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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
Details on inoculum
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): aeration tank of a wastewater plant treating predominantly domestic sewage (Wupper area water authority, WWTP Odenthal)
- Date of collection :2017-06-12
- Storage conditions: one day at room temperature under continuous stirring with aeration
- Pretreatment: 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.
- Concentration of sludge: 30 mg/L suspended solids
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Test temperature: 22 ± 1 °C
- Aeration of dilution water: yes
- Suspended solids concentration: 30 mg/L
- Continuous darkness: yes
- pH: 7.7 - 8.2 after 28 days

TEST SYSTEM
- Culturing apparatus: OxiTopControl System (WTW)
- Test volume : 250 mL
- Mixing : 1 magnetic stirrer per test vessel
- 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).
- 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.
- 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.

CONTROL AND BLANK SYSTEM
- 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.
Pre-treatment:25 mg of the reference compound were weighed out on aluminium foil and both were 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 reference compound /L..
- 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.
Pre-treatment: 25 mg of the test item and 25 mg of the reference compound were weighed out on aluminium foil and were 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
Parameter:
% degradation (O2 consumption)
Value:
35
Sampling time:
28 d
Details on results:
- The used concentration of the test item is not toxic to bacteria.
Results with reference substance:
Kinetic of reference substance ( % Degr.):
81 after 7 days
88 after 14 days
91 after 21 days
93 after 28 days
Validity criteria fulfilled:
yes
Remarks:
(-Ready biodegradability of reference compound 60 percent within 14 d -Toxicity control exhibited degradation rates > 25 % within 14 d -Replicates difference< 20% -Oxygen uptake of blank inoculum =< 60mg/L -No pH influence (6.0 and 8.5 at test end))
Interpretation of results:
not readily biodegradable
Conclusions:
Within 28 days, a degradation rate of 35 % was determined for Anthraquinone-2-sulfonic acid sodium salt monohydrate.
Executive summary:

This study was conducted in accordance with the Council Regulation (EC) No 440/2008, Method C.4-D “Manometric Respirometry Test“(2008). This test method is in all essential parts identical with OECD Guideline 301 F (adopted July 1992).

To assess the ready biodegradability Anthraquinone-2-sulfonic acid sodium salt monohydrate was stirred in a 250 ml closed flask and inoculated at a constant temperature (22 ± 1 °C) for up to 28 days under aerobic conditions in the dark. The inoculum (activated sludge) was collected from an aeration tank of a wastewater plant treating predominantly domestic sewage. 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 Anthraquinone-2-sulfonic acid sodium salt monohydrate (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 Anthraquinone-2-sulfonic acid sodium salt monohydrate 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 oxygen uptake was calculated from the readings taken at regular and frequent intervals, using the method given by the manufacturer of the equipment. After 28 days a pH of 7.7 - 8.2 was measured in the flasks. The suspended solids concentration was 30 mg/L. The concentration of Anthraquinone-2-sulfonic acid sodium salt monohydrate was 100 mg/L. The reference compound sodium benzoate showed 88 % degradation after 14 days.

Within 28 days, a degradation rate of 35 % was determined for Anthraquinone-2-sulfonic acid sodium salt monohydrate.

Anthraquinone-2-sulfonic acid sodium salt monohydrate is considered to be "Not Readily Biodegradable", but seems to be in principle biodegradable.

Description of key information

Within 28 days, a degradation rate of 35 % was determined for Anthraquinone-2-sulfonic acid sodium salt monohydrate.

Key value for chemical safety assessment

Biodegradation in water:
under test conditions no biodegradation observed

Additional information