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Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2008-07-10 to 2008-08-28
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD Guideline study report

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
2-methylbenzene-1,4-diyl bis{4-[4-(acryloyloxy)butoxy]benzoate}
EC Number:
Cas Number:
Molecular formula:
2-methylbenzene-1,4-diyl bis{4-[4-(acryloyloxy)butoxy]benzoate}
Constituent 2
Reference substance name:
4-({4-[4-(Acryloyloxy)butoxy]benzoyl}oxy)-2-methylphenyl 4-[4-(acryloyloxy)butoxy]benzoate
4-({4-[4-(Acryloyloxy)butoxy]benzoyl}oxy)-2-methylphenyl 4-[4-(acryloyloxy)butoxy]benzoate
Details on test material:
Name: Me-3K4A2
CAS No.: 132900-75-5
Chemical Name(s): Benzoic acid, 4-[4-[(1-oxo-2-propenyl)oxy]butoxy]-, 2-methyl-1.4-phenylene ester
Empirical Formula / Molecular Weight: C7H6O2 / 122.12 g/mol (based on Benzoic acid)
Theoretical Oxygen Demand: ThODNH4: 1.965 mg oxygen per mg test item (based on Benzoic acid)
Physical Appearance / Colour: Powder / beige
Stability: In water: test item is considered to be stable under test conditions
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
not applicable

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
Aerobic activated sludge, micro organisms from a domestic waste water treatment plant was supplied by the sewage plant Darmstadt Germany.
The aerobic activated sludge used for this study 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, corresponding to 1.5 g dry material per litre were mixed with test water. This suspension was used for the experiment.
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
102 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
- Test temperature: 20 +/- 2 °C
- pH: 7.6 (measured at the start of the test); 7.5–7.7 (measured at the end of the test)
- pH adjusted: no
- Suspended solids concentration: 0.1 mg/ml
- Continuous darkness: yes

- Culturing apparatus: BSB Sensomat system, Aqualytic Langen
- Number of culture flasks/concentration: 2
- Test performed in closed vessels due to significant volatility of test substance: no
- Details of trap for CO2 and volatile organics if used: not available

- Sampling frequency: daily

- Inoculum blank: yes
- Abiotic sterile control: yes
- Toxicity control: yes
Reference substance
Reference substance:
benzoic acid, sodium salt

Results and discussion

Preliminary study:
not available
Test performance:
The test item was exposed to aerobic activated sludge from the aeration tank of a domestic waste water treatment plant for 28 days. The biodegradation was followed by the oxygen uptake of the micro organisms during exposure. As a reference item sodium benzoate was tested simultaneously under the same conditions as the test item, and functioned as a procedure control. The test flasks were incubated at 20 +/- 2 °C. The pressure decrease in the reaction vessels was measured over complete experimental phase of 28 days using the BSB Sensomat system, Aqualytic Langen. The test flasks were closed gas-tight by a measuring head. Potassium hydroxide solution (45%) was used for trapping the produced carbon dioxide. The amount of O2 consumed by the activated sludge was calculated from the decrease of pressure in the reaction vessel.
% Degradationopen allclose all
% degradation (CO2 evolution)
Sampling time:
1 d
Remarks on result:
other: none
% degradation (CO2 evolution)
Sampling time:
5 d
Remarks on result:
other: none
% degradation (CO2 evolution)
Sampling time:
10 d
Remarks on result:
other: none
% degradation (CO2 evolution)
Sampling time:
14 d
Remarks on result:
other: none
% degradation (CO2 evolution)
Sampling time:
27 d
Remarks on result:
other: none
Details on results:
Time Percentage Biodegradation1
(Days) Me-3K4A2 1 Sodium Benzoate 2 Toxicity control 1, 2
Flask 1 [%] Flask 2 [%] Flask 5 [%] Flask 7 [%]
1 0 0 29 11
2 1 -1 45 21
3 0 0 70 27
4 -1 -1 78 33
5 0 0 82 35
6 0 0 88 36
7 0 0 91 36
8 0 0 94 38
9 2 0 94 38
10 1 1 95 38
11 0 0 94 38
12 0 0 97 39
13 0 0 97 39
14 2 2 100 40
15 2 2 100 40
16 2 2 103 42
17 2 2 105 42
18 1 1 104 41
19 0 0 105 42
20 2 0 105 42
21 1 1 107 42
22 0 0 105 42
23 0 0 105 42
24 0 0 105 42
25 0 0 108 42
26 2 0 108 43
27 2 2 108 43
28 2 2 108 43
1 ThODNH4 of Me-3K4A2: 1.965 mg/mg
2 ThODNH4 of sodium benzoate: 1.666 mg/mg

BOD5 / COD results

Results with reference substance:
Reference item was completely degraded within 14 days.

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
not inherently biodegradable
Substance is not readily biodegardable.
Executive summary:

The ready biodegradability of Me3K4A2 was assessed in a manometric respirometry test according to OECD Guideline 301 F over a period of 28 days. Aerobic activated sludge (micro-organisms from a domestic waste­water treatment plant) was supplied by the sewage works of,.The biodegradation was followed by the oxygen uptake of the micro-organisms during exposure. As a reference item sodium benzoate was tested simultaneously under the same conditions as the test item, and functioned as a procedure control. Degradation rate of test item calculated by the oxygen consumption of the aerobic activated sludge microorganisms after 28 days of incubation.. The Test Item Loading Rate was 104 mg/L corresponding to an oxygen demand of about 205 mg/L (ThODNH4). The criterion for ready biodegradability under the conditions of a manometric respirometry test is the 10-day window, describing the period between reaching at least 10% degradation and 60% degradation. This period should not exceed 10 days. The mean biodegradation after 28 days of Me3K4A2 was 2% (ThODNH4); the 10 day window criterion was not passed. Therefore, Me-3K4A2 is considered not to be readily biodegradable. The reference item sodium benzoate was sufficiently degraded to a mean of 100% after 14 days and to a mean of 108% after 28 days of incubation, thus confirming the suitability of the aerobic activated sludge inoculum used. In the toxicity control containing both, the test item and the reference item sodium benzoate, a mean of 40% biodegradation was noted within 14 days and 43% biodegradation after 28 days of incubation. Thus, the test item can be assumed to be not inhibitory to the aerobic activated sludge micro organisms.