ASTM E1706 - 19

    Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates

    Active Standard ASTM E1706 | Developed by Subcommittee: E50.47

    Book of Standards Volume: 11.06

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    Significance and Use

    5.1 General: 

    5.1.1 Sediment provides habitat for many aquatic organisms and is a major repository for many of the more persistent chemicals that are introduced into surface waters. In the aquatic environment, most anthropogenic chemicals and waste materials including toxic organic and inorganic chemicals eventually accumulate in sediment. Mounting evidences exists of environmental degradation in areas where USEPA Water Quality Criteria (WQC; (65)) are not exceeded, yet organisms in or near sediments are adversely affected (66) . The WQC were developed to protect organisms in the water column and were not directed toward protecting organisms in sediment. Concentrations of contaminants in sediment may be several orders of magnitude higher than in the overlying water; however, bulk sediment concentrations have not been strongly correlated to bioavailability (67). Partitioning or sorption of a compound between water and sediment may depend on many factors including: aqueous solubility, pH, redox, affinity for sediment organic carbon and dissolved organic carbon, grain size of the sediment, sediment mineral constituents (oxides of iron, manganese, and aluminum), and the quantity of acid volatile sulfides in sediment (40, 41). Although certain chemicals are highly sorbed to sediment, these compounds may still be available to the biota. Chemicals in sediments may be directly toxic to aquatic life or can be a source of chemicals for bioaccumulation in the food chain.

    5.1.2 The objective of a sediment test is to determine whether chemicals in sediment are harmful to or are bioaccumulated by benthic organisms. The tests can be used to measure interactive toxic effects of complex chemical mixtures in sediment. Furthermore, knowledge of specific pathways of interactions among sediments and test organisms is not necessary to conduct the tests (68). Sediment tests can be used to: (1) determine the relationship between toxic effects and bioavailability, (2) investigate interactions among chemicals, (3) compare the sensitivities of different organisms, (4) determine spatial and temporal distribution of contamination, (5) evaluate hazards of dredged material, (6) measure toxicity as part of product licensing or safety testing, (7) rank areas for clean up, and (8) estimate the effectiveness of remediation or management practices.

    5.1.3 A variety of methods have been developed for assessing the toxicity of chemicals in sediments using amphipods, midges, polychaetes, oligochaetes, mayflies, or cladocerans (Section 13 and 14; Annex A1 to Annex A5; (2), (4), (69), (70). Several endpoints are suggested in these methods to measure potential effects of contaminants in sediment including survival, growth, behavior, or reproduction; however, survival of test organisms in 10-day exposures is the endpoint most commonly reported. These short-term exposures which only measure effects on survival can be used to identify high levels of contamination in sediments, but may not be able to identify moderate levels of contamination in sediments (USEPA (2); Sibley et al., (54); Sibley et al., (55); Sibley et al., (71); Benoit et al., (72); Ingersoll et al., (56)). Sublethal endpoints in sediment tests might also prove to be better estimates of responses of benthic communities to contaminants in the field (18). The previous version of this standard (Test Method E1706-95b) described 10-day toxicity tests with the amphipod Hyalella azteca and midge Chironomus dilutus (formerly known as C. tentans; Shobanov et al. 1999.(1), (see Section 13 and 14). This version of the standard now outlines approaches for evaluating sublethal endpoints in longer-term sediment exposures with these two species (Annex A6 and Annex A7). The decision to conduct short-term or long-term toxicity tests depends on the goal of the assessment. In some instances, sufficient information may be gained by measuring sublethal endpoints in 10-day tests. In other instances, the 10-day tests could be used to screen samples for toxicity before long-term tests are conducted. While the long-term tests are needed to determine direct effects on reproduction, measurement of growth in these toxicity tests may serve as an indirect estimate of reproductive effects of contaminants associated with sediments (A6.4.5 and A7.4.6.2). Additional studies are ongoing to more thoroughly evaluate the relative sensitivity between lethal and sublethal endpoints measured in 10-day tests (Sections 13 and 14) and between sublethal endpoints measured in the long-term tests. Results of these studies and additional applications of the methods described in Annex A6 and Annex A7 will provide data that can be used to assist in determining where application of long-term tests will be most appropriate. Use of sublethal endpoints for assessment of contaminant risk is not unique to toxicity testing with sediments. Numerous regulatory programs require the use of sublethal endpoints in the decision-making process (Pittinger and Adams (73)) including: (1) Water Quality Criteria (and State Standards); (2) National Pollution Discharge Elimination System (NPDES) effluent monitoring (including chemical-specific limits and sublethal endpoints in toxicity tests); (3) Federal Insecticide, Rodenticide and Fungicide Act (FIFRA) and the Toxic Substances Control Act (TSCA, tiered assessment includes several sublethal endpoints with fish and aquatic invertebrates); (4) Superfund (Comprehensive Environmental Responses, Compensation and Liability Act; CERCLA); (5) Organization of Economic Cooperation and Development (OECD, sublethal toxicity testing with fish and invertebrates); (6) European Economic Community (EC, sublethal toxicity testing with fish and invertebrates); and (7) the Paris Commission (behavioral endpoints).

