Microbial responses to zinc in soil microcosms with and without a natural assemblage of enchytraeids

The presence of higher trophic levels in studies on the toxicity on soil contaminants to microbial processes increases ecological realism. This study assessed the toxicity of zinc to soil microbial processes in the presence and absence of enchytraeids (Oligochaeta, Annelida). We incubated microcosms under standard conditions without or inoculated with a natural assemblage of enchytraeid species. Total zinc concentrations of 365 to 1,360 mg/kg caused no mortality of enchytraeids during six weeks' incubation. Soil nitrate concentrations showed a negative trend under zinc addition and soil ammonium concentrations were the highest at zinc concentrations of 1,360 mg/kg, indicating impairment of ammonium oxidation. Zinc decreased bacterial carbon biomass and caused a dose-response decrease of the respiration, but this was not observed in the presence of enchytraeids. Respiration, ammonium concentrations, and soil moisture contents were increased by enchytraeids. We observed no interaction between the addition of zinc and the presence of enchytraeids. The effect of enchytraeids on soil-moisture contents and microbial processes, and the importance of enchytraeid-microbial interactions are discussed.     

Toxicity of four nitrogen-heterocyclic polyaromatic hydrocarbons (NPAHs) to soil organisms

The aims of this study were: (i) to investigate the toxicity of N-heterocyclic polyaromatic hydrocarbons (NPAHs) quinoline, acridine, phenazine, and 1,10-phenanthroline to the soil invertebrates Eisenia fetida, Enchytraeus crypticus, Folsomia candida, and Caenorhabditis elegans, (ii) to compare the toxicity of four NPAHs and the species sensitivity, and (iii) to discuss possible risks of these compounds in soils. Different toxicities were found for the tested NPAHs which might be partially explained by their structure and properties. Effect concentrations expressed as soil pore-water concentrations were related to log K_ow, which indicated narcosis as the most probable mode of toxic action. The species sensitivity decreased in the rank: springtails >enchytraeids=earthworms> nematodes. Predicted no-effect concentration (PNEC) values were calculated for all tested species giving values from 0.5 to 6.8 mg/kg. It is unlikely that there is a risk for soil organisms in natural soils where lower NPAHs concentrations are expected.     

Terrestrial model food chain and environmental chemicals : I. Transfer of sodium [C]pentachlorophenate between springtails and carabids

A model soil food chain of a ruderal ecosystem has been constructed in order to study the uptake, transfer, and accumulation of [14C]pentachlorophenate (PCP-Na). The model was based on three food levels, viz. baker's yeast, collembola, and carabid beetles, and the contaminant chemical introduced was via initial food. Continuous exposure of the organisms to the test chemical resulted in a significant uptake and transfer of radiocarbon into the food chain elements. Bioaccumulation of radiocarbon in the body tissues of the organisms was low, as large amounts taken up were quickly eliminated through the excrements. The radiocarbon level of prey animals was about 100 times higher than that of their predators, but there was only small difference in concentration between collembolas and yeast. This was probably because of a faster excretion of the chemical by the beetles than by the collembolas. During the test period no conversion of [14C]PCP-Na took place in the yeast, but the collembolas and beetles metabolized 50 and 59%, respectively. Criteria are proposed for successful implementation of food chain models.     

Interaction modification among decomposers impairs ecosystem processes in lead-polluted soil

