimination kinetics. Crayfish were exposed to sub-lethal copper concentrations (average measured concentrations of 0.031 and 0.38 mg Cu L⁻¹) for 14 d and transferred to copper-free water for another 14 d. At different time points during the uptake and elimination phases copper concentrations were measured in five organs (exoskeleton, gills, muscle, ovaries and hepatopancreas). At 0.031 mg Cu L⁻¹, copper levels in the crayfish organs were not significantly increased compared to the control animals, suggesting effective regulation. Exposure to 0.38 mg Cu L⁻¹ did lead to not significantly increased copper levels in muscles and ovaries, while the gills and exoskeleton, which are in direct contact with the water, showed significantly higher copper concentrations. In these four organs, copper showed fast uptake kinetics with equilibrium reached within 10 d of exposure. Copper accumulation was highest in the hepatopancreas; uptake in this storage organ steadily increased with time and did not reach equilibrium within the 14-d exposure period. Copper accumulation levels in the marbled crayfish found in this study were hepatopancreas > gills > exoskeleton > muscle.     

Immunotoxicity in ascidians: antifouling compounds alternative to organotins: III--the case of copper(I) and Irgarol 1051

After the widespread ban of TBT, due to its severe impact on coastal biocoenoses, mainly related to its immunosuppressive effects on both invertebrates and vertebrates, alternative biocides such as Cu(I) salts and the triazine Irgarol 1051, the latter previously used in agriculture as a herbicide, have been massively introduced in combined formulations for antifouling paints against a wide spectrum of fouling organisms. Using short-term (60 min) haemocyte cultures of the colonial ascidian Botryllus schlosseri exposed to various sublethal concentrations of copper(I) chloride (LC₅₀ = 281 μM, i.e., 17.8 mg Cu L⁻¹) and Irgarol 1051 (LC₅₀ > 500 μM, i.e., >127 mg L⁻¹), we evaluated their immunotoxic effects through a series of cytochemical assays previously used for organotin compounds. Both compounds can induce dose-dependent immunosuppression, acting on different cellular targets and altering many activities of immunocytes but, unlike TBT, did not have significant effects on cell morphology. Generally, Cu(I) appeared to be more toxic than Irgarol 1051: it significantly (p < 0.05) inhibited yeast phagocytosis at 0.1 μM (∼10 μg L⁻¹), and affected calcium homeostasis and mitochondrial cytochrome-c oxidase activity at 0.01 μM (∼1 μg L⁻¹). Both substances were able to change membrane permeability, induce apoptosis from concentrations of 0.1 μM (∼10 μg L⁻¹) and 200 μM (∼50 mg L⁻¹) for Cu(I) and Irgarol 1051, respectively, and alter the activity of hydrolases. Both Cu(I) and Irgarol 1051 inhibited the activity of phenoloxidase, but did not show any interactive effect when co-present in the exposure medium, suggesting different mechanisms of action.     

Nano-TiO2 enhances the toxicity of copper in natural water to Daphnia magna

The acute toxicity of engineered nanoparticles (NPs) in aquatic environments at high concentrations has been well-established. This study demonstrates that, at a concentration generally considered to be safe in the environment, nano-TiO₂ remarkably enhanced the toxicity of copper to Daphnia magna by increasing the copper bioaccumulation. Specifically, at 2 mg L⁻¹ nano-TiO₂, the (LC₅₀) of Cu²⁺ concentration observed to kill half the population, decreased from 111 μg L⁻¹ to 42 μg L⁻¹. Correspondingly, the level of metallothionein decreased from 135 μg g⁻¹ wet weight to 99 μg g⁻¹ wet weight at a Cu²⁺ level of 100 μg L⁻¹. The copper was found to be adsorbed onto the nano-TiO₂, and ingested and accumulated in the animals, thereby causing toxic injury. The nano-TiO₂ may compete for free copper ions with sulfhydryl groups, causing the inhibition of the detoxification by metallothioneins.     

The effect of nitrate concentration on sulfide-driven autotrophic denitrification in marine sediment

Copepods have been widely used to evaluate toxicity of metals present in marine environments. However, a technical difficulty is to understand the possible routes of metal uptake and to identify in which tissues or organs metals are being accumulated. Traditional techniques are hard to be employed once each organ has to be analyzed separately. Autoradiography is an alternative technique to circumvent this limitation, since metal distribution in tissues can be visualized and quantified, even in small organisms like copepods. In the present study, accumulation and distribution of ⁶⁴Cu in the copepod Calanus hyperboreus was studied using autoradiography. Copepods were exposed for 2 h to copper (2.3 mg L⁻¹; 1.08 MBq ⁶⁴Cu mg⁻¹ Cu) and then allowed to depurate for 2 h in clean seawater. Total ⁶⁴Cu was determined by gamma-spectrometry after a metal exposure and a depuration period. ⁶⁴Cu distribution was determined based on images generated by autoradiography. Metal accumulation was observed on all external surfaces of the copepods, being accumulated mostly on the ventral region, followed by dorsal, urossoma and internal regions. After depuration, radioactivity levels had a decrease in the sum of external body surface. Our results show that copper uptake by C. hyperboreus is fast and that a non-negligible proportion of the accumulated metal can reach internal tissues, which may lead to detrimental physiological effects. Moreover, whole-body autoradiography was demonstrated to be an efficient technique to study copper accumulation and body distribution in a very small organism such as the copepod C. hyperboreus.     

