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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23rd January 2014 to 13th February 2014
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
Deviations:
yes
Remarks:
- the temperature in the definitive test exceeded the required range by 0.5°C outlined in the study plan. The maximum temperature recorded was 22.5°C (required range from study plan = 20±2°C)
GLP compliance:
yes
Vehicle:
no
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Source: Cambridge Sewage Treatment Works, Cowley Road, Cambridge
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
3 h
Post exposure observation period:
10 to 15 minutes
Hardness:
No data
Test temperature:
21 - 22.5°C
pH:
7.89 was recorded for 1000 mg/l therefore no adjustment required and measurements for lower test concentrations not necessary.
Dissolved oxygen:
Controls: 0.19 - 8.37
Test 10 to 1000 mg/l: 1.04 - 9.13
(See attached report for full results)
Nominal and measured concentrations:
- Preliminary test: 0 (Control) 10, 100, 1000 mg/l
- Main test: 0 (Control), 10, 32, 100, 320, 1000 mg/l
Details on test conditions:
TEST SYSTEM
- Test vessel: 500ml flasks with magnetic stirrer bars and aeration and glass BOD bottles for Dissolved Oxygen measurement
- Type: closed
- Material, size, headspace, fill volume: 300ml (final test volumes)
- Aeration: Yes
- No. of vessels per concentration (replicates): 5
- No. of vessels per control (replicates): 6 (in total)
- Amount of test substance per vessel (not controls): 3 ml of 3.2 g/l test stock
- Amount of synthetic sewage feed (all vessels): 9.6 ml
- Amount of 3.75 g/l microbial inoculum (all vessels): 120 ml

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: High purity deionised water
- See attached report for details of prepared medium

OTHER TEST CONDITIONS
- Adjustment of pH: Only required for reference stock (1.0 g/1 3,5 - Dichlorophenol) - adjusted to pH 7.37.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): 3h EC20, EC50 and EC80

PRELIMINARY TEST
- Range finding study: Yes
- Test concentrations: 0 (Control), 10, 100, 1000 mg/l
- Results used to determine the test concentrations and validate the conditions for the definitive study:
Mean control respiration rate = 29.16 mg O2/hour
Mean control respiration rate as mg O2/g dry weight activated sludge/hour = 19.44
Coefficient of variation for controls = 3.49%

MAIN TEST
- Test concentrations: 0 (Control), 10, 32, 100, 320, 1000 mg/l
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol (3,5-DCP)
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
31.8 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% confidence limits: 28.5 - 56.0 mg/l
Duration:
3 h
Dose descriptor:
other: EC20
Effect conc.:
8.86 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% confidence limits: 7.07 - 12.3 mg/l
Duration:
3 h
Dose descriptor:
other: EC80
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Results with reference substance (positive control):
- Results with reference substance valid? Yes

- Relevant effect levels:
3 hour EC20 = 1.06 mg/l (95% confidence limits: 1.05 - 1.07 mg/l)
3 hour EC50 = 4.60 mg/l (95% confidence limits: 4.52 - 4.69 mg/l)
3 hour EC80 >18 mg/l

Reported statistics and error estimates:
EC values were calculated by linear interpolation in ToxCalcTM Version 5.0.

- Test type: Activated Sludge Respiration Test

- Test period: 23 to 24 January 2014 - preliminary test

                       12 to 13 February 2014 - definitive test

- Test concentrations: 10, 100 and 1000 mg/l - preliminary test; 10, 32, 100, 320 and 1000 mg/l - definitive test (Reference substance (3,5-DCP): 2, 6 and 18 mg/l for both preliminary test and definitive test).

- Definitive test results:

 

3 hour exposure period

ECx values (mg/l)

EC20

EC50

EC80

Test substance

8.86 (7.07 - 12.3)

31.8 (28.5 - 56.0)

>1000

3,5-DCP (Reference)

1.06 (1.05 - 1.07)

4.60 (4.52 - 4.69)

>18

Validity criteria fulfilled:
yes
Conclusions:
- Epofloc L-1R causes respiration inhibition to microorganisms although the inhibition is significantly less than the reference substance.
- The 3-hour EC50 was determined to be 31.8 mg/l during the definitive test.
- The validity criteria were all met for the definitive test.

Description of key information

The 3h EC50 was determined to be 31.8 mg/l

Key value for chemical safety assessment

EC50 for microorganisms:
31.8 mg/L

Additional information

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