    5.1.4 Results of toxicity tests on sediments spiked at different concentrations of chemicals can be used to establish cause and effect relationships between chemicals and biological responses. Results of toxicity tests with test materials spiked into sediments at different concentrations may be reported in terms of an LC50 (median lethal concentration), an EC50 (median effect concentration), an IC50 (inhibition concentration), or as a NOEC (no observed effect concentration) or LOEC (lowest observed effect concentration). However, spiked sediment may not be representative of chemicals associated with sediment in the field. Mixing time (74), aging (36, 75, 76), and the chemical form of the material can affect responses of test organisms in spiked sediment tests.

    5.1.5 Evaluating effect concentrations for chemicals in sediment requires knowledge of factors controlling their bioavailability. Similar concentrations of a chemical in units of mass of chemical per mass of sediment dry weight often exhibit a range in toxicity in different sediments (39, 40). Effect concentrations of chemicals in sediment have been correlated to interstitial water concentrations, and effect concentrations in interstitial water are often similar to effect concentrations in water-only exposures. The bioavailability of nonionic organic compounds in sediment is often inversely correlated with the organic carbon concentration. Whatever the route of exposure, these correlations of effect concentrations to interstitial water concentrations indicate that predicted or measured concentrations in interstitial water can be used to quantify the exposure concentration to an organism. Therefore, information on partitioning of chemicals between solid and liquid phases of sediment is useful for establishing effect concentrations (40).

    5.1.6 Field surveys can be designed to provide either a qualitative reconnaissance of the distribution of sediment contamination or a quantitative statistical comparison of contamination among sites.

    5.1.7 Surveys of sediment toxicity are usually part of more comprehensive analyses of biological, chemical, geological, and hydrographic data. Statistical correlations may be improved and sampling costs may be reduced if subsamples are taken simultaneously for sediment tests, chemical analyses, and benthic community structure.

    5.1.8 Table 3 lists several approaches the USEPA has considered for the assessment of sediment quality (77). These approaches include: (1) equilibrium partitioning, (2) tissue residues, (3) interstitial water toxicity, (4) whole-sediment toxicity and sediment-spiking tests, (5) benthic community structure, (6) effect ranges (for example, effect range median, ERM), and (7) sediment quality triad (see (78-81) for a critique of these methods). The sediment assessment approaches listed in Table 3 can be classified as numeric (for example, equilibrium partitioning), descriptive (for example, whole-sediment toxicity tests), or a combination of numeric and descriptive approaches (for example, ERM, (82). Numeric methods can be used to derive chemical-specific sediment quality guidelines (SQGs). Descriptive methods such as toxicity tests with field-collected sediment cannot be used alone to develop numerical SQGs for individual chemicals. Although each approach can be used to make site-specific decisions, no one single approach can adequately address sediment quality. Overall, an integration of several methods using the weight of evidence is the most desirable approach for assessing the effects of contaminants associated with sediment (83, 84, 85, 86). Hazard evaluations integrating data from laboratory exposures, chemical analyses, and benthic community assessments (the sediment quality triad) provide strong complementary evidence of the degree of pollution-induced degradation in aquatic communities (67, 87, 88).

    5.2 Regulatory Applications of Sediment Tests: 

    5.2.1 The USEPA has authority under a variety of statutes to manage contaminated sediments (Table 4). USEPA's Contaminated Sediment Management Strategy (89, 90) establishes the following four goals for contaminated sediments and describes actions that the Agency intends to take to accomplish these goals: (1) to prevent further contamination of sediments that may cause unacceptable ecological or human health risks; (2) when practical, to clean up existing sediment contamination that adversely affects the Nation's waterbodies or their uses, or that causes other significant effects on human health or the environment; (3) to ensure that sediment dredging and the disposal of dredged material continue to be managed in an environmentally sound manner; and (4) to develop and consistently apply methodologies for analyzing contaminated sediments. The Agency plans to employ its pollution prevention and source control programs to address the first goal. To accomplish the second goal, USEPA will consider a range of risk management alternatives to reduce the volume and effects of existing contaminated sediments, including in-situ containment and contaminated sediment removal. Finally, the Agency is developing tools for use in pollution prevention, source control, remediation, and dredged material management to meet the collective goals. These tools include national inventories of sediment quality and environmental releases of contaminants, numerical assessment guidelines to evaluate contaminant concentrations, and standardized bioassays to evaluate the bioaccumulation and toxicity potential of sediment samples.

