ASTM E1525-02(2023)

Standard Guide for Designing Biological Tests with Sediments

SIGNIFICANCE AND USE
6.1 Contaminated sediments may affect natural populations of aquatic organisms adversely. Sediment-dwelling organisms may be exposed directly to contaminants by the ingestion of sediments and by the uptake of sediment-associated contaminants from interstitial and overlying water. Contaminated sediments may affect water column species directly by serving as a source of contaminants to overlying waters or a sink for contaminants from overlying waters. Organisms may also be affected when contaminated sediments are suspended in the water column by natural or human activities. Water column species and nonaquatic species may also be affected indirectly by contaminated sediments by the transfer of contaminants through ecosystems (7, 8).  
6.2 The procedures described in this guide may be used and adapted for incorporation in basic and applied research to determine the ecological effects of contaminated sediments. These same methods may also be used in the development and implementation of monitoring and regulatory programs designed to prevent and manage sediment contamination.  
6.3 Sediment tests with aquatic organisms can be used to quantify the acute and chronic toxicity and the bioavailability of new and presently used materials. Sediment toxicity may also result from environmental processes such as ammonia generation, pH shifts, or dissolved oxygen fluctuation. In many cases, consideration of the adverse effects of sediment-associated contaminants is only one part of a complete hazard assessment of manufactured compounds that are applied directly to the environment (for example, pesticides) and those released (for example, through wastewater effluents) as by-products from the manufacturing process or from municipalities (7).  
6.4 Sediment tests can be used to develop exposure-response relationships for individual toxicants by spiking clean sediments with varying concentrations of a test chemical and determining the concentration that elicits the target response in...
SCOPE
1.1 As the contamination of freshwater and saltwater ecosystems continues to be reduced through the implementation of regulations governing both point and non-point source discharges, there is a growing emphasis and concern regarding historical inputs and their influence on water and sediment quality. Many locations in urban areas exhibit significant sediment contamination, which poses a continual and long-term threat to the functional condition of benthic communities and other species inhabiting these areas (1).2 Benthic communities are an important component of many ecosystems and alterations of these communities may affect water-column and nonaquatic species.  
1.2 Biological tests with sediments are an efficient means for evaluating sediment contamination because they provide information complementary to chemical characterizations and ecological surveys (2). Acute sediment toxicity tests can be used as screening tools in the early phase of an assessment hierarchy that ultimately could include chemical measurements or bioaccumulation and chronic toxicity tests. Sediment tests have been applied in both saltwater and freshwater environments (2-6). Sediment tests have been used for dredge material permitting, site ranking for remediation, recovery studies following management actions, and trend monitoring. A particularly important application is for establishing contaminant-specific effects and the processes controlling contaminant bioavailability(7).  
1.3 This guide is arranged as follows:    
Section  
Referenced Documents  
2  
Terminology  
3  
Application  
4  
Summary of Guide  
5  
Significance and Use  
6  
Hazards  
7  
Sediment Test Types  
8  
Biological Responses  
9  
Test Organisms  
10  
Experimental Design Considerations  
11  
Data Interpretation  
12  
Keywords  
13  
1.4 The values stated in SI units are to be regarded as the standard. The values g...

General Information

• Status: Published
• Publication Date: 31-Dec-2022
• ICS: 13.080.30 - Biological properties of soils
• Technical Committee: E50 - Environmental Assessment, Risk Management and Corrective Action
• Drafting Committee: E50.47 - Biological Effects and Environmental Fate

Relations

• Refers: ASTM D1129-13(2020)e2 - Standard Terminology Relating to Water
• Effective Date: 01-May-2020
• Refers: ASTM E1688-19 - Standard Guide for Determination of the Bioaccumulation of Sediment-Associated Contaminants by Benthic Invertebrates
• Effective Date: 01-Dec-2019
• Refers: ASTM E1706-19 - Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates
• Effective Date: 01-Apr-2019
• Refers: ASTM E1688-10(2016) - Standard Guide for Determination of the Bioaccumulation of Sediment-Associated Contaminants by Benthic Invertebrates
• Effective Date: 01-Feb-2016
• Refers: ASTM E1706-05(2010) - Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates (Withdrawn 2019)
• Effective Date: 01-Sep-2010
• Refers: ASTM E1688-10 - Standard Guide for Determination of the Bioaccumulation of Sediment-Associated Contaminants by Benthic Invertebrates
• Effective Date: 01-Apr-2010
• Refers: ASTM D1129-10 - Standard Terminology Relating to Water
• Effective Date: 01-Mar-2010
• Refers: ASTM E943-08 - Standard Terminology Relating to Biological Effects and Environmental Fate
• Effective Date: 01-Mar-2008
• Refers: ASTM E1367-03(2008) - Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Estuarine and Marine Invertebrates
• Effective Date: 01-Feb-2008
• Refers: ASTM E1391-03(2008) - Standard Guide for Collection, Storage, Characterization, and Manipulation of Sediments for Toxicological Testing and for Selection of Samplers Used to Collect Benthic Invertebrates
• Effective Date: 01-Feb-2008
• Refers: ASTM E729-96(2007) - Standard Guide for Conducting Acute Toxicity Tests on Test Materials with Fishes, Macroinvertebrates, and Amphibians
• Effective Date: 01-Oct-2007
• Refers: ASTM E1023-84(2007) - Standard Guide for Assessing the Hazard of a Material to Aquatic Organisms and Their Uses
• Effective Date: 01-Oct-2007
• Refers: ASTM D4447-10 - Standard Guide for Disposal of Laboratory Chemicals and Samples
• Effective Date: 15-Nov-2006
• Refers: ASTM D4447-06 - Standard Guide for Disposal of Laboratory Chemicals and Samples
• Effective Date: 15-Nov-2006
• Refers: ASTM D1129-06a - Standard Terminology Relating to Water
• Effective Date: 01-Sep-2006

Overview

ASTM E1525-02(2023) is the standard guide developed by ASTM International for designing biological tests with sediments. This guide plays a pivotal role in evaluating the ecological risks associated with contaminated sediments in both freshwater and saltwater environments. Through clearly outlined strategies, the standard helps professionals in environmental sciences, risk assessment, and regulatory fields to assess the impact of sediment contamination on aquatic and benthic organisms, providing essential guidance on the selection, application, and interpretation of biological sediment tests.

