User Guide to Thresholds and Classification - Environmental ...

329User Guide for Thresholds and Classificationsready biodegradability tests could be considered valid, irrespective of negative results, when the scientificquality is good and the test conditions are well documented, that is, guideline criteria are fulfilled, includingthe use of non-pre-exposed (non-adapted) inoculum. None of the various screening tests is suitable for thetesting of all types of substances, and results obtained by the use of a test procedure that is not suitable forthe specific substance should be evaluated carefully before a decision on the use is taken.Thus, there are a number of factors may explain conflicting biodegradability data from screening tests,including:inoculum;toxicity of test substance;test conditions;solubility of the test substance; andvolatilisation of the test substance.The suitability of the inoculum for degrading the test substance depends on the presence and amount ofcompetent degraders. When the inoculum is obtained from an environment that has previously beenexposed to the test substance, the inoculum may be adapted as evidenced by a degradation capacity, whichis greater than that of an inoculum from a non-exposed environment. As far as possible the inoculum mustbe sampled from an unexposed environment, but for substances that are used ubiquitously in high volumesand released widespread or more or less continuously, this may be difficult or impossible. When conflictingresults are obtained, the origin of the inoculum should be checked in order to clarify whether or notdifferences in the adaptation of the microbial community may be the reason.As mentioned above, many substances may be toxic or inhibitory to the inoculum at the relatively highconcentrations tested in ready biodegradability tests. Especially in the Modified MITI (I) test (OECD TestGuideline 301C) and the Manometric Respirometry test (OECD Test Guideline 301F) where highconcentrations (100 mg/L) are prescribed. The lowest test substance concentrations are prescribed in theClosed Bottle test (OECD Test Guideline 301D) where 2–10 mg/L is used. The possibility of toxic effectsmay be evaluated by including a toxicity control in the ready biodegradability test or by comparing the testconcentration with toxicity test data on micro-organisms, for example, the respiration inhibition tests (OECDTest Guideline 209), the nitrification inhibition test (ISO 9509) or, if other microbial toxicity tests are notavailable, the bioluminescence inhibition test (ISO 11348). When conflicting results are found, this may becaused by the toxicity of the test substance. If the substance is not inhibitory at environmentally realisticconcentrations, the greatest degradation measured in screening tests may be used as a basis forclassification. If simulation test data are available in such cases, consideration of these data may beespecially important, because a low non-inhibitory concentration of the substance may have been employed,thus giving a more reliable indication of the biodegradation half-life of the substance under environmentallyrealistic conditions.When the solubility of the test substance is lower than the concentrations employed in a test, this parametermay be the limiting factor for the actual degradation measured. In these cases, results from tests employingthe lowest concentrations of test substance should prevail, that is, often the Closed Bottle test (OECD TestJanuary 2012 EPA0109