EPA Method 2021

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EPA Method 2021:
Acute Toxicity Tests with Effluents and Receiving Waters using Daphnia. Official Name: Test Method 2021.0, Daphnia pulex and D. magna Acute Toxicity Tests with Effluents and Receiving Waters

Test type can be either static-renewal, static non-renewal, or flow-through. The selection of test type will depend on the NPDES permit requirements, the objectives of the test, the available resources, the requirements of the test organisms, and effluent characteristics such as fluctuations in effluent toxicity. Effluent acute toxicity is generally measured using a multi-concentration, or definitive test, consisting of a control and a minimum of five effluent concentrations.

This manual describes acute toxicity tests for use in the National Pollutant Discharge Elimination System (NPDES) Permits Program to identify effluents and receiving waters containing toxic materials in acutely toxic concentrations. With the exception of the Holmesimysis costata Acute test, the methods included in this manual are referenced in Table 1A, 40 CFR Part 136 regulations and, therefore, constitute approved methods for acute toxicity tests. They are also suitable for determining the toxicity of specific compounds contained in discharges. The tests may be conducted in a central laboratory or on-site, by the regulatory agency or the permittee.

Methods for Measuring the Acute Toxicity of Effluents and Receiving Waters to Freshwater and Marine Organisms. Fifth Edition, October 2002. EPA Volume 821-R-02-012

None provided

QC Requirements:
Reference toxicant testing with control charts for each combination of toxicant, test species, test condition, and endpoint.

Maximum Holding Time:


Acute Toxicity

Effluents: 5 and a control Receiving waters: 100% receiving water and a control (recommended)

Sample Prep:
Filtration, dechlorination

It is a laboratory's responsibility to demonstrate its ability to obtain consistent, precise results with reference toxicants before it performs toxicity tests with effluents for permit compliance purposes. To meet this requirement, the intra-laboratory precision, expressed as percent coefficient of variation, of each type of test to be used in a laboratory should be determined by performing five or more tests with different batches of test organisms using the same reference toxicant, at the same concentrations, with the same test conditions, and the same data analysis methods.


Revision Number:
Fifth Edition, October 2002