This ASTM standard is globally recognized and developed according to the World Trade Organization’s principles for standardization, ensuring its relevance and alignment with international regulatory and scientific expectations.

Key Topics

• Significance of Sediment Testing: Contaminated sediments can pose long-term threats to aquatic life and ecosystem health. ASTM E1525 highlights how sediments act as both a source and a sink for contaminants, directly and indirectly affecting a range of organisms through ingestion, water exposure, and ecosystem transfer.
• Test Objectives and Strategies: The guide provides a framework for designing sediment toxicity tests, including the selection of appropriate test organisms, exposure methods (whole sediment, elutriate, pore water, suspended sediment), and endpoints (such as survival, growth, and reproduction).
• Experimental Design: Recommendations include controls, replication, and statistical procedures to ensure robust, defensible results. The guide also details considerations for sampling, sample handling, and laboratory safety.
• Types of Biological Responses: Sediment tests can evaluate acute and chronic toxicity, bioavailability of contaminants, and sublethal effects. Endpoints include mortality, growth inhibition, reproductive impacts, behavioral changes, and bioaccumulation.
• Role in Regulatory and Monitoring Programs: Procedures may be adapted for ecological research, environmental monitoring, and regulatory compliance, supporting management and remediation decisions for hazardous sites.

Applications

ASTM E1525 is widely adopted in various practical scenarios, providing actionable guidance for:

• Environmental Impact Assessments: Used during site assessments and environmental remediation planning to evaluate sediment quality and its biological effects on target organisms.
• Dredging and Sediment Disposal Permitting: Essential for assessing toxicity of dredged materials prior to disposal in aquatic ecosystems, fulfilling regulatory requirements for environmental protection agencies.
• Trend Monitoring and Recovery Studies: Facilitates long-term ecological monitoring, helping track the effectiveness of pollution control and sediment remediation actions.
• Basic and Applied Research: Supports research projects aiming to determine exposure-response relationships for specific contaminants and the mechanisms controlling contaminant bioavailability.
• Development of Regulatory Standards: Informs water quality criteria, effluent monitoring, and other regulatory programs requiring sediment toxicity data, ensuring compliance with environmental standards.

Related Standards

ASTM E1525 references and complements several other standards and guides, including:

• ASTM E724 - Guide for Conducting Static Short-Term Chronic Toxicity Tests Starting with Embryos of Four Species of Saltwater Bivalve Molluscs
• ASTM E729 - Guide for Conducting Acute Toxicity Tests on Test Materials with Fishes, Macroinvertebrates, and Amphibians
• ASTM E1391 - Guide for Collection, Storage, Characterization, and Manipulation of Sediments for Toxicological Testing
• ASTM E1706 - Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates
• ASTM E1688 - Guide for Determination of the Bioaccumulation of Sediment-Associated Contaminants by Benthic Invertebrates

By integrating these related standards, users can develop comprehensive approaches to sediment testing, ensuring consistency, comparability, and scientific credibility in their environmental assessments.

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Keywords: ASTM E1525, sediment toxicity testing, biological sediment tests, contaminated sediments, aquatic toxicology, environmental standards, risk assessment, sediment contamination monitoring, dredged material assessment, environmental compliance.

Standards Content (Sample)

