Subcapitata Growth Research Articles

Polymers of low concern? Assessment of microplastic particles used in 3D printing regarding their toxicity on Raphidocelis subcapitata and Daphnia magna

The potential effects of microplastic particle exposure on environmental organisms has sparked intense research activities. Various studies have been conducted, however on a limited set of mostly pristine polymer materials. In parallel to the ongoing research activities, it is discussed to include non-natural polymers into the registration process under REACH. Currently, non-natural polymers are exempted from registration, based on the general assumption of being non-hazardous due to their high molecular weight. In addition, the extensive number of polymers exceeding the registration capacities was mentioned as a reason for exemption. Hence, relevant polymers requiring a registration shall be selected according to specific criteria that help to identify those with a concern for hazardous effects. In the line of these developments we here present the results of a systematic ecotoxicity testing of 16 microplastic particles of different polymer composition, part of which are micronized polymer powders used in 3D printing (intentionally produced primary microplastic, losing particle shape by the 3D printing). All polymer materials were systematically varied and extensively characterised with regard to their properties (e.g. particle size, cross-linking, molar mass distribution, end groups, reactivity). Most of the polymers exerted toxicity in 48 h acute Daphnia magna immobilisation assay and 72 h chronic Raphidocelis subcapitata growth inhibition assay, except one PA and one HDPE material showing toxicity in D. magna, and one HDPE material showing toxicity in green algae. From these results we conclude that none of the microplastic particles studied here, independent of their polymer properties, give raise to concern for hazardous effects.

Chronic Toxicity of Iron to Aquatic Organisms under Variable pH, Hardness, and Dissolved Organic Carbon Conditions.

A series of chronic toxicity tests was conducted exposing three aquatic species to iron (Fe) in laboratory freshwaters. The test organisms included the green algae Raphidocelis subcapitata, the cladoceran Ceriodaphnia dubia, and the fathead minnow Pimephales promelas. They were exposed to Fe (as Fe (III) sulfate) in waters under varying pH (5.9-8.5), hardness (10.3-255 mg/L CaCO3 ), and dissolved organic carbon (DOC; 0.3-10.9 mg/L) conditions. Measured total Fe was used for calculations of biological effect concentrations because dissolved Fe was only a fraction of nominal and did not consistently increase as total Fe increased. This was indicative of the high concentrations of Fe required to elicit a biological response and that Fe species that did not pass through a 0.20- or 0.45-µm filter (dissolved fraction) contributed to Fe toxicity. The concentrations frequently exceeded the solubility limits of Fe(III) under circumneutral pH conditions relevant to most natural surface waters. Chronic toxicity endpoints (10% effect concentrations [EC10s]) ranged from 442 to 9607 µg total Fe/L for R. subcapitata growth, from 383 to 15 947 µg total Fe/L for C. dubia reproduction, and from 192 to 58,308 µg total Fe/L for P. promelas growth. Toxicity to R. subcapitata was variably influenced by all three water quality parameters, but especially DOC. Toxicity to C. dubia was influenced by DOC, less so by hardness, but not by pH. Toxicity to P. promelas was variable, but greatest under low hardness, low pH, and low DOC conditions. These data were used to develop an Fe-specific, bioavailability-based multiple linear regression model as part of a companion publication. Environ Toxicol Chem 2023;00:1-15. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Freshwater alga Raphidocelis subcapitata undergoes metabolomic changes in response to electrostatic adhesion by micrometer-sized nylon 6 particles

Nylon powders are a type of microplastic (MP) used in personal care products such as cosmetics and sunscreens. To determine the effects of nylon polymers on freshwater microalgae, we investigated the effects of two types of micrometer-sized nylon polymers, i.e., powdered nylon 6 (Ny6-P) and nylon 12 (Ny12), and four other micrometer-sized MPs, i.e., low-density polyethylene, polyethylene terephthalate, polystyrene, and ultra-high-molecular-weight polyethylene, on the microalga Raphidocelis subcapitata. The results showed that Ny6-P inhibited R. subcapitata growth more than the other MPs; R. subcapitata growth was inhibited by 54.2% with 6.25 mg/L Ny6-P compared with the control. Ny6-P in the culture media adhered to R. subcapitata cells electrostatically, which may have disrupted growth and photosynthetic activity. Metabolomic analysis revealed that many metabolites related to the amino acid catabolic pathway and γ-glutamyl cycle were induced, which might trigger responses to avoid starvation and oxidative stress. Our study provides important information on the effects of Ny6-P on algae in freshwater environments.

