OECD 202: Daphnia sp., acute immobilization test

The 48-hour acute daphnid immobilisation test according to OECD 22 assesses short-term aquatic toxicity by measuring immobilisation of daphnids, usually Daphnia magna, and using those observations to derive concentration–response values. The primary reported metric is the EC50 at 48 hours, with an optional EC50 at 24 hours calculated from dose–response analysis.

The protocol is the following:

• The test begins with neonates less than 24 hours old exposed for 48 hours to at least five concentrations plus a control, with a minimum of twenty animals per concentration typically arranged in replicate groups (for example, four replicates of five).

• Immobilisation is observed and recorded at 24 and 48 hours, and the numbers or proportions immobilised at each concentration and in the control are tabulated for statistical modelling.

• Test solutions are prepared by diluting the substance into acceptable test water, avoiding solvents or emulsifiers when possible. Analytical measurements of test substance concentration, dissolved oxygen, and pH are performed at the start and end of the exposure to document conditions.

• The biological endpoint is direct observation of loss of swimming activity consistent with immobilisation, EC50 values are derived from dose–response modelling and are normally reported in concentration units such as mg/L.

The report includes the counts or proportions immobilised per concentration and control at each time point, the calculated EC50(s), and supporting water chemistry and analytical concentration data.

The method is accepted under REACH for aquatic hazard characterization in registration dossiers submitted to ECHA, where OECD TG 202 is equivalent to EU Method C.2 under Commission Regulation (EC) No. 440/2008. Under the U.S. Toxic Substances Control Act (TSCA), EPA may require daphnid acute toxicity testing to OPPTS 850.1010/OECD 202 for new chemical substances expected to be acutely toxic.