    (A) CERCLA Comprehensive Environmental Response, Compensation and Liability Act (“Superfund”)

    CWA Clean Water Act

    FIFRA Federal Insecticide, Fungicide, and Rodenticide Act

    MPRSA Marine Protection, Resources and Sanctuary Act

    NEPA National Environmental Policy Act

    TSCA Toxic Substances Control Act

    RCRA Resource Conservation and Recovery Act

    5.2.2 The Clean Water Act (CWA) is the single most important law dealing with environmental quality of surface waters in the United States. The objective of the CWA is to restore and maintain the chemical, physical, and biological integrity of the nation's waters (CWA, Section 101). Federal and state monitoring programs traditionally have focused on evaluating water column problems caused by point source dischargers. Findings in the National Sediment Quality Survey, volume I of the first biennial report to Congress on sediment quality in the U.S., indicate that this focus needs to be expanded to include sediment quality impacts (Section 1.1.2 and (92).

    5.2.3 The Office of Water (OW), the Office of Prevention, Pesticides, and Toxic Substances (OPPTS), the Office of Solid Waste (OSW), and the Office of Emergency and Remedial Response (OERR) are all committed to the principle of consistent tiered testing described in the Contaminated Sediment Management Strategy (USEPA, (90)). Agency-wide consistent testing is desirable because all USEPA programs will use standard methods to evaluate health risk and produce comparable data. It will also provide the basis for uniform cross-program decision-making within the USEPA. Each program will, however, retain the flexibility of deciding whether identified risks would trigger regulatory actions.

    5.2.4 Tiered testing refers to a structured, hierarchial procedure for determining data needs relative to decision-making that consists of a series of tiers, or levels, of investigative intensity. Typically, increasing tiers in a tiered testing framework involve increased information and decreased uncertainty (USEPA, (90)). Each EPA program office intends to develop guidance for interpreting the tests conducted within the tiered framework and to explain how information within each tier would trigger regulatory action. Depending on statutory and regulatory requirements, the program specific guidance will describe decisions based on a weight of evidence approach, a pass-fail approach, or comparison to a reference site. The following two approaches are currently being used by USEPA: (1) the Office of Water-U.S. Army Corps of Engineers dredged material testing framework and (2) the OPPTS ecological risk assessment tiered testing framework. USEPA-USACE (93) describes the dredged material testing framework and Smrchek and Zeeman (94) summarizes the OPPTS testing framework. A tiered testing framework has not yet been chosen for agency-wide use, but some of the components have been identified to be standardized. These components are toxicity tests, bioaccumulation tests, chemical criteria, and other measurements that may have ecological significance including benthic community structure, colonization rate, and in situ testing within a mesocosm (77).

    5.3 Performance-based criteria: 

    5.3.1 The USEPA's Environmental Monitoring Management Council (EMMC) recommended the use of performance-based methods in developing standards (95). Performance-based methods were defined by EMMC as a monitoring approach which permits the use of appropriate methods that meet preestablished demonstrated performance standards (see 11.2).

    5.3.2 The USEPA Office of Water, Office of Science and Technology, and Office of Research and Development held a workshop to provide an opportunity for experts in the field of sediment toxicology and staff from USEPA's Regional and Headquarters Program offices to discuss the development of standard freshwater and marine sediment testing procedures (77, 96). Workgroup participants arrived at a consensus on several culturing and testing methods. In developing guidance for culturing test organisms to be included in the USEPA's methods manual for sediment tests, it was agreed that no one method should be required to culture organisms. However, the consensus at the workshop was that success of a test depends on the health of the cultures. Therefore, having healthy test organisms of known quality and age for testing was determined to be the key consideration relative to culturing methods. A performance-based criteria approach was selected in USEPA (2) as the preferred method through which individual laboratories could use unique culturing methods rather than requiring use of one culturing method.

    5.3.3 This standard recommends the use of performance-based criteria to allow each laboratory to optimize culture methods and minimize effects of test organism health on the reliability and comparability of test results. See 13.1.2 and 14.1.2 and Annex A1 to Annex A7 for a listing of performance criteria for culturing and testing.