Microtubule integrity and cell viability under metal (Cu,Ni and Cr) stress in the seagrass Cymodocea nodosa

The effects of increasing Cu, Ni and Cr concentrations (0.5, 5, 10, 20 and 40 mg L⁻¹) on microtubule organization and the viability of leaf cells of the seagrass Cymodocea nodosa for 13 consecutive days were investigated under laboratory conditions. Increased oblique microtubule orientation, microtubule depolymerization at the 5–40 mg L⁻¹ Ni treatments after 3 d of exposure, and a complete microtubule depolymerization at all Ni treatments after 5 d were observed. Cu depolymerised microtubules after three to 7 d of exposure, while Cr caused an extensive microtubule bundling after 9 or 11 d of exposure, depending on metal dosage. Fluorescence intensity measurements further consolidated the above phenomena. Cell death, occurring at later time than microtubule disturbance, was also observed at all Cu and Ni treatments and at the 10–40 mg L⁻¹ Cr treatments and adding to the above quantification of the number of dead cells clearly showed that only a portion of the cell population studied died. The data presented, being the first assessment of microtubule disturbance in seagrasses, indicate that microtubules in seagrass leaf cells could be used as a valuable and early marker of metal-induced stress in biomonitoring programmes.     

The impact of production type and region on polychlorinated biphenyl (PCB), polychlorinated dibenzo-p-dioxin and dibenzofuran (PCDD/F) concentrations in Canadian chicken egg yolks

Chicken eggs from five different production types (conventional, omega-3 enriched, free range, organic and free run) were collected, when available, from three regions (west, central and east) of Canada to determine persistent organic pollutant (POP) concentrations. Total polychlorinated biphenyl (PCB) concentrations (∑37 congeners) in yolks from the eggs ranged from 0.162 ng g⁻¹ lipid to 24.8 ng g⁻¹ lipid (median 1.25 ng g⁻¹ lipid) while the concentration of the sum of the 6 indicator PCBs ranged from 0.100 ng g⁻¹ lipid to 9.33 ng g⁻¹ lipid (median 0.495 ng g⁻¹ lipid). Total polychlorinated dibenzo-p-dioxin/dibenzofuran (PCDD/F) concentrations ranged from 2.37 pg g⁻¹ lipid to 382 pg g⁻¹ lipid (median 9.53 pg g⁻¹ lipid). The 2005 WHO toxic equivalency (TEQ) ranged from 0.089 pg TEQ_{PCDD/F+dioxin-like[DL]-PCB} g⁻¹ lipid to 12.8 pg TEQ_{PCDD/F+DL-PCB} g⁻¹ lipid (median 0.342 pg TEQ_{PCDD/F+DL-PCB} g⁻¹ lipid). PCB and PCDD/F concentrations were significantly different (p < 0.001) in egg yolks from different regions of collection. In contrast to observations in Europe, PCB and PCDD/F concentrations in Canadian egg yolks were not impacted solely by the production type (e.g., conventional, free range, organic, etc.) used to maintain the laying chickens. Additionally, only one Canadian free range yolk from western Canada (12.8 pg TEQ_{PCDD/F+DL-PCB} g⁻¹ lipid) exceeded the European toxic equivalent concentration limits for eggs (5 pg TEQ_{PCDD/F+DL-PCB} g⁻¹ lipid). This differs from observations in Europe where free range/home produced eggs frequently have higher POP concentrations than eggs from other production types. Median PCB dietary intake estimates based on consumption of eggs were less than 10 ng d⁻¹ while median PCDD/F intakes were less than 45 pg d⁻¹.     

Evaluation of food chain transfer of the antibiotic oxytetracycline and human risk assessment

There has been recent concern regarding the possibility of antibiotics entering the aquatic food chain and impacting human consumers. This work reports experimental results of the bioconcentration of the antibiotic oxytetracycline (OTC) by the Asian watermeal plant (Wolffia globosa Hartog & Plas) and bioaccumulation of OTC in watermeal and water by the seven-striped carp (Probarbus jullieni). They show, for the first time, the extent to which OTC is able to transfer from water to plant to fish and enter the food chain. The mean bioconcentration factor (dry weight basis) with watermeal was 1.28 × 10³ L kg⁻¹. Separate experiments were undertaken to characterize accumulation of OTC by carp from water and watermeal. These showed the latter pathway to be dominant under the conditions employed. The bioconcentration and biomagnification factors for these processes were 1.75 L kg⁻¹ and 2 × 10⁻⁴ kg g⁻¹ respectively. Using an aqueous concentration range of 0.34–3.0 μg L⁻¹, hazard quotients for human consumption of contaminated fish of 1.3 × 10⁻² to 1.15 × 10⁻¹ were derived.     