Greening effect of slag cement-based concrete: Environmental and ecotoxicological impact

Materials containing ground granulated blast furnace slag (GGBFS) display a transient green–bluecolor after demolding. This greening effect has been investigated for leaching behavior and ecotoxicological impact. Color of concretes and pure pastes containing GGBFS was assessed with a portable spectrophotometer, and samples were then submitted to a tank monolith leaching test. Ecotoxicological tests were conducted on reference sample and a green concrete sample at both natural and adjusted pH of 8.1. Main results support that the temporary greening effect of GGBFS-containing materials has no particular impact neither on the chemistry of leachates, nor ecotoxicity. Additionally, alkaline leachates are the main issue of leached cement or GGBFS based materials with pH around 11.5–12.5. Alkaline pH is a preponderant factor of ecotoxicity to sensitive organisms such as Daphnia magna , immobilization assay (48 h) resulting in 5.10 Toxic Units (TU) for reference sample at pH 12.50 against 1.38 TU at pH 8.10. Furthermore, sulfides are a specific issue of GGBFS materials concentrated up to 0.94 mmol m −2 in leachates, having an ecotoxic impact on living organisms at all trophic levels. At pH 8.10, green concrete leachates have 4.85 TU for Raphidocelis subcapitata growth assay (sulfides concentration of 0.63 mmol m −2 ) against 3.0 TU for green concrete sample. However, sulfides are easily removed from natural solution by oxidation or evaporation. • Greening effect is not associated with hazardous leachates or ecotoxic effects. • Alkaline pH (> 11) is the main issue with cement or slag-based materials. • The algae R. subcapitata is not affected by high pH unlike the crustacean D. magna . • GBFS contain sulfides up to 1 wt.% release in eluates of concretes and pastes. • Sulfides are quickly neutralized in natural environment.

Remediation of acid mine drainage-affected stream waters by means of eco-friendly magnetic hydrogels crosslinked with functionalized magnetite nanoparticles

Acid mine drainage (AMD) is a global environmental challenge that represents a primary source of contamination of heavy metals in both active and abandoned mining areas. The present study explored the suitability of a carboxymethylcellulose (CMC) hydrogel cross-linked with functionalized magnetite (Fe3O4) nanoparticles, (CMC-Fe3O4), for the removal of heavy metals from AMD-affected stream waters and tested its eco-friendly nature. AMD-affected stream waters, collected in a former mining site located in the Metalliferous Hills ore district (Central Italy), were chosen as a suitable benchmark for validating the CMC-Fe3O4 hydrogel applications. The remediation capability of the CMC-Fe3O4 hydrogel was investigated on solutions of Zn(II) in deionized (DI) water and on samples of AMD-affected stream waters. The CMC-Fe3O4 hydrogel was able to adsorb Zn(II) from DI water solutions as well as to reduce significantly the concentration of Zn and other heavy metals such as Cd, Co, Cu, Ni and Pb, in AMD-affected stream waters. CMC-Fe3O4 hydrogel and treated waters (both Zn-solution and AMD-affected stream waters) were tested for their effects on the freshwater green alga R. subcapitata by using 72-h growth inhibition tests. No effects were observed on the R. subcapitata growth upon exposure to CMC-Fe3O4 hydrogel, suggesting its eco-friendly nature. Moreover, CMC-Fe3O4 hydrogel was able to reduce the toxicity of heavy metals to the green alga, both in DI and AMD-affected stream water. Based on our findings, CMC-Fe3O4 hydrogel could be considered as a valuable tool for an environmentally safe treatment of AMD-affected stream waters.