    1. Scope

    1.1 This test method covers procedures for testing freshwater organisms in the laboratory to evaluate the toxicity of contaminants associated with whole sediments. Sediments may be collected from the field or spiked with compounds in the laboratory.

    1.1.1 Test methods are described for two toxicity test organisms, the amphipod Hyalella azteca (H. azteca) (see 13.1.2) and the midge Chironomus dilutus (formerly known as C. tentans; Shobanov et al. 1999.(1) (see 14.1.2). The toxicity tests are conducted for 10 days in 300-mL chambers containing 100 mL of sediment and 175 mL of overlying water. Overlying water is renewed daily and test organisms are fed during the toxicity tests. Endpoints for the 10-day toxicity tests are survival and growth. These test methods describe procedures for testing freshwater sediments; however, estuarine sediments (up to 15 ppt salinity) can also be tested with H. azteca. In addition to the 10-day toxicity test method outlined in 13.1.2 and 14.1.2, general procedures are also described for conducting 10-day sediment toxicity tests with H. azteca (see 13.1.2) and C. dilutus (see 14.1.2).

    Note 1: Morphological comparison of populations of Chironomus (Camptochironomus) tentans(Fabricius) from Europe, Asia, and North America have confirmed cytogenetic evidence that two distinct species inhabit the Palearctic and Nearctic under this name. The Palearctic species is the true C. tentans and the Nearctic populations constitute a new species described under the name Chironomus (Camptochironomus) dilutus (Shobanov et al. 1999 (1).”

    1.1.2 Guidance for conducting sediment toxicity tests is outlined in Annex A1 for Chironomus riparius, in Annex A2 for Daphnia magna and Ceriodaphnia dubia, in Annex A3 for Hexagenia spp., in Annex A4 for Tubifex tubifex, and in Annex A5 for the Diporeia spp. Guidance is also provided in Annex A6 for conducting long-term sediment toxicity tests with H. azteca by measuring effects on survival, growth, and reproduction. Guidance is also provided in Annex A7 for conducting long-term sediment toxicity tests with C. dilutus by measuring effects on survival, growth, emergence, and reproduction. 1.6 outlines the data that will be needed before test methods are developed from the guidance outlined in Annex A1 to Annex A7 for these test organisms. General procedures described in Sections 1 – 14 for sediment testing with H. azteca and C. dilutus are also applicable for sediment testing with the test organisms described in Annex A1 to Annex A7.

    1.2 Procedures outlined in this test method are based primarily on procedures described in the United States Environmental Protection Agency (USEPA) (2-9)2, Test Method E1367, and Guides E1391, E1525 and E1688.

    1.3 Additional research and methods development are now in progress to: (1) evaluate additional test organisms, (2) further evaluate the use of formulated sediment, (3) refine sediment dilution procedures, (4) refine sediment toxicity identification evaluation (TIE) procedures (10), (5) refine sediment spiking procedures, (6) develop in situ toxicity tests to assess sediment toxicity and bioaccumulation under field conditions, (7) evaluate relative sensitivities of endpoints measured in tests, (8) develop methods for new species, (9) evaluate relationships between toxicity and bioaccumulation, and (10) produce additional data on confirmation of responses in laboratory tests with natural populations of benthic organisms. Some issues that may be considered in interpretation of test results are the subject of continuing research including the influence of feeding on bioavailability, nutritional requirements of the test organisms, and additional performance criteria for organism health. See Section 6 for additional detail. This information will be described in future editions of this standard.

    1.4 The USEPA (2) and Guide E1688 also describes 28-day bioaccumulation methods for the oligochaete Lumbriculus variegatus.

    1.5 Results of tests, even those with the same species, using procedures different from those described in the test method may not be comparable and using these different procedures may alter bioavailability. Comparison of results obtained using modified versions of these procedures might provide useful information concerning new concepts and procedures for conducting sediment tests with aquatic organisms. If tests are conducted with procedures different from those described in this test method, additional tests are required to determine comparability of results. General procedures described in this test method might be useful for conducting tests with other aquatic organisms; however, modifications may be necessary.

