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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Feb 15 - 20, 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Version / remarks:
Activated Sludge, Respiration Inhibition Test, adopted April 04, 1984
Deviations:
yes
Remarks:
Due to technical reaseons the temperature of 18-22 °C according to the study protocol was exceeded to 23 °C.
Qualifier:
according to guideline
Guideline:
EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)
Version / remarks:
88/302/EEC
Deviations:
yes
Remarks:
Due to technical reaseons the temperature of 18-22 °C according to the study protocol was exceeded to 23 °C.
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
Due to the low solubility of the test item in test water, adequate amounts of the test item were directly weighed into each test flask. One test solution with a final volume of 500 mL was tested per treatment in a glass flask (>500 mL). Samples were prepared in about 15-minute intervals.
The following test item concentrations have been used: 1000, 320, 100, 32, and 10 mg/L
Test organisms (species):
activated sludge, domestic
Details on inoculum:
Species: activated sludge, micro organisms from a domestic waste water treatment plant.
Origin: supplied by the sewage plant Darmstadt, Germany

Conditioning:
The sludge was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in tap water and again centrifuged. The latter pro¬cedure was repeated twice. An aliquot of the fmal sludge sus-pension was weighed, dried and the ratio of wet sludge (g) to its dry weight (g) determined. The washed sludge suspension was made up with tap water and 50 mL synthetic sewage feed (see 6.6) per litre was added and the sludge was kept at room temperature under continuous aeration for two days until use. Immediately before use, the dry weight of the activated sludge was determined, and diluted to 2 g/L with tap water. The pH of the activated sludge was 7.1.

Test type:
other: aerobic
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Test temperature:
20-23 °C
pH:
7-8.5
Dissolved oxygen:
7.3 mg/L
Nominal and measured concentrations:
1000, 320, 100, 32, and 10 mg/L
Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Duration:
3 h
Dose descriptor:
other: EC20
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Results with reference substance (positive control):
EC50 = 13.2 mg/L (Acceptance range: 5-30 mg/L)

Results and Discussion

Inhibition of Respiration Rate

In comparison to the inoculum controls the respiration rate of the activated sludge was slightly inhibited between 0%and 5.9 % in a concentration range from 10mg/Lup to 1000 mg/L. Concentrations exceeding 1000 mg/L nominal were not tested.

 

3-hour EC20/80and 3-hour EC50:

Based on measured inhibition rates, no 3-hour EC20/80and EC50could be quantified because up to the highest nominal test concentration of 1000 mg/L less than 10%inhibition was noted after three hours incubation. Nevertheless, the 3-hour EC20/80and EC50are clearly higher than 1000 mg/L under the present test conditions.

Validity criteria fulfilled:
yes
Conclusions:
The 3-hour EC20 and EC50 are clearly higher than 1000 mg/L under the present test conditions.
Executive summary:

Purpose
The influence of the test item on the activity of activated sludge was evaluated by measuring the respiration rate under defined conditions.

Study design

The respiration rate (oxygen consumption) of an aerobic activated sludge fed with a standard amount of synthetic sewage was measured in the presence of various concentrations (10, 32, 100, 320 and 1000 mg/L) of the test item after an incubation period of 3 hours. This study was conducted according to Good Laboratory Practice (GLP) and the method applied followed the OECD Guideline for Testing of Chemicals, Section 2, No.209: "Activated Sludge, Respiration Inhibition Test", adopted April 04, 1984.

Results

In comparison to the inoculum controls the respiration rate of the activated sludge was slightly inhibited between 0 % and 5.9 % in a concentration range from 10 mg/L up to 1000 mg/L. Concentrations exceeding 1000 mg/L nominal were not tested.

Based on measured inhibition rates, the 3-hour EC 20 and EC 50 could not be quantified because up to the highest nominal test concentration of 1000 mg/L less than 20 % inhibition was noted after three hours incubation. Nevertheless, the 3-hour EC 20 and EC 50 are clearly higher than 1000 mg/L under the present test conditions.

The 3-hour EC 50 for the positive control 3,5 -dichlorophenol, which was tested in the same way as the test item, was found to be 13.2 mg/L and is within the range of 5 - 30 mg/L recommended by the test guidelines; thus, confirming suitability of the activated sludge.

Conclusion

The 3 -hour EC20and EC50 of the test material are clearly higher than 1000 mg/L under the present test conditions.

Description of key information

Based on measured inhibition rates, no 3-hour EC20/80 and EC50 could be quantified because up to the highest nominal test concentration of 1000 mg/L less than 10% inhibition was noted after three hours incubation. Nevertheless, the 3-hour EC20/80 and EC50

are clearly higher than 1000 mg/L under the present test conditions.

Key value for chemical safety assessment

Additional information

The influence of the test material on the growth and growth rate of the green algae species Desmodesmus subspicatus was tested. Therefore, a GLP compliant study according to OECD TG 201 was undertaken. The study design included one control and one test material group with three replicates, each containing 100 mL reconstituted water or test medium and about 10,000 cells/mL at the start of the experimental phase. The study was performed as a limit test in an open static test system. The algae were exposed to a filtrate of nominal 100 mg/L. The growth of the algae was calculated after 24, 48, and 72 hours exposure in the test medium. The analysis of the saturated aqueous preparation revealed that the water solubility of the test item in reconstituted water was < 0.0114 mg/L. An exact value could not be determined. The test material concentration in the aqueous medium could not be quantified since it was below the limit of quantification.

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