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Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
28 - 29 Oct 2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes (incl. certificate)
Remarks:
Department of Health, UK
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Information provided by the Sponsor indicated that the test substance is not sufficiently soluble to prepare an aqueous solution. Therefore, at test inition weights were established by the addition of appropriate volumes (based on specific gravity, 0.99) of test substance to the test vessels containing RO water, synthetic sewage and microbial inoculum (see Table 1).
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Preparation of inoculum for exposure: A sample of activated sludge was obtained the day before the start of the test from Worlingworth Sewage Treatment Works, which treats predominantely domestic waste. In the laboratory, the sample was maintained under aerobic conditions until required. One day one of collection, an aliquot (10 mL) of the activated sludge was filtered through a dried, preweighed Whatman GF/C filter paper, which was then dried again at ~ 105 °C for at least one hour, allowed to cool in a desiccator before being reweighed. The mixed liquor suspended solids content of the activated sludge was then calculated. Synthetic sewage (50 mL/L) was added to the stock of activated sludge. The mixture was aerated overnight. The mixed liquor suspended solids content of the activated sludge was adjusted to 4 g/L by addition of tap water. Aliquots (200 mL) were added to the test vessels to give a final suspended solids concentration of 1.6 g/L.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Test temperature:
test start: 19.5 - 19.6 °C
test end: 19.3 - 19.4 °C
pH:
test start: 7.3 - 7.4
test end: 7.6 - 7.9
Nominal and measured concentrations:
Nominal: 10, 100 and 1000 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: beaker
- Type: closed (loosely covered with aluminium foil)
- Aeration: using a glass aerator connected to a laboratory supply of oil-free compressed air (~ 1 L/min.)
- No. of vessels per concentration: 1 (10, 100 mg/L); 3 (1000 mg/L)
- No. of vessels per control: 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reverse osmosis (RO) water
- Culture medium different from test medium: no, according to OECD guideline 209
- Intervals of water quality measurement: The pH and temperature were measured at the start and end of the test.

EFFECT PARAMETERS MEASURED: The respiration rate was measured after a test period of 3 h.
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
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:
EC10
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
The test substance showed no inhibitory effect on the respiration rate of activated sludge at any of the concentrations employed in the test. Respiration rates of mixtures containing the test substance showed an increase in respiration rate above that of the controls.
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Relevant effect levels: EC50 (3h) = 6.4 mg/L (4.1 - 9.6 mg/L)

Table 1: Preparation of test solutions

Treatment

Test substance [µL] or reference substance [mL]

Synthetic sewage [mL]

RO water [mL]

Microbial Inoculum [mL]

Control 1

0

16

284

200

Control 2

0

16

284

200

Test substance [mg/L]

 

 

 

 

10

5.1

16

284

200

100

50

16

284

200

1000

505

16

284

200

Reference substance [mg/L]

 

 

 

 

3

3

16

281

200

10

10

16

274

200

32

32

16

252

200

Table 2: Dissolved oxygen concentration and measurement times

Treatment

Initial dissolved oxygen conc. in culture [mg O2/L]

Initial measured dissolved oxygen conc. [mg O2/L]

Final measured dissolved oxygen conc. [mg O2/L]

Measurement time [min]

Control 1

6.9

6.5

2.5

8.5

Control 2

6.9

6.5

2.5

9.0

Test substance [mg/L]

 

 

 

 

10

4.5

4.2

1.5

5.3

100

5.0

4.8

1.5

6.3

1000

5.8

5.5

2.5

5.7

1000

4.9

4.6

1.5

5.8

1000

5.5

4.9

2.0

5.8

Reference substance [mg/L]

 

 

 

 

3

7.0

6.5

3.2

9.5

10

8.0

7.8

5.8

9.9

32

7.4

8.1

7.7

8.1

Table 3: Measurement of respiration rate

Treatment

Specific respiration rate [mg O2/g/h]

% inhibition or stimulation ()

Control 1

17.6

-

Control 2

16.7

-

Test substance [mg/L]

 

 

10

19.1

0 (11)

100

19.6

0 (14)

1000

19.7

0 (15)

1000

20.0

0 (17)

1000

18.8

0 (9)

Reference substance [mg/L]

 

 

3

13.0

24

10

7.6

56

32

1.9

89

Endpoint:
activated sludge respiration inhibition testing
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
Please refer to the analogue justification attached to IUCLID section 13.
Reason / purpose:
read-across source
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:
EC10
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Conclusions:
The key values for the toxicity to activated sluge microorganisms are a EC10 (3 h) of > 1000 mg/L (nominal), based on experimental data with one structurally and chemically closely related source substances (OECD 209, activated sludge).

Description of key information

Key value for chemical safety assessment

Additional information

Since no studies investigating the toxicity of Fatty acids, C8-10-(even numbered), esters with pentaerythritol and adipic acid (EC 948-383-1) to microorganisms are available, in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5, a read-across to the structurally related source substance Decanoic acid, mixed esters with heptanoic acid, octanoic acid, pentaerythritol and valeric acid (CAS 71010-76-9) was conducted. The source substance is representative to evaluate the toxicity of the target substance to microorganisms. This read-across is justified in detail within the analogue justification in IUCLID section 13.

The study with the source substance Decanoic acid, mixed esters with heptanoic acid, octanoic acid, pentaerythritol and valeric acid (CAS 71010-76-9) was conducted under GLP in a static test design according OECD Guideline 209 (Respiration Inhibition Test), using activated sludge from predominantly domestic sewage. Three nominal concentrations of 10, 100 and 1000 mg/L of the test substance were prepared. The respiration rate was measured after a test period of 3 h. The test substance showed no inhibitory effect on the respiration rate of activated sludge at any of the concentrations employed in the test. Therefore, effect values of EC50 (3 h) > 1000 mg/L (nominal) and EC10 (3 h) > 1000 mg/L (nominal) were determined.

Based on this result obtained from the source substance Decanoic acid, mixed esters with heptanoic acid, octanoic acid, pentaerythritol and valeric acid (CAS 71010-76-9) (in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5) which is characterized by a similar ecotoxicological profile and comparable structure, it can be concluded that the target substance Fatty acids, C8-10-(even numbered), esters with pentaerythritol and adipic acid (EC 948-383-1) is non-inhibitory of aquatic microorganisms activity and further testing for this endpoint is regarded not necessary.