Environmental pollution by toxicants is generally believed to lead both to direct toxic effects and indirect effects via altered species interactions in a stressed community. We examined how contamination of a coniferous forest soil floor by lead (from an abandoned shooting range area) may alter trophic interactions and decomposition processes. We constructed laboratory microcosms containing microbial-based food webs with various trophic structures: microbes alone; microbes with microbivorous-detritivorous enchytraeid worms (Cognettia sphagnetorum, a potential keystone species of boreal forest soil); and microbes, worms, and predatory mites (Lysigamasus spp.) together. The direct toxic effect of lead and the effectiveness of the predators in preying upon the worms were studied in separate experiments. Polluted humic soil (acid-extractable lead concentration was 2,720 +/- 349 mg/kg of dry soil [mean +/- standard error]) was toxic to enchytraeids, and predators effectively reduced worm populations. Positive density-dependent effects of worms on microbes and a trophic cascade from predatory mites to microbes, observed in the unpolluted humus, were not observed in the polluted humus, indicating pollution-induced interaction modification among organisms. Concomitantly, lead pollution altered the process of nutrient mineralization in the humic soil. However, causality and relative importance of direct toxicity and indirect effects of lead on the dynamics of the decomposer food web could not be pointed out. Examination of our results indicates that traditional food web models are incapable of predicting mechanisms that alter dynamics of pollution-stressed belowground food webs.     

The use of enchytraeids in ecological soil classification and assessment concepts

In many soils worldwide, enchytraeids (Annelida: Oligochaeta) play a major role in soil functions like the decomposition of organic matter. In addition, standardized sampling methods are available and taxonomic as well as ecological knowledge is rapidly increasing (in particular, ecological profiles of the most abundant species). For these reasons, enchytraeids were recognized as a part of ecological classification and assessment schemes. In this contribution, the most important of these schemes are presented and it is shown that enchytraeids are most valuable as part of a battery approach. By use of examples from The Netherlands (BISQ), and Germany (BBSK), the practicability of the use of enchytraeids is demonstrated in identifying the influence of different anthropogenic land use forms. In these cases, the structure of the enchytraeid community and their abundance and biomass were used as endpoints. However, automation of the identification process can still be improved.     

Effects of Zinc Contamination on a Natural Nematode Community in Outdoor Soil Mesocosms

The effect of zinc on the indigenous nematode fauna of a sandy soil was determined in an experimentally contaminated outdoor field plot. The aims of the study were to describe and quantify the changes in density of separate nematode taxa and total nematodes, and the changes in the number of taxa, species diversity, community maturation, and species composition in response to zinc exposure with time, and to compare the observed responses with benchmark concentrations for soil as derived from the species sensitivity distribution (SSD) for zinc toxicity. Speciation of zinc in pore water was considered and CaCl₂-exchangeable zinc concentrations were used as a measure of the bioavailable zinc fraction. After contamination, a reduction of total zinc and an increase of labile sorbed zinc over time occurred, concurrent with various changes in soil and biological characteristics. Data analyses on the nematode species revealed different sensitivity levels for several population and community endpoints to zinc exposure. Based on no observed effect concentration (NOEC) values, the most sensitive community-level response was obtained with principal response curve (PRC) analysis, which incorporates all density data in a single analysis. The PRC-based community NOECs were 56, 100, and 100 mg total Zn/kg dry soil after 3, 10, and 22 months of exposure, respectively. Based on 0.01 M CaCl₂-exchangeable zinc, the community response appeared to increase, as NOECs were 4.9, 4.4, and 0.67 mg exchangeable Zn/kg dry weight. Total density was least sensitive, followed by diversity of taxa and the Shannon-Wiener index. NOECs for separate species covered a broad range from sensitive to tolerant species. This range of sensitivities was similar to the one found for other species groups tested in the field plot soil. A comparison was made between benchmark concentrations HC5 and HC50 derived from the general SSD of soil organisms for zinc and the nematode response data. These comparisons roughly confirm the predictions of the SSD model, that is, the community NOEC is in agreement with the benchmark that should protect the soil ecosystem's integrity, and large adverse effects were found at the benchmark derived for setting remediation urgency. Received: 9 November 2000/Accepted: 2 August 2001     

Modeling impact of parathion and its metabolite paraoxon on the nematode Caenorhabditis elegans in soil

