Toxicity and removal of pesticides by selected aquatic plants

Pesticides are being detected in water bodies on an increasingly frequent basis. The present study focused on the phytoremediation potential of selected aquatic plants to remove phytosanitary products from contaminated water. We investigated the uptake capacity of Lemna minor (L. minor), Elodea canadensis (E. canadensis) and Cabomba aquatica (C. aquatica) on three pesticides: copper sulphate (fungicide), flazasulfuron (herbicide) and dimethomorph (fungicide). Pesticide toxicity was evaluated by exposing plants to five concentrations (0-1 mg L(-1)) in culture media for 7d using chlorophyll fluorescence as a biomarker. The toxicity of the contaminants was the same for all the aquatic plants studied and occurred in this descending order of toxicity: flazasulfuron>copper>dimethomorph. We found that L. minor had the most efficient uptake capacity, followed by E. canadensis and then C. aquatica. The maximum removal rate (microg g(-1)fresh weight d(-1)) of copper, flazasulfuron and dimethomorph was 30, 27 and 11, respectively.     

Oxidative stress in duckweed (Lemna minor L.) caused by short-term cadmium exposure

The mechanisms of plant defence against cadmium toxicity have been studied by short-term exposure of Lemna minor L. (common duckweed) to concentrations of CdCl2 ranging from 0 to 500microM. High accumulation of cadmium was observed (12,320+/-2155microgg(-1) at 500microM CdCl2), which caused a gradual decrease of plant growth, increased lipid peroxidation, and weakened the entire antioxidative defence. Total glutathione concentration decreased significantly; however, the concentration of oxidized glutathione remained stable. The responses of four antioxidant enzymes showed that catalase was the most inhibited after CdCl2 exposure, ascorbate peroxidase and guaiacol peroxidase moderately, and glutathione reductase least. The total antioxidative potential revealed an induced antioxidative network at 0.1microM CdCl2 (137+/-13.2% of the control) and its reduction to only 47.4+/-4.0% of the control at higher cadmium concentrations. The possible application of the examined biomarkers in ecotoxicological research is discussed.     

Heavy metal toxicity to Lemna minor: studies on the time dependence of growth inhibition and the recovery after exposure

Environmental concentrations of toxic substances are not necessarily constant but fluctuate over time. Periods of intense exposure might be followed by episodes with a relatively low or no exposure, in principle allowing exposed organisms to recover from toxic injury. The growth reproduction assay with the limnic vascular plant Lemna minor allows for convenient studies on the time dependence of the aquatic toxicity of chemicals. Here we report on a study with four priority metals (Zn, Cu, Ni, Cd). Aims of the study were to determine the impact of the exposure duration on the observed toxicities and to determine the potential for recovery. The bioconcentrations of the test metals were recorded during the exposure in order to analyse, whether changes in the internal concentrations are a governing factor for the dynamics of toxicity. After an exposure of 7 days, Cd and Cu showed the highest toxicity to Lemna (EC50's of 1.9 and 9.7 microM respectively), while Ni and Zn had a slightly lower toxicity (EC50's of 56.3 and 46.1 microM respectively). Additionally, Zn showed a severely delayed toxicity and the exposed plants did not recover even 7 days after the exposure had ended. This is in sharp contrast to the other test metals, for which a considerable recovery was observed. These results indicate the necessity to more thoroughly consider the dynamics of toxicity, instead of recording toxic effects only after a constant exposure over a fixed time.     

A review of the fate and toxicity of 3,4-dichloroaniline in aquatic environments

The physicochemical properties, fate and toxicity of 3,4-dichloroaniline (3,4-DCA) in aquatic environments are reviewed.     

