Sodium 6,14-diethyl-1,1,1,2,2,18,18,19,19,19-decafluoro-8,12-dioxo-10-({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy}carbonyl)-4,7,13,16-tetraoxanonadecane-9-sulfonate

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08 OCT 2012 - 07 MAY 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 (Carbon and Ammonium Oxidation))

Version / remarks:

22 July 2010

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Gewerbeaufsichtsamt Hildesheim (5 November 2013)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test item was warmed-up to 30 °C for 30 minutes. Afterwards the test item was weighed out directly in Erlenmeyer flasks, filled up with 200 mL demineralised water and stirred overnight (450 upm).
- Controls: blank control (test medium without test or reference item), positive control (reference item: Copper (II) sulphate pentahydrate)

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Name and location of sewage treatment plant where inoculum was collected: Municipal sewage treatment plant of 31137 Hildesheim, Germany
- Preparation of inoculum for exposure/ Pretreatment: The sludge was washed twice with tap water and adjusted to a dry sludge concentration of 3.0 g/L ± 10 %. The sludge was used within 24 h after sampling.

Test type:

static

Water media type:

freshwater

Remarks:

demineralised water

Limit test:

yes

Total exposure duration:

3 h

Remarks on exposure duration:

none

Post exposure observation period:

none

Test temperature:

21.1 °C

pH:

pH-value of the activated sludge: 7.43
pH-value of the synthetic waste water: 7.45

Nominal and measured concentrations:

Nominal concentrations 100, 320, 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 500 m L Erlenmeyer flasks, DIN 12380
- Material, size, headspace, fill volume: glass, 500 mL, DIN 12380
- Aeration: Permanent, to keep the dissolved oxygen concentration above 60 - 70 % saturation and to maintain the sludge flakes in suspension
- No. of vessels per concentration (replicates): 5 replicates per concentration
- No. of vessels per control (replicates): Triplicates
- No. of vessels per abiotic control (replicates): Triplicates per concentration
- Sludge concentration (weight of dry solids per volume): dry sludge concentration 3.17 g/L, corresponding to 1.59 g/L suspended solids in the test vessel
- Nutrients provided for bacteria: 16 mL Synthetic waste water acc. to OECD TG 209
- Nitrification inhibitor used: none

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Demineralised water

OTHER TEST CONDITIONS
- Adjustment of pH: The pH value of the activated sludge was determined prior to test start, the pH value of the synthetic waste water was determined prior to use. Adjustment to 7.5 ± 0.5 was not necessary.

EFFECT PARAMETERS MEASURED: NOEC, EC50

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Range finding study : Concentrations were based on a non-GLP range finding test (100 - 500 - 1000 mg/L).
- Test concentrations: 100 - 320 - 1000 mg/L

Reference substance (positive control):

yes

Remarks:

Copper (II) sulphate pentahydrate was used as reference item and the reference toxicity was determined.

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

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

320 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks on result:

other: No statistically significant effect (p < 0.05), ANOVA, Dunnett's Method

Details on results:

- Blank controls oxygen uptake rate: 34.5 mg 02/g·h
- Coefficient of variation of oxygen uptake rate in control replicates: 8.1 %

for detailed results please refer to attached table under 'Attached background material'

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: EC50 (3h) = 104 mg/L (95% Confidence lnterval = 102 - 107 mg/L). This is in the recommended range of validity of 53 - 155 mg/L.

Reported statistics and error estimates:

The NOEC was determined by calculation of statistical significance of the inhibition of respiration in comparison to the control. One Way Analysis of Variance (ANOVA) was used for NOEC calculations. When running a One Way Analysis of Variance a Normality test and an Equal Variance test were done first. P-values for both Normality and Equal Variance test are 0.05. The a-value (acceptable probability of incorrectly concluding that there is a difference) is a=0.05.
The EC50-value of the reference item was calculated by linear regression using software GraphPad Prism. Calculation of the confidence intervals for the EC-values was carried out using standard procedures.

Validity criteria fulfilled:

yes

Conclusions:

The NOEC of the test item is 320 mg/L.
The test item is not toxic up to the concentration of 320 mg/L to activated sludge of a municipal sewage treatment plant.

Executive summary:

A Respiration Inhibition Test with activated sludge according to OECD Guideline No. 209 was carried out with the test item in compliance with GLP. Test system was activated sludge of a municipal treatment plant. The test was carried out under static conditions with the substance concentrations of 100 - 320 - 1000 mg/L. The respiration rates of the control, reference and test item replicates were measured after a contact time of three hours, and the inhibitory effects of the test and reference item were determined in comparison to the control respiration rates. The mean inhibition of respiration for the test item replicates were in the range of 2 % to 36 %.

In order to check the activity of the test system and the test conditions a reference test was carried out with copper (II) sulphate pentahydrate as reference item and the reference toxicity was determined. The EC₅₀-value for the reference item was 104 mg/L. The NOEC of the test item is 320 mg/L.

The test item is not toxic up to the concentration of 320 mg/L to activated sludge of a municipal sewage treatment plant.

Description of key information

NOEC (3h) = 320 mg/L and EC₅₀ (3h) > 1000 mg/L for activated sludge based on respiration inhibition (OECD 209, GLP)

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

320 mg/L

Additional information

Respiration inhibition test with activated sludge, OECD 209

A Respiration Inhibition Test with activated sludge according to OECD Guideline No. 209 was carried out with the test item in compliance with GLP. Test system was activated sludge of a municipal treatment plant. The test was carried out under static conditions with the substance concentrations of 100 - 320 - 1000 mg/L. The respiration rates of the control, reference and test item replicates were measured after a contact time of three hours, and the inhibitory effects of the test and reference item were determined in comparison to the control respiration rates. The mean inhibition of respiration for the test item replicates were in the range of 2 % to 36 %.

In order to check the activity of the test system and the test conditions a reference test was carried out with copper (II) sulphate pentahydrate as reference item and the reference toxicity was determined. The EC₅₀ value for the reference item was 104 mg/L.

The NOEC of the test item is 320 mg/L. The test item is not toxic to activated sludge of a municipal sewage treatment plant up to the concentration of 320 mg/L.