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.
Designation: E1525 − 02 (Reapproved 2023)
Standard Guide for
Designing Biological Tests with Sediments
This standard is issued under the fixed designation E1525; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope*
Test Organisms 10
Experimental Design Considerations 11
1.1 As the contamination of freshwater and saltwater eco-
Data Interpretation 12
systems continues to be reduced through the implementation of Keywords 13
regulations governing both point and non-point source
1.4 The values stated in SI units are to be regarded as the
discharges, there is a growing emphasis and concern regarding
standard. The values given in parentheses are for information
historical inputs and their influence on water and sediment
only.
quality. Many locations in urban areas exhibit significant
1.5 This standard does not purport to address all of the
sediment contamination, which poses a continual and long-
safety concerns, if any, associated with its use. It is the
term threat to the functional condition of benthic communities
responsibility of the user of this standard to establish appro-
and other species inhabiting these areas (1). Benthic commu-
priate safety, health, and environmental practices and deter-
nities are an important component of many ecosystems and
mine the applicability of regulatory limitations prior to use.
alterations of these communities may affect water-column and
For specific hazard statements, see Section 7.
nonaquatic species.
1.6 This international standard was developed in accor-
1.2 Biological tests with sediments are an efficient means
dance with internationally recognized principles on standard-
for evaluating sediment contamination because they provide
ization established in the Decision on Principles for the
information complementary to chemical characterizations and
Development of International Standards, Guides and Recom-
ecological surveys (2). Acute sediment toxicity tests can be
mendations issued by the World Trade Organization Technical
used as screening tools in the early phase of an assessment
Barriers to Trade (TBT) Committee.
hierarchy that ultimately could include chemical measurements
2. Referenced Documents
or bioaccumulation and chronic toxicity tests. Sediment tests
have been applied in both saltwater and freshwater environ-
2.1 ASTM Standards:
ments (2-6). Sediment tests have been used for dredge material
D1129 Terminology Relating to Water
permitting, site ranking for remediation, recovery studies
D4447 Guide for Disposal of Laboratory Chemicals and
following management actions, and trend monitoring. A par-
Samples
ticularly important application is for establishing contaminant-
E724 Guide for Conducting Static Short-Term Chronic Tox-
specific effects and the processes controlling contaminant
icity Tests Starting with Embryos of Four Species of
bioavailability(7).
Saltwater Bivalve Molluscs
E729 Guide for Conducting Acute Toxicity Tests on Test
1.3 This guide is arranged as follows:
Materials with Fishes, Macroinvertebrates, and Amphib-
Section
ians
Referenced Documents 2
Terminology 3
E943 Terminology Relating to Biological Effects and Envi-
Application 4
ronmental Fate (Withdrawn 2023)
Summary of Guide 5
E1023 Guide for Assessing the Hazard of a Material to
Significance and Use 6
Hazards 7
Aquatic Organisms and Their Uses
Sediment Test Types 8
E1367 Test Method for Measuring the Toxicity of Sediment-
Biological Responses 9
Associated Contaminants with Estuarine and Marine In-
vertebrates
This guide is under the jurisdiction of ASTM Committee E50 on Environmental
Assessment, Risk Management and Corrective Action and is the direct responsibil-
ity of Subcommittee E50.47 on Biological Effects and Environmental Fate. For referenced ASTM standards, visit the ASTM website, www.astm.org, or
Current edition approved Jan. 1, 2023. Published March 2023. Originally contact ASTM Customer Service at [e-mail]. For Annual Book of ASTM
approved in 1993. Last previous edition approved in 2014 as E1525 – 02(2014). Standards volume information, refer to the standard’s Document Summary page on
DOI: 10.1520/E1525-02R23. the ASTM website.
2 4
The boldface numbers in parentheses refer to the list of references at the end of The last approved version of this historical standard is referenced on
this standard. www.astm.org.
*A Summary of Changes section appears at the end of this standard
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
E1525 − 02 (2023)
E1383 Guide for Conducting Sediment Toxicity Tests with 3.2.3 control sediment—a sediment that is essentially free of
Freshwater Invertebrates (Withdrawn 1995) contaminants and is used routinely to assess the acceptability
E1391 Guide for Collection, Storage, Characterization, and of a test.
Manipulation of Sediments for Toxicological Testing and
3.2.4 elutriate—the water and soluble portion extracted
for Selection of Samplers Used to Collect Benthic Inver-
from the sediment.
tebrates
3.2.5 exposure—contact with a chemical or physical agent.
E1563 Guide for Conducting Short-Term Chronic Toxicity
3.2.6 overlying water—the water placed over the solid
Tests with Echinoid Embryos
phase of a sediment in the test chamber for the conduct of the
E1611 Guide for Conducting Sediment Toxicity Tests with
biological test; this may also include the water used to
Polychaetous Annelids
manipulate the sediments. In field situations, the water column
E1676 Guide for Conducting Laboratory Soil Toxicity or
above the sediment/water interface.
Bioaccumulation Tests with the Lumbricid Earthworm
Eisenia Fetida and the Enchytraeid Potworm Enchytraeus
3.2.7 pore water/interstitial water—water occupying space
albidus between sediment or soil particles.
E1688 Guide for Determination of the Bioaccumulation of
3.2.8 reference sediment—a whole sediment near the area of
Sediment-Associated Contaminants by Benthic Inverte-
concern used to assess sediment conditions exclusive of
brates
material(s) of interest.
E1706 Test Method for Measuring the Toxicity of Sediment-
3.2.9 sediment—(1) particulate material that usually lies
Associated Contaminants with Freshwater Invertebrates
below water and (2) formulated paticulate matter that is
IEEE/ASTM SI-10 Standard for Use of the International
intended to lie below water in a test.
System of Units (SI): The Modern Metric System
3.2.10 spiked sediment—a sediment to which a material has
2.2 Other Standards:
been added for experimental purposes.
Title 29 Code of Federal Regulations 1910.132 (f)
3.2.11 suspension—a slurry of sediment and water.
3. Terminology 3.2.12 toxicity—the property of a material or combination of
materials to affect organisms adversely.
3.1 Definitions:
3.2.13 whole sediment—sediment and associated pore water
3.1.1 The words “must,” “should,” “may,” “can,” and
that has had minimal manipulation following collection or
“might” have very specific meanings in this guide. “Must” is
formulation.
used to express an absolute requirement, that is, to state that the
test ought to be designed to satisfy a specific condition, unless
4. Application
the purpose of the test requires a different design. “Must” is
4.1 An ASTM guide outlines a series of options or instruc-
used only in connection with the factors that apply directly to
