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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:
2010-06-08 till 2010-07-20
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 E (Ready biodegradability: Modified OECD Screening Test)
Version / remarks:
(1992)
GLP compliance:
yes (incl. certificate)
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic (adaptation not specified)
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, STP Odenthal)
- Date of collection: 2010-06-08
- 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.
− 1.049 g 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.:
ca. 100 mg/L
Based on:
test mat.
Details on study design:
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

Pre-treatment of the test item:
- 25 mg of test item were weighed out, added to the test flasks and filled with 200 mL of mineral medium
- 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 reference compound (sodium benzoate (Sigma-Aldrich))
- 25 mg of the reference compound were weighed out, added to the test flasks and filled with 200 mL of mineral medium
- 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.

Pre-treatment of the toxicity control
- 25 mg of the test item and 25 mg of the reference compound were weighed out, added to the test flasks and filled with 200 mL of mineral medium
- 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
Remarks:
Sigma-Aldrich, purity: 99.7 %, batch-no.: 1438955
Parameter:
% degradation (O2 consumption)
Value:
72
Sampling time:
28 d
Details on results:
37 % degradation after 7 days
54 % degradation after 14 days
67 % degradation after 21 days
Results with reference substance:
The reference compound sodium benzoate showed 79 % degradation after 14 days.

With an overall degradation rate of 72 % within 28 days, and about 50 % COD removal within the 10-d-window, the substance clearly exhibits its potential to become degraded by aerobic bacteria under environmental conditions. The pass levels for ready biodegradability, as defined by OECD 301, have only marginally been failed to reach. It appears that, with more favourable conditions or a slightly elongated exposure period, the substance would be easily degraded to the levels required by OECD 301.

The degradation curves are given in figure 1.

The pH-values at the end were between 7.5 and 8.1.

Validity criteria fulfilled:
yes
Remarks:
reference compound reached the level of ≥ 60 %, toxicity control exhibited degradation rates > 25 %, oxygen uptake of the inoculum blank was ≤ 60 mg/L
Interpretation of results:
readily biodegradable, but failing 10-day window
Conclusions:
The substance is considered as readily biodegradable but failing the 10d-window.
Executive summary:

To assess the ready biodegradability of the substance, a 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).

A solution of 100 mg/L substance in a mineral medium 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 determinations of the biodegradation of the test item were carried out in duplicate.

The endogenous activity of the inoculum was checked running parallel blanks with inoculum but without test item in triplicate. A reference compound (sodium benzoate) was run in duplicate to check the operation of the procedures.

A toxicity control (test item and reference compound mixed) was run in parallel, to ensure that the chosen concentration of the test item was not inhibitory to microorganisms.

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 to be 7.5 - 8.1.

For the substance a degradation of 72 % after 28 days was observed. Due to the degradation within the 10-day window was less than 60 % removal, the substance is considered to be “Readily Biodegradable but failing 10-day window“.

Description of key information

To assess the ready biodegradability of the substance, a study was conducted in accordance with the Council Regulation (EC) No 440/2008, Method C.4-D “Manometric Respirometry Test“ (2008). Since the degradation was 72 % after 28 days but the degradation within the 10-d window was less than 60 % removal, the substance is considered to be “Readily Biodegradable, but failing 10day-window“.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable but failing 10-day window

Additional information

A study on inherent biodegradability supports the information above as a degradation of 78 % after 28 days was observed for the substance in a study which was conducted in accordance with the OECD Guideline 302C “Inherent Biodegradability: Modified MITI Test (II)” (1981). Thus the substance a is considered to be "Inherently Biodegradable".