Eco-friendly profile of pegylated nano-graphene oxide at different levels of an aquatic trophic chain

Nanographene oxide (nGO) has been recently proposed as a new antitumoral therapeutic agent, drug delivery carrier and gene transfection vehicle, among others. Treatment is carried out by hyperthermia induced by infrared irradiation. After treatment, the nanosystems will be inevitably excreted and released to the environment. To understand the potential impacts of pegylated nGO (nGO-PEG), three key species from different trophic levels were used: the green micro-algae Raphidocelis subcapitata (growth inhibition test), the cladocera Daphnia magna (acute and chronic tests), and the fish Danio rerio (fish embryo test). Besides a regular standard procedure to assess toxicity, and considering the mode of action of nGO-PEG in cancer treatment, a simultaneous infrared lamp exposure was carried out for D. magna and D. rerio. Additionally, and taking advantage of the phenotypic transparency of D. magna, nGO-PEG was fluorescently tagged to evaluate the potential uptake of nGO-PEG. The R. subcapitata growth inhibition test showed effects during the first 48 h, recovering till the end of the test (96 h). No acute or chronic effects were observed for D. magna, under standard or infrared light exposures although confocal microscope images showed nGO-PEG uptake. Very small percentages of mortality and abnormalities were observed in D. rerio exposed with and without the infrared lamp. Although low hazard may be expected for nGO-PEG in aquatic ecosystems, further studies with species with different life traits should be accomplished, in order to derive more accurate conclusions.

Ecotoxicity and antibiotic resistance of a mixture of hospital and urban sewage in a wastewater treatment plant.

Hospital and urban effluents are a source of diverse pollutants such as organic compounds, heavy metals, detergents, disinfectants, pharmaceuticals, and microorganisms resistant to antibiotics. Usually, these two types of effluent are mixed in the sewage network, but a pilot site in France now allows studying them separately or mixed to understand more about their characteristics and the phenomena that occur following their mixing. In this study, their ecotoxicity (Daphnia magna mobility, Pseudokirchneriella subcapitata growth, Brachionus calyciflorus reproduction, and SOS Chromotest) and antibiotic resistance (integron quantification) were assessed during mixing and treatment steps. The main results of this study are (i) the ecotoxicity and antibiotic resistance potentials of hospital wastewater are higher than in urban wastewater and (ii) mixing two different effluents does not lead to global synergistic or antagonistic effects on ecotoxicity and antibiotic resistance potential. The global additivity effect observed in this case must be confirmed by other studies on hospital and urban effluents on other sites to improve knowledge relating to this source of pollution and its management.

Effects of diuron and carbofuran and their mixtures on the microalgae Raphidocelis subcapitata

In aquatic environments, organisms are often exposed to mixtures of several pesticides. In this study, the effects of carbofuran and diuron and their mixtures on the microalgae Raphidocelis subcapitata were investigated. For this purpose, toxicity tests were performed with the single compounds (active ingredients and commercial formulations) and their combinations (only active ingredients). According to the results, the toxicity of active ingredients and their commercial formulations to R. subcapitata was similar. In the single exposures, both carbofuran and diuron inhibited significantly the R. subcapitata growth and caused physiological (chlorophyll a content) and morphological (complexity and cell size) changes in cells, as captured by flow cytometry single-cell properties. Regarding the mixture toxicity tests, data fitted to both reference models, concentration addition (CA) and independent action (IA), and evidenced significant deviations. After the CA fitting, dose-ratio dependent deviation had the best fit to the data, demonstrating synergism caused mainly by diuron and antagonism caused mainly by carbofuran. After fitting the IA model, a synergistic deviation represented the best fit for the diuron and carbofuran mixtures. In general, the two reference models indicated the occurrence of synergism in the mixtures of these compounds, especially when diuron was the dominant chemical in the combinations. The increased toxicity caused by the mixture of these pesticides could pose a greater environmental risk for phytoplankton. Thus, exposure to diuron and carbofuran mixtures must also be considered in risk assessments, since the combination of these compounds may result in more severe effects on algae population growth than single exposures.

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).

Occurrence, environmental impacts and removal of legacy and emerging contaminants from two wastewater and one water treatment plant in Southern Ontario. Part II: environmental impacts

This is part two of a paper about the potential environmental impacts of treated effluent from a wastewater treatment plant (WWTP) discharging to the Detroit River in Windsor, Ontario, Canada. The WWTP uses conventional activated sludge with nitrification. The assessment was conducted over six months using a variety of established tests, including in vitro cell-based screening assays, as well as acute, chronic and full-life cycle in vivo exposures. Effluent monitoring included pharmaceutically active compounds and endocrine disrupting compounds. No tests reported significant toxicity. However, enhanced algal growth was observed in a Pseudokerchneriella subcapitata growth inhibition test. In full life-cycle fathead minnow exposure, liver-somatic index changes were noted in exposed fish – increases for males, decreases for females – and production of viable fry decreased. Neither alteration is thought biologically significant. Because the effluent is diluted substantially by the receiving water, the level of risk posed to aquatic receptors and the environment is probably negligible.

