Toxicity Identification Evaluation Procedures Research Articles

Using the Sediment Quality Triad to characterize baseline conditions in the Anacostia River, Washington, DC, USA

The Sediment Quality Triad (SQT) consists of complementary measures of sediment chemistry, benthic community structure, and sediment toxicity. We applied the SQT at 20 stations in the tidal portion of the Anacostia River from Bladensburg, MD to Washington, DC to establish a baseline of conditions to evaluate the effects of management actions. Sediment toxicity was assessed using 10-day survival and growth tests with the freshwater amphipod, Hyalella azteca and the midge, Chironomus dilutus. Triplicate grabs were taken at each station for benthic community analysis and the Benthic Index of Biotic Integrity (B-IBI) was used to interpret the data. Only one station, #92, exhibited toxicity related to sediment contamination. Sediments from this station significantly inhibited growth of both test species, had the highest concentrations of contaminants, and had a degraded benthic community, indicated by a B-IBI of less than 3. Additional sediment from this station was tested and sediment toxicity identification evaluation (TIE) procedures tentatively characterized organic compounds as the cause of toxicity. Overall, forty percent of the stations were classified as degraded by the B-IBI. However, qualitative and quantitative comparisons with sediment quality benchmarks indicated no clear relationship between benthic community health and contaminant concentrations. This study provides a baseline for assessing the effectiveness of management actions in the Anacostia River.

Development of a Toxicity Identification Evaluation Protocol Using Chlorophyll-a Fluorescence in a Marine Microalga

Growth inhibition bioassays with the microalga Nitzschia closterium have recently been applied in marine Toxicity Identification Evaluation (TIE) testing. However, the 48-h test duration can result in substantial loss of toxicants over time, which might lead to an underestimation of the sample toxicity. Although shorter-term microalgal bioassays can minimize such losses, there are few bioassays available and none are adapted for marine TIE testing. The acute (5-min) chlorophyll-a fluorescence bioassay is one alternative; however, this bioassay was developed for detecting herbicides in freshwater aquatic systems and its suitability for marine TIE testing was not known. In this study, a chlorophyll-a fluorescence bioassay using the marine microalga Isochrysis galbana was able to detect contaminants other than herbicides at environmentally relevant concentrations and tolerated the physical and chemical manipulations needed for a Phase I TIE. Phase I TIE procedures were successfully developed using this chlorophyll-a fluorescence bioassay and used to identify all classes of contaminants present in a synthetic mixture of known chemical composition. In addition, TIEs with both the acute fluorescence bioassay and the standard growth inhibition bioassay identified the same classes of toxicants in a sample of an unknown complex effluent. Even though the acute chlorophyll-a fluorescence end point was less sensitive than the chronic cell division end point, TIEs with the chlorophyll-a fluorescence bioassay provided a rapid and attractive alternative to longer-duration bioassays.

TOXICITY AND METAL SPECIATION IN ACID MINE DRAINAGE TREATED BY PASSIVE BIOREACTORS

Sulfate-reducing passive bioreactors treat acid mine drainage (AMD) by increasing its pH and alkalinity and by removing metals as metal sulfide precipitates. In addition to discharge limits based on physicochemical parameters, however, treated effluent is required to be nontoxic. Acute and sublethal toxicity was assessed for effluent from 3.5-L column bioreactors filled with mixtures of natural organic carbon sources and operated at different hydraulic retention times (HRTs) for the treatment of a highly contaminated AMD. Effluent was first tested for acute (Daphnia magna and Oncorhynchus mykiss) and sublethal (Pseudokirchneriella subcapitata, Ceriodaphnia dubia, and Lemna minor) toxicity. Acute toxicity was observed for D. magna, and a toxicity identification evaluation (TIE) procedure was then performed to identify potential toxicants. Finally, metal speciation in the effluent was determined using ultrafiltration and geochemical modeling for the interpretation of the toxicity results. The 10-d HRT effluent was nonacutely lethal for O. mykiss but acutely lethal for D. magna. The toxicity to D. magna, however, was removed by 2 h of aeration, and the TIE procedure suggested iron as a cause of toxicity. Sublethal toxicity of the 10-d HRT effluent was observed for all test species, but it was reduced compared to the raw AMD and to a 7.3-d HRT effluent. Data regarding metal speciation indicated instability of both effluents during aeration and were consistent with the toxicity being caused by iron. Column bioreactors in operation for more than nine months efficiently improved the physicochemical quality of highly contaminated AMD at different HRTs. The present study, however, indicated that design of passive treatment should include sufficient HRT and posttreatment aeration to meet acute toxicity requirements.

