§ 797.1300 Daphnid acute toxicity test.
(a) Purpose. This guideline is intended for use in developing 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 an acute toxicity test in which daphnids (Daphnia magna or D. pulex) are exposed to a chemical in static and flow-through systems. The United States Environmental Protection Agency will use data from this test in assessing the hazard a chemical may present in the aquatic environment.
(b) Definitions. The definitions in section 3 of the Toxic Substances Control Act (TSCA) and part 792—Good Laboratory Practice Standards of this chapter apply to this test guideline. In addition, the following definitions apply to this guideline:
(1) Brood stock means the animals which are cultured to produce test organisms through reproduction.
(2) EC50 means that experimentally derived concentration of test substance in dilution water that is calculated to affect 50 percent of a test population during continuous exposure over a specified period of time. In this guideline, the effect measured is immobilization.
(3) Ephippium means a resting egg which develops under the carapace in response to stress conditions in daphnids.
(4) Flow-through means a continuous or an intermittent passage of test solution or dilution water through a test chamber or culture tank with no recycling.
(5) Immobilization means the lack of movement by the test organisms except for minor activity of the appendages.
(6) Loading means the ratio of daphnid biomass (grams, wet weight) to the volume (liters) of test solution in a test chamber at a point in time, or passing through the test chamber during a specific interval.
(7) Static system means a test system in which the test solution and test organisms are placed in the test chamber and kept there for the duration of the test without renewal of the test solution.
(c) Test procedures—(1) Summary of the test. (i) Test chambers are filled with appropriate volumes of dilution water. In the flow-through test, the flow of dilution water through each chamber is adjusted to the rate desired. The test chemical is introduced into each treatment chamber. The addition of test chemical in the flow-through system is conducted at a rate which is sufficient to establish and maintain the desired concentration in the test chamber. The test is started within 30 minutes after the test chemical has been added and uniformly distributed in static test chambers or after the concentration of test chemical in each flow-through test chamber reaches the prescribed level and remains stable. At the initiation of the test, daphnids which have been cultured and acclimated in accordance with the test design are randomly placed into the test chambers. Daphnids in the test chambers are observed periodically during the test, the immobile daphnids removed, and the findings recorded.
(ii) Dissolved oxygen concentration, pH, temperature, the concentration of test chemical and other water quality parameters are measured at specified intervals in selected test chambers. Data are collected during the test to develop concentration-response curves and determine EC50 values for the test chemical.
(2) [Reserved]
(3) Range-finding test. (i) A range-finding test should be conducted to establish test solution concentrations for the definitive test.
(ii) The daphnids should be exposed to a series of widely spaced concentrations of the test chemical (e.g., 1, 10, 100 mg/1, etc.), usually under static conditions.
(iii) A minimum of five daphnids should be exposed to each concentration of test chemical for a period of 48 hours. The exposure period may be shortened if data suitable for the purpose of the range-finding test can be obtained in less time. No replicates are required and nominal concentrations of the chemical are acceptable.