3种草本植物对Pb-Cd污染水体的修复研究

采用水培实验方法,研究3种水生草本植物对Pb-Cd污染水体的生长反应及对重金属离子的修复效果。结果表明,在去除率实验中3种植物对Pb2+-Cd2+的去除率均有提高,对Pb2+的去除率分别为浮萍47.88%,水蕹菜63.89%,凤眼莲77.27%,Cd2+的去除率分别为浮萍48.32%,水蕹菜58.96%,凤眼莲76.87%。植物各器官对Pb2+-Cd2+的富集规律为:Pb2+Cd2+,根茎叶,凤眼莲水蕹菜浮萍。     

Role of plant activity and contaminant speciation in aquatic plant assimilation of 2,4,5-trichlorophenol

Aquatic plants uptake, transform and sequester organic contaminants and are used as a bioremediation strategy for the removal of pollutants from wastewaters. A better understanding of factors affecting rate of uptake of contaminants by aquatic plants is needed to improve engineered systems for removal of pollutants from wastewaters. This work focused on delineating sorption to plant surfaces and understanding effects of plant metabolic activity, inhibition, and media pH on the uptake of the ionizable contaminant 2,4,5-trichlorophenol (TCP) by aquatic plant Lemna minor. During L. minor exposure to TCP (0.5-13.9 mg l(-1)), a range of plant metabolic activities was measured using oxygen production rate (0-18.4 micromol h(-1)). A positive correlation was shown between contaminant uptake rate and plant activity. Contaminant uptake was examined at a range of media pH values (6-9) and uptake rates were linearly correlated to fraction of contaminant in protonated form. These results demonstrated a link between plant activity and uptake of contaminant by plants and stress the importance of incorporating plant metabolic activity and contaminant speciation in development of natural and engineered phytoremediation systems. This research also indicates that aquatic plants can actively accumulate trace-organic contaminants and may ultimately serve as a sink for these materials in the natural environment.     

The fate of isoproturon in a freshwater microcosm with Lemna minor as a model organism

Degradation, bioaccumulation and volatile loss of the 14C-labeled phenylurea herbicide isoproturon (IPU) was examined in a freshwater microcosm with the free floating macrophyte species Lemna minor during a 21-day exposure time. Isoproturon volatilisation was very low with 0.13+/-0.01% of the initially applied herbicide. Only a minor amount of the herbicide was completely metabolised, presumably by rhizosphere microorganisms and released as 14CO2. In total, about 9% isoproturon was removed from the aquatic medium during 21 days. The major portion of the pesticide was removed by bioaccumulation of Lemna minor (5.0+/-0.8%) and the bioconcentration factor (BCF) based on freshweight was 15.8+/-0.2. However, this study indicated a high persistence of IPU in freshwater ecosystems and a potential hazard due to bioaccumulation in non-target species. The novel experimental system of this study, developed for easy use and multiple sampling abilities, enabled quantitatively studying the fate of isoproturon and showed high reproducibility with a mean average (14)C-recovery rate of 97.1+/-0.7%.     

Metabolic fate of [14C]-2,4-dichlorophenol in macrophytes

The metabolic fate of 2,4-dichlorophenol (DCP) was investigated in six macrophytes representing different life forms. Salvinia natans and Lemna minor were chosen as surface-floating plants, Glyceria maxima and Mentha aquatica as emergent species and Myriophyllum spicatum and Hippuris vulgaris as submerged aquatic plants. After uptake of a [U-phenyl-14C]-DCP solution followed by a 48 h water chase, whole plants (L. minor, S. natans) or excised shoots were harvested and aqueous extracts were analysed by high performance liquid chromatography (HPLC). Metabolites were then isolated, submitted to enzymatic or chemical hydrolyses and characterised by electrospray ionisation-mass spectrometric analyses. Whereas DCP monoglucosides or more complex monoglucoside esters, either malonyl or acetyl, were found in most species, an unusual glucosyl-pentose conjugate was identified as the DCP major metabolite in L. minor and G. maxima. Our results showed for the first time the ability of five macrophytes to uptake and metabolise DCP and the characterisation of their metabolic pathways of DCP biotransformation.     

