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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:
18 Jan - 15 Feb 2011
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
The oxygen depletion in blank was 2.31 mg O2/L (slightly higher than specified in guideline 1.5 mg/L, but seemed to be acceptable).
according to guideline
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Version / remarks:
GLP compliance:
yes (incl. QA statement)
Niedersächsisches Umweltministerium, Hannover, Germany
Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: Municipal sewage treatment plant, Hildesheim
- Pretreatment: filtered with folded filter. First 200 mL of the filtrate were disposed. The second filtrate was used for inoculation
- Colony forming units in test vessels: 10^4 - 10^6 CFU/L
- Water filtered: yes
Duration of test (contact time):
28 d
Initial conc.:
3 mg/L
Initial conc.:
8.01 other: mg O2/L
Based on:
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
- Composition of medium: mineral nutrient solution according to OECD 301 D
- Test temperature: 20 - 21 °C
- O2 of demineralized water after aeration: 9.09 mg/L
- pH: 6.93 - 7.44
- Aeration of dilution water: yes

- Culturing apparatus: BOD bottles with glass stoppers (volume 300 mL)
- Number of culture flasks/concentration: 10
- Method used to create aerobic conditions: 1 day before test start, demineralized water for the test medium was aerated until oxygen saturation and then left at room temperature for at least 20 h.
- Measuring equipment: pH: pH-Meter, "CORNING" pH 240, Oxygen concentration: Oximeter, "WTW" Oxi 530

- Sampling frequency: the oxygen concentration was measured in the vessels on day 0, 8, 14, 21 and 28
- Other: temperature in the incubator was recorded once every working day. The pH value of the solutions was determined at the beginning and at the end of the test.

- Inoculum control: 10
- Reference control: 10
- Toxicity control: 10 (1.5 mg/L test item + 5.0 mg/L reference substance)
- Other: test series was accompanied by a complete parallel series for the determination of O2 demand of the inoculated control medium (C1, C2)

Reference substance:
acetic acid, sodium salt
10 mg/L (ThOD: 0.78 mg O2/mg)
Key result
% degradation (O2 consumption)
Sampling time:
28 d
Details on results:
10-day window criterion fulfilled.
Results with reference substance:
71% degradation after 28 d.

The oxygen demand in the inoculum control reached a maximum of 2.31 mg O2/L after 28 days.

The biodegradation rate of the toxicity control reached 65% after 14 days. The test item did not inhibit the biodegradation of the reference substance.

Table 1: Oxygen determination at each sampling point and % biodegradation.

Sampling date [d] Test item Toxicity control Reference control
O2 mg/L BOD mg/L % degr. O2 mg/L BOD mg/L % degr. O2 mg/L BOD mg/L % degr.
0 9.41  -  - 9.38  -   -  9.56  -   - 
8 3.23 4.93 61 3.78 4.35 55 2.96 5.35 69
14 2.91 4.97 62 2.71 5.14 65 2.21 5.82 74
21 1.94 5.52 69 2.36 5.07 64 2.06 5.55 72
28 2.13 4.97 62 2.59 4.48 57 1.76 5.49 71
Validity criteria fulfilled:
The oxygen depletion in blank was 2.31 mg O2/L (slightly higher than specified in guideline 1.5 mg/L, but seemed to be acceptable).
Interpretation of results:
readily biodegradable
This study indicates that the substance is readily biodegradable (62% after 28 d (O2 consumption), OECD 301 D). Furthermore the test showed that the substance is not inhibitory ot microorganisms at the tested concentration of 1.5 mg/L.

Description of key information

Readily biodegradable (62% in 28 d, 14-d window passed, OECD 301 D)

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

There is one GLP study available investigating the biodegradability of the substance in an aerobic, aqueous medium according to the OECD guideline 301 D.

In a closed bottle test a nominal concentration of 3.0 mg/L (8.01 mg O2/L) test substance was inoculated with non-adapted activated domestic sludge for 28 d under controlled conditions. A functional, toxicity, and inoculum control were run in parallel. Degradation was followed by measuring the oxygen concentrations in regular intervals. % biodegradation was expressed in terms of theoretical oxygen demand (ThOD) and biological oxygen demand (BOD).

The test substance reached the 10% level (begin of degradation) after 2 d. In the 10-d window a mean biodegradation rate of 62% was reached. The biodegradation reached the pass level of > 60% after 8 d and came to a maximum of 69% after 21 d. The toxicity control containing both test and reference item reached a maximum degradation rate of 65% within 14 d. The degradation of the reference item was not inhibited by the test substance. The oxygen depletion in blank was 2.31 mg O2/L, it is slightly higher than specified in guideline 1.5 mg/L, but seemed to be acceptable.