A SCREENING METHOD FOR TOXICITY IDENTIFICATION EVALUATION ON AN INDUSTRIAL EFFLUENT USING CHELEX®-100 RESIN AND CHELATORS FOR SPECIFIC METALS

Toxicity of an industrial effluent was characterized using a toxicity identification method established at our research institute. Toxicity was evaluated using the Ceriodaphnia dubia survival test. The industrial effluent chosen in the present study had toxic effects on C. dubia before treatment. The effluent was first filtered to remove suspended solids (F-treatment). Activated charcoal was then added to the effluent to adsorb organic substances (AC-treatment), followed by treatment of the supernatant with Chelex-100 resin to remove di- and trivalent cations, including transition metals (C100-treatment). The effluent exhibited a toxic effect on C. dubia even after the AC-treatment, but it was no longer toxic after the C100-treatment, indicating that the effluent contained cations that affect C. dubia survival. To further identify the metals responsible for the toxicity, benzoin-alpha-oxime or dimethylglyoxime was added to the effluent to chelate specific cations. The toxicity of the effluent was eliminated by addition of dimethylglyoxime but not significantly so by addition of benzoin-alpha-oxime, suggesting that the toxicants in the effluent strongly formed complexes with dimethylglyoxime. These results combined with those of metal analysis strongly suggest that nickel might be the noxious agent. The toxicity identification method described in this paper is effective for the identification of metal toxicants in industrial effluents. The method using insoluble chelating resins and chelators for specific metals would serve as a useful addition to the standard toxicity identification evaluation procedure.

Whole‐sediment toxicity identification evaluation tools for pyrethroid insecticides: II. Esterase addition

An esterase enzyme capable of catalyzing the hydrolysis of the ester bond common to all pyrethroid insecticides has been proposed as a toxicity identification evaluation procedure for the compounds when present in water samples. It appeared to show promise in previous applications to pore water and in one published application to bulk sediment. The present study was designed to provide the additional validation of the technique when applied to whole sediment, demonstrating its efficacy and specificity to pyrethroids. When added to the overlying water in a standard sediment toxicity test with the amphipod Hyalella azteca, the enzyme reduced toxicity of pyrethroid-spiked laboratory sediments. The technique had no effect on toxicity of DDT or chlorpyrifos, but it dramatically reduced the toxicity of cadmium, presumably by complexation of the cadmium with dissolved organic matter rather than by enzymatic activity. Esterase addition consistently reduced pyrethroid-related toxicity in 12 tests with field-collected sediments. The procedure, however, also results in substantial changes in water quality, provides an undesirable nutritional benefit to the test organisms exposed to the esterase, and achieves a substantial fraction of the protective capability simply by complexation of the pyrethroids with the esterase or the microbial growth that the esterase promotes. The esterase addition technique has potential for whole-sediment application, but further development of procedures and confirmation by independent lines of evidence are needed to reduce the potential for misinterpretation of results.