Ecological Risk Assessment of a Metal-Contaminated Area in the Tropics. Tier II: Detailed Assessment.

This study presents data on the detailed evaluation (tier 2) of a site-specific ecological risk assessment (ssERA) in a former smelter area contaminated with metals (Santo Amaro, Bahia, Brazil). Combining information from three lines of evidence (LoE), chemical (ChemLoE), ecotoxicological (EcotoxLoE) and ecological (EcoLoE), in the Triad approach, integrated risk values were calculated to rank sites and confirm the potential risk disclosed with tier 1. Risk values were calculated for the habitat and for the retention functions in each sampling point. Habitat function included the ChemLoE calculated from total metal concentrations. The EcotoxLoE was based on reproduction tests with terrestrial invertebrates (Folsomia candida, Enchytraeus crypticus, Eisenia andrei), shoot length and plant biomass (Avena sativa, Brassica rapa). For the EcoLoE, ecological parameters (microbial parameters, soil invertebrate community, litter breakdown) were used to derive risk values. Retention function included the ChemLoE, calculated from extractable metal concentrations, and the EcotoxLoE based on eluate tests with aquatic organisms (Daphnia magna reproduction and Pseudokirchneriella subcapitata growth). Results related to the habitat function indicated that the metal residues are sufficient to cause risk to biota, while the low metal levels in extracts and the general lack of toxicity in aquatic tests indicated a high soil retention capacity in most sampling points. Integrated risk of tier 2 showed the same trend of tier 1, suggesting the need to proceed with remediation actions. The high risk levels were related to direct toxicity to organisms and indirect effects, such as failure in the establishment of vegetation and the consequent loss of habitat quality for microorganisms and soil fauna. This study shed some light on the selection of tools for the tier 2 of an ssERA in tropical metal-contaminated sites, focusing on ecological receptors at risk and using available chemical methods, ecological surveys and ecotoxicity tests.

The aquatic impact of ionic liquids on freshwater organisms

Ionic liquids (ILs), also known as liquid electrolytes, are powerful solvents with a wide variety of academic and industrial applications. Bioassays with aquatic organisms constitute an effective tool for the evaluation of ILs’ toxicity, as well as for the prediction and identification of possible moieties that act as toxicophores.In this work, the acute toxicity of six ILs and two commonly used organic solvents was evaluated using freshwater organisms: Daphnia magna, Raphidocelis subcapitata and Hydra attenuata. The bioassays were performed by exposing the organisms to increasing concentrations of the ILs and observing D. magna immobilization, R. subcapitata growth inhibition, and the morphological or mortality effects in H. attenuata. The results demonstrate that the tested organisms are not equally susceptible to the ILs, e.g., bmpyr [BF4] was the least toxic compound for R. subcapitata, N1,1 [N1,1,1OOH] for D. magna and emim [Tf2N] for H. attenuata. This highlights the importance of applying a battery of assays in toxicological analysis. Additionally, Hydra proved to be the most tolerant species to the tested ILs. According to their hazard rankings, the tested ILs are considered practically harmless or moderately toxic, except (Hex)3(TDec)P [Cl], which was classified as highly toxic. The ILs were revealed to be more harmful to aquatic systems than the tested organic solvents, reaffirming the need to analyze carefully the (eco)toxicological impact of these compounds. The present study provides additional data in the evaluation of the potential hazard and the impact of ILs in the environment.