Biodegradation and removal of 3,4-dichloroaniline by Chlorella pyrenoidosa based on liquid chromatography-electrospray ionization-mass spectrometry

3,4-Dichloroaniline (3,4-DCA), widely used in the synthesis of dyes, textile and herbicides, is toxic to living organisms. The purpose of this study was to investigate the capability of green algae in degrading and removing 3,4-DCA in water. An environmentally ubiquitous green alga Chlorella pyrenoidosa was isolated from fresh aquatic environment. Then unicellular alga was incubated with 3,4-DCA at a concentration of 4.6 μg/?mL in water. The residual concentration of 3,4-DCA in the medium and the metabolites were analyzed. A removal percentage of 78.4 % was obtained over a 7-day period. Two major metabolites with less toxicity were identified as 3,4-dichloroformanilide and 3,4-dichloroacetanilide from the liquid chromatography-electrospray ionization-mass spectrometry analysis. The application of microalga C. pyrenoidosa may have potential for removing the environmental pollutant in aquatic environment.     

Comparison of different physiological parameter responses in Lemna minor and Scenedesmus obliquus exposed to herbicide flumioxazin

The sensitivity of different physiological parameters in Scenedesmus obliquus and Lemna minor exposed to herbicide (flumioxazin) was investigated to indicate the most convenient and sensitive parameter. To assess toxicity of flumioxazin, we used a panel of biomarkers: pigment contents, chlorophyll fluorescence parameters and antioxidative enzyme activities. Algae and duckweed were exposed to 48-h IC50 for growth rate. In L. minor, the sensitivity of the parameters was as follows: QN > Oxygen emmision > phiS(PSII) > QP > phi(PSII) > CAT, GR > Pigment> APO > Growth. For S. obliquus, this ranking was as follows: CAT > Oxygen emission > QP > APO > GR > Pigment > phiS(PSII) > Growth > phi(PSII) > QN (from the greatest to the least sensitive). The results demonstrated that the observed toxicity is related not only to interspecific variations but also to the selected parameter.     

Phytotoxicity and antioxidative enzymes of green microalga (Desmodesmus subspicatus) and duckweed (Lemna minor) exposed to herbicides MCPA, chloridazon and their mixtures

In this study, we evaluate the toxicity of MCPA (auxin-like growth inhibitor), chloridazon (CHD) (PSII-inhibitor) and their mixtures to floating plants and planktonic algae. Toxicity of MCPA (4-chloro-2-methylphenoxyacetic acid) and CHD (5-amino-4-chloro-2-phenyl-3(2H)-pyridazinone) was first assessed in two growth inhibition tests with Lemna minor (ISO/DIS 20079) and Desmodesmus subspicatus (ISO 8692). Next, herbicide mixtures at concentrations corresponding to the EC values were used to assess their interactive effects, and the biomarkers were: for duckweed fresh weight, frond area, chlorophyll content and number of fronds, and for algae cell count and cell volume. The 3d EC?? and EC?? values using cell counts of D. subspicatus were 142.7 and 529.1 mg/L for MCPA and 1.7 and 5.1 mg/L for CHD. The 7d EC?? and EC?? values using frond number of L. minor amounted to 0.8 and 5.4 mg/L for MCPA and 0.7 and 10.4 mg/L for CHD. Higher sensitivity of reproductive (number of cells/fronds) than growth processes (cell volume/frond area) to herbicides applied individually and in mixtures was especially pronounced in the responses of Desmodesmus. Herbicide interactions were assessed by the two-way ANOVA and Abbott's formula. Generally, an antagonistic interaction with Lemna was revealed by MCPA and chloridazon, whereas additive effect of both herbicides was observed for Desmodesmus. A significant stimulation of SOD and APX activity by binary mixtures was noted in algal cells mainly after 24 and 48 hours of exposure. The extremely high stimulation of the activity of both enzymes was induced by the combination EC??CHD + EC??MCPA (48 h). Presumably due to oxidative stress, the treatment with CHD at concentration EC?? after 72 h was lethal for algae grown in aerated cultures, in contrast to standardized test conditions. Taking into account the consequences of risk assessment for herbicide mixtures we can state that a relatively low toxicity, as well as the lack of significant synergy between MCPA and CHD to non-target plants appears to be the most important result.     