Parathion is an insecticide of a group of highly toxic organophosphorous compounds. In vivo, it is activated to the toxic metabolite paraoxon. Laboratory experiments have shown that a single relationship between the variable (concentration x time of application) and the percentage of paralyzed nematodes is relevant. Aqueous (0.01 M CaCl2) extracts from soil that had received a dose of parathion as used in practice during an incubation experiment had no effect on nematodes, because sorption and biodegradation of the pesticide decreased the pesticide concentration in the soluble phase. To predict the toxicological effects of parathion and paraoxon on nematodes under various soil conditions during a simulation period of 20 d, we used a model predicting the concentrations of parathion and paraoxon over time in the soil liquid phase. In this model, sorption and biodegradation of both parathion and paraoxon were taken into account, and the results indicated that sorption effects were dominant and determined the differential toxicological risks between soils. Variable effects were predicted for short times (typically <5 d), and critical toxicological conditions were predicted for longer duration (typically >10-15 d), in all cases.     

Individual and joint toxic effects of pentachlorophenol and bisphenol A on the development of zebrafish (Danio rerio) embryo

Investigation of the toxicological effects of pentachlorophenol (PCP) and bisphenol A (BPA) alone and in combination was carried out following the method of the early life stage (ELS) test on zebrafish embryos. Both chemicals revealed lethal and sub-lethal effects, such as no blood flow, cardiac edema, delayed hatching, and tail malformations. According to their median effective concentrations (EC₅₀ values) in the single exposure, the toxic level of PCP was about two orders of magnitude higher than that of BPA. Result of the joint action modes varied depending on different endpoints. Synergistic action was observed based on the endpoint of 24 h mortality and antagonistic effect displayed based on the endpoint of 72 h cardiac edema. It was also found that the toxicity of PCP would be enhanced with the addition of BPA even below its no observed effect concentration (NOEC) level at the endpoint of 32 h with no blood flow, and the level of the increase was influenced by the toxic unit (TU) ratio.     

Acute and chronic toxicity of 2,3,4,6-tetrachlorophenol and pentachlorophenol to Daphnia and rotifers

The toxicities of 2,3,4,6-tetrachlorophenol (TeCP) and pentachlorophenol (PCP) were determined in standardized, single-species laboratory toxicity tests using daphnids and rotifers. For Daphnia magna, 48-h LC50 estimates with neonates indicated that PCP was more toxic than TeCP. The commercial TeCP formulation DIATOX^®, containing a 6.5:1 ratio of TeCP:PCP, exhibited intermediate toxicity. Mean LC50 estimates were 1.23, 2.66, and 2.07 mg/L, respectively. All three LC50 estimates were significantly different (p<0.05). Daphnia galeata mendotae was more sensitive to TeCP than D. magna displaying a mean 48-h LC50 value of 0.58 mg/L. A 21-d partial life cycle test with D. magna revealed no effects at 0.25 mg/L TeCP, but a significant (p<0.05) increase in adult mortality was noted at 0.50 mg/L TeCP. There was no effect on reproductive success of surviving adults at any test concentration.Rotifer tests, conducted with a commercially available species (Brachionus calyciflorus) and a field collected species (Keratella cochlearis), also indicated that PCP was more acutely toxic than TeCP. LC50 estimates for B. calyciflorus were highly variable depending upon the source of resting eggs (cysts) and the test conditions. LC50 estimates (24-h) ranged from 2.09 to 7.76 mg/L for PCP and from 2.31 to >16.00 mg/L for TeCP. In general, B. calyciflorus was less sensitive to TeCP and PCP than D. magna and D. g. mendotae. Assays with K. cochlearis were only marginally successful, but the mean 12-h LC50 estimate for TeCP of 0.96 mg/L suggested that this species may be more sensitive to TeCP than B. calyciflorus.     

