Registration Dossier

Environmental fate & pathways

Biodegradation in water: screening tests

Currently viewing:

Administrative data

Link to relevant study record(s)

Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10-DEC-2008 to 28-JAN-2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
This study was performed according to OECD Guideline 301F with GLP certificate. All validity criteria were fulfilled and no deviation was observed. This report is clear and the study clearly explained.
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. QA statement)
Remarks:
Inspected on: 23 bis 25 August 2006 / Signed on 21 March 2007
Specific details on test material used for the study:
- Water solubility (20°C, pH 7): 100 µg/L
- Partition coefficient (log Pow): 6.57 (measured)
- Density (20°C): ca. 0.93 g/cm3
- Storage condition of test material: Room temperature
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): sewage treatment plant Ruhrverband Kläranlage, Sunthelle 6, 57392 Schmallenberg, Germany, which is mainly fed with municipal wastewater
- Laboratory culture: NA
- Method of cultivation: NA
- Storage conditions: The samples were washed once with mineral medium after the arrival at the laboratory and kept aerobic until use.
- Storage length: no data
- Preparation of inoculum for exposure: no data
- Pretreatment: no data
- Concentration of sludge: 30.0 mg dry mass/litre (7.50 mg dry mass/250 mL)
- Initial cell/biomass concentration: no data
- Water filtered: no data
- Type and size of filter used, if any: no data
Duration of test (contact time):
>= 28 d
Initial conc.:
>= 20 mg/L
Based on:
test mat.
Initial conc.:
>= 100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Composition of medium: ( a ) KH2PO4 8.50 g/L, K2HPO4 21.75 g/L, Na2HPO4 x 2 H2O 33.40 g/L, NH4Cl 0.50 g/L ( b ) CaCl2 x 2 H2O 36.40 g/L ( c ) MgSO4 x 7 H2O 22.50 g/L ( d ) FeCl3 x 6 H2O 0.25 g/L. The mineral medium applied in the test contained 10 mL/L and 1 mL/L of the mineral stock solution a and b–d, respectively.
- Additional substrate: NA
- Solubilising agent (type and concentration if used): NA
- Test temperature: 22°C
- pH: 7.5-7.6 at test initiation
- pH adjusted: no
- Aeration of dilution water: The suspension was stirred during the whole test
- Suspended solids concentration: 30.0 mg dry mass inoculum per litre mineral medium
- Continuous darkness: yes

TEST SYSTEM
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: stirring
- Method used to create anaerobic conditions: NA
- Measuring equipment: The measurement and recording of the oxygen demand was carried out continuously using a SAPROMAT respirometer (VOITH Inc.)
- Test performed in closed vessels due to significant volatility of test substance: no
- Test performed in open system: no
- Details of trap for CO2 and volatile organics if used: Evolved carbon dioxide is absorbed in a suitable absorbent (not used for measurement

SAMPLING
- Sampling frequency: continuously
- Sampling method: NA
- Sterility check if applicable: No data
- Sample storage before analysis: No data

CONTROL AND BLANK SYSTEM
- Inoculum blank: duplicate
- Abiotic sterile control: none
- Toxicity control: duplicate

STATISTICAL METHODS: none
Reference substance:
benzoic acid, sodium salt
Preliminary study:
NA
Test performance:
The degradation of the reference substance sodium benzoate had reached 81 % within the first 14 days. The difference of extremes of replicate values of the removal of the test item at the end of the 10-day-window is less than 20%. Therefore, the test can be considered as valid.
Parameter:
% degradation (O2 consumption)
Value:
63.2
St. dev.:
3.5
Sampling time:
14 d
Remarks on result:
other: mean value at 100 mg/L of test item
Key result
Parameter:
% degradation (O2 consumption)
Value:
79.9
St. dev.:
2.8
Sampling time:
28 d
Remarks on result:
other: mean value at 100 mg/L of test item
Parameter:
% degradation (O2 consumption)
Value:
71.4
St. dev.:
5.8
Sampling time:
14 d
Remarks on result:
other: mean value at 20 mg/L of test item
Key result
Parameter:
% degradation (O2 consumption)
Value:
81.3
St. dev.:
7
Sampling time:
28 d
Remarks on result:
other: mean at 20 mg/L of test item
Details on results:
The biodegradation of the test substance in the static test was found to be 81 % for a concentration of 20 mg test item/L and 80 % for a concentration of 100 mg test item/L after 28 days. The biodegradation within the 10-day-window was 71 % and 63 %, respectively. The 10-day window started at day three (20 mg/L) and four (100 mg/L), respectively. No clear lag phase / adaptation phase was noticeable since degradation in the procedural control also started on day four. The biodegradation of the item mixture in the toxicity control was found to be 76 % for a concentration of 20 mg test item/L and 64 % for a concentration of 100 mg test item/L after 14 days of incubation. Thus, the demanded threshold value of 25 % is exceeded and the test item can be identified as non-toxic in a ready biodegradability test.
Results with reference substance:
The degradation of the reference substance sodium benzoate had reached 81 % within the first 14 days.

