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Diss Factsheets

Administrative data

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

Data source

Reference Type:
study report

Materials and methods

Test guideline
according to guideline
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:
other: mud
Details on test material:
Activated sludge was collected and preferably used on day of collection, if is not possible the mud was
used no later than two days feeding it with the medium.
If in the sludge there were present coarse particles they were removed by settling for a period of 10
minutes. Then the mud samples, before their use, were washed and analyzed to prepare the inoculum.
The sludge was continuously aerated at the test temperature.
The suspended solids in the mud was about 2-4 g/l to have in the final mixture a concentration of 1.5 g/L.

Test organisms

Test organisms (species):
activated sludge

Study design

Test type:
Water media type:
Limit test:
Total exposure duration:
3 h

Test conditions

Test temperature:
20°C ± 0.5°C
Dissolved oxygen:
the vessels were aerated to keep the dissolved oxygen concentration above 60-70 % saturation
and to maintain the sludge floes in suspension.
Nominal and measured concentrations:
Test item was directly weighted in to bottles given because it was poorly soluble in water. Bottles were
prepared with water, synthetic sewage feed and 3 different concentrations of test item for the range
finding test (1 g/I, 100 mg/I and 10 mg/I).
Details on test conditions:
The incubation is considered to begin with the initial contact of the activated sludges with the other
constituent of the final mixture. Each incubation was started in turn at intervals of about 5-10 minutes.
Then the vessels were aerated to keep the dissolved oxygen concentration above 60-70 % saturation
and to maintain the sludge floes in suspension.
The bottles prepared were incubated at 20°C+ 2°C for three hours.
After the exposure period each sample will be transferred from the aeration vessel to a bottle fitted with a
magnetic stirrer (see figure below). An oxygen probe was inserted into the neck of the bottle to measure
the concentration of dissolved oxygen.
The concentration of dissolved oxygen was recorded in turn continuously for a period of 5-10 minutes or
until the oxygen concentration falls below 2 mg/L.
The probe was washed with deionised water between measurements.
Reference substance (positive control):
3,5 Dichlorophenol (Aldrich Company)

Results and discussion

Effect concentrations
3 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
respiration rate
Details on results:
Quality criteria of the test are satisfied
the blank controls oxygen uptake rate: 1093 mg oxygen per one gram of activated sludge in an
the coefficient of variation of oxygen uptake rate in control replicates: 1,62 %
ECso of 3,5 - Dichlorophenol 17,20 mgtl for total respiration.
pH synthetic sewage feed: 7,18
pH reference substance stock solution: 7,08
oxygen consumption rate for abiotic control: 74 mgAh
Results with reference substance (positive control):
EC50 17.200mg/L

Applicant's summary and conclusion