    1.6 Selection of Toxicity Testing Organisms: 

    1.6.1 The choice of a test organism has a major influence on the relevance, success, and interpretation of a test. Furthermore, no one organism is best suited for all sediments. The following criteria were considered when selecting test organisms to be described in this standard (Table 1 and Guide E1525). A test organism should: (1) have a toxicological data base demonstrating relative sensitivity and discrimination to a range of chemicals of concern in sediment, (2) have a database for interlaboratory comparisons of procedures (for example, round-robin studies), (3) be in contact with sediment [e.g., water column vs benthic organisms], (4) be readily available through culture or from field collection, (5) be easily maintained in the laboratory, (6) be easily identified, (7) be ecologically or economically important, (8) have a broad geographical distribution, be indigenous (either present or historical) to the site being evaluated, or have a niche similar to organisms of concern, (for example, similar feeding guild or behavior to the indigenous organisms), (9) be tolerant of a broad range of sediment physico-chemical characteristics (for example, grain size), and (10) be compatible with selected exposure methods and endpoints. The method should also be (11) peer reviewed and (12) confirmed with responses with natural populations of benthic organisms (see 1.6.8).

    S = survival, G = Growth, B = Bioaccumulation, A = avoidance
     R = Reproduction, M = Maturation, E = Emergence

    1.6.2 Of the criteria outlined in Table 1, a data base demonstrating relative sensitivity to contaminants, contact with sediment, ease of culture in the laboratory, interlaboratory comparisons, tolerance of varying sediment physico-chemical characteristics, and confirmation with responses of natural benthos populations were the primary criteria used for selecting H. azteca and C. dilutus to be described as test methods in the current version of this standard (see Sections 13 and 14). Procedures for conducting sediment tests with organisms in accordance with Annex A1 to Annex A7 do not currently meet all the required selection criteria listed in Table 1. A similar data base must be developed before these or other test organisms can be included as standard test methods instead of as guidance in future versions of these this method.

    1.6.3 An important consideration in the selection of specific species for test method development is the existence of information concerning relative sensitivity of the organisms both to single chemicals and complex mixtures. A number of studies have evaluated the sensitivity of H. azteca, C. dilutus, and L. variegatus, relative to one another, as well as other commonly tested freshwater species. For example, Ankley et al (11) found H. azteca to be as, or slightly more, sensitive than Ceriodaphnia dubia to a variety of sediment elutriate and pore-water samples. In that study, L. variegatus were less sensitive to the samples than either the amphipod or the cladoceran. West et al (12) found the rank sensitivity of the three species to the lethal effects of copper in sediments from the Keweenaw Waterway, MI was (from greatest to least): H. azteca > C. dilutus > L. variegatus. In short-term (48 to 96 h) exposures, L. variegatus generally was less sensitive than H. azteca, C. dubia, or Pimephales promelas to cadmium, nickel, zinc, copper, and lead (13). Of the latter three species, no one species was consistently the most sensitive to the five metals. In a study of contaminated Great Lakes sediment, H. azteca, C. dilutus, and C. riparius were among the most sensitive and discriminatory of 24 organisms tested (14-17). Kemble et al (18) found the rank sensitivity of four species to metal-contaminated sediments from the Clark Fork River, MT to be (from greatest to least): H. azteca > C. riparius > Oncorhynchus mykiss (rainbow trout) > Daphnia magna. Relative sensitivity of the three endpoints evaluated in the H. azteca test with Clark Fork River sediments was (from greatest to least): length > sexual maturation > survival. In 10-day water-only and whole-sediment tests, Hyalella azteca and C. dilutus were more sensitive than D. magna to fluoranthene-spiked sediment (19). Ten-day, water-only tests also have been conducted with a number of chemicals using H. azteca, C. dilutus, and L. variegatus ((19) and Table 2). These tests all were flow-through exposures using a soft natural water (Lake Superior) with measured chemical concentrations that, other than the absence of sediment, were conducted under conditions (for example, temperature, photoperiod, feeding) similar to those being described for the standard 10-day sediment test in 13.1.2. In general, H. azteca was more sensitive to copper, zinc, cadmium, nickel, and lead than either C. dilutus or L. variegatus. Chironomus dilutus and H. azteca exhibited a similar sensitivity to several of the pesticides tested. Lumbriculus variegatus was not tested with several of the pesticides; however, in other studies with whole sediments contaminated by dichlorodiphenyltrichloroethane (DDT) and associated metabolites, and in short-term (96-h) experiments with organophosphate insecticides (diazinon, chlorpyrifos), L. variegatus has proved to be far less sensitive than either H. azteca or C. dilutus. These results highlight two important points germane to these test methods. First, neither of the two test species selected for estimating sediment toxicity (H. azteca, C. dilutus) was consistently most sensitive to all chemicals, indicating the importance of using multiple test organisms when performing sediment assessments. Second, L. variegatus appears to be relatively insensitive to most of the test chemicals, which perhaps is a positive attribute for an organism used for bioaccumulation testing (9).

