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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: Municipal sewage treatment plant, D-31137 Hildesheim.
- Storage at test facility: Refrigerator 7 +- 2°C, protected from light and moisture
- Pretreatment: The activated sludge was filtered with foided filter. The first 200 mL of the filtrate are not used. The second filtrate effluent from the domestic waste water is used to initiate inoculation.
- Concentration of sludge: 10^4 - 10^6 CFU/L
Duration of test (contact time):
28 d
Initial conc.:
4 mg/L
Based on:
test mat.
Remarks:
corresponding to ThOD of 7.72 mg/L
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST ITEM:
- Dispersion treatement: Agitation
- Test concentration: 4.0 mg/L test material
- ThOD: 1.93 mg O2/mg test item
- ThOD in test vessel: 7.72 mgO2/L

CONTROL AND BLANK SYSTEM
- Inoculum blank: sodium acetate
- Test concentration: 10 mg/L
- ThOD: 0.78 mg O2/mg

INOCULUM CONTROL:
Nutrient solution and inoculum

TOXICITY CONTROL:
2.0 mg/L test item + 0.5 mg/L functional control item + nutrient solution and inoculum.

TEST CONDITIONS
- Duration: 28 days
- Frequency and duration of the application: Once per setting up over 28 d
- Test volume: 300 mL
- Composition of medium: Mineral nutrient solution according to OECD 301 D

PERFORMANCE OF THE TEST:
- One day before the test started the demineralized water for the test medium was aerated until oxygen saturation and then left at room temperature for at least 20 h.
- The test series was accompanied by a complete parallel series for the determination of the O2 demand of the inoculated control medium (C1, C2).
- All test solutions were made as stock solutions. 0.2 mL of the inoculum was given in each BOD bottle. The stock solution were filled to the BOD bottles with a siphon. The incubation vessels were closed without air bladders.
- At all measuring point the oxygen concentration of duplicates of the inoculum control, functional control, test item and toxicity control was determined.

TYPE AND FREQUENCY OF MEASUREMENTS:
- The temperature in the incubator was documented once every working day.
- At the beginning and the end of the test the pH value of the solution was determine
- The oxygen concentration was measured in the incubation vessels on day 0, 7, 14, 21 and 28.
- Test temperature: 20 °C
- pH measurement: yes

VALIDITY:
- The percentage degradation of the functional control must reach the pass level of 60% by day 14.
- The oxygen depletion in the inoculum control should not exceed 1.5 mg O2/L after 28 day.
- The residual concentrationof oxygen in the test bottles should not fall below 0.5 mg/L at any time.
- The differente of extremes of replicate values of removal of the test item at the end of the test, at the plateau or at the end of the 10-d-window as appropriate must be less than 20 %, if the test item is readily biodegradable.
- If in the toxicity test containing both test item and reference item, less than 25 % degradation occurred within 14 days, the test can be considered to be inhibitory.

TEST SYSTEM
- Culturing apparatus: Based on the ThOD of the test item the test concentration of 4.0 mg/L corresponding to a oxygen demand of 7.72 mgO2/L in the vessel was selected.
- Number of culture flasks/concentration: BOD bottles with glass stoppers (volume 300 mL)
- 10 bottles containing test item and inoculum (test suspension)
- 10 bottles containing only inoculum (inoculum control)
- 10 bottles containing reference item and inoculum (functional control)
- 10 bottles containing test and reference item and inoculum (toxicity control)


EVALUATION METHOD:
For the calculation of the BODn and the biodegradation of the test item, functional control and toxicity control the oxygen depletion of the control was taken into account. (please see detail under section "Any other information on materials and method" section)
Reference substance:
acetic acid, sodium salt
Remarks:
sodium acetate
Key result
Parameter:
% degradation (O2 consumption)
Value:
71
Sampling time:
28 d
Parameter:
% degradation (O2 consumption)
Value:
58
Sampling time:
7 d
Parameter:
% degradation (O2 consumption)
Value:
62
Sampling time:
14 d
Parameter:
% degradation (O2 consumption)
Value:
65
Sampling time:
21 d
Details on results:
- Results of the toxicity control: 69% degradation after 14 days, indicating that the reference item is not inhibiting the activated sludge.
- Results inoculum control: oxygen demans came to 1.4 mg O2/L
- Results of functional control: pass level of > 60 % was reached after 6 days, fulfilling the validity criteria.
- Results with test item: pass level of 10 % was reached after 1 d and was 62% after 14 days - the 14-day window criteria was met.

Results of the biodegradation trial, % degradation from 2 replicates

day test item  toxicity control
0 - 0
7 58 66
14 62 69
21 65 75
28 71 76
Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test item was found to be ready biodegradable, with a maximum degradation of 71 % after 28 days.
Executive summary:

The study was performed according to OECD guideline 301 D and GLP principles, as a Closed Bottle Test was determined with a non adapted activated sludge for the test item. The test item concentration selected as appropriate was 4.0 mg/L, corresponding to a ThOD 7.72 mg O2/L in the test vessel.

The oxygen depletion in the inoculum control reached 1.40 mg O2/L until day 28. The residual concentration of oxygen in the test bottles did not fall below 0.5 mg/L at any time.

In order to check the activity of the study system sodium acetate was used as functional control. The functional control reached the pass level > 60% after 6 days. After 14 days a biodegradation rate of 69 % was reached.

In the toxicity control containing both test and reference item 69% degradation occured within 14 days. The degradation came to a maximum of 76% after 28 days. The degradation of the reference item was not inhibited by the test item.

The test item reached the 10% level after 1 day. The biodegradation came to 62 % after 14 days to a maximum of 71 % after 28 days.

According to CLP criteria, the test item must be regarded to be readily biodegradable

Description of key information

readily biodegradable (71% after 28 days; OECD 301 D)

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

The study was performed according to OECD guideline 301 D and GLP principles, as a Closed Bottle Test was determined with a non adapted activated sludge for the test item. The test item concentration selected as appropriate was 4.0 mg/L, corresponding to a ThOD 7.72 mg O2/L in the test vessel.

The oxygen depletion in the inoculum control reached 1.40 mg O2/L until day 28. The residual concentration of oxygen in the test bottles did not fall below 0.5 mg/L at any time.

In order to check the activity of the study system sodium acetate was used as functional control. The functional control reached the pass level > 60% after 6 days. After 14 days a biodegradation rate of 69 % was reached.

In the toxicity control containing both test and reference item 69% degradation occured within 14 days. The degradation came to a maximum of 76% after 28 days. The degradation of the reference item was not inhibited by the test item.

The test item reached the 10% level after 1 day. The biodegradation came to 62 % after 14 days to a maximum of 71 % after 28 days.

According to CLP criteria, the test item must be regarded to be readily biodegradable