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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
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, 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: 2012-08-22
- Concentration of inoculum: 30 mg/L suspended solids
Pre-treatment of the inoculum:
- 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.
- Before use, the inoculum was stored for one day at room temperature under continuous shaking with aeration.
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Details on study design:
A suspension of 100 mg/L active ingredient of the test test item (= 283.7 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 chemical oxygen demand (COD).
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) was not run in parallel, because 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 degradation rate of the test item was set in relation to its experimentally determined COD. As the COD implicitly covers the oxygen demand for the nitrification process, there was no need to take additional correction measures into account.
Exposure conditions:
Test volume : 250 mL
Test apparatus : OxiTopControl System (WTW)
Mixing : 1 magnetic stirrer per test vessel
Incubation time : 28 days
Incubation temperature : 22 ± 1 °C

Reference substance:
benzoic acid, sodium salt
Remarks:
Purity : 99.7 %
Parameter:
% degradation (O2 consumption)
Value:
29
Sampling time:
7 d
Parameter:
% degradation (O2 consumption)
Value:
99
Sampling time:
14 d
Parameter:
% degradation (O2 consumption)
Value:
100
Sampling time:
21 d
Parameter:
% degradation (O2 consumption)
Value:
100
Sampling time:
28 d
Details on results:
Biodegradation started on day 6 with 20% degradation and reached 100% on day 15.
Parameter:
COD
Value:
0.728 mg O2/g test mat.
Results with reference substance:
The reference compound sodium benzoate showed 88 % degradation after 14 days.
Validity criteria fulfilled:
yes
Remarks:
Ready biodegradability of reference compound ≥ 60 percent within 14 days. -In the toxicity control degradation rates > 25 % within 14 days. -Replicates difference< 20% . - Oxygen uptake of blank inoculum=< 60mg/l. -No pH influence
Interpretation of results:
readily biodegradable
Conclusions:
A test on ready biodegradability was conducted according to OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test). After 28 days showed 100 % degradation fulfilling the 10 day window.

Executive summary:

This study was designed to assess the ready biodegradability of BAYSCRIPT Blaukomponente (unkonserviert, filtriert). It 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). A suspension of 100 mg/L active ingredient of the test test item (= 283.7 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 chemical oxygen demand (COD). 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) was not run in parallel, because 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 degradation rate of the test item was set in relation to its experimentally determined COD. As the COD implicitly covers the oxygen demand for the nitrification process, there was no need to take additional correction measures into account. The 28-day degradation was 100 % and for the positive control (with reference substance) the biodegrdation was 88 %. Therefore, the substance is considered to be “Readily Biodegradable, fulfilling the 10 day window“.

Description of key information

A test on ready biodegradability was conducted according to OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test). After 28 days showed 100 %  degradation fulfilling the 10 day window. 

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

In accordance with REACH Regulation, Art. 3, and ECHA "Guidance for identification and naming of substances under REACH and CLP" v1.2, March 2012, substances have to be separated from solvents if this is not affecting the stability of the substance or changing its composition.

In this case, separation of water would only be possible by thermical damaging of the substance. Furthermore, it was not possible to spray dry the pigment solution as it stuck to and partly melted on the conus dryer.