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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-09-08 to 2010-09-28
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
Volume of the 500 mL test medium
Composition of 1 mg/L:test medium
200 mL inoculum
16 mL synthetic waste water
234 mL autoclaved tap water
50 mL test item stock solution

3.2 mg/L:
200 mL inoculum
16 mL synthetic waste water
124 mL autoclaved tap water
160 mL test item stock solution

10 - 320 mg/L:
200 mL inoculum
16 mL synthetic waste water
34 mL autoclaved tap water
Test item suspension (250 mL)
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
A fresh sample of activated sludge taken from the sewage plant Sarstedt was used for the test procedure. It was washed twice and adjusted with autoclaved tap water to a dry sludge concentration of 4.0 g/L ± 10
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
3 h
Test temperature:
21°C
pH:
pH value of the activated sludge 6.67
Dry sludge concentration [g/L] 4.19
Details on test conditions:
TEST METHOD Static
Test duration 3 hours
Replicates One per concentration and two controls
Test vessels 500 mL Erlenmeyer flasks, DIN 12380
Nutrient solution Synthetic waste water according to OECD Guideline 209
Volume of the 500 mL test medium
Aeration Permanent
Reference substance (positive control):
yes
Remarks:
Copper (II) sulphate pentahydrate p.a.
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
29.9 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Remarks on result:
other: 95% CL
Duration:
3 h
Dose descriptor:
EC10
Effect conc.:
2.65 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
VALIDITY CRITERIA The difference between the two control respiration rates should be not more than 15 %.

The EC50 of the reference item should be in the range of
54 - 156 mg/L. The range of validity (EC50  3 SD) of copper (II) sulphate pentahydrate is calculated from the mean of all reference tests (n = 298) from 1991 - 2010.
Results with reference substance (positive control):
EC50 = 135 mg/L
This is in the recommended confidence range of 54 - 156 mg/L.
Reported statistics and error estimates:
The EC-values and confidence intervals of the test item were calculated by linear regression using software GraphPad Prism5, GRAPHPAD SOFTWARE, INC.

The EC50-value of the reference item was calculated with probit analysis using software SIGMAPLOT (Windows), SPSS CORPORATION. Probit values were taken from WEBER (1986).

Table2:    Inhibitions in the Preliminary Tests

Nominal Test Item Concentration

Inhibition
[%]

 

1ststudy

2ndstudy

3rdstudy

[mg/L]

No stirring

24 h stirring

1 h stirring

1

- 7

10

17

2

52

100

45

41

81

1000

- 5

34

2

Table3:   Respiration Rates and Inhibition of Respiration of Test and Reference Item Concentrations

 

Concentration

Respiration rate

Inhibition

 

[mg/L]

[mgO2/(L×h)]

[%]

Test item

320

9.2

42

 

100

4.4

72

 

32

8.0

49

 

10

10.8

32

 

3.2

14.0

11

 

1

16.4

-4

Control 1

16.4

Control 2

15.2

Reference item

180

5.6

65

 

100

10.8

32

 

58

12.8

19

 

EC-values

Confidence interval

 

[mg/L]

P = 95 [%]

[mg/L]

EC20

4.85

-

EC50

29.9

23.6 - 39.3

EC80

n.d.

-

Solubility of the Test Item

 

After stirring for 1 h all test item concentrations were dissolved. The colours of the solutions were slightly brown (10 mg/L stock solution and 10 mg/L), brown (32 mg/L and 100 mg/L) and colourless to slightly yellow (320 mg/L).

Validity criteria fulfilled:
yes
Conclusions:
The EC 50 of the test item p-Aminophenol is 29.9 mg/L and EC10 is 2.65 mg/L to activated sludge of a municipal sewage treatment plant.

Description of key information

One key experimental study was reported,  Respiration Inhibition Test with Activated Sludge with EC50 of 29.9 mg/L.

Key value for chemical safety assessment

EC50 for microorganisms:
29.9 mg/L
EC10 or NOEC for microorganisms:
2.65 mg/L

Additional information

For short-term toxicity to micro-organisms, there is one key study conducted in 2010, Respiration Inhibition Test with Activated Sludge according to OECD 209 with an EC50 of 29.9 mg/L.