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l~~D 1\~TM ~~l~ Designation: E 729 - 80 4lo~1 AMERICAN SOCIETY FOR TESTING AND MATERIALS 1916 Race St., Philadelphia, Pa. 19103 Reprinted from the Annual Book of ASTM Standards, Copyright ASTM If not listed in the current combined index, will appear in the next edition. Standard Practice for CONDUCTING ACUTE TOXICITY TESTS WITH FISHES , MACROINVERTEBRA TES, AND AMPHIBIANS' This standard is issued under the fixed designation E 729; the number immediately following the designation indicates the year of ongmal adoptIOn or, III the case of reVlSlon, the year of last revision. A number in parentheses indicates the year of last reapproval. 1. Scope 1.1 This practice (1)2 describes procedures for obtaining information concerning the acute toxicity of chemical compounds, either individ- ually or in formulations, commercial products, or known mixtures, to freshwater and saltwater fishes, macroinvertebrates, and amphibians. Two tests are described: the static acute toxicity test and the flow-through acute toxicity test. 1.2 Selection of the test to be used in a specific situation will depend upon the needs of the investigator and upon available re- sources. Static tests provide the most easily obtained measure of toxicity, but should not last longer than 96 h. Flow-through tests may last longer than 96 h because the dissolved oxygen and toxicant concentrations and pH are maintained at desired levels and degradation and metabolic products are removed. Because the results of static tests on some kinds of chemicals have limited utility, flow-through tests should usually be conducted on toxicants that have a high biochemical oxygen demand, are highly volatile, are unstable in aqueous solution, are readily biodegradable, or are re- moved from test solutions in significant quan- tities by test organisms. Shell deposition tests with bivalve molluscs must always be flow- through tests. Although use of the recirculation and renewal techniques may be justified in some cases, the flow-through technique should generally be used when a technique other than the static is desired. 1.3 These procedures are applicable to nearly all chemical compounds. However, the results of tests on formulations and commercial products that are complex mixtures of sub- stances of diverse solubilities and toxicities may depend greatly on the procedure used to pre- pare the test solutions. With slight modification these procedures can be used to test other kinds of toxic agents, such as petroleum hydrocar- bons, temperature, and particulate matter. 1.4 In developing these procedures an at- tempt was made to balance scientific and prac- tical considerations and to ensure that the re- sults will be accurate and precise enough for the majority of situations in which they will be used. The requirements for acceptable static tests are less stringent than those for flow- through tests because of the inherent advan- tages ofthe latter technique (see 1.2). Although both tests can be improved by using more organisms, longer acclimation times, etc., the requirements presented herein should usually be sufficient. A decision to conduct one ofthese tests in a more stringent manner will usually be based more on personal perference than on a practical need for more precise results. 1.5 Using procedures that meet specific needs is more important than following pre- scribed procedures. However, either the static or the flow-through test presented herein should be applicable to most situations. When- ever toxicity tests. are conducted with aquatic organisms, these procedures should be followed as closely as possible. The results of all properly conducted tests are valid for the procedures 1 This practice is under the jurisdiction of ASTM Com- mittee E-35 on Pesticides and is the direct responsibility of Subcommittee E 35.23 on Safety to Aquatic Organisms. Current edition approved March 3, 1980. Published May 1980. 2 Boldface numbers in parentheses refer to the list of references at the end of this standard.