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Determination of the inhibition of the gas production of anaerobic bacteria

OECD 224 respectively Council Regulation (EC) No 440/2008, C.34 and ISO 13641-1


Assessing the potential toxicity of a test material by determining the production of biogas from the anaerobic digestion of sewage sludge.


Aliquots of a mixture of anaerobically digesting sludge and a degradable substrate solution are incubated alone and simultaneously with a range of concentrations of the test material in sealed vessels for up to 3 days. The amount of gas (methane plus carbon dioxide) produced is measured by the increase in pressure in the bottles. The percentage inhibition of gas production is calculated from the amounts produced in the respective test and control bottles. The EC50 and other effective concentrations are calculated from plots of percentage inhibition against the concentration of the test chemicals or its logarithm.

Test substance properties

The test is applicable to substances which are soluble or insoluble in water, including volatile substances, as well as wastewaters, sludges or other environmental samples.