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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:
21 February, 2018 - 21 March, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 835.3110 (Ready Biodegradability)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: DSP-001
- Expiration date of the lot/batch: 15th August 2018.
- Purity test date: 97 %w/w.
Oxygen conditions:
aerobic
Inoculum or test system:
sewage, predominantly domestic, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: Obtained on 21 February 2018 from the final effluent stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK.
The sample of effluent was filtered through coarse filter paper (first approximate 200 mL discarded) and the filtrate maintained on continuous aeration in a temperature controlled room at approximately 21 °C prior to use.
Duration of test (contact time):
28 d
Initial conc.:
4 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Composition of medium:
Solution a:
KH2PO4 - 8.50 g/L
K2HPO4 - 21.75 g/L
Na2HPO4.2H2O - 33.40 g/L
NH4Cl - 0.50 g/L
pH = 7.4
Solution b: CaCl 2 - 27.50 g/L
Solution c: MgSO4.7H2O - 22.50 g/L
Solution d: FeCl3.6H2O - 0.25 g/L

- Test temperature: 20 - 21 °C
- pH: 7.3 - 7.5
- pH adjusted: No
- Aeration of dilution water: The ‘dilution water’ was aerated for at least 20 minutes and allowed to stand for approximately 20 hours at the test temperature to give a dissolved oxygen concentration of approximately 9 mg O2/L.

TEST SYSTEM
The following test preparations were prepared and inoculated in 250 mL BOD bottles (darkened glass) with ground glass stoppers:
- An inoculum control, 28 bottles, consisting of inoculated mineral medium.
- The procedure control containing the reference item (sodium benzoate), 28 bottles, in inoculated mineral medium to give a concentration of 3.0 mg/L.
- The test item, 28 bottles, in inoculated mineral medium to give a concentration of 4.0 mg/L.
- The test item (4.0 mg/L) plus the reference item (1.5 mg/L), 10 bottles, in inoculated mineral medium to act as a toxicity control.
Test media were inoculated with sewage treatment micro-organisms at a rate of 1 drop of inoculum per liter.
The BOD bottles were incubated in darkness in a temperature controlled water bath at temperatures of between 20 and 21 °C.

SAMPLING
- Sampling frequency: Dissolved oxygen concentrations in duplicate samples from the control, reference and test item series were determined, on days 0, 2, 5, 7, 9, 14, 18, 21 and 28.
- Sampling method: YSI 54A dissolved oxygen meter and BOD Probe

TOXICITY CONTROL
An aliquot (12 mL) of the 1000 mg/L test item stock solution plus an aliquot (4.5 mL) of the sodium benzoate stock solution were dispersed in a final volume of 3 liters of mineral medium to give a final concentration of 4.0 mg test item/L plus 1.5 mg sodium benzoate/L.
Reference substance:
benzoic acid, sodium salt
Remarks:
An initial stock solution of 1000 mg/L was prepared by dissolving the reference item directly in mineral medium, and an aliquot (24 mL) dispersed in a final volume of 8 liters of inoculated mineral medium to give a test concentration of 3.0 mg/L.
Test performance:
The oxygen depletion of the inoculated control did not exceed 1.5 mg O2/L after 28 days, the residual oxygen concentration in the test bottles remained at 3.3 mg O2/L or greater in all test vessels and the difference between the extremes of replicate oxygen depletion values at the end of the test was less than 20% in all vessels thereby satisfying the validation criteria.
The test item attained 65% biodegradation after 28 days and, therefore, can be considered as readily biodegradable under the strict terms and conditions of OECD Test Guideline 301D.
Variation in biodegradation rates on different sampling days was considered to be due to normal biological variation in respiration rates between control and test vessels.
The toxicity control attained 61% biodegradation after 14 days and 71% biodegradation after 28 days therefore confirming that the test item was not toxic to the sewage treatment micro-organisms used in the test.
The reference item, sodium benzoate, attained 63% biodegradation after 14 days with greater than 60% degradation being attained in a 10-Day window. After 28 days 69% biodegradation was attained. These results confirmed the suitability of the inoculum and test
conditions and satisfied the validation criterion given in the OECD Test Guidelines.
Key result
Parameter:
% degradation (O2 consumption)
Value:
65
Sampling time:
28 d
Parameter:
% degradation (O2 consumption)
Value:
65
Sampling time:
21 d
Parameter:
% degradation (O2 consumption)
Value:
62
Sampling time:
18 d
Parameter:
% degradation (O2 consumption)
Value:
62
Sampling time:
14 d
Parameter:
% degradation (O2 consumption)
Value:
47
Sampling time:
9 d
Parameter:
% degradation (O2 consumption)
Value:
54
Sampling time:
7 d
Parameter:
% degradation (O2 consumption)
Value:
37
Sampling time:
5 d
Parameter:
% degradation (O2 consumption)
Value:
20
Sampling time:
2 d
Details on results:
Mean biodegradation for the test item after 28 days was 65%. O2 depletion was 2.25 and 2.3 mg O2/L in the replicate 1 and 2, respectively.
Key result
Parameter:
COD
Value:
>= 0.87 - <= 0.89 other: mg O2/mg
Remarks on result:
other: Stock solutions of 50 and 100 mg/L.
Parameter:
ThOD
Value:
5.01 other: mg O2/L
Remarks on result:
other: Estimated oxygen depletion for 3.0 mg/L.
Results with reference substance:
Mean COD of the reference item was 1.67 mg O2/mg (Percentage ThOD (1.67 mg O 2 /mg) = 100%).
Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test item attained 65% biodegradation after 28 days and is therefore considered to be readily biodegradable.
Executive summary:

A GLP compliant ready biodegradability test was performed on the substance in accordance with OECD Test Guideline 301D (Closed Bottle Test).

The test item, at a concentration of 4.0 mg/L, was exposed to sewage treatment micro-organisms in sealed vessels in the dark, between 20 - 21 °C for 28 days.

The degradation was assessed by the % oxygen consumption and observed to achieve 65% biodegradation after 28 days.

The toxicity control attained 61% biodegradation after 14 days and 71% biodegradation after 28 days, confirming that the test item was not toxic to the sewage treatment micro-organisms used in the test.

The performance of the inoculum and reference item controls also satisifed the validation criteria.

Chemical Oxygen Demand (COD) was calculated to be 0.88 mg O2/L (mean of six values) and Theoretical Oxygen Demand (ThOD) was calculated to be 5.01 mg O2/L.

Under the conditions of the test and considering the substance is a UVCB, 1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-coco acyl derivs., hydroxides, inner salts can be considered as readily biodegradable.

Description of key information

A GLP compliant ready biodegradability test was performed on the UVCB substance in accordance with OECD Test Guideline 301D (Closed Bottle Test).

The test item, at a concentration of 4.0 mg/L, was exposed to sewage treatment micro-organisms in sealed vessels in the dark, between 20 - 21°C for 28 days.

The degradation was assessed by the % O2 consumption and observed to achieve 65% biodegradation after 28 days.

The toxicity control attained 61% biodegradation after 14 days and 71% biodegradation after 28 days, confirming that the test item was not toxic to the sewage treatment micro-organisms used in the test.

The performance of the inoculum and reference item controls also satisifed the validation criteria.

Chemical Oxygen Demand (COD) was calculated to be 0.88 mg O2/L (mean of six values) and Theoretical Oxygen Demand (ThOD) was calculated to be 5.01 mg O2/L.

Under the conditions of the test and considering the substance is a UVCB, 1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-coco acyl derivs.,hydroxides, inner salts can be considered as readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

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