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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:
05 June 2018 to 06 September 2018
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Version / remarks:
GLP compliance:
Specific details on test material used for the study:
Purity: UVCB; solid matter 41.4%
Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: Waterschap Aa en Maas', 's-Hertogenbosch, The Netherlands
- Storage conditions: continuous aeration
- Storage length: freshly obtained
- Concentration of sludge: 7.2 mg/L suspended solids
- Type and size of filter used, if any: before use, coarsely sieved (1 mm)
Duration of test (contact time):
28 d
Initial conc.:
12 mg/L
Based on:
Initial conc.:
60.5 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
- Test temperature: 22 – 23 °C
- pH: 7.5 – 7.6
- pH adjusted: no
- Aeration of dilution water: the test media were aerated and stirred continuously
- Suspended solids concentration: 7.2 mg/L
- Continuous darkness: yes

- Number of culture flasks/concentration:
Test suspension: containing test item and inoculum (2 bottles).
Inoculum blank: containing only inoculum (2 bottles)
Procedure control: containing reference item and inoculum (1 bottle).
Toxicity control: containing test item, reference item and inoculum (1 bottle)
- Details of trap for CO2 and volatile organics if used: barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl

- Sampling frequency: Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until day 28, for the inoculum blank and test item. Titrations for the procedure and toxicity control were made over a period of at least 14 days.
Reference substance:
acetic acid, sodium salt
% degradation (CO2 evolution)
Sampling time:
28 d
% degradation (CO2 evolution)
Sampling time:
28 d
Details on results:
In the toxicity control, more than 25% biodegradation occurred within 14 days (48%, based on ThCO2). Therefore, the test item was assumed not to inhibit microbial activity.
Results with reference substance:
In the procedure control, 73% biodegradation occurred within 14 days.

The reference item was biodegraded by at least 60% (actual result: 73%) within 14 days.


The difference of duplicate values for %-degradation of the test item was always less than 20 (actual result: ≤ 6%).


The total CO2 release in the blank at the end of the test did not exceed 40 mg/L (48.7 mg CO2 per 2 litres of medium, corresponding to 24.4 mg CO2/L).


The Inorganic Carbon content (IC) of the test item (suspension) in the mineral medium at the beginning of the test was less than 5% of the Total Carbon content (TC). Since the test medium was prepared in tap-water purified by reverse osmosis, IC was less than 5% of TC (mainly coming from the test item, 12 mg TOC/L).


Since all criteria for acceptability of the test were met, this study was considered to be valid.

Validity criteria fulfilled:
Interpretation of results:
inherently biodegradable
In conclusion, 1-Octanol, reaction products with phosphorus oxide (P2O5), potassium salts was not readily biodegradable under the conditions of the modified Sturm test presently performed. However, since the results of the present test indicate that the pass level criterion was almost fulfilled the results can be used to indicate inherent biodegradability.

Description of key information

The biodegradation of the test item was assessed following OECD 301 B (CO2 Evolution Test). The test item was tested in duplicate at a target concentration of 60.5 mg/L, corresponding to 12 mg TOC/L. The relative biodegrading values calculated from the measurements performed during the test period revealed 51% and 45% biodegradation of test Item (based on ThCO2), for the duplicate vessels tested. These results indicate that the test item is not readily biodegradable. However, since the results of the present test indicate that the pass level criterion was almost fulfilled the results can be used to indicate inherent biodegradability.

Key value for chemical safety assessment

Biodegradation in water:
inherently biodegradable
Type of water:

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