Comparison of approaches towards ecotoxicity evaluation for the application of dredged sediment on soil

The toxicity of 36 dredged sediments from the Czech Republic was investigated using a large battery of bioassays. The aim of this study was to investigate the feasibility of ecotoxicity testing in general and of individual bioassays more specific and to investigate how the results of bioassays are determined by the physicochemical properties of sediment samples and/or sediment contamination. In 2008 and 2010, 36 sediment samples were collected from rivers and ponds and from sediment heaps in different parts of the Czech Republic. Both their physicochemical properties and their levels of contamination with POPs and heavy metals were analyzed. The ecotoxicities of the sediments were evaluated using the four bioassays from the new Czech directive 257/2009 Coll. concerning the application of dredged sediments on agricultural land (Enchytraeus crypticus reproduction, Folsomia candida reproduction, Lactuca sativa root elongation, and potential ammonium oxidation). The results of the four directive bioassays were compared with the results of other soil bioassays (Caenorhabditis elegans mortality, Eisenia fetida avoidance and reproduction) and eluate bioassays (Daphnia magna immobilization, Pseudokirchneriella subcapitata growth inhibition test, and Vibrio fischeri luminescence). We demonstrate that the battery suggested in Czech directive 257/2009 Coll. is highly effective in identifying toxic samples; these bioassays clearly revealing different types of toxicity and different exposure routes. Shorter alternative bioassays may be added especially when fast toxicity identification is needed. Eluate bioassays identified samples potentially hazardous to aquatic ecosystems. Their inclusion into the assessment scheme should be considered if the goal of assessment is also the protection of aquatic ecosystems. The results of our multivariate analysis show that specific physicochemical properties and contamination may affect bioassay responses. C. elegans was the most sensitive bioassay to physicochemical properties and also to organic contamination, while eluate bioassays were sensitive to heavy metal pollution. Most effects detected by the bioassays could not be explained by the levels of toxicants measured or by the natural characteristics of sediments. Our results show that bioassays are irreplaceable in dredged sediment risk assessment because they complement information provided by chemical analyses.

Assessment of sediment quality and pore water ecotoxicity in Kebir Rhumel basin (NE-Algeria): a combined approach

The objectives of this study are to use different approaches to assess the current pollution status in the wadis of the Kebir Rhumel basin. First, sediment trace metal contents were measured by flame atomic absorption spectroscopy. Then, sediment quality was assessed on the basis of contamination assessment indexes such as: Geoaccumulation Index (Igeo), Contamination factor (C(f)), Contamination degree (C(d)), Sediment Pollution Index (SPI) and SEQ guidelines (Consensus Sediment Quality Guidelines). In addition, several toxicity tests (Daphnia magna mobility inhibition acute test-48 h, Aliivibrio fischeri luminescence inhibition acute test - 15/30 mn and Pseudokirchneriella subcapitata growth inhibition chronic test - 72 h) were conducted to assess sediment pore water ecotoxicity. Trace metal concentrations followed the order: Mn > Zn > Pb > Cr > Cu > Ni > Co > Cd. Indexes used indicate varying degrees of sediment quality. Igeo, C(f), C(d) and SPI reveal a polymetallic contamination dominated by two or more elements in which Cd, Cu and Pb are of greatest concern. SEQ guidelines showed that biological effects on fauna would likely be observed occasionally and/or frequently for Cd, Cr, Cu, Pb and Zn contents. Test organisms exposed to sediment pore water showed that the algal P. subcapitata test was more sensitive than the D. magna and A. fischeri tests. Hence, algal growth inhibition proved to be the most sensitive response to contaminants present in sediment extracts but a significant relationship with trace metal contents was not demonstrated.

Oxidation of triclosan by ferrate: Reaction kinetics, products identification and toxicity evaluation

The oxidation of triclosan by commercial grade aqueous ferrate (Fe(VI)) was investigated and the reaction kinetics as a function of pH (7.0–10.0) were experimentally determined. Intermediate products of the oxidation process were characterized using both GC–MS and RRLC–MS/MS techniques. Changes in toxicity during the oxidation process of triclosan using Fe(VI) were investigated using Pseudokirchneriella subcapitata growth inhibition tests. The results show that triclosan reacted rapidly with Fe(VI), with the apparent second-order rate constant, kapp, being 754.7M−1s−1 at pH 7. At a stoichiometric ratio of 10:1 (Fe(VI):triclosan), complete removal of triclosan was achieved. Species-specific rate constants, k, were determined for reaction of Fe(VI) with both the protonated and deprotonated triclosan species. The value of k determined for neutral triclosan was 6.7(±1.9)×102M−1s−1, while that measured for anionic triclosan was 7.6(±0.6)×103M−1s−1. The proposed mechanism for the oxidation of triclosan by the Fe(VI) involves the scission of ether bond and phenoxy radical addition reaction. Coupling reaction may also occur during Fe(VI) degradation of triclosan. Overall, the degradation processes of triclosan resulted in a significant decrease in algal toxicity. The toxicity tests showed that Fe(VI) itself dosed in the reaction did not inhibit green algae growth.