Lethal Body Residues for Pentachlorophenol in Zebra Mussels (Dreissena polymorpha) under Varying Conditions of Temperature and pH

Pentachlorophenol (PCP) toxicity was measured in the zebra mussel under varying conditions of pH (6.5, 7.5, or 8.5) and temperature (10, 17, or 25°C). Toxicity decreased significantly with increasing pH at all temperatures. At a given pH level, toxicity increased significantly with increasing temperature. PCP was most toxic at pH 6.5, 25°C and least toxic at pH 8.5, 10°C. Toxicokinetic parameters were determined at trace PCP concentrations under each combination of pH and temperature. Increasing temperature generally increased the PCP uptake clearance (k_u) although elimination rate constants (k_d) were unaffected. The effect of pH on toxicokinetic parameters was inconsistent but k_u tended to decrease as pH and ionization of PCP increased. Lethal body residues (LR₅₀s), estimated from kinetic parameters determined at trace PCP concentrations and the LC₅₀ values, varied by a factor of 122 as a function of environmental conditions while LC₅₀s varied by a factor of 381. LR₅₀s were also estimated from the measured PCP tissue concentrations and varied by a factor of 8 across conditions. Calculated LR₅₀s were always higher than measured LR₅₀s, determined under identical conditions, by at least a factor of five. However, when LR₅₀ values were recalculated using k_u values measured at the LC₂₅ concentration, the resulting adjusted LR₅₀s varied only by a factor of 2.5 across the range of conditions studied and were more consistent with measured LR₅₀ values. Thus, variance in the PCP concentration required to produce toxicity is reduced when LR₅₀s are used in place of LC₅₀s. Further, the method by which lethal residues (LR₅₀ values) are determined can significantly affect the results and their interpretation.     

The influence of soil properties on the toxicity of molybdenum to three species of soil invertebrates

Mo toxicity to earthworms (Eisenia andrei), Collembola (Folsomia candida) and enchytraeids (Enchytraeus crypticus) was determined in 10 European soils and a standard artificial soil, freshly spiked with Na₂MoO₄, after 28 days exposure. Mo affected survival only in three low pH sandy soils; in all other soils LC50 was >3200 mg Mo/kg dry soil. EC50 values for the reproduction toxicity of Mo were 129-2378 mg/kg for earthworms, 72->3396 mg/kg for Collembola, and 301->2820 mg/kg for enchytraeids. Variation in toxicity among soils could not be explained by differences in available (pore water, water and 0.01 M CaCl₂ extractable) Mo concentrations. Clay content best predicted the EC50 for Mo toxicity to earthworms, while toxicity of Mo for enchytraeids was best described by soil pH. For Collembola no relationships could be derived due to the absence of toxicity in most soils. Soil properties had a strong but species-specific effect on Mo toxicity to soil invertebrates.     

Carbofuran effects in soil nematode communities: using trait and taxonomic based approaches

This work intends to implement the use of native soil nematode communities in ecotoxicological tests using a model pesticide and two geographically nematode communities (Mediterranean and sub-tropical) in order to obtain new perspectives on the evaluation of the toxic potential of chemical substances.The environmental condition of the nematode communities was described using a trait-based approach (grouping the organisms according to their feeding traits) and a traditional taxonomic method (identification to family level). Effects on total nematode abundance, number of families and abundance of nematode feeding groups as well as potential shifts in both trophic and family structure were assessed.Agricultural soils from Curitiba (Brazil) and Coimbra (Portugal) were sampled and the corresponding nematode communities were extracted. Part of the collected soil was defaunated and spiked with four doses of a carbofuran commercial formulation. Afterwards each of the replicates was inoculated with a nematode suspension containing ≈200 or 300 nematodes. After 14 and 28 d of exposure the nematodes were extracted, counted and identified at family level and separately classified according to their feeding traits. The patterns of nematode responses revealed a decrease in the total abundance and a reduction in the number of families. Despite the similar effects observed for both communities, statistically significant toxic effects were only found within the Portuguese community. The total nematode abundance was significantly reduced at the highest carbofuran concentrations and significant shifts in the family structure were detected. However, the trophic structure, i.e., the contribution of each feeding group for the overall community structure, did not significantly change along the contamination gradient. Results showed that using such a trait-based approach may increase the ecological relevance of toxicity data, by establishing communalities in the response to a chemical from two different taxonomic communities, although with potential loss of information on biodiversity of the communities.     