tions and does not recommend a specific course of action. The
the acceptability of the test. “Should” is used to state that the
purpose of a guide is to offer guidance, based on a consensus
specified conditions are recommended and ought to be met in
of viewpoints, but not to establish a fixed procedure. A guide is
most tests. Although a violation of one “should” is rarely a
intended to increase the awareness of the user to available
serious matter, violation of several will often render the results
techniques in a given subject area and to provide information
questionable. Terms such as “is desirable,” “is often desirable,”
from which subsequent evaluation and standardization can be
and “might be desirable” are used in connection with less
derived.
important factors. “May” is used to mean “is (are) allowed to,”
“can” is used to mean“ is (are) able to,” and “might” is used to 4.2 This guide provides general interpretative guidance on
the selection, application, and interpretation of biological tests
mean “could possibly.” Thus, the classic distinction between
“may” and“ can” is preserved, and “might” is never used as a with sediments. As such, this guide serves as a preface to other
ASTM documents describing methods for sediment collection,
synonym of either “may” or “can.”
storage, and manipulation (Guide E1391); and toxicity or
3.1.2 For definitions of terms used in this guide, refer to
bioaccumulation tests with sediment ( Guides E724, E1367,
Guide E729, Terminologies D1129 and E943, and Guide
E1391, E1611, E1563, E1688, and Test Method E1706). Much
E1023. For an explanation of the units and symbols, refer to
of the guidance presented in this standard is also applicable to
IEEE/ASTM SI-10.
toxicity testing of soils (Guide E1676). This guide serves as an
3.2 Definitions of Terms Specific to This Standard:
introduction and summary of sediment testing and is not meant
3.2.1 bioaccumulation—the net uptake of a material by an
to provide specific guidance on test methods. Rather, its intent
organism from its environment through exposure by means of
is to provide information necessary to accomplish the follow-
water and food.
ing:
3.2.2 concentration—the ratio of the weight or volume of
4.2.1 Select a sediment exposure strategy appropriate to the
test material(s) to the weight or volume of test sample.
assessment need. For example, a suspended phase exposure is
relevant to the evaluation of dredged sediments for disposal at
a dispersive aquatic site. (See Annex A1).
4.2.2 Select the test organism and biological endpoints
Available from Superintendent of Documents, U.S. Government Printing
Office, Washington DC 20402. appropriate to the desired exposure and aquatic resources at
E1525 − 02 (2023)
risk. For example, the potential for water quality problems and 6.4 Sediment tests can be used to develop exposure-
subsequent effects on oyster beds may dictate the use of response relationships for individual toxicants by spiking clean
sediment elutriate exposures with bivalve larvae (Guide E724). sediments with varying concentrations of a test chemical and
4.2.3 Establish an experimental design consistent with the determining the concentration that elicits the target response in
objectives of the sediment evaluation. The use of appropriate the test organism (Guide E1391). Sediment tests can also be
controls is particularly important for evaluating sediment designed to determine the effects that the physical and chemi-
contamination (see Section 11). cal properties of sediments have on the bioavailability and
4.2.4 Determine which statistical procedures should be toxicity of compounds.
applied to analysis of the data, and define the limits of
6.5 Sediment tests can provide valuable information for
applicability of the resultant analyses in data interpretation
making decisions regarding the management of contaminated
(Test Method E1706).
sediments from hazardous waste sites and other contaminated
areas. Biological tests with sediments can also be used to make
5. Summary of Guide
defensible management decisions on the dredging and disposal
5.1 This guide provides general guidance and objectives for
of potentially contaminated sediments from rivers and harbors.
conducting biological tests with sediments. Detailed technical
((7, 8), Test Method E1706.)
information on the conduct and evaluation of specific sediment
tests is included in other documents referenced in this guide.
7. Hazards
5.2 Neither this guide nor any specific test methodology can
7.1 General Precautions:
adequately address the multitude of technical factors that must
7.1.1 Development and maintenance of an effective health
be considered when designing and conducting a specific
and safety program in the laboratory requires an ongoing
investigation. The intended use of this document is therefore
commitment by laboratory management and includes: (1) the
not to provide detailed guidance, but rather to assist the
appointment of a laboratory health and safety officer with the
investigator in developing technically sound and environmen-
responsibility and authority to develop and maintain a safety
tally relevant biological tests that adequately address the
program, (2) the preparation of a formal, written health and
questions being posed by a specific investigation.
safety plan, which is provided to each laboratory staff member,
6. Significance and Use
(3) an ongoing training program on laboratory safety, and (4)
regular safety inspections.
6.1 Contaminated sediments may affect natural populations
of aquatic organisms adversely. Sediment-dwelling organisms 7.1.2 Collection and use of sediments may involve substan-
may be exposed directly to contaminants by the ingestion of tial risk to personal safety and health. Chemicals in field-
sediments and by the uptake of sediment-associated contami- collected sediment may include carcinogenics, mutagens, and
nants from interstitial and overlying water. Contaminated other potentially toxic compounds. Inasmuch as sediment
sediments may affect water column species directly by serving testing is often started before chemical analysis can be
as a source of contaminants to overlying waters or a sink for completed, worker contact with sediment needs to be mini-
contaminants from overlying waters. Organisms may also be mized by (1) using gloves, laboratory coats, safety glasses, face
affected when contaminated sediments are suspended in the shields and respirators as appropriate, (2) manipulating sedi-
water column by natural or human activities. Water column ments under a ventilated hood or in an enclosed glove box, and
species and nonaquatic species may also be affected indirectly (3) enclosing and ventilating the exposure system. Personal
by contaminated sediments by the transfer of contaminants collecting sediment samples and conducting tests should take
through ecosystems (7, 8). all safety precautions necessary for the prevention of bodily
injury and illness which might result from ingestion or invasion
6.2 The procedures described in this guide may be used and
of infectious agents, inhaltion or absorption of corrosive or
adapted for incorporation in basic and applied research to
toxic substances through skin contact, and asphixiation be-
determine the ecological effects of contaminated sediments.
cause of lack of oxygen or precense of noxious gases.
