The Zahn-Wellens/EMPA Test which is used to determine inherent biodegradability.

A mixture containing the non-volatile and water soluble test substance, mineral nutrients and a relatively large amount of activated sludge in aqueous medium is agitated and aerated at 20-25ºC in the dark or in diffuse light, for up to 28 days.

Blank controls, containing activated sludge and mineral nutrients but no test substance, are run in parallel. The functional capability of the activated sludge is tested using a reference compound (ethylene glycol, diethylene glycol, lauryl sulfonate or aniline). In a typical run 1 or 2 vessels for the test suspension and for the inoculum blank, 1 for procedure control are used.

The biodegradation process is monitored by determination of DOC, Dissolved Organic Carbon, (or COD, Chemical Oxygen Demand) in filtered samples, taken at daily or other time intervals. It is mandatory to follow DOC in the test suspension and inoculum blanks in parallel.

The ratio of eliminated DOC (or COD), corrected for the blank, after each time interval, to the initial DOC value is expressed as the percentage biodegradation at the sampling time. The percentage biodegradation is plotted against time to give the biodegradation curve.

The test is considered valid if the procedural control shows the removal of the reference compound by at least 70% within 14d and if DOC (or COD) in the test suspension is removed relatively gradually over days or weeks, since this indicates biodegradation.

OECD (1992), Test No. 302B: Inherent Biodegradability: Zahn-Wellens/ EVPA Test, OECD Guidelines for the Testing of Chemicals, Section 3, OECD Publishing, Paris, https://doi.org/10.1787/9789264070387-en.