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Toxicity to microorganisms

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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2011-12-07 - 2012-01-30
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The GLP study was conducted according to an internationally accepted guideline. All study parameters are given in detail.
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Details on sampling:
In the control vessels 16 mL nutrient solution were mixed with 234 mL water. The positive control and the treatments were prepared by putting the appropriate amount of positive control solution respectively test item into the test vessel, adding 16 mL nutrient solution and water to give 250 mL. Then, 250 mL inoculum was added in five minute intervals and the mixture was aerated.

After three hours, the content of the first vessel was poured in a 250 mL narrow-neck bottle and the respiration rate was determined by measurement of the 02-concentration over a period of five minutes1. The following vessels were measured likewise in five minute intervals.
Vehicle:
no
Details on test solutions:
Chemicals and Nutrient Solution

All chemicals used in the test were "analytical grade" or "for use in microbiology".

Composition of nutrient solution (synthetic sewage):

Peptone 16.0 g

Meat Extract 11.0 g

Urea 3.0 g

NaCI 0.7 g

CaCI2*2H20 0.4 g

MgS04*7H20 0.2 g

K2HP04 2.8 g

Deionised water ad 1000 mL

The measured pH of the solution was 7.1. Therefore no pH adjustment to pH 7.5 ± 0.5 was necessary.
The nutrient solution was freshly prepared; the inoculum was taken from its source, washed, aerated and the dry matter was determined. The stock solutions of the test item and the positive control were prepared.
Concentrations of test and positive control were calculated using the concentration of the respective stock solution and the dilution factor.
The test item was added to the test vessels directly.
Test organisms (species):
activated sludge, domestic
Details on inoculum:
The sludge was taken from the activation basin of the ESN (Stadtentsorgung Neustadt) sewage treatment plant D-67435 NW-Lachen-Speyerdorf and aerated during transport.

Upon arrival in the test facility, the sludge was filtrated, washed with tap water and re-suspended in tap water. The activated sludge was aerated until usage in the test and fed daily with 50 ml /L synthetic sewage feed.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Hardness:
1.08 mmol/l
Test temperature:
18.1-18.8 "C
pH:
7.9 - 8.1
Nominal and measured concentrations:
Nominal:
1008, 1068, 1036, 1086, 1040, 101.4, 11.2, 1 mg/l
Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol
Key result
Duration:
3 h
Dose descriptor:
other: EC20
Remarks:
EC20
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Key result
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:
NOEC
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:
see below
Results with reference substance (positive control):
see below

Vessel

Content

Concentration

02consump­

02consump­

Inhibition

pH

No.

 

in mg/L

tion in

tion in

in%

 

 

 

 

mg/(L*min.)

mg/(L*h)

 

 

1

Control

0

0.4495

26.970

 

8.0

2

Control

0

0.4290

25.738

8.0

3

Positive Control

5

0.3011

18.063

34.5

8.1

4

Positive Control

10

0.1958

11.748

57.4

8.1

5

Positive Control

20

0.1201

7.205

73.9

8.1

6

Positive Control

40

0.0683

4.097

85.1

8.2

7

Control

0

0.4705

28.230

8.0

8

Control

0

0.4625

27.748

 

8.0

9

Test Item

1008

0.4706

28.235

-2.4

8.0

10

Test Item

1068

0.4832

28.991

-5.1

7.9

11

Test Item

1036

0.4849

29.095

-5.5

7.9

12

Test Item

1086

0.4732

28.394

-2.9

7.9

13

Test Item

1040

0.4872

29.231

-6.0

7.9

14

Test Item

101.4

0.4800

28.798

-4.4

7.9

15

Test Item

11.2

0.4662

27.973

-1.4

7.9

16

Test Item

1.0

0.4765

28.590

-3.6

8.0

17

Control

0

0.4738

28.431

8.0

18

Control

0

0.4733

28.400

 

8.0

Validity criteria fulfilled:
yes
Conclusions:
The following results for the test item Ukanol FR 70 were determined:

3h NOEC = 1000 mg/L
3h EC20 > 1000 mg/L
3h EC50 > 1000 mg/L
Executive summary:

The toxicity of Ukanol FR 70 towards microorganism was assessed according to OECD test method 209. One experiment was performed. As the EC50 was found as > 1000 mg/L and the highest test concentration of 1000 mg/l showed no significant inhibition, no further testing was done. All validity criteria were met. For the estimation of the EC50 of the positive control 3,5-dichlorophenole, the fit showed good statistical correspondence of the data with the dose-response-equation. The positive control gave an EC50 of 8.3 mg/L which is within the recommended range of 2 - 25 mg/L. The coefficient of variation of oxygen uptake rate in control replicates was below 30% at the end of the test. In the all treatments (range 1000 - 1 mg/L), no inhibition could be observed. The O2-consumption in all replicates of treatment 1000 mg/L was higher than the mean of the controls. Therefore 1000 mg/L can be stated as NOEC without an additional experiment for statistical determination.

Description of key information

The following results for the test item Ukanol FR 70 were determined:

3h NOEC = 1000 mg/L

3h EC20 > 1000 mg/L

3h EC50 > 1000 mg/L

Key value for chemical safety assessment

EC50 for microorganisms:
1 000 mg/L

Additional information