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Environmental fate & pathways

Biodegradation in water: screening tests

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biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2006-06-14 to 2006-06-21
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)
Version / remarks:
according to guideline
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Version / remarks:
GLP compliance:
yes (incl. QA statement)
Oxygen conditions:
Inoculum or test system:
sewage, predominantly domestic, non-adapted
Details on inoculum:
aeration tank of a wastewater plant treating predominantly domestic sewage (Wupper area water authority, STP Odenthal)
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
- Test temperature: 22 °C
- pH: 7.4
- pH adjusted: no

Pre-treatment of the test item
25 mg test item were weighed out on aluminium foil. This substance inclusive the aluminium foil was added to the test vessels to give a test concentration of 100 mg test item/1.

Exposure conditions
Test volume: 250 mL
Test apparatus: Oxi Top System (WTW)
Mixing: 1 magnetic stirrer per test vessel
Incubation time: 28 days
Incubation temperature: 22 ± 1 °C

Pre-treatment of the inoculum:
- activated sludge is washed twice with mineral medium; separation of the sludge by centrifugation
- Concentration of inoculum: 30 mg/L ss
Reference substance:
benzoic acid, sodium salt
Key result
% degradation (O2 consumption)
Sampling time:
28 d
Details on results:
The reference compound sodium benzoate showed 88 % degradation after 14 days.
Results with reference substance:
The reference compound sodium benzoate showed 88 % degradation after 14 days.

Validity criteria for the measurement of the biodegradation in water:

Target condition according to guideline:Actual condition according to the study:Validity criteria met:
In order to check the procedure, reference compounds which meet the criteria for ready
biodegradability are tested by setting up an appropriate vessel in parallel as part of normal test runs. Suitable compounds are aniline (freshly distilled), sodium acetate and sodium benzoate.
Benzoic acid, sodium salt was used as a reference substance.Yes
A test is considered valid if the difference of extremes of replicate values of the removal of
the test chemical at the plateau, at the end of the test or at the end of the 10-d window, as appropriate, is less than 20% and if the percentage degradation of the reference compound has reached the pass levels by day 14.

% degradation of the three measured values of the test substance at the end of the test (28 d) range between 25 - 29%

The reference compound has reached the level for ready biodegradability by 14 days.

If in a toxicity test, containing both the test substance and a reference compound, less than
35% degradation (based on total DOC) or less than 25% (based on total ThOD or ThCO2) occurred within 14 days, the test substance can be assumed to be inhibitory. The test series should be repeated, using a lower concentration of test substance (if this can be
done without seriously impairing the accuracy of the DOC determination) and/or a higher concentration of inoculum, but not greater than 30 mg solids/l.
No toxicity of the test item has been observed in the toxicity control.Yes
The oxygen uptake of the inoculum blank is normally 20-30 mg O2/L and should not be greater than
60 mg/L in 28 d.
The oxygen uptake of the inoculum blank is < 60 mg/L.Yes
If the pH value is outside the range 6 - 8.5 and the oxygen consumption by the test chemical is less than 60 %, the test should be repeated with a lower concentration of test chemical.The pH is between 6.0 and 8.5 in the test vessels at the end of the test (if degradation is less than 60 %).Yes


Validity criteria fulfilled:
Interpretation of results:
under test conditions no biodegradation observed
The substance is considered to be "Not Readily Biodegradable".
Executive summary:

A study was performed to assess the ready biodegradability of the substance.

The study was conducted in accordance with the Council Directive 92/69/EEC Method C.4-D"Manometlic Respirometry Test". This test method is in all essential parts identical with OECD Guideline 301 F.

A solution of the substance in a mineral medium was inoculated and incubated for 28 d under aerobic conditions. During this period, the ready biodegradability is determined.


The substance showed:

0 % degradation after 4 days.

0 % degradation after 6 days.

0 % degradation after 8 days.

1 % degradation after 12 days.

1 % degradation after 14 days.

1 % degradation after 18 days.

0 % degradation after 20 days.

1 % degradation after 22 days.

1 % degradation after 26 days.

2 % degradation after 28 days.


Therefore, the substance is considered to be "Not Readily Biodegradable".  

Description of key information

The substance is considered to be "Not Readily Biodegradable".

Key value for chemical safety assessment

Biodegradation in water:
not biodegradable
Type of water:

Additional information