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:
no
Qualifier:
according to
Guideline:
other: EEC L 133 s. 118-122 (30. May 1988)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
not specified

Test organisms

Test organisms (species):
activated sludge
Details on inoculum:
Activated sludge collected from the sewage treatment plant of CH-4153 Reinach on November 3, 1999.
The pH after collection was 7.7.
The preparation was carried out according to the method in the guideline.
The sludge was separated from the aqueous layer by settling instead of centrifugation.
The pH of the sludge before use was 7.9.
The sludge concentration in the test bottles was 1.75 g suspended solids/L.

Study design

Test type:
static
Total exposure duration:
3 h

Test conditions

Test temperature:
20 ± 2 °c
pH:
The pH of sludge was 7.7 after collection and 7.9 after preparation (before use).
The pH of the mixtures in the test bottles was 8.2 – 8.3 at the end of exposure time.
Nominal and measured concentrations:
Stock solution:
No stock solution was prepared.
27.4, 7.1 and 1.8 mg of the test item for concentrations of 136, 35.4 and 8.98 mg/L were directly given into the test vessels.

Test concentrations :
Test item: 136.0, 35.4 and 8.98 mg/L (3.2 and 1.0 mg/L were not tested)
Reference item: 32.0, 10.0 and 3.2 mg/L
Details on test conditions:
Vessels: 250 ml BOD flasks with gas inlet
Water: De-chlorinated drinking water
Temperature: 20 ± 2 °C
Duration: 3 hours.


Stock solution:
No stock solution was prepared.
27.4, 7.1 and 1.8 mg of the test item for concentrations of 136, 35.4 and 8.98 mg/L were directly given into the test vessels.

Test concentrations :
Test item: 136.0, 35.4 and 8.98 mg/L (3.2 and 1.0 mg/L were not tested)
Reference item: 32.0, 10.0 and 3.2 mg/L

Calculated amounts of the test item to produce the desired test concentrations were mixed with 114 ml water and 6.4 ml nutrient solution (= test solution). Then 80 ml of the sludge were added.
The test item, 6.4 ml of the nutrient solution, 114 ml water and 80 ml of the sludge were mixed in 250 ml BOD flasks and aerated immediately.

Measurements:
Oxygen consumption per hour in mg/L with a ORION – Electrode Type 97-08, ORION Microprocessor Ionalizer 901 and plotted on a recorder.

Calculations:
The inhibitory values were calculated on the basis of the measured time dependant oxygen consumption of a blank and test solution.

Nutrient Solution:
16g Peptone
11g Meat extract
3.0g Urea
0.7g NaCL
0.4g CaCl2.2H2O
0.2g MgSO4.7H2O
2.8g K2HPO4
Were dissolved and the volume made up to one litre with de-chlorinated drinking water.

Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol

Results and discussion

Effect concentrationsopen allclose all
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
> 136 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Duration:
3 h
Dose descriptor:
other: EC20
Effect conc.:
> 136 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Duration:
3 h
Dose descriptor:
other: EC80
Effect conc.:
> 136 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Details on results:
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:

The test item CA 2219 A was tested at nominal concentrations of 8.98, 35.4 and 136 mg test item/L after a contact time of 3 hours. The calculation of the following effect concentrations was based on these nominal values.

EC50 (3h) : > 136 mg/L

EC20 (3h) : > 136 mg/L
EC80 (3h) : > 136 mg/L


Conclusion:
The EC50 (3h) for CA 2218 A is > 136 mg/L. Based on the inherent toxicity of CA 2219 A in this test alone, the test item can be classified as: “not toxic to bacteria”.


Results with reference substance (positive control):
For the reference item (tested nominal concentrations) 3.2, 10 and 32 mg/L:

EC50 (3h) : > 11.4 mg/L

EC20 (3h) : > 4.1 mg/L
EC80 (3h) : > 31.4 mg/L

Test Validity;
The deviation of the respiration rates in controls was 8 %. The EC 50 (3h) of the reference item was 11.4 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.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The EC 50 (3h) for CA 2219 A is > 136 mg/L.
Based on the inherent toxicity of CA 2219 A in this test alone, the test item can be classified as:
“not toxic to bacteria”
Executive summary:

The respiration rate of an activated sludge fed with 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 8.98, 35.4 and 136 mg test item/L and 3.2, 10 and 32 mg/L reference item, respectively. No stock solutions for the test item were prepared. The test was not performed with test concentrations of 3.2 and 1.0 mg/L, because no formulations were possible. 

The sludge concentration in the test bottles was 1.75 g suspended solids/L. 

 

The test with CA 2219 A the temperature was maintained constant at 20 ± 1°C. 

The pH of sludge was 7.7 after collection and 7.9 after preparation (before use). 

The pH of the mixtures in the test bottles was 8.2 – 8.3 at the end of exposure time. 

 

The EC50 (3h) of the reference item 3,5-Dichlorophenol was 11.4 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) : > 136 mg/L

 

EC20 (3h) : > 136 mg/L

EC80 (3h) : > 136 mg/L

 

Conclusion:

The EC 50 (3h) for CA 2219 A is > 136 mg/L. 

Based on the inherent toxicity of CA 2219 A in this test alone, the test item can be classified as:

“not toxic to bacteria”