    1 50 % mortality at highest concentration tested.

    2 70 % mortality at lowest concentration tested.

    3 NT, not tested. Using the data from Table 2, sensitivity of H. azteca, C. dilutus, and L. variegatus can be evaluated relative to other freshwater species. For this analysis, acute and chronic toxicity data from water quality criteria (WQC) documents for copper, zinc, cadmium, nickel, lead, DDT, dieldrin, and chlorpyrifos, and toxicity information from the AQUIRE data base (20) for 1,1,dichloro-2,2-bis(p-chlorophenyl)ethane (DDD) and dichlorodiphenyldichloroethylene (DDE), were compared to assay results for the three species (19). The sensitivity of H. azteca to metals and pesticides, and C. dilutus to pesticides was comparable to chronic toxicity data generated for other test species. This was not completely unexpected given that the 10-day exposures used for these two species are likely more similar to chronic partial life-cycle tests than the 48 to 96-h exposures traditionally defined as acute in the WQC documents. Interestingly, in some instances (for example, dieldrin and chlorpyrifos), LC50 data generated for H. azteca or C. dilutus were comparable to or lower than any reported for other freshwater species in the WQC documents. This observation likely is a function not only of the test species, but of the test conditions; many of the tests on which early WQC were based were static, rather than flow-through, and report unmeasured contaminant concentrations. Measurable concentrations of ammonia are common in the pore water of many sediments and have been found to be a common cause of toxicity in pore water (21 , 22, 23). Acute toxicity of ammonia to H. azteca, C. dilutus, and L. variegatus has been evaluated in several studies. As has been found for many other aquatic organisms, the toxicity of ammonia to C. dilutus and L. variegatus has been shown to be dependent on pH. Four-day LC50 values for L. variegatus in water-column (no sediment) exposures ranged from 390 to 6.6 mg/L total ammonia as pH was increased from 6.3 to 8.6 Schubauer-Berigan et al.(24). For C. dilutus, 4-day LC50 values ranged from 370 to 82 mg/L total ammonia over a similar pH range (Schubauer-Berigan et al.) (24). Ankley et al. (25) reported that the toxicity of ammonia to H. azteca (also in water-only exposures) showed differing degrees of pH-dependence in different test waters. In soft reconstituted water, toxicity was not pH dependent, with 4-day LC50 values of about 20 mg/L at pH ranging from 6.5 to 8.5. In contrast, ammonia toxicity in hard reconstituted water exhibited substantial pH dependence with LC50 values decreasing from >200 to 35 mg/L total ammonia over the same pH range. Borgmann and Borgmann ( 26) later showed that the variation in ammonia toxicity across these waters could be attributed to differences in sodium and potassium content, which appear to influence the toxicity of ammonia to H. azteca.

    (1) Although these studies provide benchmark concentrations that may be of concern in sediment pore waters, additional studies by Whiteman et al. (27) indicated that the relationship between water-only LC50 values and those measured in sediment exposures differs among organisms. In sediment exposures, the 10-day LC50 for L. variegatus and C. dilutus occurred when sediment pore water reached about 150 % of the LC50 determined from water-only exposures. However, experiments with H. azteca showed that the 10-day LC50 was not reached until pore water concentrations were nearly 10× the water-only LC50, at which time the ammonia concentration in the overlying water was equal to the water-only LC50. The authors attribute this discrepancy to avoidance of sediment by H. azteca. Thus, it appears that water-only LC50 values may provide suitable screening values for potential ammonia toxicity, higher concentrations may be necessary to actually induce ammonia toxicity in sediment exposures, particularly for H. azteca. Further, these data underscore the importance of measuring the pH of pore water when ammonia toxicity may be of concern. Ankley Schubauer-Bergian (28) and Besser et al. (29) describe procedures for conducting toxicity identification evaluations (TIEs) for pore-water or whole-sediment samples to determine if ammonia is contributing to the toxicity of sediment samples.

    1.6.4 Relative species sensitivity frequently varies among chemicals; consequently, a battery of tests including organisms representing different trophic levels may be needed to assess sediment quality (14, 17, 30-33). For example, Reish (34) reported the relative toxicity of six metals (arsenic, cadmium, chromium, copper, mercury, and zinc) to crustaceans, polychaetes, pelecypods, and fishes and concluded that no one species or group of test organisms was the most sensitive to all of the metals. Sensitivity of a species to chemicals is also dependent on the duration of the exposure and the endpoints evaluated. Annex A6 and Annex A7 describe results of studies which demonstrate the utility of measuring sublethal endpoints in sediment toxicity tests with the amphipod H. azteca and the midge C. dilutus.

