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Environmental fate & pathways

Biodegradation in water: screening tests

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biodegradation in water: inherent biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2018-11-21 to 2018-12-20
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
OECD Guideline 302 C (Inherent Biodegradability: Modified MITI Test (II))
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
batch number: 05347/MA
Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
Test system Non-adapted activated sludge
Receipt: 2018-11-19
Activated sludge from the sewage plant at Hildesheim is well suited as it receives predominantly municipal sewage and hardly any industrial chemical waste.
Municipal sewage treatment plant, 31137 Hildesheim, Germany

The fresh sampled sludge was washed twice with tap water. The sludge was maintained in an aerobic condition by aeration with CO2-free air until test start (2 days).
The amount of inoculum used to initiate inoculation was 29.3 mL/L (100 mg/L dry weight) and 8.8 mL/L (30 mg/L dw) for the functional control.

Duration of test (contact time):
28 d
Initial conc.:
80.9 mg/L
Based on:
Initial conc.:
28 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
- Composition of medium: Mineral salts medium acc. to OECD 302 C

- Solubilising agent for a better bioavailability: Silicon oil (Wacker AR 20 Salben Silikonöl)
- Test temperature: 25 ± 2 °C
- pH: Start: 7.36; End prior to Acidification: 6.65 - 6.75
- pH adjusted: no
- Continuous darkness: Dark, in an incubator (brown glass bottles)

- Culturing apparatus and test procedure: The test solution were prepared in measuring flasks and were placed in brown glass bottles as incubation vessels (inoculum control and functional control) or were prepared directly in brown glass bottles (inoculum control with silicone oil, functional control with silicone oil, test item and blank)
- Number of culture flasks/concentration: 3 with test substance, 2 for inoculum control, 2 for inoculum control with silicon oil, 1 functional control, 1 functional control with silicon oil, 1 blank

- Sampling frequency: daily

- Inoculum blank: yes (in duplicate)
- Toxicity control: yes
- Other: reference substance used was sodium benzoate

The theoretical production of oxygen (ThOD) is calculated.

ThOD = 2.89 mgO2/mg

Reference substance:
benzoic acid, sodium salt
Key result
% degradation (O2 consumption)
< 5
Sampling time:
28 d
Details on results:
Not inherently biodegradable
Results with reference substance:
The percentage degradation of sodium benzoate reached the pass level of > 40% within 2 days and > 65% within 3 days, with a maximum biodegradation of 100 % on day 15.


 Study Day

Functional Control [%]

Functional Control

with silicon oil [%]

Test item [%]


 1  17  15  0
 2  62  60  0
 3  70  68  0
 4  77  77  0








































 15  100  93  3
 16  100  93  4
 17  -  93  2
 18  -  94  2
 19  -  95  2
 20  -  94  2
 21  -  96  2
 22  -  96  2
 23  - 96  2
 24  - 96  3
 25  -  97  3
 26  -  97  3
 27  -  98  3
 28  -  98  2


Validity criteria fulfilled:
Interpretation of results:
not inherently biodegradable
The test item Reaction product of guerbet alcohols, C24-26, branched and cyclic with 1,2,4-benzenetricarboxylic acid is not inherently biodegradable.
Executive summary:

The inherent, ultimate biodegradability of the test item was determined in the Modified MITI Test (II) with non-adapted inoculum for a period of 28 days. The study was conducted according to OECD Guideline 302 C at the test facility. The test item concentration selected as appropriate was 28 mg/L, corresponding to a ThOD of 80.9 mg O2/L. The oxygen was depleted by respiration of bacteria and the degradation was followed by measuring the oxygen concentration. The biodegradation is therefore expressed as the percentage BOD depletion and was calculated for each study day.

The mean oxygen depletion in the inoculum control was 28.2 mg O2/L on day 28 (25.9 mg O2/L for the inoculum control with silicone oil).

In order to check the activity of the test system Sodiurn Benzoate was used as functional control in separate replicates with and without silicone oil. The functional control Sodium Benzoate and the functional control with silicone oil reached the pass level of > 65% within 3 days. The biodegradation reached a rnaximum of 100% degradation on day 28 (98 % for the functional control with silicone oil).

The test item replicates did not reach the 10% level (beginning of biodegradation) within the 28 day period of the study. The pass level > 70 % for evidence of inherent, ultimate biodegradability was not reached within the 28 days.

Description of key information

The inherent, ultimate biodegradability of the substance Esterification products of Guerbet alcohols, C24-26, branched and cyclic with benzene-1,2,4-tricarboxylic acid 1,2-anhydride was determined in the Modified MITI Test (II) according OECD guideline 302C with non-adapted inoculum for a period of 28 days. Based on the result the substance cannot be considered inherently biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
under test conditions no biodegradation observed
Type of water:

Additional information