This Test Guideline is designed to assess the effects of a substance on micro-organisms from activated sludge of waste-water treatment plants by measuring their respiration rate (carbon and/or ammonium oxidation) as oxygen consumption. The test results may also serve as an indicator of suitable non-inhibitory concentrations of test substances to be used in biodegradability tests. The test allows the determination of ECx and/or NOEC values of the test substance. The inhibition of three different oxygen uptakes may be determined, i.e. total, heterotrophic only, and that due to nitrification in the absence and presence of N-allylthiourea, a specific nitrification inhibitor. For obtaining both NOEC and ECx, six controls and five treatment concentrations in a geometric series with five replicates are recommended. In each test vessel, test mixtures containing water, synthetic sewage feed and the test substance are incubated at the pH of 7.5 ±0.5 and the temperature within 20±2°C under forced aeration to keep the dissolved oxygen concentration above 60-70% saturation. The oxygen consumption is measured after 3 hours of exposure and additional measurements at 30 minutes of exposure can be performed in the case that the test substance is rapidly degraded.