These same methods may also be used in the development and
7.1.3 Before beginning sample collection and laboratory
implementation of monitoring and regulatory programs de-
work, personnel should determine that all the required safety
signed to prevent and manage sediment contamination.
equipment and materials have been obtained and are in good
6.3 Sediment tests with aquatic organisms can be used to
condition.
quantify the acute and chronic toxicity and the bioavailability
7.2 Safety Equipment:
of new and presently used materials. Sediment toxicity may
also result from environmental processes such as ammonia 7.2.1 Personal Safety Gear—Personnel should use safety
generation, pH shifts, or dissolved oxygen fluctuation. In many equipment, such as, rubber aprons, laboratory coats,
cases, consideration of the adverse effects of sediment- respirators, gloves, safety glasses, face shields, hard hats, and
associated contaminants is only one part of a complete hazard safety shoes. Before beginning sample collection and labora-
assessment of manufactured compounds that are applied di- tory work, personnel should be properly trained in the follow-
rectly to the environment (for example, pesticides) and those ing: (1) when and what personal protective equipment (PPE) is
released (for example, through wastewater effluents) as by- necessary, (2) How to properly wear PPE, (3) limitations to the
products from the manufacturing process or from municipali- PPE, and proper care maintenance, useful life, and (4) disposal
ties (7). of PPE (29 CFR 1910.132(f) ).
E1525 − 02 (2023)
7.2.2 Laboratory Safety Equipment—Each laboratory pertinent regulations applicable in the State in which you are
should be provided with safety equipment such as first-aid kits, operating. Refer to (11) for the citations of the Federal
fire extinguishers, fire blankets, emergency showers, and eye requirements.
wash stations. Mobile laboratories should be equipped with a
8. Sediment Test Types
telephone to enable personnel to summon help in case of
8.1 Many methods for assessing the toxicity of saltwater
emergency.
and freshwater sediments to benthic organisms have been
7.3 General Laboratory and Field Operations:
reported. Those methods are provided in Table 1 for saltwater
7.3.1 Special handling and precautionary guidance in Ma-
tests and in Table 2, for freshwater tests, respectively.
terial Safety Data Sheets (MSDS) should be followed for
8.2 The selection of a specific toxicity test type is intimately
reagents and other chemicals purchased from supply houses.
related to the objectives of the sediment evaluation program.
7.3.2 Work with some sediments may require compliance
These assessments, whether they be for monitoring, regulatory,
with rules pertaining to the handling of hazardous material.
or research purposes, should be guided by a set of null
Personnel collecting samples and performing tests should not
hypotheses that define the appropriate exposure route and the
work alone.
endpoint of interest.
7.3.3 It is adviseable to wash the exposed parts of the body
8.3 Organism exposure methods most commonly employ
with bacterial soap and water immediately after collecting or
the whole sediment in the bedded phase (solid phase), but pore
manipulating sediment samples.
water, suspended and elutriate phase exposures have also been
7.3.4 Strong acids and volatile organic solvents should be
used (7).
used in a fume hood or under an exhaust canopy over the work
8.4 Programs seeking to characterize or rank sediments on a
area.
basin-wide or regional scale typically use whole sediment,
7.3.5 An acidic solution should not be mixed with a
solid-phase exposures. Regulatory or permitting programs for
hypochlorite solution because hazardous fumes might be
dredged material disposal at a containment site may also
produced.
evaluate this exposure route (8, 12). Disposal at a dispersive
7.3.6 To prepare and dilute acid solutions, concentrated acid site, or concerns over the resuspension and transport of
should be added to water, not vise versa. Opening a bottle of in-place sediments, would suggest the use of suspended phase
concentrated acid and adding concentrated acid to water should or elutriate exposures (Annex A1).
be preformed only under a fume hood.
8.5 Methods have been developed to isolate and test the
7.3.7 Use of ground-fault systems and leak detectors is
toxicity of elutriates (99) or sediment interstitial water (100) to
strongly recommended to help prevent electrical shocks. Elec-
aquatic organisms. The elutriate test was developed for assess-
trical equipment or extension cords not bearing the approval of
ing the potential acute effects of open-water disposal of
Underwriter Laboratories should not be used. Ground-Fault
dredged material. Tests with elutriate samples are used to
interrupters should be installed in all “wet” laboratories where
estimate the water-soluble constituents that may be released
electrical equipment is used.
from sediment to the water column during disposal operations
(101). Toxicity tests of the elutriate with water column organ-
7.3.8 All containers should be adequately labeled to indicate
isms have generally indicated that little toxicity is associated
their contents.
with the discharge material (4). However, elutriates have been
7.3.9 A clean well-organized work place contributes to
reportedly more toxic than interstitial water samples (102).
safety and reliable results.
8.5.1 For many benthic invertebrates, the toxicity and bio-
7.4 Disease Prevention—Personnel handling samples which
accumulation of sediment-associated contaminants, such as
are known or suspected to contain human wastes should be
metals and non-ionic organic contaminants, may be correlated
immunized against hepatitis B, tetanus, typhoid fever and
with the concentration of these chemicals in the interstitial
polio. Thorough washing of exposed skin with bacterial soap
water (100, 103). The sediment interstitial water toxicity test
should follow handling of samples collected in the field.
was developed for assessing the potential in situ effects of
contaminated sediment on aquatic organisms. Once the inter-
7.5 Safety Manuals—For further guidance on safe practices
stitial water (or elutriate) has been isolated from the whole
when handling sediment samples and conducting toxicity tests,
sediment, the toxicity testing procedures are similar to effluent
check with the permittee and consult general industrial safety
toxicity testing with non-benthic species. If benthic species are
manuals including (9, 10).
used as test animals, they may be stressed by the absence of
7.6 Pollution Prevention, Waste Management and Sample
sediment (4).
Disposal—Guidelines for the handling and disposal of hazard-
8.5.2 The examination of organic extracts may have specific
ous material should be strictly followed (Guide D4447). The
uses. However, caution should be exercised in the use of
Federal Government has published regulations for the manage-
organic extracts since the availability of sediment contaminants
ment of hazardous waste and has given the States the option of
to organisms may have been altered (7).
either adopting those regulations or developing their own. If
9. Biological Responses
States develop their own regulations they are required to be as
