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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23.05.2018 - 24.05.2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Version / remarks:
1981
Deviations:
no
GLP compliance:
no
Remarks:
accredited by DAkkS
Analytical monitoring:
no
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
Activated sludge from an urban municipal waste water treatment facility was used as inoculum. It was washed with tap water in the laboratory. For this purpose, the activated sludge was transferred into a large beaker and was allowed to settle for a few minutes. The aquous supernatant was removed by decanting. Subsequently, slurrying of the residue was conducted with tap water. This process was repeated for 3 times and served as purification of the activated sludge from possible inhibitors. After the last decantation, no slurrying was performed so that a higher concentration was achieved. Subseqently, the dry matter was determined and adjusted to 3,2% by a renewed addition of tap water (the value was determined by a renewed measurement of the dry matter). The prepared activated sludge like this was used as inoculum and supplied with oxygen during the whole test duration by permanent stirring.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Test temperature:
20 +/- 2°C
pH:
see table 2
Dissolved oxygen:
see table 2
Nominal and measured concentrations:
6,4 mg/l, 16 mg/l, 40 mg/l, 100 mg/l, 250 mg/l
Details on test conditions:
The test was performed in single samples with a test volume of 500 ml. Solely, the control sample was performed in dupilcate. All samples contained 16 ml synthetic wastewater (for the composition see table 1) and 200 ml inoculum with a dry matter of 3.2%. In the control samples 284 ml tap water was added and in the test samples 284 ml methlydroquinone in different concentrations. The reference sample contained tap water and mercury chloride in a concentrion of 10 mg/l.
Reference substance (positive control):
yes
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
22.15 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
EC10
Effect conc.:
4.58 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
other: EC20
Effect conc.:
6.79 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
other: EC80
Effect conc.:
72.26 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Results with reference substance (positive control):
see table 2 and 3

Table 2: Oxygen content and pH-value of control samples, test samples and reference samples

t (min)

O2 content in mg/l

control samples

test samples

reference sample

C1

C2

6,4 mg/l

16 mg/l

40 mg/l

100 mg/l

250 mg/l

0,0

7,96

7,80

6,40

8,14

8,09

8,51

8,27

8,48

0,5

7,25

7,51

6,03

7,96

7,97

8,48

8,21

8,46

1,0

6,81

7,22

5,74

7,80

7,84

8,46

8,14

8,45

1,5

6,47

6,91

5,49

7,66

7,75

8,31

8,06

8,43

2,0

6,19

6,65

5,24

7,54

7,65

8,21

8,00

8,41

2,5

5,91

6,37

4,97

7,41

7,57

8,17

7,94

8,37

3,0

5,65

6,06

4,72

7,27

7,37

8,12

7,87

8,36

3,5

5,39

5,78

4,45

7,14

7,38

8,06

7,80

8,34

4,0

5,13

5,49

4,19

7,01

7,30

8,02

7,74

8,31

4,5

4,86

5,20

3,92

6,86

7,19

7,97

7,66

8,30

5,0

4,59

4,93

3,66

6,73

7,11

7,91

7,62

8,27

5,5

4,32

4,64

3,39

6,61

7,03

7,87

7,54

8,25

6,0

4,06

4,35

3,13

6,47

6,94

7,82

7,48

8,24

6,5

3,78

4,06

2,86

6,34

6,84

7,77

7,41

8,20

7,0

3,53

3,75

2,60

6,20

6,76

7,72

7,35

8,19

7,5

3,26

3,47

2,33

6,06

6,68

7,67

7,27

8,17

8,0

2,98

3,19

2,07

5,91

6,60

7,62

7,22

8,13

8,5

2,71

2,90

1,81

5,78

6,52

7,57

7,15

8,11

9,0

2,45

2,58

1,57

5,64

6,43

7,52

7,09

8,09

9,5

2,18

2,32

1,32

5,50

6,34

7,48

7,03

8,07

10,0

1,92

2,02

1,06

5,36

6,27

7,43

6,97

8,05

 

pH

7,28

7,48

7,49

7,72

7,74

7,74

7,52

7,74

Table 3: Oxygen consumption of control samples, test samples and reference samples in mg O2/l*h and %

Sample

concentration (mg/l)

Oxygen consumption

mg O2/l*h

mean

%

control

0

36,24

35,46

100

0

34,68

test sample

6,4

32,04

 

90,36

16

16,68

47,04

40

10,92

30,80

100

6,48

18,27

250

7,8

22,00

mercury chloride

10

2,58

7,28

Validity criteria fulfilled:
yes
Remarks:
The validity of the test is given since the oxygen consumption in the control samples only deviate by 4.3%. Moreover, the respiration inhibition in the reference sample reached a value of 93%.
Conclusions:
The respiration inhibition of activated sludge for methylhydroquinone was investigated using a respirometer in a study according to OECD test guideline 209. The 3 h EC50 was determined to be 22.15 mg/L (nominal).
Executive summary:

The OECD 209 activated sludge, respiration inhibition test was performed as a method for assessing the potential impact of the test substance on wastewater treatment system. Optimal contact between the test substance and test medium was ensured applying continuous aeration during the 3-hour exposure period. Thereafter, oxygen consumption was recorded for 10 minutes. Five concentrations, ranging from 6.4 to 250 mg/l were tested.

A statistically significant inhibition of the respiration rate of the activated sludge was recorded and an EC50 of 22.15 mg/l was determined by linear regression.

Description of key information

The respiration inhibition of activated sludge for methylhydroquinone was investigated using a respirometer in a study according to OECD test guideline 209. The EC50 (3 h) was determined to be 22.15 mg/L.

Key value for chemical safety assessment

EC50 for microorganisms:
22.15 mg/L

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