Effects of procymidone, fludioxonil and pyrimethanil on two non-target aquatic plants.

Procymidone, fludioxonil, and pyrimethanil are widely used to control the pathogenic fungus Botrytis cinerea in Champagne's vineyards. These fungicides may end up in surface waters and present potential risks for aquatic vascular plants and algae. Therefore, their toxicity was evaluated on Lemna minor and Scenedesmus acutus in six-day or 48-h tests, respectively. Based on growth and chlorophyll (Chl) content of L. minor and S. acutus cultures, the results showed that the alga was the most sensitive to the fungicides. Among the fungicides, pyrimethanil was the most toxic for L. minor, its nominal IC50 was 46.16 mg l(-1) and that of the other two was >100 mg l(-1). In contrast, pyrimethanil appeared the least toxic for S. acutus at low concentration, nominal IC50 were 22.81, 4.85, and 4.55 mg l(-1) for pyrimethanil, fludioxonil, and procymidone, respectively. Fate of the fungicides in the media was also investigated and acute toxicity of the agrochemicals is discussed in regard to concentration in the culture media. Poor solubility of procymidone and fludioxonil appeared to be partly responsible for the low toxicity of these fungicides. Based on these toxicity data and the concentrations found in ponds collecting vineyard runoff water, these pesticides should not impair the establishment of pioneer plants.     

Acceleration of catalase and peroxidase activities in Lemna minor L. and Allium cepa L. in response to low levels of aquatic mercury

The purpose of this study was to assess certain physiological responses of Lemna minor L. (duckweed) and Allium cepa L. (onion) to aquatic mercury at low concentrations. Following a 96-h exposure of plants to nutrient medium contaminated with known levels of mercuric chloride (HgCl(2)), 0.001 to 4 mg litre(-1) (0.0007 to 2.95 mg Hg litre(-1)) or methyl mercuric chloride (MeHgCl(2)), 0.0001 to 0.1 mg litre(-1) (0.0007 to 0.07 mg Hg litre(-1)), the physiological endpoints measured were the growth of fronds (Lemna minor) or roots (Allium cepa), and catalase and peroxidase activities in both plant assays. The EC(50) for HgCl(2) on the basis of the growth curve of Lemna minor was found to be 2.1 mg litre(-1). HgCl(2) and MeHgCl(2) were lethal to L. minor at concentrations of 4 and 0.01 mg litre(-1), respectively. The range of low concentrations that accelerated growth as well as enzymic activities in L. minor was 0.004 to 0.04 mg litre(-1) for HgCl(2) and 0.001 mg litre(-1) for MeHgCl(2). HgCl(2) and MeHgCl(2) induced maximum enzymic activity in Lemna fronds at concentrations of 0.008 and 0.0005 mg litre(-1), respectively. In Allium roots, catalase activity was accelerated at all the concentrations of HgCl(2) (0.001-2 mg litre(-1)) and MeHgCl(2) (0.0001-0.1 mg litre(-1)) tested. The activity of peroxidase was, however, accelerated by HgCl(2) at concentration range 0.01-1.0 mg litre(-1), or by MeHgCl(2) at 0.001 mg litre(-1). The concentrations of HgCl(2) and MeHgCl(2) that induced the highest enzymic activity in Allium roots were 0.05 mg litre(-1) and 0.001 mg litre(-1), respectively.     

Phytotoxicity of diuron alone and in combination with copper or folpet on duckweed (Lemna minor)

The photosystem II-herbicide diuron is widely used for weed control in Champagne's vineyards. Its important use and its relative persistence make it of particular interest for ecotoxicological studies. Toxicity of diuron was assessed on Lemna minor L., a representative aquatic macrophyte regularly used for toxicological studies. Toxicity assessments were based on inhibition of growth and total chlorophyll content of L. minor cultures after 7 days. Growth was inhibited and IC(50) and IC(90) were, respectively, 25 and 60 microg l(-1), but chlorophyll content of L. minor increased in response to the herbicide. When diuron was combined with copper, growth inhibition of L. minor depended on the concentrations of both chemicals. For some concentrations, combination of these chemicals resulted in a slight (but non-significant) antagonism. Additivity was observed for all other mixtures. When diuron was combined with folpet, growth and chlorophyll content of L. minor only depended on the concentration of the herbicide. Diuron was also found to prevent the copper-induced decrease of chlorophyll content when it was combined with this metal. A multifactorial model was found more appropriate to characterize interactions between pesticides than Abott's model.     