stringent as the Federal regulations. As a handler of hazardous 9.1 Toxicity endpoints in sediment tests range from
materials, it is your responsibility to know and comply with the lethality, growth, reproductive impairment, and physiological
E1525 − 02 (2023)
TABLE 1 Organisms Used in Assessing the Toxicity of Saltwater TABLE 2 Organisms Used in Assessing the Toxicity of
A A
Sediments Freshwater Sediments
Taxa Exposure Reference Taxa Exposure Reference
Mortality Mortality
B
Amphipods So (12,13, 14, 15-20), Guide Amphipods El (70)
E1367 So (5,6,8,71,70-73) Test Method
C
Su (21, 20-24) E1706
Bivalves So (16, 20) Guide E724 Cladocerans El (70)
Su (20,25,26) So (5,70,72,74-84) Test Method
Copepods So (15) E1706
Su (15) Su (82)
Crab Su (26) El (70)
Cumaceans So (14, 17-19 So (70,72, 74-77)
D
Fish El (27,28) Insect larvae El (70)
So (20,29) So (5,8,85, 70-81, 86) Test Method
Su (20,25) E1706
Isopods So (74-77)
Isopods So (15)
Su (15) Oligochaetes So (87-89) Guide E1688
Lobster Su (26) Growth/reproduction
Mysids So (20) Amphipods So (5,6,71) Test Method E1706
Su (20-24) Bacteria El (90)
Polychaetes So (16,30,31) Guide E1611 So (90)
Phytoplankton El (32) Cladocerans El (90) Test Method E1706
Shrimp So (15, 30-34) So (5,90)
Su (15,28,33,34) Fish El (90)
So (90)
Tunicate Su (26)
Avoidance/behavior Insect larvae So (85,86,91,92) Test Method
E1706
Amphipods So (35,36)
Bivalves So (35,37,38-40) Nematodes El (93)
Crab So (35,36) Physiology
Echinoderm So (35) Oligochaetes El (94,95)
Fish So (37,41) Genetic damage
Fish El (2,57,58,94,95)
Lobster So (35)
Polychaetes So (37,42) Nematodes El (93)
Bacterial activity
Shrimp So (35,37)
Growth/reproduction/life cycle Bacteria El (96,97)
Amphipods Su (24) Behavior
Bivalves Su (43) Guide E724 Oligochaetes So (98)
Copepods So (44)
A
Many of these species have a salinity tolerance and therefore may be suitable for
Fish Su (45)
testing estuarine sediments.
Mysids Su (22,23,46)
Nematodes So (47)
Polychaetes So (45,48,49) Guide E1611
Su (45,48,49)
Sea urchin El (50) Guide E1563
Pathology
responses to alterations in community levels of organization
Amphipods So (51)
Su (51) (Table 1 and Table 2). Selection of the proper toxic endpoint is
Bivalves So (51)
predicated largely on the objectives of the evaluation program
Su (51)
and the available resources, time, and available methods.
Fish So (27,52,53)
Su (52) Several endpoints are suggested in published methods to
Oyster So (52)
measure the potential effects of contaminants in sediment
Su (52)
including, survival, growth, behavior, or reproduction;
Polychaetes So (51)
Su (51) however, survival of test organisms in 10–d exposures is the
Physiology
endpoint most commonly reported (Table 1 and Table 2). These
Fish Su (54)
short-term exposures which only measure effects on survival
Oligochaetes El (55)
Polychaetes So (48) can be used to identify high levels of contamination on
Su (48,56)
sediments, but may not be able to identify moderate levels of
Shrimp Su (56)
contamination in sediments (Test Method E1706, (8)). Sub-
Chromosome damage
Fish El (57-59) lethal endpoints in sediment tests might also prove to be better
Polychaetes Su (60)
estimates of reponses if benthic communities to contaminates
Bacterial activity
in the field (85-106).
Bacteria El (61,62)
Community recolonization
9.2 The decision to conduct short-term or long-term toxicity
Macrobenthos So (63-69)
tests depends on the goal of the assessment. In some instances,
A
Many of these species have a wide salinity tolerance and therefore may be
sufficient information may be gained by measuring sublethal
suitable for testing estuarine sediments.
B
So—solid-phase sediment exposure.
endpoints in 10-d tests. In other instances, the 10-d test could
C
Su—suspended sediment exposure.
be used to screen samples for toxicity before long-term tests
D
El—elutriate, extract, pore water exposure.
are conducted. While the long-term tests are needed to deter-
mine direct effects on reproduction, measurement of growth in
these toxicity tests may serve as an indirect estimate of
E1525 − 02 (2023)
reproductive effects of contaminates associated with sediments a greater concern relative to bioaccumulation potential. With
(Test Method E1706, (8)). the exception of some saltwater amphipods, few test species
have broad sediment toxicity databases. Additionally, many
9.3 Use of sublethal endpoints for assessment of contami-
species can be maintained in the laboratory long enough for
nate risk is not unique to toxicity testing with sediments.
acclimation to test conditions, but very few are cultured easily.
Numerous regulatory programs require the use of sublethal
Widespread toxicity testing will require cultured organisms or
endpoints in the decision-making process (7) including: (1)
the use of standard source populations that can be transported
Water Quality Criteria (and State Standards), (2) National
without experiencing excessive stress.
Pollution Discharge Elimination System (NPDES) effluent
monitoring (including chemical-specific limits and sublethal
10.3 Toxicity is related to the species-specific physiological
endpoints in toxicity tests); (3) Federal Insecticide, Rodenti-
and biochemical response to a toxicant and the degree of
cide and Fungicide Act (FIFRA) and the Toxic Substances
contact between the sediment and the organism. Feeding
Control Act (TSCA, tiered assessment includes several sub-
habits, including the type of food and feeding rate, will
lethal endpoints with fish and aquatic invertebrates); (4)
influence the exposure of contaminants from sediment (108).
Superfund (Comprehensive Environmental Response, Com-
Infaunal deposit-feeding species can receive an exposure of
pensation and Liability Act, CERCLA); (5) Organization of
sediment contaminants by means of three exposure routes:
Economic Cooperation and Development (OECD, sublethal
interstitial water, sediment particles, and overlying water.
toxicity testing with fish an invertebrates); (6) European
Benthic invertebrates may selectively consume particles with
Economic Community (EC, sublethal toxicity testing with fish
higher organic carbon and higher contaminant concentrations.
and invertebrates); and (7) the Paris Commission, (behavioral
Organisms in direct contact with sediment may also accumu-
endpoints).
late contaminants by direct adsorption to the body wall or
exoskeleton, or by absorption through the integument (109).
10. Test Organisms
Estimates of bioavailability will thus be more complex for
10.1 Once the exposure routes and endpoints of interest
epibenthic animals that inhabit both the sediment and the water
have been established, several criteria should be considered
column. Some benthic species are exposed primarily by detrital
when selecting appropriate species (3, 8, 107 ) and Test
feeding (110). Detrital feeders may not receive most of their
Method E1706 for which tests can be conducted that have
body burden directly from interstitial water. For certain higher
ecologically relevant endpoints. Ideally, the test species should
Kow compounds, uptake by the gut can exceed uptake across
meet the following criteria:
the gill (111, 112). However, for many benthic invertebrates,
10.1.1 Have a toxicological (sediment) database demon-
the toxicity and bioaccumulation of sediment-associated con-
strating sensitivity to a range of contaminants or the contami-