TOXICITY AND METAL SPECIATION IN ACID MINE DRAINAGE TREATED BY PASSIVE BIOREACTORS

Sulfate-reducing passive bioreactors treat acid mine drainage (AMD) by increasing its pH and alkalinity and by removing metals as metal sulfide precipitates. In addition to discharge limits based on physicochemical parameters, however, treated effluent is required to be nontoxic. Acute and sublethal toxicity was assessed for effluent from 3.5-L column bioreactors filled with mixtures of natural organic carbon sources and operated at different hydraulic retention times (HRTs) for the treatment of a highly contaminated AMD. Effluent was first tested for acute (Daphnia magna and Oncorhynchus mykiss) and sublethal (Pseudokirchneriella subcapitata, Ceriodaphnia dubia, and Lemna minor) toxicity. Acute toxicity was observed for D. magna, and a toxicity identification evaluation (TIE) procedure was then performed to identify potential toxicants. Finally, metal speciation in the effluent was determined using ultrafiltration and geochemical modeling for the interpretation of the toxicity results. The 10-d HRT effluent was nonacutely lethal for O. mykiss but acutely lethal for D. magna. The toxicity to D. magna, however, was removed by 2 h of aeration, and the TIE procedure suggested iron as a cause of toxicity. Sublethal toxicity of the 10-d HRT effluent was observed for all test species, but it was reduced compared to the raw AMD and to a 7.3-d HRT effluent. Data regarding metal speciation indicated instability of both effluents during aeration and were consistent with the toxicity being caused by iron. Column bioreactors in operation for more than nine months efficiently improved the physicochemical quality of highly contaminated AMD at different HRTs. The present study, however, indicated that design of passive treatment should include sufficient HRT and posttreatment aeration to meet acute toxicity requirements.

Identification of chlorfenvinphos toxicity in a municipal effluent in Sydney, New South Wales, Australia

Acute toxicity in a municipal sewage treatment plant in Sydney, New South Wales, Australia, was traced to chlorfenvinphos, an organophosphorous pesticide. Toxicity identification evaluation procedures led to the tentative identification of chlorfenvinphos as the toxic contaminant in the sample. Subsequent analytical verification revealed 0.95 microg/L of chlorfenvinphos in the effluent sample, and spiking studies confirmed that it accounted for the observed toxicity. The 48-h median lethal concentration of chlorfenvinphos to Ceriodaphnia dubia averaged 0.28 microg/L (n = 4). Source-control measures were effective at eliminating chlorfenvinphos and associated toxicity from the discharge.

Toxicity evaluation by using intact sediments and sediment extracts

The toxicity of intact sediments and sediment extracts, from both an uncontaminated site and a site contaminated by pulp-mill effluents, was tested in a five months study. The deposit-feeding amphipod Monoporeia affinis was exposed in soft-bottom flow-through water microcosms. To examine potential toxicity a set of reproduction endpoints was used including fecundity and different embryo aberrations such as malformed eggs. Among extracts, the aliphatic/monoaromatic and diaromatic fractions along with the total extract were shown to cause the highest toxicity measured as malformed eggs, while the polyaromatic fraction caused toxicity at background levels. A comparison between sediment extracts and pulp mill contaminated intact sediment, however, showed no toxicity of the intact sediment. Thus, the extraction procedure seems to increase bioavailability and subsequently toxicity as compared to the intact sediments in situ. In toxicity testing using fractionated extracts of sediments in a toxicity identification evaluation (TIE) procedures, caution should therefore be taken when assessing bioavailable contaminants in contaminated areas. This should be taken in account both in determining remediation priorities as well as in ecological risk assessments.

Development of toxicity identification evaluation procedures for pyrethroid detection using esterase activity

Recent agrochemical usage patterns suggest that the use of organophosphate (OP) pesticides will decrease, resulting in a concomitant increase in pyrethroid usage. Pyrethroids are known for their potential toxicity to aquatic invertebrates and many fish species. Current toxicity identification evaluation (TIE) techniques are able to detect OPs, but have not been optimized for pyrethroids. Organophosphate identification methods depend upon the use of piperonyl butoxide (PBO) to identify OP-induced toxicity. However, the use of PBO in TIE assays will be confounded by the co-occurrence of OPs and pyrethroids in receiving waters. It is necessary, therefore, to develop new TIE procedures for pyrethroids. This study evaluated the use of a pyrethroid-specific antibody, PBO, and carboxylesterase activity to identify pyrethroid toxicity in aquatic toxicity testing with Ceriodaphnia dubia. The antibody caused significant mortality to the C. dubia. Piperonyl butoxide synergized pyrethroid-associated toxicity, but this effect may be difficult to interpret in the presence of OPs and pyrethroids. Carboxylesterase activity removed pyrethroid-associated toxicity in a dose-dependent manner and did not compromise OP toxicity, suggesting that carboxylesterase treatment will not interfere with TIE OP detection methods. These results indicate that the addition of carboxylesterase to TIE procedures can be used to detect pyrethroids in aquatic samples.

