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activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Aluminiumchlorid wasserfrei gemahlen (PBG 10000464)
Test substance N°: 00/0813-2
Batch n°: 32254175L0
Homogenous; expery date october 2008
Analytical monitoring:
Details on test solutions:
The test substance was weighed in the required amounts for the test concentrations directly to the test vessels. Than 180 mL demineralized water were added. The mixtures were stirred for 20 ± 2 hours at room temperature.
Aliquot of 50 mL of this suspension were added to the test vessels to obtain a sludge concentration of 1.5 g/L dry substance.
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
Activated sludge from the municipal wastewater treatment plant of Mannheim/Baden Württemberg, Germany was collected on 14 May 2008 from the aeration tank of the plant. The activated sludge suspension was washed 3 times with tap water and sieved with a fine woven mesh (mesh size about 1 mm). This suspension was adjusted to a concentration of 7.5 g/L dry weight and pre-aerated over night. After the pre-aeration the lost water of the suspension was replaced.
Test type:
Water media type:
Limit test:
Total exposure duration:
3 h
Post exposure observation period:
Duration of the measurement of oxygen consumption: 8-10 minutes
Test temperature:
22 +/_ 2 °C
7.2 - 7.3 (corrected before addition of inocumum; pH before correction was situated between 3.3 and 5.6)
pH-values after incubation time: 7.8 - 8.1
Dissolved oxygen:
oxygen concentration during aeration:> 2.5 mg/L
oxygen concentration immediately before measurement: > 6.5 mg/L
Nominal and measured concentrations:
Nominal: 62,4, 125, 250, 500, 1000 mg/L test substance
Details on test conditions:
Test vessesls: Erlenmeyer vessels (nominal value 250 mL)
Test volume: 250 mL
Synthetic medium: 8 mL/vessel 100-fold concentrated OECD medium
Key result
3 h
Dose descriptor:
Effect conc.:
1 000 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
3 h
Dose descriptor:
Effect conc.:
> 200 mg/L
Nominal / measured:
Conc. based on:
Basis for effect:
inhibition of total respiration
Results with reference substance (positive control):
EC50 of 3,5 dichlorophenol is in the range of 5-30 mg/L (validity criteria has been met)

Assay identification  Blanc#1  Blanc#2  1000 mg/L  500 mg/L  250 mg/L  125 mg/L  62.5 mg/L

O2 conc at start


 8.0  7.6 7.5  8.1   8.1 8.1  7.7 

O2 conc. after 6 min


 6.5  6.0 6.0   6.5  6.5 6.5  6.0 

 O2 consumption rate

(mg/L in 6 min)

 1.5  1.6 1.5  1.6  1.6   1.6  1.7

O2 consumption rate

(mg/L x h)

 15  16 15  16  16   16 17 

Specific O2 consumption


(mg/g x h)

 10  11 10  11   11 11  11 

Calculation of


respiration (%)

--  --  -3  -3  -3   -10
Validity criteria fulfilled:
In conclusion the 180 minutes EC10 and EC20 values for Aluminiumchlorid wasserfrei gemahlen (PBG 10000464) in the Activated Sludge Respiration Inhibition Test was >1000 mg/L. The EC50 was >1000 mg/L.
No signification inhibition was noted in any of the tested concentration levels.

Description of key information

One activated sludge respiration inhibition test is available with a soluble aluminium salt. This test was performed with aluminum chloride CAS RN 7446-70-0 according to standard guideline. No effects were seen up to the highest test concentration of 1000 mg/l test material. The test substance contained 20 g Al/100 g and the test concentrations can be converted to mg/l Al. 1000 mg/l test material is equal to 200 mg/l Al. See Table # "Toxicity to aquatic micro-organisms" (above).

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
200 mg/L

Additional information

Table #: Toxicity to aquatic micro-organisms



set up

result (mg/l)


Aluminum chloride

Activated sludge, domestic