Bioaccumulation and depuration of anthracene in Penaeus monodon (Fibricius) through food ingestion

Understanding on the bioaccumulation and depuration of PAHs (polycyclic aromatic hydrocarbons) in Penaeus monodon is important in seafood safety because it is one of the most popular seafood consumed worldwide. In this study, we used anthracene as the precursor compound for PAHs accumulation and depuration in the shrimp. Commercial feed pellets spiked with anthracene were fed to P. monodon. At 20 mg kg⁻¹ anthracene, P. monodon accumulated 0.1% of the anthracene from the feed. P. monodon deputed the PAH two times faster than its accumulation. The shrimp reduced its feed consumption when anthracene content in the feed exceeded 20 mg kg⁻¹. At 100 mg kg⁻¹ anthracene, P. monodon started to have necrosis tissues on the posterior end of their thorax. The bioaccumulation factor (BAF), uptake rate constant (k₁) and depuration rate constant (k₂) of anthracene in P. monodon were 1.15 × 10⁻³, 6.80 × 10⁻⁴ d⁻¹ and 6.28 × 10⁻¹ d⁻¹, respectively. The depuration rate constant is about thousand times higher than the uptake rate constant and this indicated that this crustacean is efficient in depurating hydrocarbons from their tissue.     

Effects of dioxin exposure in Eisenia andrei: integration of biomarker data by an Expert System to rank the development of pollutant-induced stress syndrome in earthworms

A battery of biomarkers has recently been developed in the earthworm Eisenia andrei. In this study, different biomarkers (i.e. Ca²⁺-ATPase activity, lysosomal membrane stability-LMS, lysosomal lipofuscin and neutral lipid content) were utilized to evaluate the alterations in the physiological status of animals, induced by exposure for 3 d to different sublethal concentrations of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) (1.5 × 10⁻³, 1.5 × 10⁻², 1.5 × 10⁻¹ ng mL⁻¹) utilizing the paper contact toxicity test. Lysosome/cytoplasm volume ratio and DNA damage were also evaluated as a biomarker at the tissue level and as a biomarker of genotoxicity, respectively. Moreover, the NR retention time assay conditions were optimized for the determination of in vivo LMS in earthworm coelomocytes. The results demonstrate that LMS and Ca²⁺-ATPase activity were early warning biomarkers able to detect the effects of minimal amounts of TCDD and that biomarkers evaluated at the tissue level are important for following the evolution of the stress syndrome in earthworms. To evaluate the health status of the animals, an Earthworm Expert System (EES) for biomarker data integration and interpretation was developed. The EES proved to be a suitable tool able to rank, objectively, the different levels of the stress syndrome in E. andrei induced by the different concentrations of TCDD.     

Biodegradable chelate enhances the phytoextraction of copper by Oenothera picensis grown in copper-contaminated acid soils

Oenothera picensis plants (Fragrant Evening Primrose) grow in the acid soils contaminated by copper smelting in the coastal region of central Chile. We evaluated the effects of the biodegradable chelate MGDA (methylglycinediacetic acid) on copper extraction by O. picensis and on leaching of copper through the soil profile, using an ex situ experiment with soil columns of varying heights. MGDA was applied in four rates: 0 (control), 2, 6 and 10 mmol plant⁻¹. MGDA application significantly increased biomass production and foliar concentration, permitting an effective increase in copper extraction, from 0.09 mg plant⁻¹ in the control, to 1.3 mg plant⁻¹ in the 6 and 10 mmol plant⁻¹ treatments. With 10 mmol plant⁻¹ rate of MGDA, the copper concentration in the leachate from the 30 cm columns was 20 times higher than in the control. For the 60 cm columns, copper concentration was 2 times higher than the control. It can be concluded that at increased soil depths, copper leaching would be minimal and that MGDA applications at the studied rates would not pose a high risk for leaching into groundwater. It can thus be stated that applications of MGDA are an effective and environmentally safe way to improve copper extraction by O. picensis in these soils.     

Distribution of Se and its species in Myriophyllum spicatum and Ceratophyllum demersum growing in water containing Se (VI)

The uptake of Se (VI) by two aquatic plants, Myriophyllum spicatum L. and Ceratophyllum demersum L., and its effects on their physiological characteristics have been studied. Plants were cultivated outdoors under semi-controlled conditions and in two concentrations of Na selenate solution (20 μg Se L⁻¹ and 10 mg Se L⁻¹). The higher dose of Se reduced the photochemical efficiency of PSII in both species, while the lower dose had no effect on PSII. Addition of Se had no effect on the amounts of chlorophyll a and b. The concentration of Se in plants grown in 10 mg Se L⁻¹, averaged 212 ± 12 μg Se g⁻¹ DM in M. spicatum (grown from 8-13 d), and 492 ± 85 μg Se g⁻¹ DM in C. demersum (grown for 31 d). Both species could take up a large amount of Se. The amount of soluble Se compounds in enzyme extracts ranged from 16% to 26% in control, and in high Se solution from 48% to 36% in M. spicatum and C. demersum, respectively. Se-species were determined using HPLC-ICP-MS. The main soluble species in both plants was selenate (∼37%), while SeMet and SeMeSeCys were detected at trace levels.