Responses of serum transaminases in Notopterus notopterus chronically exposed to phenolic compounds and their combinations

Specimens of Notopterus notopterus were exposed to sublethal concentrations (10, 15, and 20 times diluted 96-hr LC₅₀) of phenol (P), dinitrophenol (DNP), pentachlorophenol (PCP), and their three combinations—additive [(DNP + P)/PCP], antagonistic [(PCP + DNP)/P], and synergistic [(P + DNP)/PCP]—for 30 days and their effects on serum transaminases (SGOT and SGPT) were studied. Significant (P < 0.05, P < 0.01, P < 0.001) enzymatic alterations were observed after treatment. The maximum stimulation (127.76%) was observed in SGOT at the highest concentration of the (P + DNP)/PCP combination and the minimum stimulation (6.67%) was observed in SGPT at the 20 times dilution concentration of the (PCP + DNP)/P combination after 30 days. However, inhibition in enzyme activity at the lowest concentration of P, DNP, and the (PCP + DNP)/P combination was also occasionally observed.     

Benzo(a)pyrene shows low toxicity to three species of terrestrial plants, two soil invertebrates, and soil-nitrifying bacteria

We examined the toxicity of benzo(a)pyrene (BaP) to several standard test organisms including the seed emergence and early life-stage growth of three terrestrial plants (Trifolium pratense, Lolium perenne, and Brassica alba), the survival and reproduction of enchytraeids (Enchytraeus crypticus), and the nitrifying ability of soil bacteria. To also have a look at possible food-chain effects, we included a two-species reproduction test with predatory mites (Hypoaspis aculeifer) and collembolan (Folsomia fimetaria) prey. No effect or only weak effects even at very high BaP concentrations were observed for all tests. None of the soil invertebrates were affected within the concentration range tested (up to 947 mg kg⁻¹). For soil-nitrifying bacteria, significant effects were recorded at 977 mg kg⁻¹, leaving a no observable effect concentration (NOEC) of 293 mg kg⁻¹. BaP did not affect seed emergence for any of the plants, but the growth of B. alba was significantly reduced at the highest concentration tested (375 mg kg⁻¹), leaving a NOEC of 69 mg kg⁻¹. Compared to a number of other polycyclic aromatic compounds previously tested in the same soil type, BaP is generally less toxic.     

Application of Plant and Earthworm Bioassays to Evaluate Remediation of a Lead-Contaminated Soil

Earthworm acute toxicity, plant seed germination/root elongation (SG/RE) and plant genotoxicity bioassays were employed to evaluate the remediation of a lead-contaminated soil. The remediation involved removal of heavy metals by a soil washing/soil leaching treatment process. A portion of the soil after remediation was rinsed with water in order to simulate exposure to rainfall. The bioassay results showed that the soils before treatment (BT) and after treatment plus water rinsing (RT) were not toxic to earthworms in a 14-day exposure, while after treatment (AT) showed significant toxicity. The LC50 values for Eisenia fetida and Lumbricus terrestris were 44.04 and 28.83 (as % AT soil supplemented in artificial soil), respectively. The phytotoxicity data indicated that all three test soils significantly inhibited lettuce SG/RE in a dose-related manner, with AT being the most phytotoxic. In oats, RT had no effect on SG/RE and AT was more toxic than BT. For the two local site grass seeds tested (blue grama and sideoat grama), the AT soil was the most phytotoxic followed by BT and RT. In Allium cepa (common onion), BT and AT induced similar levels of genetic damage to root tip cells, whereas RT was not genotoxic. High salt levels generated during the remediation process appeared to be responsible for the increased toxicity of AT soil for both plants and earthworms. The rinsing of the AT soil with water effectively removed both acutely toxic and genotoxic components of the soil. Received: 20 May 1996/Revised: 7 August 1996     

