N-ethyl-4-[[4-(ethylamino)-m-tolylphenyl][4-(ethylimino)-3-methylcyclohexa-2,5-dien-1-ylidene]methyl]-m-toluidine monoacetate

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2016-02-17 to 2016-04-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study performed according to OECD 301B.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

ISO DIS 9439 (Ultimate Aerobic Biodegradability - Method by Analysis of Released Carbon Dioxide)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

N-ethyl-4-[[4-(ethylamino)-m-tolylphenyl][4-(ethylimino)-3-methylcyclohexa-2,5-dien-1-ylidene]methyl]-m-toluidine monoacetate

EC Number:

305-170-2

EC Name:

N-ethyl-4-[[4-(ethylamino)-m-tolylphenyl][4-(ethylimino)-3-methylcyclohexa-2,5-dien-1-ylidene]methyl]-m-toluidine monoacetate

Cas Number:

94349-52-7

Molecular formula:

C28H38N3.C2H3O2

IUPAC Name:

[4-[bis[4-(ethylamino)-3-methyl-phenyl]methylene]-2-methyl-cyclohexa-2,5-dien-1-ylidene]-ethyl-ammonium;acetate

Details on test material:

- Chemical name: N-Ethyl-4-[[4-(ethylamino)-m-tolylphenyl][4-(ethylimino)-3-methylcyclohexa-2,5-dien-1-ylidene]methyl]-m-toluidinemonoacetate
- Physical state: solid / green
- Expiration date of the lot/batch: 01 July 2020
- Water solubility: miscible
- Storage conditions: Storage at room temperature; avoid direct sun light
- Total Organic Carbon: 734 mg/g

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Municipal activated sludge from the wastewater treatment plant of Mannheim, Germany.
- Preparation of inoculum for exposure: A suitable aliquot of the activated sludge suspension was sieved by a finely woven mesh with a mesh size about 1 mm. To reduce the content of inorganic carbon in the blank controls the activated sludge was aerated with carbon dioxide free air for about 48 hours at 22 ± 2° C. At the day of exposure the suspension was washed one time with drinking water. Therefore the aeration was stopped and the sludge was allowed to settle. After settling the supernatant was discarded and the remaining sludge suspension was filled up with drinking water.
- Concentration of sludge: 6 g/L dry weight
- Water filtered: yes
- Type and size of filter used, if any: Finely woven mesh with a size about 1 mm

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

15 mg/L

Based on:

test mat.

Initial test substance concentration 2

Initial conc.:

10 mg/L

Based on:

other: TOC nominal

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: The used mineral medium complies with the test guidline OECD 301B.
- Test temperature: 22 +/- 2 °C
- pH: 7.4 +/- 0.2
- pH adjusted: Yes
- Concentration of activated sludge: 30 mg/L dw

TEST SYSTEM
- Culturing apparatus: The test was performed in 2 L incubation bottles filled up to a volume of 1.5 L. The bottles were connected to two serial scrubbing bottles (total volume 250 mL) filled with 100 mL 0.05 mol sodium hydroxide solution for the adsorption of carbon dioxide from biodegradation processes.
- Number of culture flasks/concentration: 2 blank controls and 2 flasks per test substance concentration.
- Method used to create aerobic conditions: The aeration was performed with carbon dioxide free air at a flow of approximately 800 mL per hour for about 24 hours.
- Measuring equipment: TOC-analyser equipped with an auto sampler (Shimadzu TOC-5000A and/or TOC-L (CSH/CSN))
- Details of trap for CO2: The test bottles were connected to two serial scrubbing bottles (total volume of 1.5 L). The bottles were connected to two serial scrubbing bottles (total volume 250 mL) filled with 100 mL 0.05 mol sodium hydroxide solution for the adsorption of carbon dioxide from biodegradation processes.

SAMPLING
- Sampling frequency and method:
The TIC- and DOC-analyses were performed as repeat determination, using a TOCanalyzer equipped with an auto sampler (Shimadzu TOC-5000A and/or TOC-L CSH/CSN). The samples for the DOC-analysis were centrifuged for about 15 minutes at 4000 rpm. Measurement of Total Inorganic Carbon (TIC) of the adsorption solution was performed on day 2, 5, 8, 12, 14, 16, 19, 23, 26, 28 and used for the calculation of the produced carbon dioxide. Measurement of Dissolved Organic Carbon (DOC) was performed at beginning and at the end of exposure.
The DOC was calculated using the following formula:

TC - IC = TOC (since samples were centriuged TOC = DOC)
TC: total carbon
IC: inorganic carbon
TOC: total organic carbon

CONTROL AND BLANK SYSTEM
- Inoculum blank: Yes
- Inhibition control: Yes
- Reference substance control: Yes

Reference substance

Reference substance:

aniline

Results and discussion

Preliminary study:

No data

% Degradation

Parameter:

% degradation (CO2 evolution)

Value:

10 - 20

Sampling time:

28 d

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

other: not readily biodegradable

Conclusions:

The test substance was not readily biodegradable under the conditions of this test.