3-methyl-6-(p-toluidino)-3H-dibenz[f,ij]isoquinoline-2,7-dione

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Version / remarks:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Type : mixed population of aquatic microorganisms (activated sludge)
Origin : aeration tank of a wastewater plant treating predominantly domestic sewage (Wupper area water authority, WWTP Odenthal)
Date of collection : 2017-03-06
Pre-treatment of the inoculum
− Before use, the inoculum was stored for two days at room temperature under continuous stirring with aeration.
− The sludge was washed twice by adding mineral medium and centrifuging for 10 min at 2000 rpm and 20 °C and decanting off the supernatant.
− An aliquot of the wet sludge was dried in order to determine the wet weight / dry weight ratio of the sludge and to prepare a stock suspension (activated sludge) of 3 g dw/L.
− The calculated amount of sludge, needed to achieve 300 mL of this stock suspension, was dissolved in mineral medium and then filled up to a defined end volume.
Sludge concentration: 30 mg/L suspended solids

Duration of test (contact time):

28 d

Initial conc.:

100 mg/L

Based on:

test mat.

Details on study design:

25 mg of the test item were weighed out and added to the test flasks, filled with 200 mL of mineral medium. Afterwards the volume was made up to 250 mL with mineral medium containing the inoculum to give a test concentration of 100 mg test item/L.
Pre-treatment of the toxicity control
25 mg of the test item and 25 mg of the reference compound were weighed out and added to the test flasks, filled with 200 mL of mineral medium. Afterwards the flask volume was made up to 250 mL with mineral medium containing the inoculum to give a test concentration of 100 mg test item and reference compound/L.

Reference substance:

benzoic acid, sodium salt

Test performance:

The oxygen demand was measured daily. Between 0 and 28 days all resultes were 0% biodegradation.

Key result

Parameter:

% degradation (O2 consumption)

Value:

0

Sampling time:

28 d

Results with reference substance:

Reference substance benzoic acid Na salt showed 90% degradation after 28 days. In the toxicity control 39% degradation were found indicating that no relevant intoxication occurred.

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

In a ready biodegradation test according to OECD 301F no biodegradation (0%) was found after 28 days. The reference substance sodium benzoate was degraded by 90% after 28 days.

Executive summary:

In accordance with the Council Regulation (EC) No 440/2008, Method C.4-D “Manometric Respirometry Test“(2008), which is in all essential parts identical with OECD Guideline 301 F (adopted July 1992), a suspension of 100 mg/L test item in a mineral medium, equalling to 50-100 mg ThOD or COD/Litre as the nominal sole source of organic carbon, was stirred in a closed flask and inoculated at a constant temperature (22 ± 1 °C) for up to 28 days under aerobic conditions in the dark. The consumption of oxygen (BOD) was determined by measuring the drop in pressure in the automated respirometer flasks. Evolved carbon dioxide was absorbed in sodium hydroxide. The amount of oxygen taken up by the test item (corrected for uptake by blank inoculum, run in parallel) was expressed as a percentage of theoretical oxygen demand (ThOD). The endogenous activity of the inoculum was checked running parallel blanks with inoculum but without test item. A reference compound (sodium benzoate) was run in parallel to check the operation of the procedures. A toxicity control (test item and reference compound mixed, one replicate) was run in parallel, to ensure that the chosen concentration of the test item was not inhibitory to microorganisms. Degradation was followed by the determination of oxygen uptake and measurements were taken at frequent intervals to allow the identification of the beginning and end of biodegradation and the slope of the biodegradation curve. The test lasted for 28 days. Because of the nature of biodegradation and of the mixed bacterial populations used as inoculum, determinations of test item and inoculum blank were carried out in triplicate and of reference compound in duplicate. The oxygen uptake was calculated from the readings taken at regular and frequent intervals, using the method given by the manufacturer of the equipment. At the end of incubation, the pH was measured in the flasks. The test item is an N-containing substance. Therefore, the concentration of nitrite and nitrate was determined at the start of the test. The oxygen consumed by nitrification was not determined after 28 days, because no degradation of the test item was observed. No biodegradation (0%) was found after 28 days of exposure. The reference substance sodium benzoate was degraded by 90% after 28 days. The toxicity control revealed no relevant sign of intoxication. Therefore the test is considered to be reliable.

Description of key information

In a ready biodegradation test according to OECD 301F no biodegradation (0%) was found after 28 days of exposure. The reference substance sodium benzoate was degraded by 90% after 28 days.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

The toxicity control revealed no relevant sign of intoxication. Therefore the test is considered to be reliable.