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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
Study period:
03 Jul 2017 - 04 Aug 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Version / remarks:
Adopted July 2017, 1992
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
C11H17NO2 (Molecular formula used for calculating ThOD)
Oxygen conditions:
aerobic
Inoculum or test system:
sewage, predominantly domestic (adaptation not specified)
Details on inoculum:
- Source of inoculum/activated sludge: secondary effluent freshly obtained from a municipal sewage treatment plant: 'Waterschap Aa en Maas', 's-Hertogenbosch, The Netherlands, receiving predominantly domestic sewage.
- Preparation of inoculum for exposure: Secondary effluent was filtered through a coarse filter paper, the first 200 mL was discarded. The filtrate was kept aerated until inoculation.
- Inoculation: 4 mL filtrate of secondary effluent per litre of final volume
Duration of test (contact time):
28 d
Initial conc.:
1 mg/L
Based on:
test mat.
Initial conc.:
3 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
PREPARATION OF TEST SOLUTIONS
Preparation of the test media started with a clear and colourless stock solution of 100 mg/L by adding 50.1 mg test item to 500 mL of mineral medium. Magnetic stirring (17 min.) and sonication (21 min.) was used to accelerate dissolution of the test item in mineral medium. Exact volumes of the stock solution corresponding to the test concentrations and microbial organisms were added to the test medium.

TEST CONDITIONS
- Composition of medium: Mineral medium according to OECD 301D
- Test temperature: 22.0 - 23.1°C.
- pH: 7.4-7.5
- pH adjusted: no
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 250-300 mL BOD bottles with glass stoppers.
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: aeration of filtered secondary effluent until inoculation.
- Measuring equipment: WTW oxygen meter supplied with a WTW CellOx 325 oxygen electrode, electrolyte type ELY/G.

SAMPLING
- Sampling frequency: In duplicate; immediately at the start of the oxygen concentration experiment (day 0), and at day 7, 14, 21 and 28.
- Sampling method: oxygen concentration measurement with oxygen meter

CONTROL AND BLANK SYSTEM
- Inoculum blank: containing only inoculum (no test item)
- Abiotic sterile control: not included.
- Toxicity control: containing test item at the lowest concentration, reference item and inoculum
- Procedure control: containing reference item and inoculum
Reference substance:
acetic acid, sodium salt
Remarks:
at 2 mg/L
Key result
Parameter:
% degradation (O2 consumption)
Value:
-4
Sampling time:
28 d
Remarks on result:
other: Test item concentration 1 mg/L
Key result
Parameter:
% degradation (O2 consumption)
Value:
-1
Sampling time:
28 d
Remarks on result:
other: Test item concentration 3 mg/L
Details on results:
The ThOD of the test item was calculated to be 2.54 mg O2 per mg, with nitrogen eliminated as nitrate.
The ThOD of sodium acetate (reference item) was calculated to be 0.78 mg O2/mg.

The relative biodegradation values calculated from the O2 measurements performed during the test period of 28 days revealed no significant biodegradation of test item at both concentrations. See Table 1 in 'Any other information on results' for details on the biodegradation percentages at different points in time.

In the toxicity control more than 25% biodegradation occurred within 14 days (36%, based on ThOD). Thus, the toxicity control showed that the test item did not inhibit microbial activity.
Results with reference substance:
The reference substance was degraded by 70% on day 14.

Table 1: Biodegradation Percentages at Different Points in Time

Test medium

Concentration

% Biodegradation after x days(1)

(mg/L)

0

7

14

21

28

 

 

 

 

 

 

 

Procedure control A(2)

2

0

85

68

Procedure control B(2)

2

0

83

71

Procedure control mean A and B

2

0

84

70

Difference between duplicate values

2

0

3

4

Test item low A(3)

1

0

-4

-6

-6

-5

Test item low B(3)

1

0

-1

-3

0

-3

Test item low mean A and B

1

0

-3

-5

-3

-4

Difference between duplicate values

1

0

3

4

6

2

Test item high A(3)

3

0

-1

1

-1

0

Test item high B(3)

3

0

0

-2

0

-1

Test item high mean A and B

3

0

0

-1

0

-1

Difference between duplicate values

3

0

1

3

1

1

Toxicity control A

2 + 1(4)

0

27

33

 

 

Toxicity control B

2 + 1(4)

0

26

39

 

 

Toxicity control mean A and B

2 + 1(4)

0

26

36

 

 

Difference between duplicate values

2 + 1(4)

0

1

6

 

 

(1): Except for the percentages biodegradation, all calculations were performed without rounding off.

(2): ThOD positive control sodium acetate: 0.78 mg O2/mg

(3): ThOD test item: 2.54 mg O2/mg

(4): Toxicity control contains sodium acetate and test item.

Validity criteria fulfilled:
yes
Remarks:
See "Overall remarks" for details on validity criteria
Interpretation of results:
not readily biodegradable
Conclusions:
The test itemwas not readily biodegradable under the conditions of the closed bottle test presently performed.
Executive summary:

The ready biodegradation of the substance under the conditions of the closed bottle test was investigated according to OECD guideline 301 D and GLP principles. Test concentrations of 1 and 3 mg/L were tested in duplicate during 28 days. The relative biodegradation values calculated from the O2 measurements performed during the test period of 28 days revealed no significant biodegradation of the substance. Since biodegradation of at least 60% was not reached within 14 days of biodegradation exceeding 10%, the substance was concluded to be not readily biodegradable. In the toxicity control, the substance was found not to inhibit microbial activity. The criteria for acceptability of the test were met and the study is considered to be reliable without restriction. In conclusion, the substance was determined to be not readily biodegradable under the conditions of the closed bottle test.

Description of key information

The substance was determined to be not readily biodegradable under the conditions of the closed bottle test (OECD 301D). The substance was found not to inhibit microbial activity.

Key value for chemical safety assessment

Biodegradation in water:
under test conditions no biodegradation observed
Type of water:
freshwater

Additional information

The ready biodegradation of the substance under the conditions of the closed bottle test was investigated according to OECD guideline 301 D and GLP principles. Test concentrations of 1 and 3 mg/L were tested in duplicate during 28 days. The relative biodegradation values calculated from the O2 measurements performed during the test period of 28 days revealed no significant biodegradation of the substance. Since biodegradation of at least 60% was not reached within 14 days of biodegradation exceeding 10%, the substance was concluded to be not readily biodegradable. In the toxicity control, the substance was found not to inhibit microbial activity. The criteria for acceptability of the test were met and the study is considered to be reliable without restriction. In conclusion, the substance was determined to be not readily biodegradable under the conditions of the closed bottle test.