Sorption of the neutral and charged forms of pentachlorophenol on soil: evidence for different mechanisms

Laboratory soil sorption experiments have been conducted on pentachlorophenol (PCP) at different pH values in an attempt to elucidate differences in sorption mechanisms between the charged and neutral species. Sorption of PCP on soil was investigated by maintaining pH 4 or 8 in batch sorption experiments. Pre-equilibration of the soil was necessary to maintain a constant pH over the course of the experiments. Additionally, a CaCO₃-CO₂/N₂ buffered solution was necessary to maintain a pH of 8. Sorption of the neutral PCP species conformed to a linear isotherm model, while a Langmuir model provided the best fit for the charged species. Desorption of the neutral form was completely reversible over the sorption times studied but the charged species exhibited some resistance to desorption. Temperature effects on the distribution coefficients (K_d) were investigated and thermodynamic parameters were calculated. The ionized species showed a clear decrease in K_d with increasing temperature while the protonated species showed no apparent trend. Enthalpies (ΔH°), entropies (ΔS°), and free energies (ΔG°), support the conclusions that the neutral form of PCP partitions by hydrophobically binding to the soil while the charged form sorbs by a more specific exothermic adsorption reaction. Received: 30 May 2000/Accepted: 20 November 2000     

Toxicological assessment of biodegraded pentachlorophenol: Microtox® and fish embryos

A Gram-negative bacterium,Pseudomonas sp. strain SR3, was isolated from soil at a former wood treatment plant in north central Florida. The ability of this bacterium to degrade pentachlorophenol (PCP) was confirmed by growing cells in a basal salts medium in which PCP was the only source of carbon and energy. Degradation from a measured concentration of 39–40 g PCP/ml to 0.0006 g PCP/ml was observed within 120 h of incubation in the presence of PCP-induced cells ofPseudomonas sp. strain SR3. The initial cell density in these cultures was 6 x 10⁶ cfu/ml. Microtox^® 5 min EC50 toxicity tests revealed that aqueous solutions of PCP, measured concentrations 39–40 p /ml were toxic but that final biodegraded samples, 0.0006 /g PCP/ml were nontoxic. However, bioassays with embryonic inland silversides,Menidia beryllina, showed that the biodegraded samples were embryotoxic or teratogenic. Water containing added PCP at concentrations up to 30 times higher than measured in the final biodegraded samples was less toxic/teratogenic. These results indicate that while biodegradation of PCP was nearly complete, intermediate metabolites of the degradation process or undegraded impurities in PCP were toxic or teratogenic. Thus, theM. beryllina bioassay allows extremely sensitive assessment of toxicity associated with biodegraded environmental pollutants and may be a useful criterion for determining whether bioremediated water or soil is safe for discharge back into the environment.Contribution No. 777 of the Gulf Breeze Environmental Research Laboratory.     

Tungsten effects on survival, growth, and reproduction in the earthworm, Eisenia fetida

To provide basic toxicity data for formulating risk characterization benchmarks, the effects of tungsten on survival, growth, and reproduction were investigated in the earthworm Eisenia fetida. Parallel studies with lead as a reference toxicant also were conducted. Although sodium tungstate (Na2WO4) was less acutely toxic than lead nitrate (Pb(NO3)2) in 14-d spiked field soil acute toxicity assays (lethal concentrations for 50% of organisms: W, 6,250 mg/kg; Pb, 2,490 mg/kg), tungstate completely inhibited reproduction in 28- and 56-d assays at all tested tungsten concentrations (> or = 704 mg/kg). By comparison, cocoon production was not significantly reduced for lead concentrations until concentrations reached 766 mg/kg, and cocoon production was still observed at the highest concentration tested (1,650 mg/kg). These data indicate that tungsten is a reproductive toxicant for earthworms and that, by comparison, its sublethal toxicity is greater than that of lead. Toxicity data for other soil invertebrate species are required to fully establish benchmark levels/ecological soil screening levels for tungsten.     

