4,4-dimethyl-3,5,8-trioxabicyclo[5.1.0]octane

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:

21 Mar 2002

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study was performed according to OECD 209 (1984) and fulfilled validation criteria according to the guideline. The two control respiration rates differ not more than 15% and the EC50 (3h) of the reference substance (copper (II) sulphate pentahydrate) were in the range of 54-146 mg/L. However, validity criteria according OECD 209 (adopted 2010) were not fully met since oxygen uptake rate in the control was less than 20 mg oxygen per one gram of activated sludge (dry weight).

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Version / remarks:

1984

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Remarks:

The test item and reference substance concentrations are not confirmed by analytical methods as they are directly weighed into the test vessels.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: test substance and reference substance were weighted directly into flasks and stirred for 24 hours.
- Controls: Yes, blank control and positive control (reference substance) :
- Test concentration separation factor: 1.8

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 Sarstedt
- Preparation of inoculum for exposure: at test begin 16 mL synthetic waste water, 34 mL autoclaved tap water and 200 mL activated sludge were added to 250 mL test solution containing the test substance in different concentrations.
- Pretreatment: sewage sludge was collected one day before the start of the experiment, the sewage sludge was washed twice with autoclaved tap water and kept under aerobic conditions.
- Initial biomass concentration: 4.05 g/L (dry sludge concentration).

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Test temperature:

20.5°C

pH:

6.6

Dissolved oxygen:

2.92 to 5.52 mg O2/L

Nominal and measured concentrations:

100, 180, 320, 580, 1000 mg/L (nominal concentration)

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flask
- Material, size, headspace, fill volume: 0.5 L
- No. of vessels per concentration (replicates): 1 replicate
- No. of vessels per control (replicates): 2 replicates (blank control), 1 replicate (positive control).
- Sludge concentration (weight of dry solids per volume): 4.05 g/L
- Nutrients provided for bacteria: synthetic waste water

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

EFFECT PARAMETERS MEASURED
- respiration inhibition after 3 hours of exposure.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Test concentrations: 100, 180, 320, 580, 1000 mg/L (nominal concentration).

Reference substance (positive control):

yes

Remarks:

Copper (II) sulphate pentahydrate

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

Details on results:

- Blank controls oxygen uptake rate: 23 mg O2/L*h

Results with reference substance (positive control):

- Results with reference substance valid? yes, all validity criteria are fulfilled
- Relevant effect levels: EC50 (3h)=120 mg/L

Validity criteria for the measurement of the bacterial toxicity

Target condition according to guideline:	Actual condition according to the study:	Validity criteria met:
The blank controls (without the test substance or reference substance) oxygen uptake rate should not be less than 20 mg oxygen per one gram of activated sludge (dry weight of suspended solids) in an hour. If the rate is lower, the test should be repeated with washed activated sludge or with the sludge from another source. The coefficient of variation of oxygen uptake rate in control replicates should not be more than 30% at the end of definitive test.	The oxygen uptake rate of the controls were 21.2 and 24.8 mg/L*h (1.6 g/L suspended solids). The study was performed according to OECD 209 (adopted 1984) and fulfilled validation criteria according to the guideline. The two control respiration rates differ not more than 15% and the EC50 (3h) of the reference substance (copper (II) sulphate pentahydrate) were in the range of 54-146 mg/L. However, validity criteria according OECD 209 (adopted 2010) were not fully met since oxygen uptake rate in the control was less than 20 mg oxygen per one gram of activated sludge (dry weight).	No
Reference substances In a 2004 international ring test organized by ISO using activated sludge derived from domestic sewage, the EC50 of 3,5-dichlorphenol was found to lie in the range 2 mg/L to 25 mg/L for total respiration, 5 mg/L to 40 mg/L for heterotrophic respiration and 0.1 mg/L to 10 mg/L for nitrification respiration. If the EC50 of 3,5-DCP does not lie in the expected range, the test should be repeated with activated sludge from another source. The EC50 of copper (II) sulphate pentahydrate should lie in the range of 53-155 mg/L for the total respiration.	The EC50 value for the reference substance (Copper (II) sulphate pentahydrate; total respiration) was 120 mg/L (nominal concentration).	Yes
The test should be performed at a temperature within the range 20±2°C.	During the test the temperature in the test vessels was maintained at 20.5 C°.	Yes

Validity criteria fulfilled:

no

Remarks:

See 'Any other information on results incl. tables' for further explanation.

Conclusions:

The respiration of bacteria was not inhibited in any of the test concentrations, thus the test substance is not toxic in concentrations

Executive summary:

A study was performed to assess the toxicity of the test substance to bacteria. The test was conducted in accordance with the OECD Guideline for Testing of Chemicals No. 209 'Activated Sludge, Respiration Inhibition Test (Carbon and Ammonium Oxidation)'. The activated sludge was exposed to the test substance at different concentration (100, 180, 320, 580, 1000 mg/L; nominal concentration). The respiration rate of each mixture was determined after a exposure period of 3 hours. The test substance showed 0% respiration inhibition of activated sludge at test substance concentrations up to 1000 mg/L (nominal concentration).

Description of key information

A study was performed to assess the toxicity of the test substance to bacteria. The test was conducted in accordance with the OECD Guideline for Testing of Chemicals No. 209 'Activated Sludge, Respiration Inhibition Test (Carbon and Ammonium Oxidation)'. The activated sludge was exposed to the test substance at different concentration (100, 180, 320, 580, 1000 mg/L; nominal concentration). The respiration rate of each mixture was determined after a exposure period of 3 hours. The test substance showed 0% respiration inhibition of activated sludge at test substance concentrations up to 1000 mg/L (nominal concentration).