cis-2-(2,4-dichlorophenyl)-2-(1H-imidazol-1-ylmethyl)-1,3-dioxolane-4-ylmethyl methanesulphonate monohydrochloride

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:

2015-06-11 to 2015-07-03

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:

no

Qualifier:

according to guideline

Guideline:

EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO 8192 (Water quality - Test for inhibition of oxygen consumption by activated sludge for carbonaceous and ammonium oxidation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: I14KB4717
- Expiration date of the lot/batch: 206-11-25 (retest date)
- Purity test date: 2014-12-13 (certificate of analysis release date)

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature
- Stability under test conditions: no data

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stock solutions of 10 g/L were prepared. The test item (5.00 g) was added to 500 mL of Milli-RO water. Magnetic stirring for 30-36 minutes was applied to accelerate dissolution and to ensure homogeneity. Volumes of the clear and colorless stock solution (with a small foaming layer - pH3 3.4) corresponding to the test concentration were then added to the test media (16 mL synthetic medium and made up to 250 mL with Milli-RO). Thereafter, 250 mL of activated sludge was added resulting in the required concentrations (3.2, 10, 32, 100, 320 and 1000 mg/L). Optimal contact was ensured applying continuous aeration and stirring.
- Controls:blank controls: test medium without test item and treated in the same way as the test item solutions.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Laboratory culture: predominantly domestic sewage treatment plant, "Waterschap Aa en Maas", 's-Hertogenbosch, The Netherlands.
- Preparation of inoculum for exposure: coarsely sieved (1 mm) and allowed to settle. The supernatant was removed and ISO-medium was added. A small amount of the sludge was weighed and dried overnight at ~105°C to determine the amount of suspended solids
- Pretreatment: The batch of sludge was used one day after collection; therefore 50 mL of synthetic medium was added per L of activated sludge at the end of the collection day. The sludge was kept aerated at test temperature until use.
- Initial biomass concentration: 3.0 g/L of sludge

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

After 3 h of exposure, oxygen consumption was recorded for approximately 10 minutes.

Hardness:

not applicable

Test temperature:

20-22°C

pH:

Before addition of sludge: 5.2-7.6.
After the 3-h exposure period: 7.4-8.3

Dissolved oxygen:

The aeration was adjusted in such a way that the dissolved oxygen concentration at the start was above 60-70% saturation (60% of air saturation is > 5 mg/L at 20°C) and to maintain the sludge flocs in suspension.

Salinity:

not applicable

Nominal and measured concentrations:

Nominal concentrations: Control, 3.2, 10, 32, 100, 320, 1000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: All glass open bottles/vessels
- Aeration: The aeration was adjusted in such a way that the dissolved oxygen concentration at the start was above 60-70% saturation (60% of air saturation is > 5 mg/L at 20°C) and to maintain the sludge flocs in suspension..
- Number of replicates: 5
- Number of control replicates: 6
- Biomass loading rate: initial loading ca. 3.0 g dw/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
Adjusted ISO medium, formulated using RO-water (tap-water purified by reverse osmosis; GEON Waterbehandeling, Berkel-Enschot, The Netherlands) with the following composition:
CaCl2.2H2O: 211.5 mg/L
MgSO4.7H2O: 88.8 mg/L
NaHCO3: 46.7 mg/L
KCl: 4.2 mg/L

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): the oxygen consumption was recorded for a period of ~10 min. Determination of oxygen was performed with multiple oxygen probes connected to a BlueBox (GO-Systemelektronik GmbH, Germany), a multichannel measuring and controlling system.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Range finding study
- Test concentrations: 10.9, 109, 1090 mg/L
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

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

Key result

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:

- Effect concentrations exceeding solubility of substance in test medium: no data

Results with reference substance (positive control):

3-h EC50 = 4.5 mg/L.
This is within the recommended range of 2-25 mg/L confirming suitability of the activated sludge used.

Reported statistics and error estimates:

The 3-h EC50 values were based on Probit Analysis.
The NOEC determination was based on the STUDENT-t test for Homogeneous Variances with a Bonferroni-Holm Adjustment in the final test, alpha=., one sided, smaller).

Validity criteria fulfilled:

yes

Conclusions:

An activated sludge respiration inhibition test (according OECD guideline 209) with activated sludge from a predominantly domestic sewage treatment plant resulted in a 3-h EC50 > 1000 mg T001202/L. The results of the study can be considered reliable without restriction.

Description of key information

The study of Soeter (2015), investigating the toxicity of T001202 to microorganisms according to OECD guideline 209, was considered as the key study for endpoint coverage. The 3-h EC50 was > 1000 mg/L. The 3-h NOEC was 1000 mg/L.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

1 000 mg/L