Temporal pattern of toxicity in runoff from the Tijuana River Watershed

Samples were collected from the Tijuana River under both dry weather (baseflow) conditions and during wet weather, and tested for toxicity using Ceriodaphnia dubia tests. Toxicity of waters in the Tijuana River was generally low under baseflow conditions, but increased markedly during high flow runoff events. In order to determine the temporal pattern of toxicity during individual rain events, sequential grab samples were collected using an autosampler at 5–7 h intervals after the start of the rain event, and tested for acute toxicity. In all cases, peak toxicity values (ranging from 2.8 to 5.8 TU) for each storm occurred within the first 1–2 h of initiation of the rain event, and were statistically higher (using the 95% CL) for each of the pre-storm base flow values. However, there was no statistically significant correlation ( p<0.05) between flow rate and toxicity when all storm data was pooled. Additionally, we used toxicity identification evaluation (TIE) procedures to attempt to identify the classes of chemicals that account for this early storm toxicity. Solid phase extraction was the only treatment that showed consistent and significant ( P<0.05) removal of toxicity. These TIEs, conducted on the most toxic sample of the river's flow during runoff events, suggest that non-polar organics may be responsible for such toxicity. The temporal pattern of toxicity, both during a given storm event and seasonally, indicates that wash-off from the watershed by rainfall may deplete the supply of toxicity available for wash-off in subsequent events, so that a clearly consistent relationship between flow and toxicity was not evident.

Application of toxicity identification evaluation procedures on wastewaters and sludge from a municipal sewage treatment works with industrial inputs

A comparison of influent and effluent wastewater toxicities to Daphnia magna was undertaken to determine the most representative aqueous fraction for future toxicity identification evaluation (TIE) studies. Wastewater samples originated from a wastewater treatment plant which receives mainly domestic wastewater and industrial effluents. The TIE results reveal that the key toxicant contained in the influent was 2-propylbezaldehyde oxime. Results suggest that greater attention needs to be given to the receiving influent containing 2-propylbezaldehyde oxime to avoid damaging the bacterial populations in the primary activated sludge process. In addition, in order to evaluate the applicability of the reuse of activated sludge as a fertilizer in agriculture, TIE studies were also performed on water extracts from surplus sludge.

Toxicity characterisation of sediment porewaters collected from UK estuaries using a Tisbe battagliai bioassay

The lethal toxicity of sediment porewater isolated from samples collected from six United Kingdom estuaries was tested using a Tisbe battagliai bioassay. A selection of Phase 1 toxicity identification evaluation (TIE) procedures was then used to characterise the substances responsible for the measured toxicity. In samples collected in 2000, cationic metals, ammonia, organic compounds were identified as the cause of toxicity in the most toxic sample collected. Some of the toxicity remained uncharacterised. It was shown that the toxicity in samples collected from the same locations in 2001 was due to ammonia. The successful application of the T. battagliai bioassay demonstrates that it is a suitable assay for marine sediment porewater TIE and that combined with simple characterisation procedures allow an initial assessment to be made on the type of substances responsible for the measured toxicity.

Toxicity identification evaluation of leachates from municipal solid waste landfills: a multispecies approach

The toxicity of leachates from two municipal solid waste (MSW) landfills in Southern Italy was characterized using a toxicity identification evaluation procedure. The chemical and physical fractionation techniques were: pH adjustment, pH adjustment/filtration, pH adjustment/C18 solid phase extraction, graduated pH and EDTA chelation. All the samples exhibited acute toxicity towards the bacterium Vibrio fischeri, the freshwater rotifer Brachionus calyciflorus and the freshwater crustaceans Thamnocephalus platyurus and Daphnia magna. Statistical techniques were used to determine the discriminatory power and the toxicity detection capacity of the different assays and to choose a minimal battery of tests for the toxicity identification of leachates. Toxicity was closely associated with pH, generally increasing at higher pH levels and decliming at lower ones. Furthermore, results showed that toxicants could be characterized as cations, basic chemicals, suspended solids and apolar compounds.

