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Diss Factsheets

Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-11-10 to 2016-12-09
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Version / remarks:
July 1992
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:
solid: particulate/powder

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: municipal wastewater treatment plant Breisgauer Bucht Germany, 600,000 inhabitant equivalents, predominantly domestic wastewater.
- Concentration of sludge: 4.2 g/L after drying at 105°C
- Initial cell/biomass concentration: 30 mg/L
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
20 mg/L
Based on:
calculated based on molecular structure
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
- Composition of medium: mineral medium according to guideline
- Additional substrate: no
- Solubilising agent: no
- Test temperature: 20.5 - 23.1°C
- pH: not registered
- pH adjusted: no
- Aeration of dilution water: 2.5 to 4.3 bubbles/second or 30-100 mL/min with CO2-free air
- Suspended solids concentration: 30 mg dry solids/L
- Continuous darkness: no, diffuse light

- Number of culture flasks/concentration: 3
- Method used to create aerobic conditions: aeration with CO2-free air
- Measuring equipment: Total organic carbon analyser TOC-5050A with autosampler ASI 5000A, Shimadzu.
- Details of trap for CO2 and volatile organics if used: two 250 mL glass wash bottles in series each filled with 200 mL 0.2 M NaOH, sampling through lateral connecting pieces, through butyl rubber septum using 5 mL PE syringes

- Sampling frequency: on day 4, 7, 11, 14, 21 and 28
- Sampling method: 4 mL of NaOH sampled from 1st CO2-absorber flasks
- Sample storage before analysis: samples immediately closed with sealing film
- Final sampling: on day 28, 2mL of 4M HCl is added to the reactor to release CO2 dissolved in water. IC-analysis on day 29.

- Inoculum blank: 3 replicates
- Abiotic sterile control: no
- Toxicity control: yes, 1 replicate at 40.1 mg/L TOC
- Reference substance: 3 replicates at 30 mg/L

Reference substance
Reference substance:
benzoic acid, sodium salt

Results and discussion

% Degradation
Key result
% degradation (CO2 evolution)
St. dev.:
Sampling time:
29 d
Remarks on result:
other: measured after acidification of the reactor flasks to release CO2 dissolved in water.
mean of 3 replicates
Details on results:
The 10-day window started on day 7. The maximal degradation observed was 16.4 %, hence, the 10-day window criteria was not met. See below for details on results.

BOD5 / COD results

Results with reference substance:
The reference substance was rapidly biodegraded, with maximal degradation of 87.1 %.

Any other information on results incl. tables

% biodegradation in the different test vessels:

Reactor day 0 day 4 day 7 day 11 day 14 day 21 day 28 day 29
Test item, rep 1 0 5.3 9.9 14.4 14.2 11.8 16.4 16.1
Test item, rep 2 0 4.0 10.9 13.6 19.7 10.8 14.1 14.0
Test item, rep 3 0 7.1 12.1 16.9 19.8 11.5 14.9 14.9
Reference, rep 1 0 71.0 80.8 79.2 83.0 77.1 82.4 84.0
Reference, rep 2 0 77.6 79.1 80.5 85.0 80.2 86.0 87.1
Reference, rep 3 0 68.5 77.6 77.1 84.1 76.7 82.6 84.6
Toxicity control 0 39.7 41.9 46.9 48.6 48.9 53.6 54.6

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
not readily biodegradable
The biodegradability of the test item was examined in a CO2-evolution test according to OECD 301B. The test item was found to be not readily biodegradable with on average 15.0 % biodegradation after 28 days.
Executive summary:

The biodegradability of the test item was examined in a CO2-evolution test according to OECD 301B. After 28 days, the ultimate biodegradation was found to be 15.0 % (average of 3 replicates). The 10 -day window criteria was not fulfilled and the test item should be considered to be not readily biodegradable.

Up to 54.6 % degradation was observed in the toxicity control containing sodium benzoate and the test item. This result shows that the test item was not inhibitory to the inoculum.