This International Standard specifies a method for the determination of the chronic toxicity to rotifer Brachionus calyciflorus, based on population growth inhibition in 48 h. The method is applicable to: a) chemical substances which are soluble or which can be maintained as stable suspensions or dispersions under the conditions of the test; b) industrial or sewage effluents, treated or untreated, if appropriate after decantation, filtration or centrifugation; c) fresh waters; d) aqueous extracts. This International Standard is not applicable to the testing of unstable chemicals (hydrolysing, absorbing, etc.) in water unless exposure concentration is measured, nor to the testing of aquatic samples from the estuarine or marine environment