§ 797.1400 Fish acute toxicity test.
(a) Purpose. This guideline may be used to develop data on the acute toxicity of chemical substances and mixtures (“chemicals”) subject to environmental effects test regulations under the Toxic Substances Control Act (TSCA) (Pub. L. 94-469, 90 Stat. 2003, 15 U.S.C. 2601 et seq.). This guideline prescribes tests to be used to develop data on the acute toxicity of chemicals to fish. The United States Environmental Protection Agency (EPA) will use data from these tests in assessing the hazard of a chemical to the environment.
(b) Definitions. The definitions in section 3 of the Toxic Substances Control Act (TSCA), and the definitions in part 792—Good Laboratory Practice Standards of this chapter apply to this test guideline. The following definitions also apply to this guideline:
(1) Acclimation means the physiological compensation by test organisms to new environmental conditions (e.g., temperature, hardness, pH).
(2) Acute toxicity test means a method used to determine the concentration of a substance that produces a toxic effect on a specified percentage of test organisms in a short period of time (e.g., 96 hours). In this guideline, death is used as the measure of toxicity.
(3) Carrier means a solvent used to dissolve a test substance prior to delivery to the test chamber.
(4) Conditioning means the exposure of construction materials, test chambers, and testing apparatus to dilution water or to test solutions prior to the start of a test in order to minimize the sorption of the test substance onto the test facilities or the leaching of substances from the test facilities into the dilution water or test solution.
(5) Death means the lack of opercular movement by a test fish.
(6) Flow-through means a continuous or an intermittent passage of test solution or dilution water through a test chamber, or a holding or acclimation tank with no recycling.
(7) Incipient LC50 means that test substance concentration, calculated from experimentally-derived mortality data, that is lethal to 50 percent of a test population when exposure to the test substance is continued until the mean increase in mortality does not exceed 10 percent in any concentration over a 24-hour period.
(8) LC50 means that test substance concentration, calculated from experimentally-derived mortality data, that is lethal to 50 percent of a test population during continuous exposure over a specified period of time.
(9) Loading means the ratio of fish biomass (grams, wet weight) to the volume (liters) of test solution in a test chamber or passing through it in a 24-hour period.
(10) Static means the test solution is not renewed during the period of the test.
(11) Test solution means the test substance and the dilution water in which the test substance is dissolved or suspended.
(c) Test procedures—(1) Summary of the test. (i) Test chambers are filled with appropriate volumes of dilution water. If a flow-through test is performed, the flow of dilution water through each chamber is adjusted to the rate desired.
(ii) The test substance is introduced into each test chamber. In a flow-through test, the amount of test substance which is added to the dilution water is adjusted to establish and maintain the desired concentration of test substance in each test chamber.
(iii) Test fish which have been acclimated in accordance with the test design are introduced into the test and control chambers by stratified random assignment.
(iv) Fish in the test and control chambers are observed periodically during the test; dead fish are removed at least twice each day and the findings are recorded.
(v) The dissolved oxygen concentration, pH, temperature and the concentration of test substance are measured at intervals in selected test chambers.
(vi) Concentration-response curves and LC50 values for the test substance are developed from the mortality data collected during the test.
(2) [Reserved]
(3) Range finding test. If the toxicity of the test substance is not already known, a range finding test should be performed to determine the range of concentrations to be used in the definitive test. The highest concentration of test substance for use in the range finding test should not exceed its solubility in water or the permissible amount of the carrier used.
(4) Definitive test. (i) A minimum of 20 fish should be exposed to each of five or more test substance concentrations. The range of concentrations to which the fish are exposed should be such that in 96 hours there are at least two partial mortality exposures bracketing 50 percent survival.