taminants such as metals, kepone, fluoranthene, and organo-
nant of interest, and be taxonomically identified;
chlorines are highly correlated with the concentration of these
10.1.2 Be readily available through field collection or cul-
chemicals in the interstitial water (100).
ture;
10.1.3 Be easily maintained in the laboratory;
10.4 The saltwater test species include a broad spectrum of
10.1.4 Be ecologically or economically important;
taxa and feeding types including crustaceans, bivalves,
10.1.5 Have a broad geographical distribution, or be indig-
polychaetes, and fish (Table 1). Tests using amphipods have
enous to the site being evaluated or have a similar niche, be in
received a great deal of attention because of their overall
the same feeding guild, or be similar in behavior to an
sensitivity and because they are often absent from contami-
inhabitant (species);
nated sites (13). This sensitivity has led to the development of
10.1.6 Be tolerant to a broad range of sediment physico-
routine methods using the burrowing amphipod Rheopoxynius
chemical characteristics (for example, organic carbon and
abronius. This 10-day acute toxicity test has recently been
grain size);
adapted for use with other amphipod species and has been
10.1.7 Be compatible with selected exposures and end-
established (Guide E1367, (14,12)). Since 1977, the U.S. Army
points; and
Corps of Engineers dredging permit program has routinely
10.1.8 Be tolerant of a range of different water quality
required tests with three species: a bivalve, a polychaete, and a
characteristics.
fish or shrimp, incorporating both species that burrow into the
10.2 Of these criteria, demonstrated sensitivity to
sediment and those which inhabit the water column. Broad
contaminants, ecological relevance, and tolerance to varying
applications of these protocols reveal that these tests are not as
sediment physico-chemical characteristics are the most impor-
sensitive as those with amphipods, and the latter have recently
tant. The sensitivity of a species to contaminants should be
been recommended for permit programs.
balanced with the concept of discrimination. Species responses
10.5 Freshwater sediment tests use a number of different
may need to provide discrimination between different levels of
species, including amphipods, midges, mayflies, cladocerans,
contamination. Additionally, insensitive species may be pre-
and oligochaetes (Table 2). Whole sediment tests with the
ferred for determining bioaccumulation potential. The use of
amphipod Hyalella azteca generally start with juvenile animals
indigenous species that are ecologically important and col-
lected easily is often very straightforward; however, many and are Typically conducted for 10 to 14–d with measurement
of survival or growth (Test Method E1706 , (8,71)). Methods
indigenous species at a contaminated site may be insensitive to
contaminants (Guide E1688). Indigenous species might present for conducting 42-d tests with H. azteca have been described in
E1525 − 02 (2023)
Test Method E1706 and (8). Endpoints measured in these be the Rheopoxynius abronius survival 10-day acute test, and
long-term tests with H. azteca include survival, growth, and for freshwater tests it should be Hyalella azteca survival and
reproduction. growth in 28-d exposures (122). While chronic tests with
whole sediments have been described for a variety of freshwa-
10.6 Tests with midge Chironomus tentans are generally
ter tests, research is ongoing to describe chronic tests with
started with second instar larvae (10 to 14 days old) and
marine amphipods.
continued for 10 to 17 days until the fourth instar; larval
survival or growth is the measure of toxicity (Test Method 10.10 Multispecies and microcosm tests can also be used to
E1706 (8, 85)). Methods for conducting 60–d tests with C. evaluate potential ecosystem responses to contaminated sedi-
tentans have been described in Test Method E1706 and (8). ments. The use of multi-species tests may provide toxicity
Exposures start with first instar C. tentans and endpoints information not available from single-species tests since rela-
measured in these long-term tests include survival, growth, tive species sensitivity may vary among contaminants (6).
emergence, reporduction, and egg hatching. Whole sediment However, results from multi-species or microcosm tests are
testing procedures with the midge C. riparius are started with more difficult to interpret due to interactions and limited
1 to 3-day-old larvae and may continue through pupation and reference literature (123, 124).
adult emergence ((6) Test Method E1706). Midge exposures
started with older larvae may underestimate midge sensitivity 11. Experimental Design Considerations
to toxicants. For instance, first instar C. tentans larvae were 6
11.1 Sampling Methods:
to 27 times more sensitive than fourth instar larvae to acute
11.1.1 Sampling methods are dependent on the purpose and
copper exposure (5, 113), and first instar C. riparius larvae
design of the study. The probable source and type of contami-
were 127 times more sensitive than second instar larvae to
nation and the objectives of the study should be evaluated
acute cadmium exposure (114).
before developing a sediment sampling regime. The number
10.7 Sediment toxicity tests with mayflies and cladocerans
and type of samples taken depends on the objectives of the
are generally conducted for up to 10 days (5, 115, 116) and Test
study (125-128).
Method E1706. Survival and molting frequency are the toxicity
11.1.2 The number of replicate samples taken at a site
endpoints monitored in the mayfly tests, and survival, growth,
should be determined based on the objectives of the study and
and reproduction are monitored in the cladoceran tests. While
a preliminary survey of sediment variability at the site.
cladocerans are not in direct contact with the sediment, they are
Information from the preliminary survey and the objectives of
frequently in contact with the sediment surface and are
the study can be used to determine the minimum number of
probably exposed to both water-soluble and particulate bound
replicates that should be sampled at each site (126, 127).
contaminants in the overlying water and surface sediment (Test
11.1.3 In general, both toxicity and bioaccumulation tests
Method E1706). Cladocerans are also one of the more sensitive
require at least two exposures: a control and one or more test
groups of species used in aquatic toxicity testing.
treatments (see 11.3.12). The experimental unit for each test is
the exposure chamber. A sediment sample is typically split into
10.8 The most frequently described sediment testing proce-
four or more test chambers. Individual observations obtained
dures for oligochaetes are acute toxicity testing methods (98 ,
from within an individual chamber should not be used as
8) also see, Guide E1688. However, methods for conducting up
replicate observations. Replicate chambers for a particular
to 500-day oligochaete exposures, with growth and reproduc-
sediment provide an estimate of the variability within the test
tion as the toxicity endpoints, have been described (117). A
system and are not considered sediment sample or location
shorter 28-d test starting with sexually mature Tubifex tubifex
replicates.
