2,2-dimethylpropane-1,3-diyl dibenzoate

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study without restriction

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge, domestic

Details on inoculum:

- Type: mixed population of aquatic microorganisms (activated sludge)
- Origin: aeration tank of a domestic sewage treatment plant (Municipal STP Cologne-Stammheim)
- Date of collection: 2014-07-07
- The sludge was settled and the supernatant was decanted. After centrifuging the sludge (15 min at 4000 rpm and 20°C) the supernatant was
decanted again. Approximately 1 g of the wet sludge was dried in order to calculate the amount of wet sludge to achieve a concentration of activated sludge of 3 g/L (dry weight) suspended solids. The calculated amount of sludge was dissolved in synthetic medium and then filled
up to a defined volume with deionised water.
- Storage of sludge: aeration of the activated sludge at 20°C ± 2°C, daily fed with synthetic medium
- pH of the suspension before application: 7.5
- Synthetic sewage feed : A synthetic waste water feed was prepared by dissolving the following amounts of substance per 1 litre of water
- pH of the synthetic sewage feed: 7.5 ± 0.5

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Hardness:

No data

Details on test conditions:

TEST SYSTEM:
- Before use the wet weight/dry weight relationship of the activated sludge was determined by drying 10 mL of sludge suspension. Subsequently, a sludge suspension of 2 g (dry weight)/L was prepared. The pH of this suspension was measured and adjusted to 6-8. 8 mL of the synthetic medium and 100 mL of activated sludge were added to the dissolved test item. The mixture was filled up with deionised water to 250 mL and aerated at 20 ± 2 °C.
- The exposure medium with the reference substance was prepared by adding 8 mL of the synthetic medium, 100 mL of activated sludge and a defined amount of the stock solution to achieve the test concentrations, and was filled up with deionised water to 250 mL and aerated at 20 ± 2°C. Control vessels (inoculated sample without test item) were prepared the same way.
- Additional vessels to determine the physico-chemical oxygen consumption were prepared containing the test item, and the synthetic medium, but no activated sludge.
- Oxygen consumption was measured and recorded after an aeration time of 3 hours in all these vessels starting with control 1. Thereafter, temperature and pH were measured as well. Then the other test vessels were measured. Control 2 terminated the measurements.

PRE-TREATMENT OF TEST- AND REFERENCE SUBSTANCE:
- Direct weighings were prepared to give the different test item concentrations
- The test item was added into Erlenmeyer flasks (incubation vessels) to about 130 mL deionised water and was stirred before testing (equilibration phase) for 1 hour.
- The pH was measured and ranged between pH 6.1-6.5. The pH was adjusted to pH 7.2-7.5 with NaOH.
- For the reference compound a stock solution at a concentration of 500 mg/L was prepared by dissolving 250 mg 3,5-dichlorophenol in 5 mL of 1 N NaOH and diluting to 0.5 litre with deionised water. The pH was adjusted to pH 7 ± 0.5 with HCl.

EXPOSURE CONDITIONS:
- Test item concentration/s: 1000 mg/L, 3 replicates
- Control : 6 replicates
- Test item concentration in physico-chemical oxygen consumption control: 1000 mg/L
- Concentration of reference compound 3,5-dichlorophenol: 2.5, 5, 10, 20 and 40 mg/L
- The test item and reference compound concentrations were not confirmed by analytical methods, they were based on nominal concentrations.
- Test vessels: 300 mL glass Erlenmeyer flasks
- Method of application: direct weighing
- Test concentration of the activated sludge: 800 mg/L suspended solids
- Test temperature: 20 ± 2°C
- Stirring period of the test item before start of incubation time: 1 hour
- Incubation time: 3 hours with permanent aeration

Reference substance (positive control):

yes

Remarks:

[3,5-Dichlorophenol (Acros Organics) (purity 99.9 %, batch number A0330535)]

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

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

estimated

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

EC10

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:

The effect value relates to a nominal concentration, since no analytical monitoring was performed.

Results with reference substance (positive control):

3 h-EC50: 6.963 mg/L (n)

Validity criteria fulfilled:

yes

Remarks:

(Respiratory rates of the 2 controls differ less than 15 % from each other. The EC50 of the reference compound 3,5-dichlorophenol is in the range 5–30 mg/L.)

Conclusions:

2,2-Dimethylpropan-1,3-diyl dibenzoate showed 5 % respiration inhibition of activated sludge at a test item concentration of 1000 mg/L, and hence the EC50 is higher than 1000 mg/L and the NOEC equal or higher than 1000 mg/L. The effect value relates to a nominal concentration.

Executive summary:

A study was performed to assess the toxicity of 2,2-dimethylpropan-1,3-diyl dibenzoate to bacteria.

The study was conducted in accordance with OECD TG 209 (Carbon and Ammonium Oxidation) (2010) and considered the Question-and-Answer Document by the German Federal Environment Agency (Version 2012-03-02). This test method is in most essential parts equal to Council Regulation (EC) No 440/2008, Method C.11 ”Biodegradation: Activated Sludge Respiration Inhibition Test” (2008).

The activated sludge was exposed to 2,2-dimethylpropan-1,3-diyl dibenzoate at a limit test item concentration of 1000 mg/L. The respiration rate of each mixture was determined after aeration periods of 3 hours.

2,2-Dimethylpropan-1,3-diyl dibenzoate showed 5% respiration inhibition of activated sludge at a test item concentration of 1000 mg/L. Hence the EC50 is higher than 1000 mg/L and the NOEC equal or higher than 1000 mg/L. This toxicity study is classified as acceptable and satisfies the guideline requirements for the toxicity study to microorganisms.

Description of key information

2,2-Dimethylpropan-1,3-diyl dibenzoate showed 5 % respiration inhibition of activated sludge at a test item concentration of 1000 mg/L, and hence the EC50 is higher than 1000 mg/L and the NOEC equal or higher than 1000 mg/L. The effect value relates to a nominal concentration. 

Key value for chemical safety assessment

EC50 for microorganisms:

1 000 mg/L

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

Should read: "EC50 >1000 mg/L" and "NOEC >=1000 mg/L"