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EC number: 701-396-4 | CAS number: -
Biodegradation in water: screening tests
Administrative data
Link to relevant study record(s)
- Endpoint:
- biodegradation in water: ready biodegradability
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 08 - 29 Sep 2016
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 310 (Ready Biodegradability - CO2 in Sealed Vessels (Headspace Test)
- Version / remarks:
- 2014
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- ISO 14593:1999 (Water quality - Evaluation of ultimate aerobic biodegradability of organic compounds in aqueous medium - Method by analysis of inorganic carbon in sealed vessels (CO2 headspace test))
- Version / remarks:
- 1999
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 835.3140 - Ready Biodegradability – CO2 in Sealed Vessels (Headspace Test)
- Version / remarks:
- 2008
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Hess. Ministerium für Umwelt, Energie, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany
- Oxygen conditions:
- aerobic
- Inoculum or test system:
- activated sludge, domestic, non-adapted
- Details on inoculum:
- - Source of inoculum/activated sludge: aeration tank of a municipal sewage treatment plant, Rossdorf, Germany
- Storage conditions: The washed sludge suspension was adjusted to 4 g dry material per litre and aerated overnight.
- Storage length: overnight
- Preparation of inoculum for exposure: The activated sludge was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in tap water and centrifuged again. This procedure was done three times. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to its dry weight was determined. Based on this ratio, calculated aliquots of washed sludge suspension were mixed with test water corresponding to 4 g dry material per litre and aerated overnight.
- Concentration of sludge: 4 mg/L (final concentration) - Duration of test (contact time):
- 28 d
- Initial conc.:
- 19.8 mg/L
- Based on:
- TOC
- Initial conc.:
- 29.5 mg/L
- Based on:
- test mat.
- Parameter followed for biodegradation estimation:
- CO2 evolution
- Details on study design:
- TEST CONDITIONS
- Composition of medium: Reconstituted water composed of analytical grade salts in pure water.
- Solubilising agent: acetone
- Test temperature: 20 ± 2 °C
- pH: 7.6
- pH adjusted: no
- Aeration of dilution water: Yes, the test water was aerated with CO2-free air prior to test start.
- Suspended solids concentration: 4 mg/L dw
- Continuous darkness: yes
TEST SYSTEM
- Culturing apparatus: Glass bottles with screw-caps and Teflon-coated septa.
- Number of culture flasks/concentration: 3 replicates per treatment and per sampling
- Method used to create aerobic conditions: Aeration of the activated sludge and the test water with CO2-free air prior to test start.
- Measuring equipment: TOC-Analzyer, TOC-V CPH, Shimadzu
- Test performed in closed vessels due to significant volatility of test substance: The test vessel was sealed with gas-tight septum caps.
- Details of trap for CO2 and volatile organics if used:
- Other: The headspace to liquid ratio was 1:2.
SAMPLING
- Sampling frequency: Test item and Inoculum Control (blank): Days 0, 3, 6, 10, 14, 17, 20, 28; Procedure Control and Toxicity Control: Days 0, 6, 14, 28
- Sampling method: The produced CO2 was converted into carbonate using 7 M NaOH solution (0.8 mL to 85 mL test medium) injected into the test vessels at each sampling. The alkalised medium was shaken for 1 h and subjected to total inorganic carbon (TIC) analysis. TIC consists of the carbon contained in carbonates and in carbon dioxide dissolved in water. By acidification all carbonates are converted to carbon dioxide. Carbon dioxide and dissolved carbon dioxide in the sample are volatilized by bubbling CO2-free air through the sample. The TIC-content was analyzed automatically by the TOC-Analyzer using the sealed test vessels as sampling vials.
- Sample storage before analysis: The analysis was done immediately after sampling or the alkalised samples were stored in the refrigerator (4 ± 4°C) for a maximum of 4 d. Samples of Day 0 were stored deep frozen at < -20 °C for 4 d before analysis.
CONTROL AND BLANK SYSTEM
- Inoculum blank: Yes, consisting of 0.5 mL acetone (evaporated overnight), activated sludge (4 mg/L) in test water (2500 mL).