Structural effects of the bioavailable fraction of pesticides in soil: Suitability of elutriate testing

This study focused the ecotoxicological evaluation of four different pesticides (chlorpyrifos, glyphosate, vinclozolin, endosulfan), sprayed into an agricultural soil, using a standard battery of aquatic bioassays for testing of soil elutriates: Vibrio fischeri -Microtox®; Pseudokirchneriella subcapitata growth inhibition; Daphnia magna acute and chronic toxicity. Despite relevant pesticide residues were recovered from the soil matrix (concentrations higher than 1000μgkg−1), much lower concentrations could be retrieved from elutriates (highest records for endosulfan of ca. 250ngL−1and 1400ngL−1; dissolved and particulate concentration, respectively) and little effects were generally found in the bioassays. Lethal effects (D. magna 48h-EC50 of 36.8%) could be noticed following exposure to the endosulfan elutriate. Elutriates induced no toxicity on V. fischeri; algal growth was generally inhibited at high elutriate dilutions and stimulated at the lower elutriate dilutions; and no overall impairment of D. magna life-history was noticed. Results revealed that cross-contamination during field application, input of organic matter and nutrients by elutriates in test solutions, and choice of test species and endpoints may constrain the ecotoxicological assessment. Suitability of established aquatic bioassay test batteries for these purposes, and questioning on whether direct assays with soil organisms could be more protective tools is discussed.

The Combined Toxic and Genotoxic Effects of Chromium and Volatile Organic Contaminants to Pseudokirchneriella subcapitata

In this report, the toxic effect of TCE (trichloroethylene), PCE (tetrachloroethylene), and potassium dichromate on P. subcapitata was investigated. The test was conducted at different concentrations of pollutants, starting from the European Community limit values defined for each analysed contaminant. Mixtures of pollutants were also tested to verify the combined effect of algae cells. Results suggest that both TCE and PCE were able to reduce P. subcapitata growth and metabolism starting from 0.05 and 0.02 mg L−1 of contaminant, respectively. PCE seems to be substantially more toxic than TCE. Chromium produces a clear effect on algae growth and esterase activity only starting from 1 mg L−1 of potassium dichromate; this result confirms the suitability of EU limit value. AFLP analysis showed that all tested pollutants produce DNA mutations probably due to oxygen radicals. Generally, chromium, at high concentrations, is more toxic and genotoxic that TCE or PCE. Test performed with a mixture of pollutants showed a synergic effect of chromium and organic compounds suggesting that the membrane damage induced from organic substances should increase the chromium cellular access.

Growth Stimulation and Inhibition Effects of 4-Hydroxybenzoic Acid and Some Related Compounds on the Freshwater Green Alga Pseudokirchneriella subcapitata

4-hydroxybenzoic acid (4-HBA) exhibited low algal toxicity with the 72-h median inhibition concentration (IC50) of 9.9 mmol/L in the standard growth inhibition test using the freshwater green alga Pseudokirchneriella subcapitata. In contrast, it stimulated the algal growth at lower concentrations ranging from 0.1 to 1.0 mmol/L. Comparative studies with benzoic acid and 2- and 3-hydroxybenzoic acids (2-HBA and 3-HBA) indicated that 2-HBA was the most toxic, giving a 72-h IC50 of 0.172 mmol/L, and 4-HBA was the least toxic and that only 4-HBA had the pronounced growth stimulation activity. In a semicontinuous exposure to 4-HBA (0.15 and 0.3 mmol/L), algae maintained increased cell growth compared with controls during up to 10 times consecutive batch cultures, without any indication of adaptive responses to the growth enhancing effect of 4-HBA. Return to the clean standard medium of the exposed cells resulted in the quick recovery from the stimulant effect. Furthermore, 4-HBA (0.3 mmol/L) was found to diminish the toxicity of 2-HBA (<1.0 mmol/L). The presence of such compounds in the complex mixture might interact with other toxicants to alter the whole toxicity when assessed by the algal growth inhibition test. The effects of 4-HBA on P. subcapitata growth observed in the present study are not expected for planktonic algae in the aquatic environments, because known environmental concentrations are far below the effective concentration range.