1-(3-chloropropyl)-1,3-dihydro-2H-benzimidazol-2-one

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:

2015-06-18 to 2015-07-24

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)

Remarks:

see study report

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No. of test material: A14JB3442
- Expiration date of the lot/batch: 2016-09-30 (retest date)
- Purity test date: 2015-02-18

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature

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 not sufficiently soluble to allow the preparation of a 10 g/L stock solution in water. Therefore, 1 L test bottles were filled with 200 mL of test item mixtures in Milli-RO water with initial loading rates of 2.5 times the final loading rate. These mixtures were stirred in closed dark brown bottles for ~24 hours. Subbsequently, 16 mL synthetic medium made up to 50 mL with Milli-RO water and 250 mL sludge were added resulting in the required loading rates. 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:

- combined limit/range-finding test: 17-23 °C. The temperature was slightly outside the range prescribed by the protocol (20 ± 2°C) during the exposure phase. Evaluation: The validity criteria for the controls and reference item were met and therefore this deviation is considered not to have influenced the sludge.
- final test: 18-22 °C

pH:

- combined limit/range-finding test:
* Before addition of sludge: 7.5-7.7
* After the 3-h exposure period: 7.4-8.4
- final test:
* Before addition of sludge: 7.6-7.7
* After the 3-h exposure period: 7.5-8.2

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:
- Range-finding test: 10, 100, 1000 mg/L
- Final test: 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 (final test)
- 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)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): 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: yes
- Test concentrations range finding study: loading rates of 10, 100, 1000 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:

other: ELR10

Effect conc.:

36 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: ELR50

Effect conc.:

540 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:

other: NOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- Blank controls oxygen uptake rate: The mean blank control oxygen uptake rate exceeded 20 mg O2/g.h (30 and 25 mg O2/g.L for the combined limit/range finding and final test respectively).

Results with reference substance (positive control):

3-h EC50 = 2.1 and 2.4 mg/L (for the range-finding and limit test respectively).
This is within the recommended range of 2-25 mg/L confirming suitability of the activated sludge used.

Reported statistics and error estimates:

The EC50 values were based on Probit Analysis using linear maximum likelihood regression with the percentage of respiration inhibition versus the logarithms of the corresponding loading rates of the test item.
The NOELR determination was based on the Welch-t test for inhomogeneous variances with Bonferroni-Holm Adjustment, alpha=0.05, 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 = 540 mg T001036/L. In conclusion, JNJ-119717-AAA (T001036) was not toxic to activated sludge at or below a loading rate of 100 mg/L. The results of the study can be considered reliable without restriction.

Description of key information

The study of Desmares-Koopmans (2016), investigating the acute toxicity of T001036 to microorganisms according to OECD guideline 209, was considered as the key study for endpoint coverage.  The 3-h ELR50 was 540 mg/L. The 3-h NOELR was 100 mg/L.

Key value for chemical safety assessment

EC50 for microorganisms:

540 mg/L

EC10 or NOEC for microorganisms:

100 mg/L

Additional information

One key study investigating the toxicity to aquatic microorganisms was available (Desmares-Koopmans, 2016). This study exposed activated sludge of predominantly domestic sewage to a series of test substance concentrations (10, 32, 100, 320 and 1000 mg/L) during 3 hours in a static test design, at a temperature of 20°C. The test item was not sufficiently soluble to allow the preparation of a 10 g/L stock solution in water. Therefore, the test item – Milli-RO water mixtures were magnetically stirred for a period of approximately 24h. The criteria for a valid test were fulfilled. The inhibitory effect of the test item T001036 on the respiration rate of aerobic wastewater microorganisms of activated sludge resulted in a 3-h ELR50 of 540 mg/L. The 3-h NOELR and 3-h ELR10 were determined to be 100 and 36 mg/L respectively.