- Toxicity control: Yes, consisting of the test item and the reference item in test water (1500 mL).
- Procedure Control: Yes, consisting of 0.5 mL acetone (evaporated overnight), reference item, activated sludge (4 mg/L) and test water (1500 mL). - Reference substance:
- benzoic acid, sodium salt
- Parameter:
- % degradation (CO2 evolution)
- Value:
- 60.7
- Sampling time:
- 14 d
- Key result
- Parameter:
- % degradation (CO2 evolution)
- Value:
- 60.4
- Sampling time:
- 28 d
- Details on results:
- The 10 d window started after 3 d of incubation, when the mean biodegradation of the test item was 10.4% (ThIC). No sample sample was taken after 13 d, therefore the results of day 14 sampling are used to determine the ready biodegradability of the test item. After 14 d of incubation, the mean biodegradation of the test item was 60.7% (ThIC). Afterwards the degradation remained almost constant in the plateau phase and ranged between degradation values of 60.4 - 72.3% (ThIC). Formally, the 10 d window criterion was not passed. However, it is considered justified to use the degradation value after 14 d since the testing regime as stipulated in OECD 310 does not allow a continuous measurement of biodegradation comparable to OECD 301B where a 14 d window is applicable.
- Results with reference substance:
- The reference item sodium benzoate was sufficiently degraded to 84.3% after 14 days and to 66.0% after 28 days of incubation.
- Validity criteria fulfilled:
- yes
- Interpretation of results:
- readily biodegradable
- Conclusions:
- The degradation rate of the test item did reach 60% within the 10-day window and after 28 days.
Therefore, LZ 53478 is considered to be readily biodegradable.The reference item sodium benzoate was sufficiently degraded to 84.3% after 14 days and to 66.0% after 28 days of incubation.
The percentage biodegradation of the reference item confirms the suitability of the used aerobic activated sludge inoculum.
Reference
The reference item (sodium benzoate) was sufficiently degraded to 84.3% after 14 d and to 66.0% after 28 d of incubation (Table 1). The percentage of the reference item confirms the suitability of the used aerobic activated sludge inoculum.
In the toxicity control containing both, the test item and the reference item, 77.8% biodegradation was noted within 14 d and 65.6% biodegradation after 28 d of incubation (Table 1). Therefore, the test item is not inhibitory.
Table 1. Biodegradation of Test Item, Reference Item and Toxicity Control during the Test Period of 28 Days.
|
Day |
Day |
||||||||||||||||
Treatment |
C-content mg/L |
replicate |
0 |
3 |
6 |
10 |
14 |
17 |
20 |
28 |
0 |
3 |
6 |
10 |
14 |
17 |
20 |
28 |
TICcorrected[mg/L]1 |
% biodegradation [net measured TIC/initial C x 100] |
|||||||||||||||||
Reference substance |
19.824 |
1 |
0.00 |
-- |
7.87 |
-- |
16.71 |
-- |
-- |
13.24 |
0.0 |
-- |
39.7 |
-- |
84.3 |
-- |
-- |
66.8 |
2 |
0.00 |
-- |
7.69 |
-- |
16.76 |
-- |
-- |
12.95 |
0.0 |
-- |
38.8 |
-- |
84.5 |
-- |
-- |
65.3 |
||
3 |
0.72 |
-- |
8.17 |
-- |
16.69 |
-- |
-- |
12.91 |
3.6 |
-- |
41.2 |
-- |
84.2 |
-- |
-- |
65.1 |
||
4 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
13.18 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
66.5 |
||
5 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
13.19 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
66.5 |
||
Mean |
0.24 |
|
7.91 |
|
16.72 |
|
|
13.09 |
1.2 |
|
39.9 |
|
84.3 |
|
|
66.0 |
||
Test substance |
19.812 |
1 |
0.00 |
2.31 |
3.28 |
9.82 |
11.70 |
14.36 |
10.72 |
12.52 |
0.0 |
11.6 |
16.5 |
49.6 |
59.1 |