    1.6.5 The sensitivity of an organism to chemicals should be balanced with the concept of discrimination (14). The response of a test organism should provide discrimination between different levels of contamination. However, insensitive organisms may be preferred for determining bioaccumulation. The use of indigenous organisms that are ecologically important and easily collected is often very straightforward; however, indigenous organisms at a site may be insensitive to the chemicals of concern. Indigenous organisms might be more important for evaluation of bioaccumulation (9). See Guides E1525, E1688, and E1850 for additional detail on selection of test organisms.

    1.6.6 Sensitivity of an organism is related to route of exposure and biochemical sensitivity to chemicals. Sediment-dwelling organisms can receive a dose from three primary sources: interstitial water, sediment particles, and overlying water. Food type, feeding rate, assimilation efficiency, and clearance rate will control the dose of chemicals from sediment (Guide E1688). Benthic invertebrates often selectively consume different particle sizes (35) or particles with higher organic carbon concentrations which may have higher chemical concentrations. Detrital feeders may receive most of their body burden directly from sediment ingestion. In amphipods (36) and clams (37) uptake through the gut can exceed uptake across the gills for certain hydrophobic compounds. Organisms in direct contact with sediment may also accumulate chemicals by direct adsorption to the body wall or by absorption through the integument (38).

    1.6.7 Despite the potential complexities in estimating the dose that an animal receives from sediment, the toxicity and bioaccumulation of many chemicals in sediment such as chlordecone, fluoranthene, organochlorines, and metals have been correlated with either the concentration of these chemicals in interstitial water or in the case of nonionic organic chemicals, concentrations of an organic-carbon basis (39, 40). The relative importance of whole sediment and interstitial water routes of exposure depends on the test organism and the specific contaminant (35, 38). Because benthic communities contain a diversity of organisms, many combinations of exposure routes may be important. Therefore, behavior and feeding habits of a test organism can influence its ability to accumulate contaminants from sediment and should be considered when selecting test organisms for sediment testing.

    1.6.8 The response of H. azteca and C. dilutus in laboratory toxicity studies has been compared to the response of natural populations of benthic organisms to potentially contaminated sediments. Chironomids were not found in sediment samples that decreased the growth of C. dilutus by 30 % or more in 10-day laboratory toxicity tests (41). Wentsel et al (42-44) reported a correlation between effects on C. dilutus in laboratory tests and the abundance of C. dilutus in metal-contaminated sediments. Canfield et al. (45,46,47) evaluated the composition of benthic invertebrate communities in sediments for the following areas: (1) three Great Lakes Areas of Concern (AOC; Buffalo River, NY: Indiana Harbor, IN: Saginaw River, MI), (2) the upper Mississippi River, and (3) the Clark Fork River located in Montana. Results of these benthic community assessments were compared to sediment chemistry and toxicity (28-day sediment exposures with H. azteca which monitored effects on survival, growth, and sexual maturation). Good concordance was evident between measures of laboratory toxicity, sediment contamination, and benthic invertebrate community composition in extremely contaminated samples. However, in moderately contaminated samples, less concordance was observed between the composition of the benthic community and either laboratory toxicity test results or sediment contaminant concentration. Laboratory sediment toxicity tests better identified chemical contamination in sediments compared to many of the commonly used measures of benthic invertebrate community composition. Benthic measures may reflect other factors such as habitat alteration in addition to responding to contaminants. Canfield et al. (45, 46, 47) identified the need to better evaluate non-contaminant factors (i.e., TOC, grain size, water depth, habitat alteration) in order to better interpret the response of benthic invertebrates to sediment contamination. Results from laboratory sediment toxicity tests were compared to colonization of artificial substrates exposed in situ to Great Lakes sediment (14) Burton et al. (17) Survival or growth of H. azteca and C. dilutus in 10–28-day laboratory exposures were negatively correlated to percent chironomids and percent tolerant taxa colonizing artificial substrates in the field. Schlekat et al (48) reported general good agreement between sediment toxicity tests with H. azteca and benthic community responses in the Anacostia River in Washington, DC. Sediment toxicity with amphipods in 10-day toxicity tests, field contamination, and field abundance of benthic amphipods were examined along a sediment contamination gradient of DDT (48). Survival of Eohaustorius estuarius, Rhepoxynius abronius, and H. azteca in laboratory toxicity tests was positively correlated to abundance of amphipods in the field and negatively correlated to DDT concentrations. The threshold for 10-day sediment toxicity in laboratory studies was about 300 μg DDT (+metabolites)/g organic carbon. The threshold for abundance of amphipods in the field was about 100 μg DDT (+metabolites)/g organic carbon. Therefore, correlations between toxicity, contamination, and field populations indicate that short-term sediment toxicity tests can provide reliable evidence of biologically adverse sediment contamination in the field, but may be underprotective of sublethal effects.

