5.1 An acute effluent toxicity test is conducted to obtain information concerning the immediate effects on test organisms of a short-term exposure to an effluent under specific experimental conditions. One can directly examine acute effects of complex mixtures of chemicals as occurs in effluents and some ambient waters. Acute effluent toxicity tests can be used to evaluate the potential for designated-use or aquatic life imperiment in the receiving stream, lake, or estuary. An acute toxicity test does not provide information about whether delayed effects will occur, although a post-exposure observation period, with appropriate feeding if necessary, might provide such information.

5.2 Results of acute effluent tests might be used to predict acute effects likely to occur on aquatic organisms in field situations as a result of exposure under comparable conditions, except that (1) motile organisms might avoid exposure when possible, (2) toxicity to benthic species might be dependent on sorption or settling of components of the effluent onto the substrate, and (3) the effluent might physically or chemically interact with the receiving water.

5.3 Results of acute effluent tests might be used to compare the acute sensitivities of different species and the acute toxicities of different effluents, and to study the effects of various environmental factors on results of such tests.

5.4 Acute tests are usually the first step in evaluating the effects of an effluent on aquatic organisms.

5.5 Results of acute effluent tests will depend on the temperature, composition of the dilution water, condition of the test organisms, exposure technique, and other factors.

This guide covers procedures for obtaining laboratory data concerning the adverse effects of aqueous ambient samples and effluents on certain species of freshwater and saltwater fishes, macroinvertebrates, and amphibians, during a short-term exposure, depending on the species, using the static, renewal, and flow-through techniques. These procedures will probably be useful for conducting acute toxicity tests on aqueous effluents with many other aquatic species, although modifications might be necessary. Static tests might not be applicable to effluents that have a high oxygen demand, or contain materials that (1) are highly volatile, (2) are rapidly biologically or chemically transformed in aqueous solutions, or (3) are removed from test solutions in substantial quantities by the test chambers or organisms during the test. Results of acute toxicity tests should usually be reported in terms of a median lethal concentration (LC50) or median effective concentration (EC50). An acute toxicity test does not provide information about whether delayed effects will occur. Specified requirements involving the following are detailed: (1) hazards; (2) apparatus: facilities, special requirements, construction materials, metering system, test chambers, cleaning, and acceptability; (3) dilution water requirements, source, treatment, and characterization; (4) effluent sampling point, collection, preservation, treatment, and test concentrations; (5) test organism species, age, source, care and handling, feeding, disease treatment, holding, acclimation, and quality; (6) procedure: experimental design, dissolved oxygen, temperature, loading, beginning the test, feeding, duration of test, biological data, and other measurements; (7) analytical methodology; (8) acceptability of test; (9) calculation of results; and (1) report of results.

1.1 This guide covers procedures for obtaining laboratory data concerning the adverse effects of an aqueous effluent on certain species of freshwater and saltwater fishes, macroinvertebrates, and amphibians, usually during 2 to 4-day exposures, depending on the species, using the static, renewal, and flow-through techniques. These procedures will probably be useful for conducting acute toxicity tests on aqueous effluents with many other aquatic species, although modifications might be necessary.

1.2 Other modifications of these procedures might be justified by special needs or circumstances. Although using appropriate procedures is more important than following prescribed procedures, results of tests conducted using unusual procedures are not likely to be comparable to results of many other tests. Comparison of results obtained using modified and unmodified versions of these procedures might provide useful information concerning new concepts and procedures for conducting acute toxicity tests on aqueous effluents.

1.3 This guide is based in large part on Guide E729. The major differences between the two guides are (1) the maximum test concentration is 100 % effluent or ambient sample, (2) testing is not chemical specific, and (3) the holding time of effluent and ambient samples is often considerably less than that for chemicals and other test materials. Because the sample is often a complex mixture of chemicals, analytical tests cannot generally be used to confirm exposure concentrations.

1.4 Selection of the technique to be used in a specific situation will depend upon the needs of the investigator and upon available resources. Static tests provide the most easily obtained measure of acute toxicity, but should not last longer than 48 h. Renewal and flow-through tests may last longer than 48 h because the pH and concentrations of dissolved oxygen and effluent are maintained at desired levels and degradation and metabolic products are removed. Static tests might not be applicable to effluents that have a high oxygen demand, or contain materials that (1) are highly volatile, (2) are rapidly biologically or chemically transformed in aqueous solutions, or (3) are removed from test solutions in substantial quantities by the test chambers or organisms during the test. Flow-through tests are generally preferable to renewal tests, although in some situations a renewal test might be more cost-effective than a flow-through test.

1.5 In the development of these procedures, an attempt was made to balance scientific and practical considerations and to ensure that the results will be sufficiently accurate and precise for the applications for which they are commonly used. A major consideration was that the common uses of the results of acute tests on effluents do not require or justify stricter requirements than those set forth in this guide. Although the tests may be improved by using more organisms, longer acclimation times, and so forth, the requirements presented in this guide should usually be sufficient.

1.6 Results of acute toxicity tests should usually be reported in terms of a median lethal concentration (LC50) or median effective concentration (EC50). In some situations, it might be necessary only to determine whether a specific concentration is acutely toxic to the test species or whether the LC50 or EC50 is above or below a specific concentration.

1.7 This guide is arranged as follows:

Section

Referenced Documents

  2

Terminology

  3

Summary of Guide

  4

Significance and Use

  5

Hazards

  7

Apparatus

  6

 Facilities

  6.1

 Special Requirements

  6.2

 Construction Materials

  6.3

 Metering System

  6.4

 Test Chambers

  6.5

 Cleaning

  6.6

 Acceptability

  6.7

Dilution Water

  8

 Requirements

  8.1

 Source

  8.2

 Treatment

  8.3

 Characterization

  8.4

Effluent

  9

 Sampling Point

  9.1

 Collection

  9.2

 Preservation

  9.3

 Treatment

  9.4

 Test Concentration(s)

  9.5

Test Organisms

  10

 Species

  10.1

 Age

  10.2

 Source

  10.3

 Care and Handling

  10.4

 Feeding

  10.5

 Disease Treatment

  10.6

 Holding

  10.7

 Acclimation

  10.8

 Quality

  10.9

Procedure

  11

 Experimental Design

  11.1

 Dissolved Oxygen

  11.2

 Temperature

  11.3

 Loading

  11.4

 Beginning the Test

  11.5

 Feeding

  11.6

 Duration of Test

  11.7

 Biological Data

  11.8

 Other Measurements

  11.9

Analytical Methodology

  12

Acceptability of Test

  13

Calculation or Results

  14

Report

  15

1.8 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 and health practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 7.