Application of toxicity identification evaluation procedures to an effluent from a nitrogenous fertilizer plant in China

The integrated method combining chemistry and toxicology, toxicity identification evaluation (TIE), was conducted to identify key toxicants in an effluent from a nitrogen fertilizer plant in China. Toxicity characterization, phase I of TIE, revealed that the suspected toxicant in the effluent was an anion that could be changed into a volatile acid. The results of toxicity identification and confirmation procedures indicated potassium cyanide to be the primary toxicant in the effluent.

Tannery wastewater characterization and toxicity effects on Daphnia spp.

Tannery wastewater contains large quantities of organic and inorganic compounds, including toxic substances such as sulfides and chromium salts. The evaluation of wastewater quality in Chile nowadays is based on chemical specific measurements and toxicity tests. The goal of this research was to characterize tannery wastewater and to relate its physical/chemical parameters with its acute toxicity effect on Daphnia pulex. To distinguish the most important toxic compounds, physical/chemical techniques were applied to a grab sample of a final effluent based on the Phase I toxicity identification evaluation (TIE) procedure. In addition, the toxicity of a beamhouse effluent after an activated sludge reactor treatment was investigated on Daphnia magna (introduced species) and D. pulex (native species). Effluent from different tannery processes (soaking, beamhouse, tanning and final) demonstrated high values of chemical organic demand (COD; 2840-27,600 mg L(-1)), chloride (1813-16,500 mg L(-1)), sulfate (230-35,200 mg L(-1)), and total solids (8600-87,100 mg L(-1)). All effluents showed extremely toxic effects on D. pulex, with 24-h mean lethal values (LC(50)) ranging from 0.36% to 3.61%. The Phase I TIE profile showed that toxicity was significantly reduced by air stripping, filtration, and a cationic exchange resin, with toxicity reductions ranging between 46% and 76%. The aerobically treated beamhouse effluent showed significantly less toxicity for both species (43%-74%). The chemical parameters demonstrated that the remaining toxicity of the treated beamhouse effluent was associated with its ammonia (120 mg N-NH(3) L(-1)) and chloride (11,300 mg Cl(-) L(-1)) contents.

Bioassay-directed chemical analysis of River Elbe surface water including large volume extractions and high performance fractionation

A bioassay-directed fractionation and identification (toxicity identification evaluation procedure) was performed on extracts of 10 l River Elbe water samples. The experimental method included a SDB-1 solid phase extraction followed by RP-HPLC fractionation and subfractionation. Chemical analysis by GC–MS as well as acute toxicity testing using a luminescent bacteria assay were conducted in the respective fractions. Many substances were identified, among which were pesticides and pharmaceuticals, but many compounds remained unknown.

Characterization of estrogenic compounds in water samples collected from United Kingdom estuaries

AbstractThis report describes the identification of important estrogenic compounds in surface and sediment pore‐water samples collected from the Tyne and Tees estuaries (United Kingdom) through the application of toxicity identification evaluation (TIE) procedures. The Tyne and Tees estuaries represent estuaries that have been historically impacted by industrial activities and continue to receive treated domestic sewage and industrial effluent. In 1998, Dabholm Gut on the Tees received a mixture of treated and untreated effluent, while Howdon sewage treatment works (STW) discharged primary treated effluents. An estrogenically active water sample collected from Howdon STW on the Tyne was shown to contain 17β‐estradiol, androsterone, and an unknown estrogenic compound(s). Most of the activity contained in a sample collected from the Dabholm Gut combined discharges on the Tees was also due to 17β‐estradiol with additional activity from nonylphenol and (tentatively) bis(2‐ethylhexyl)phthalate. The only sediment pore‐water sample to demonstrate estrogenic activity was collected from Dabholm Gut.