The effect of thallium on the growth of Lemna minor and plant tissue concentrations in relation to both exposure and toxicity

The effect of thallium on frond production and the rates of increase in surface area and fresh weight in Lemna minor were examined in terms of concentrations of the element in the growth medium and in the plant. The toxicity indices, EC(50) and threshold concentration, indicate that curtailment of frond expansion occurs at a lower thallium exposure than inhibition of frond multiplication. Production of smaller fronds, rather than chlorosis, was the most notable initial response to thallium stress and, below the threshold of toxicity, enhanced multiplication rates were recorded. Subsequent recovery in thallium-free medium was significant in the region of EC(50) exposures and below, but very little recovery occurred at higher exposures. Lemna minor has a remarkable capacity to accumulate thallium, and accumulation factors of 88 x 10(3) at low exposures, and 6 x 10(3) at high exposures, were recorded, which would suggest some form of active uptake.     

Degradation of 3,4-dichloro- and 3,4-difluoroaniline by Pseudomonas fluorescens 26-K

3,4-Dichloro- and 3,4-difluoroanilines were degraded by Pseudomonas fluorescens 26-K under aerobic conditions. In the presence of glucose strain degraded 170 mg/L of 3,4-dichloroaniline (3,4-DCA) during 2-3 days. Increasing of toxicant concentration up to 250 mg/L led to degradation of 3,4-DCA during 4 days and its intermediates during 5-7 days. Without cosubstrate and nitrogen source degradation of 3,4-DCA took place too, but more slowly--about 40% of toxicant at initial concentration 75 mg/L was degraded during 15 days. 3,4-Difluoroaniline (3,4-DFA) (initial concentration 170 mg/L) was degraded by Pseudomonas fluorescens 26-K during 5-7 days. The strain was able to completely degrade up to 90 mg/L of 3,4-DFA, without addition of cosubstrate and nitrogen during 15 days. Degradation of fluorinated aniline was accompanied by intensive defluorination. Activity of catechol 2,3-dioxygenase (C2,3DO) (0.230 micromol/min/mg of protein) was found in the culture liquid of the strain, grown with 3,4-DCA and glucose. This fact, as well as, the presence of 3-chloro-4-hydroxyaniline as a metabolite suggested that 3,4-DCA degradation pathway includes dehalogenation and hydroxylation of aromatic ring followed by its subsequent cleaving by C2,3DO. On the contrary, activity of catechol 1,2-dioxygenase (C1,2DO) (0.08 micromol/min/mg of protein) was found in the cell-free extract of biomass grown on 3,4-DFA. 3-Fluoro-4-hydroxyaniline as intermediate was found in this cell-free extract.     

Phytotoxicity of silver nanoparticles to Lemna minor L

The use of silver nanoparticles (AgNPs) in commercial products has increased significantly in recent years. Although there has been some attempt to determine the toxic effects of AgNPs, there is little information on aquatic plants which have a vital role in ecosystems. This study reports the use of Lemna minor L. clone St to investigate the phytotoxicity of AgNPs under modified OECD test conditions. AgNPs were synthesised, characterised and subsequently presented to the L. minor. Results showed that inhibition of plant growth was evident after exposure to small (∼20 nm) and larger (∼100 nm) AgNPs at low concentrations (5 μg L⁻¹) and this effect became more acute with a longer exposure time. There was a linear dose-response relationship after 14 d exposure. Using predicted environmental concentrations for wastewaters it was found that AgNPs may pose a significant potential risk to the environment.     