- Test suspension A: 2 vessels containing test item (100 mg/L) and inoculum

- Test suspension B: 2 vessels containing test item (20 mg/L) and inoculum

- Inoculum blank: 2 vessels containing only inoculum

- Procedural control: 2 vessels containing reference item (100 mg/L) and inoculum

- Toxicity control A: 2 vessels containing test item (100 mg/L), reference item (100 mg/L) and inoculum

- Toxicity control B: 2 vessels containing test item (20 mg/L), reference item (100 mg/L) and inoculum

Table 5.2.1/1: Oxygen consumption. Cumulated consumption (mg O2/L) after 14 days of incubation. Single and mean values of the parallel test vessels and standard deviation.

vessel

Test suspension A

Test suspension B

Inoculum blank

Toxicity control A

Toxicity control B

Procedural control

1

203

59

18

318

191

153

2

218

64

18

/

/

153

Mean

211

62

18

318

191

153

SD

11

4

0

/

/

0

Table 5.2.1/2: Percent degradation. Degradation (%) after 14 days of incubation. Single and mean values of the parallel test vessels and standard deviation.

vessel

Test suspension A

Test suspension B

Inoculum blank

Toxicity control A

Toxicity control B

Procedural control

1

60.7

67.3

/

63.7

76.1

81.1

2

65.7

75.5

/

/

/

81.1

Mean

63.2

71.4

/

63.7

76.1

81.1

SD

3.5

5.8

/

/

/

0

Table 5.2.1/3: Oxygen consumption. Cumulated consumption (mg O2/L) after 28 days of incubation. Single and mean values of the parallel test vessels and standard deviation.

vessel

Test suspension A

Test suspension B

Inoculum blank

Toxicity control A

Toxicity control B

Procedural control

1

260

69

22

400

211

169

2

272

75

23

/

/

170

Mean

266

72

23

400

211

170

SD

8

4

1

/

/

1

Table 5.2.1/4: Percent degradation. Degradation (%) after 28 days of incubation. Single and mean values of the parallel test vessels and standard deviation.

vessel

Test suspension A

Test suspension B

Inoculum blank

Toxicity control A

Toxicity control B

Procedural control

1

78.0

76.3

/

80.1

82.9

88.0

2

81.9

86.2

/

/

/

88.6

Mean

79.9

81.3

/

80.1

82.9

88.3

SD

2.8

7.0

/

/

/

0.4

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
In an OECD 301F test, the test substance showed readily biodegradation meeting the 10-day window requirements and is considered non-toxic for microorganisms up to 100 mg/L.
Executive summary:

The ready biodegradation of the test substance has been assessed in a manometric respirometry test OECD 301F. It is compliant with Good Laboratory Practices. Two test groups of 20 mg/L and 100 mg/L have been inoculated with activated sludge from a primary domestic sewage treatment plant. Sodium benzoate was used as reference substance at 100 mg/L and a toxicity control was set up as an inoculum blank group. O2 consumption by bacteria was measured continuously while evolving CO2 was adsorbed to maintain a constant inner pressure in the vessels.

Biodegradation started on day 3 or 4 with a very short lag phase. Test group 100 mg/L and 20 mg/L showed 63.2 ± 3.5% and 71.4 ± 5.8% of biodegradation respectively at the end of the 10-day window (on day 14). After 28 days, test group 100 mg/L and 20 mg/L showed respectively 79.9 ± 2.8% and 81.3 ± 7.0% of biodegradation. The standard deviation in replicates in each test group was lower than 20% and pH at the end of the test remained in the 6-8.5 range. The reference substance showed ready biodegradation after 14 days and no toxicity to bacteria occurred in the toxicity control group. The test is valid and the results are considered as reliable.

Finally in an OECD 301F test, the test substance showed readily biodegradation meeting the 10-day window requirements.

In addition, the biodegradation of the item mixture in the toxicity control was found to be 76 % for a concentration of 20 mg test item/L and 64 % for a concentration of 100 mg test item/L after 14 days of incubation. Thus, the demanded threshold value of 25 % is exceeded and the test item can be identified as non-toxic in a ready biodegradability test.

Description of key information

OECD Guideline 301F, GLP, key study, validity 1:

79.9 ± 2.8% and 81.3 ± 7.0% biodegradation after 28 days, for 100 and 20 mg/L of test substance, respectively, within the 10-day window.

Readily biodegradable.

Non-toxic to microorganisms up to 100 mg/L.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable
Type of water:
freshwater

Additional information

One valid key study is available (Fraunhofer, 2009), to assess the readily biodegradation of the registered substance.

This study was performed on the registered substance according to OECD Guideline 301F with GLP certificate. Briefly, two test groups at 20 mg/L and 100 mg/L was inoculated with activated sludge from a primary domestic sewage treatment plant. Sodium benzoate was used as reference substance. The test was valid. Biodegradation started on days 3 and 4 with a very short lag phase. The 10-day window ended on day 14 showing more than 60% in each test group. After 28 days test groups showed 79.9 ± 2.8% and 81.3 ± 7.0% of biodegradation, for 100 and 20 mg/L of test substance respectively. The substance is considered readily biodegradable, within the 10-day window. In addition, the biodegradation of the item mixture in the toxicity control was found to be 64 % for a concentration of 100 mg/L and 76 % for a concentration of 20 mg/L after 14 days of incubation. Thus, the demanded threshold value of 25 % is exceeded and the test substance can be identified as non-toxic for microorganisms up to 100 mg/L.