    1.7 Limitations—While some safety considerations are included in this standard, it is beyond the scope of this standard to encompass all safety requirements necessary to conduct sediment tests.

    1.8 This standard is arranged as follows: 





    Referenced Documents




    Summary of Standard


    Significance and Use




    Reagents and Materials




    Facilities, Equipment, and Supplies


    Sample Collection, Storage, Manipulation, and Characterization


    Quality Assurance and Quality Control


    Collection, Culturing, and Maintaining Test Organisms


    Procedure 1: Conducting a 10-day Sediment Toxicity Test with Hyalella azteca


    Procedure 2: Conducting a 10-day Sediment Toxicity Test with Chironomus dilutus






    Precision and Bias




    A1. Guidance for Conducting Sediment Toxicity Tests with Chironomus riparius

    A2. Guidance for Conducting Sediment Toxicity Tests with Daphnia magna and Ceriodaphnia dubia

    A3. Guidance for Conducting Sediment Toxicity Tests with Hexagenia spp.

    A4. Guidance for Conducting Sediment Toxicity Tests with Tubifex tubifex

    A5. Guidance for Conducting Sediment Toxicity Tests with Diporeia spp.

    A6. Guidance for Conducting a Hyalella Azteca 42-day Test for Measuring Effects of Sediment-Associated Contaminants on Survival, Growth, and Reproduction

    A7. Guidance for Conducting a Life-Cycle Test for Measuring Effects of Sediment-Associated Contaminants on Chironomus dilutus.

    A8. Food Preparation

    A9. Feeding Rate for the 10-day Sediment Toxicity Test Method with Chironomus dilutus


    1.9 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 8.

    1.10 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

    2. Referenced Documents (purchase separately) The documents listed below are referenced within the subject standard but are not provided as part of the standard.

    ASTM Standards

    D1129 Terminology Relating to Water

    D4387 Guide for Selecting Grab Sampling Devices for Collecting Benthic Macroinvertebrates

    D4447 Guide for Disposal of Laboratory Chemicals and Samples

    E29 Practice for Using Significant Digits in Test Data to Determine Conformance with Specifications

    E105 Practice for Probability Sampling of Materials

    E122 Practice for Calculating Sample Size to Estimate, With Specified Precision, the Average for a Characteristic of a Lot or Process

    E141 Practice for Acceptance of Evidence Based on the Results of Probability Sampling

    E177 Practice for Use of the Terms Precision and Bias in ASTM Test Methods

    E178 Practice for Dealing With Outlying Observations

    E456 Terminology Relating to Quality and Statistics

    E691 Practice for Conducting an Interlaboratory Study to Determine the Precision of a Test Method

    E729 Guide for Conducting Acute Toxicity Tests on Test Materials with Fishes, Macroinvertebrates, and Amphibians

    E943 Terminology Relating to Biological Effects and Environmental Fate

    E1193 Guide for Conducting Daphnia magna Life-Cycle Toxicity Tests

    E1241 Guide for Conducting Early Life-Stage Toxicity Tests with Fishes

    E1295 Guide for Conducting Three-Brood, Renewal Toxicity Tests with Ceriodaphnia dubia

    E1325 Terminology Relating to Design of Experiments

    E1367 Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Estuarine and Marine Invertebrates

    E1391 Guide for Collection, Storage, Characterization, and Manipulation of Sediments for Toxicological Testing and for Selection of Samplers Used to Collect Benthic Invertebrates

    E1402 Guide for Sampling Design

    E1525 Guide for Designing Biological Tests with Sediments

    E1688 Guide for Determination of the Bioaccumulation of Sediment-Associated Contaminants by Benthic Invertebrates

    E1847 Practice for Statistical Analysis of Toxicity Tests Conducted Under ASTM Guidelines

    E1850 Guide for Selection of Resident Species as Test Organisms for Aquatic and Sediment Toxicity Tests

    UNSPSC Code

    UNSPSC Code 77101501(Risk or hazard assessment)

    Referencing This Standard
    Link Here
    Link to Active (This link will always route to the current Active version of the standard.)

    DOI: 10.1520/E1706-19

    Citation Format

    ASTM E1706-19, Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates, ASTM International, West Conshohocken, PA, 2019,

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