Comparing growth development of <Emphasis Type="Italic">Myriophyllum</Emphasis> spp. in laboratory and field experiments for ecotoxicological testing

BACKGROUND, GOALS AND SCOPE: Risk assessment of herbicides and the evaluation of contaminated sediments based on algae and the macrophyte Lemna sp. alone may underestimate the potential hazard of certain compounds. Therefore, various test systems with Myriophyllum spp. have been developed recently to assess the phytotoxicity in surface waters and natural sediments. In the present study, experiments investigating the growth development of Myriophyllum spp. were performed in the laboratory under defined conditions and in mesocosms under environmentally realistic exposure conditions to evaluate the suitability of these species as potential standard test organisms in ecotoxicological testing. This study provides data on the endpoints biomass, plant length and root development. MATERIALS AND METHODS: Six independent experiments were performed to investigate the plant development of Myriophyllum spp. under control conditions. The main difference in the experiments was the complexity of the test systems ranging from simple laboratory experiments to complex outdoor mesocosm studies. At the start of each experiment, uniform cuttings of Myriophyllum spp. were placed in vessels with or without sediments to reduce variability between replicates. The endpoints considered in this investigation were biomass (fresh weight of the whole plant), length of the main shoot, length of the side shoots, total length of the plant (calculated from the length of the main and side shoots) and root formation. Root to shoot ratios were calculated as a further measure for plant development. Relative growth rates (RGR) based on plant length (RG(L)R) and on biomass (RG(B)R) were calculated. RESULTS: Despite the various experimental conditions, comparable growth was obtained in all test systems and the variability of endpoints, such as total length and biomass of plants, was low. It was observed that the RGR of M. spicatum in the simple laboratory test system with sediment were comparable to growth data obtained for M. verticillatum and M. spicatum grown in indoor and outdoor mesocosms, thus indicating that Myriophyllum growth tends to increase by the addition of sediment. High variability was determined for the endpoints length of the side shoots, total root length and biomass of roots. DISCUSSION: One challenge for a test design to investigate phytotoxicity on aquatic plants is to obtain good growth of the plants. From the results, it can be concluded that the experimental conditions in the various test systems were suitable to study the plant development of Myriophyllum spp. because obtained growth rates were comparable between laboratory and field investigations. Another challenge for developing a plant biotest system is the definition of sensitive endpoints. Low variability is preferred to detect minor effects of chemicals or polluted sediments on plant development. In our studies, the variability of the endpoints biomass and total length of plant was low and, therefore, they have much potential as endpoints for assessing toxicity. CONCLUSIONS: The methodologies presented in this study have applications within the risk assessment for aquatic plants and have the advantage of assessing effects taking into account the relevant exposure pathways via water and/or sediment for compounds under investigation. RECOMMENDATIONS AND PERSPECTIVES: Setting safe quality criteria for surface water and sediments is one of the challenges authorities are facing today. Myriophyllum spp. is recommended as suitable test species to investigate phytotoxicity in surface water and sediments. These results, thus, might serve as a basis for the compilation of a new harmonised guideline for ecotoxicological testing with aquatic macrophytes.     

Xenobiotics removal from polluted water by a multifunctional constructed wetland

Removal efficiencies on xenobiotics from polluted water in a twin-shaped constructed wetland consisting of a vertical flow chamber with the crop plant Colocasia esculenta L. Schott and a reverse vertical flow one with Ischaemum aristatum var. glaucum Honda, were assessed by chemical analysis and bioassays. After a four-month period of application, removal efficiencies of the applied pesticides parathion and omethoate were 100% with no detectable parathion and omethoate in the effluent. For the applied herbicides, the decontamination was less efficient with removal efficiencies of 36% and 0% for 4-chloro-2-methyl-phenoxyacetic acid and dicamba, respectively. As shown by toxicity assay with duckweed Lemna minor L., growth retardation may occur if the water treated for herbicide removal is used in irrigation of sensitive cultivars in agriculture or horticulture. In contrast to I. aristatum var. glaucum Honda, the crop C. esculenta L. Schott has a high yield in biomass production as a valuable source of renewable energy.