Toxicity and Bioaccumulation of Copper to Black Bindweed (Fallopia convolvulus) in Relation to Bioavailability and the Age of Soil Contamination

The use of ecotoxicity test results obtained in the laboratory for prediction of effects of potentially toxic concentrations of chemicals in the field is hampered by several factors differing between the laboratory and the field situations. One important factor is the binding of test chemicals to soil, which is affected by the age of the contamination and soil type. The present study investigated the effect of contamination age by introducing an aging period of 1 to 12 weeks between mixing the test substance, copper sulfate, in with soil and introducing the test plant, Fallopia convolvulus (L.) A. Löve. Copper accumulation, emergence of cotyledons, and growth were assessed and related to total (boiling with HNO₃) and 0.01 M CaCl₂- and DTPA-extractable soil copper concentrations. Aging of the copper-contaminated soil had only small effects on bioaccumulation of copper, copper toxicity, and extractable soil copper fractions. Soil copper had no effect on emergence of cotyledons. Estimated EC₅₀ values for shoot and root growth averaged 280 mg Cu/kg. Effects on growth in these laboratory-treated soils were much more severe than in a study performed in soil from an old copper-contaminated field site. Neither CaCl₂- nor DTPA-extractable copper fractions could explain all of the differences in effects between aged spiked soil and field soil. The accumulation pattern for roots and shoots of F. convolvulus indicated that excessive copper was accumulated and adsorbed mainly by the fine roots, whereby the copper concentrations of other plant parts were kept low until the plant was no longer able to maintain this regulation. An internal threshold for effects on growth of about 20 mg Cu/kg shoot dry weight was estimated, coinciding with a soil copper concentration of approximately 200 mg/kg.     

Acute toxicity of pure pentachlorophenol and a technical formulation to three species ofDaphnia

The acute toxicity of a technical formulation of pentachlorophenol (PCP) and pure pentachlorophenol to three age classes ofDaphnia magna, and adultD. pulex andD. galeata mendotae was determined by static toxicity tests. The influence of a number of factors on toxicity of PCP was also examined. The 48-hr LC50 estimates for adult daphnids of the three species exposed to pure PCP were 1.78, 4.59 and 0.51 mg/L, respectively, while those for the technical formulation were 2.57, 3.66 and 0.33 mg/L, respectively. There was little difference in toxicity between the technical and pure PCP; however, toxicity of both forms of PCP was influenced by duration of exposure, age (and/or size) and species of test organism and pH of the test solution.Pentachlorophenol caused a toxic response over a very narrow range of concentrations, with the greatest response occurring immediately between 0 and 24 hr. Pure PCP was equally toxic to all age classes ofD. magna but susceptibility to technical PCP decreased with maturation.D. g. mendotae was ten times more sensitive thanD. pulex to PCP. Pure PCP was significantly more toxic toD. magna at pH 5.5 than 7.0 with mean 48-hr LC50 values of 0.082 and 1.78 mg PCP/L, respectively.At 12°C, the toxicity of both forms of PCP toD. g. mendotae andD. pulex did not differ significantly from that at 20°C; however, technical PCP was significantly more toxic toD. magna at 12°C for an exposure duration of 48 hr. There was no effect of test container size (100, 250, 600 and 1,000 mL) on the toxicity of PCP toD. magna at 20°C with the lower pH of 5.5, suggesting that adsorption to glassware was not a factor in availability of PCP to test organisms. Beaker size had no effect on the toxicity of PCP toD. pulex at 20°C with test solutions having a pH of 7.0-8.0.