Registration Dossier

Administrative data

Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
yes
Remarks:
Settled sludge used instead of a centrifuged sludge
Qualifier:
according to
Guideline:
other: EEC L133 s. 118-122 (30 May 1988)
Deviations:
yes
Remarks:
Settled sludge used instead of a centrifuged sludge
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test organisms

Test organisms (species):
activated sludge
Details on inoculum:
Activated sludge collected from the sewage treatment plant of CH-4153 Reinach on September 20, 2000.
The pH after collection was 7.6.
The sludge was separated from the aqueous layer by settling instead of centrifugation.
The pH of the sludge before use was 7.2.
The sludge concentration in the test bottles was 1.72G suspended solids/L.

Study design

Total exposure duration:
3 h

Test conditions

Test temperature:
The test with CA 2814 B was conducted at a temperature of 20 ± 1°C.
pH:
The pH of sludge was 7.6 after collection and 7.2 after preparation (before use).
The pH of the mixtures in the test bottles was 8.3 – 8.5 at the end of exposure time.
Nominal and measured concentrations:
The respiration rate of an activated sludge fed with a standard amount of synthetic sewage feed was measured after a contact time of 3 hours.
The respiration rate of the same activated sludge was also measured in the presence of 1.5, 5.2, 23.5, 40.9 and 109.3 mg test item/L and 3.2, 10 and 23. mg/L reference item, respectively.
The sludge concentration in the test bottle was 1.72 g/L suspended solids/L.
Details on test conditions:
Vessels: 250 ml BOD flasks with gas inlet
Water: Dechlorinated drinking water
Temperature: 20 ± 2°C
Duration: 3 hours
Stock solution: No stock solution prepared due to lack of solubility
Test item solutions: Calculated amounts of the test item to produce the desired test concentrations were mixed with 120 ml medium
(114 ml water and 6.4 ml nutrient solution) in 250 ml BOD flasks.
Then 80 ml of the sludge were added, mixed and aerated immediately.
Nutrient solution: 16 g Peptone; 11g Meat extract; 3.0 g Urea; 0.7 g NaCl; 0.4 g CaCl2.2H2O; 0.5 g MgSO4.7H2O; K2HPO4,
were dissolved and the volume made up to 1 litre with dechlorinated drinking water.

Reference substance (positive control):
yes

Results and discussion

Details on results:
The test item CA 2814 B was tested at nominal concentrations of 1.5, 5.2, 23.5, 40.9 and 109.3 mg test item/L after a contact time of 3 hours.
The test item was not homogeneously distributed in the test vessels.
At the end of exposure small drops were sticking on the surface of the test vessels.

The calculation of the following effect levels was based on nominal values.
EC50 (3h): > 100 mg/L

EC20 (3h): > 100 mg/L
EC80 (3h): > 100 mg/L


Results with reference substance (positive control):
For the reference item (tested nominal concentrations: 3.2, 10 and 23. mg/L reference item)
EC50 (3h): 9.0 mg/L

EC20 (3h): 3.0 mg/L
EC80 (3h): 26.7 mg/L
Reported statistics and error estimates:
The deviation of the respiration rates in controls was 4%.
The EC 50 (3h) of the reference item was 9.0 mg/L.
Since the deviation of the respiration rates of the controls was <15% and the EC 50 (3h) of the reference item was in the range of 5 to 30 mg/L, the test was valid in terms of the given validity criteria.

Any other information on results incl. tables

Oxygen consumption rate and inhibitory effects of the test item and reference

 

Sample

Concentration (mg/L)

Consumption (mg/L)

Inhibition (%)

pH

Blank 1

Blank 2

0.0

0.0

41.4

39.7

-2

2

8.3

8.4

Reference 1

Reference 2

Reference 3

32.0

10.0

3.0

6.9

17.3

32.7

83

57

19

8.4

8.4

8.5

Test conc. 1

Test conc. 2

Test conc. 3

Test conc. 4

Test conc. 5

109.3

40.9

23.5

5.2

1.5

46.0

41.2

48.5

38.8

40.1

-14

-2

-20

5

1

8.5

8.4

8.5

8.4

8.5

 

The pH was measured after 3 h aeration. 

Negative inhibition results, when obtained in a test, are a consequence of the comparison of oxygen consumption values obtained in the blank, and the higher oxygen consumption values obtained in the presence of the test item, which are due to biological variations between the various test samples. 

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The EC50 (3h) for CA 2814 B is > 100 mg/L.
Based on the inherent toxicity of CA 2814 B in this test alone, the test item can be classified as :
“not toxic to bacteria”.
Executive summary:

Findings:

The test with CA 2814 B was conducted at a temperature of 20 ± 1°C. 

The pH of sludge was 7.6 after collection and 7.2 after preparation (before use). The pH of the mixtures in the test bottles was 8.3 – 8.5 at the end of exposure time. 

The EC50 (3h) of the reference item 3,5-Dichlorophenol was 9.0 mg/L. 

 

The inhibitory effect of the test item was expressed as a percentage of the mean respiration rates of two controls. Following values were calculated based on nominal concentrations:

 

EC50 (3h): > 100 mg/L

 

EC20 (3h): > 100 mg/L

EC80 (3h): > 100 mg/L

 

 

Conclusion:

TheEC50 (3h) for CA 2814 B is > 100 mg/L. 

Based on the inherent toxicity of CA 2814 B in this test alone, the test item can be classified as :

“not toxic to bacteria”.