has been described (118). Effects on growth and reproduction
11.1.4 There are several acceptable methods of sampling
are monitored in this shorter test, and the duration of the
sediments, for example, corers and grabs or dredges. Grabs or
exposure makes the test more useful for routine sediment
dredges (for example, Ponar or Ekman) are appropriate when
toxicity assessments with oligochaetes (Test Method E1706).
sediments are known to be unstratified with respect to the
Many oligochaetes have complex life cycles and reproductive
contaminants of concern. If the contaminants are in strata, or if
strategies, and therefore laboratory culturing requirements
their accumulation rates are of interest, one of several core
have prohibited their use in toxicity testing (119). However,
210 137
samplers should be used. Pb or Cs dating can be
culturing procedures have been described for Lumbriculus
performed on cores to identify the thickness of the mixed layer
variegatus and Tubifex tubifex (8, 120,121) (See also, Test
Method E1706 and Guide E1688). (125, 128). See Guide E1391 for additional details.
11.2 Sample Handling:
10.9 Because of the database that has been developed with
existing tests, it is recommended that, for whole sediment 11.2.1 Sample handling and preservation are discussed in
exposures, either phoxocephalid, ampeliscid, or haustoriid Guide E1391 and Test Method E1706, and depend on the type
amphipods be used in saltwater tests. For freshwater of chemical characterization that will be performed. Any
applications, hyalellid amphipods, midge larvae, or mayfly sediment disturbance may alter the chemical characterization
larvae would be appropriate. As new methods are developed, it of that sediment from in situ conditions. The use of clean
will be important to establish the sensitivity of each method sampling devices and sample containers is essential to ensure
relative to a benchmark procedure for comparative purposes the accurate determination of sediment contamination (126,
(2). The whole sediment benchmark for saltwater tests should 128).
E1525 − 02 (2023)
11.2.2 Physical and chemical characterization of sediments often if (1) such measurements have not been determined
is highly dependent on the needs of the investigator, but it may semiannually for at least two years or (2) surface water is used.
include loss on ignition, percent water, grain size, total organic
11.3.3 A natural overlying water is considered to be of
carbon, total phosphorus, nitrogen forms, trace metals and
uniform quality if the monthly ranges of hardness and alkalin-
organic compounds, pH, total volatile solids, biological oxygen
ity are less than 5 mg/L or 10 % of their respective averages,
demand, chemical oxygen demand, cation exchange capacity,
whichever is higher, and if the monthly range of pH is less than
Eh, pE, total inorganic carbon, acid volatile sulfides, and
0.4 units. Natural overlying waters should be obtained from an
ammonia (125, 127, 128). Many times, a sediment of concern
uncontaminated well or spring, if possible, or from a surface
has some historical data that are used as a basis for selection.
water source. If surface water is used, the intake should be
11.2.3 Indigenous organisms may be present in field-
positioned to minimize fluctuations in quality and the possi-
collected sediments. An abundance of the same organism or
bility of contamination and maximize the concentration of
organisms taxonomically similar to the test organism in the
dissolved oxygen and to help ensure low concentrations of
sediment sample may make interpretation of treatment effects
sulfide and iron. For sediment studies with saltwater, the range
difficult. Previous investigators have inhibited the biological
of salinity should be less than 10 % of the average. In addition,
activity of sediment with sieving, heat, mercuric chloride,
the ion concentrations of the water should be within 10 % of
antibiotics, or gamma irradiation. (Guide E1391.) However,
the ion concentrations (adjusted for the salinity) listed in Guide
further research is needed to determine effects on contaminate
E729. Chlorinated water should not be used for, or in the
bioavailability or other modifications of sediments from treat-
preparation of, overlying water because residual chlorine and
ments such as those used to remove or destroy indigenous
chlorine-produced oxidants are toxic to many aquatic animals
organisms.
and dechlorination is often incomplete.
11.2.4 Field-collected sediment samples tend to settle dur-
11.3.4 For certain applications, the experimental design
ing shipment. As a result, water above the sediment should not
might require the use of water from the test sediment collection
be discarded, but should be mixed back into the sediment
site.
during homogenization (Test Method E1706). Sediment
samples should not be routinely sieved to remove indigenous
11.3.5 Reconstituted fresh and salt water is prepared by
organisms unless there is a good reason to believe they will
adding specified amounts of reagent grade chemicals to high-
influence the response of the test organisms. Large indigenous
quality distilled or deionized water (see Guide E729 and Test
organisms and large debris can be removed using forceps.
Method E1706). Acceptable water can be prepared using
Reynoldson et al. (129), observed reduced growth of
deionization, distillation, or reverse-osmosis units.
amphipods, midges, and mayflies in sediments with elevated
Conductivity, pH, hardness, and alkalinity should be measured
numbers of oligochaetes and recommended sieving sediments
on each batch of reconstituted water. If the water is prepared
suspected to have high numbers of indigenous oligochaetes.
from a surface water, the total organic carbon or chemical
One approach might be to sieve an aliquot of each sediment
oxygen demand should be measured on each batch. Filtration
before the start of a test. If potential predators are recovered
through sand, rock, bag, or depth-type cartridge filters may be
from a sediment, it may be desirable to sieve all of that sample
used to keep the concentration of particulate matter acceptably
before the start of the test. Depending on the objective of the
low. The reconstituted water should be intensively aerated
test, it may be necessary to sieve all sediments or run a sieved
before use, except that buffered soft fresh waters should be
and un-sieved treatment in parallel to account for potential
aerated before, but not after, the addition of buffers. Problems
affects of sieving on test results and subsequent comparisons.
have been encountered with some species in some fresh
The size of the sieve used will depend on the size of the
reconstituted waters, but these problems can be overcome by
organisms in the sediment sample. If a sediment must be
aging the reconstituted water for one or more weeks (Guide
sieved, it is desirable to analyze a sample before and after
E729).
sieving (for example, measure pore-water metals, dissolved
11.3.6 Materials used to construct test chambers may in-
organic carbon (DOC), acid volatile sulfide (AVS), total
organic carbon (TOC)) to document the influence of sieving on clude glass, stainless steel, silicone, plastics, and fiberglass that
sediment chemistry. have been prepared properly and tested for toxicity (Guides
E1367 and Test M
...