Toxicity characterisation of organic contaminants in stormwaters from an agricultural headwater stream in South East England

The transient movement of pesticides at biologically active concentrations during storm events is considered to be a cause of biological impoverishment in some headwater streams. The programme of work described sought to identify compounds that are the cause of toxic effects during such events. Along with targeted pesticide analysis, toxicity identification evaluation (TIE) procedures were used to identify compounds with a demonstrated toxic effect. These procedures were specifically directed towards isolating and attributing toxicity to classes of organic contaminants in samples collected from an English headwater stream during a storm event. The organic load was isolated by means of solid-phase extraction (SPE). Bioassay of the SPE extract at ×100 whole water concentrations confirmed that the samples contained substances toxic to Daphnia magna, although the raw samples were not toxic. Targeted pesticide analysis identified simazine and diuron as the major pesticides present and, using a toxicity unit (TU) approach, were shown to be responsible for a significant amount of the observed concentrate toxicity during a runoff event. However, they were not present in sufficient quantities to be totally responsible for a more toxic later event. By simplification of the SPE isolate using reverse-phase HPLC, fractions from which were tested for toxicity, the cause of concentrate toxicity in the later event was isolated to two discrete fractions. GC–MS analysis of these fractions identified nonylphenol, endosulfan sulphate and pendimethalin as present, with the majority of toxicity attributed to nonylphenol (NP). The main advantage of the TIE approach is that it allows biological active compounds with a demonstrated effect to be identified that may not be selected by more traditional techniques.

Characterization of estrogenic compounds in water samples collected from United Kingdom estuaries.

This report describes the identification of important estrogenic compounds in surface and sediment pore-water samples collected from the Tyne and Tees estuaries (United Kingdom) through the application of toxicity identification evaluation (TIE) procedures. The Tyne and Tees estuaries represent estuaries that have been historically impacted by industrial activities and continue to receive treated domestic sewage and industrial effluent. In 1998, Dabholm Gut on the Tees received a mixture of treated and untreated effluent, while Howdon sewage treatment works (STW) discharged primary treated effluents. An estrogenically active water sample collected from Howdon STW on the Tyne was shown to contain 17beta-estradiol, androsterone, and an unknown estrogenic compound(s). Most of the activity contained in a sample collected from the Dabholm Gut combined discharges on the Tees was also due to 17beta-estradiol with additional activity from nonylphenol and (tentatively) bis(2-ethylhexyl)phthalate. The only sediment pore-water sample to demonstrate estrogenic activity was collected from Dabholm Gut.

Current approaches and future directions for contaminant-related impact assessments in coastal environments: Brazilian perspective

Coastal ecosystems are subject to the discharge of contaminants via sewage, industrial effluents, storm water runoff, dredged material and accidental chemical spills. The majority of contaminants reaching the ocean tend to be adsorbed to particulate matter and eventually settle on the ocean floor, where they can deleteriously affect the sediment-associated community. The degree to which a receiving body is impacted is usually assessed by the analysis of the sediments from the area of concern. If chemical data generated by monitoring programs are available, they can be compared to Sediment Quality Guidelines to help identify a potential problem. A more cost-effective approach involves the use of a sensitive toxicity test to identify areas of concern, with comprehensive chemical analyses conducted at a later stage and focusing on the toxic sites identified in the initial screening survey. More complex studies involve the assessment of the benthic communities, in addition to toxicity tests and chemical analyses can be conducted. While methodologies for chemical and benthic community assessments are relatively well established, several approaches can be used to determine sediment toxicity, including acute or chronic tests, conducted in the laboratory or in situ, on the whole sediment or the liquid phase. Several factors should be considered when interpreting toxicity test results, such as the enhancement of toxicity by the photo-oxidation of organic chemicals by ultraviolet light (UV), and confounding factors such as salinity, ammonia, sulfides, particle size distribution, organic matter content, and acid volatile sulfides (AVS). Once an impact has been identified, toxicity identification evaluation (TIE) procedures can be applied to help establish the chemical or classes of chemicals responsible for the observed adverse effects. The international and Brazilian scientific literatures are reviewed to illustrate these approaches.