72.5 |
54.1 |
63.2 |
2 |
0.00 |
2.39 |
3.25 |
10.12 |
12.04 |
14.03 |
12.86 |
11.97 |
0.0 |
12.1 |
16.4 |
51.1 |
60.8 |
70.8 |
64.9 |
60.4 |
||
3 |
0.00 |
1.49 |
2.96 |
9.72 |
12.33 |
14.59 |
12.79 |
12.39 |
0.0 |
7.5 |
15.0 |
49.1 |
62.2 |
73.6 |
64.6 |
62.5 |
||
4 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
11.36 |
-- |
|
-- |
-- |
-- |
-- |
-- |
57.3 |
||
5 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
11.62 |
-- |
|
-- |
-- |
-- |
-- |
-- |
58.7 |
||
Mean |
0.00 |
2.06 |
3.16 |
9.88 |
12.02 |
14.32 |
12.13 |
11.97 |
0.0 |
10.4 |
16.0 |
49.9 |
60.7 |
72.3 |
61.2 |
60.4 |
||
Toxicity Control |
19.824 |
1 |
0.00 |
-- |
5.89 |
-- |
15.60 |
-- |
-- |
12.83 |
0.0 |
-- |
29.7 |
-- |
78.7 |
-- |
-- |
64.7 |
2 |
0.06 |
-- |
5.51 |
-- |
15.52 |
-- |
-- |
13.08 |
0.3 |
-- |
27.8 |
-- |
78.3 |
-- |
-- |
66.0 |
||
3 |
0.23 |
-- |
5.62 |
-- |
15.14 |
-- |
-- |
12.86 |
1.2 |
-- |
28.4 |
-- |
76.4 |
-- |
-- |
64.9 |
||
4 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
13.06 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
65.9 |
||
5 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
13.20 |
-- |
-- |
-- |
-- |
-- |
-- |
-- |
66.6 |
||
Mean |
0.10 |
|
5.68 |
|
15.42 |
|
|
13.01 |
0.5 |
|
28.6 |
|
77.8 |
|
|
65.6 |
||
1: corrected by the inoculum control
--: not applicable
Description of key information
Readily biodegradable: 60.4% after 28 d, 10 d-window fulfilled (OECD 310, activated sludge)
Key value for chemical safety assessment
- Biodegradation in water:
- readily biodegradable
Additional information
One GLP study is available, which assessed the ready biodegradability of Polyaldo 2-1-IS (CAS 73296-86-3) according to the OECD guideline 310.
Non-adapted activated sludge from a domestic waste water treatment plant was inoculated with a nominal concentration of 29.5 mg test item/L, corresponding to 19.8 mg carbon/L (based on organic carbon content) for 28 d in controlled, aerobic conditions in the dark. An inoculum, procedural and toxicity control were run in parallel. The biodegradation of the test substance was determined by following the CO2evolution of the test item in the sealed incubation flasks during exposure. The degradation rate of the test item was calculated by the CO2evolution of the aerobic activated sludge microorganisms by measuring the inorganic carbon in the test flasks corrected by control vessels.
The reference substance was sufficiently degraded to 84.3% after 14 d and to 66.0% after 28 d incubation, thus confirming the suitability of the aerobic activated sludge inoculum used. The toxicity control indicated that the test item is not inhibitory to the aerobic activated sludge microorganisms. The 10 d window started after 3 d of incubation, when the mean biodegradation of the test item was 10.4% (ThIC). No sample sample was taken after 13 d, therefore the results of day 14 sampling are used to determine the ready biodegradability of the test item. After 14 d of incubation, the mean biodegradation of the test item was 60.7% (ThIC). Afterwards the degradation remained almost constant in the plateau phase and ranged between degradation values of 60.4 - 72.3% (ThIC). Formally, the 10 d window criterion was not passed. However, it is considered justified to use the degradation value after 14 d since the testing regime as stipulated in OECD 310 does not allow a continuous measurement of biodegradation comparable to OECD 301B where a 14 d window is applicable. Therefore, the test substance is considered to be ready biodegradable.
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