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Environmental fate & pathways

Biodegradation in water: screening tests

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biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Analysis certificate was provided allong with GLP accreditation this substance can therefore be considered reliable without restrictions.
according to guideline
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Ammonium chloride was omitted from the medium to prevent nitrification.
Principles of method if other than guideline:
This test has been performed according to slightly modified EEC/OECD test guidelines and in conformity with the OECD Principles of Good Laboratory Practice (GLP). Ammonium chloride was omitted from the medium to prevent nitrification. The closed bottle test was prolonged by measuring the course of the oxygen decrease in the bottles after day 28 using a special funnel.The sludge was diluted to a concentration in the BOD bottles of 2 mg DWl/itre.- The test compound may be toxic to micro-organism and for that reason silica gel was used to decrease the concentration of Aromox T/112 in the water phase.
GLP compliance:
Oxygen conditions:
Inoculum or test system:
other: Secondary Activated Sludge.
Details on inoculum:
Instead of an effluent / extract / mixture, activated sludge was used as an inoculum.The inoculum was taken from an activated sludge plant. The sludge was preconditioned to reduce the endogenous respiration rates.Secondary activated sludge was obtained from the RWZI Nieuwgraaf in Duiven. The RWZI Nieuwgraaf is an activated sludge plant treating predominantly domestic wastewater.
Duration of test (contact time):
28 d
Initial conc.:
2 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
The test was performed in 280 mL BOD (Biological Oxygen Demand) bottles. (Data minimal)
Reference substance:
other: Sodium Acetate
Preliminary study:
No Data
Test performance:
The validity of the test is shown by the oxygen consumption in the control bottle with sodium acetate and anendogenous respiration of 0.6 mg/L. The pH of the medium at the end of the test period is 7.4.
Key result
% degradation (O2 consumption)
Sampling time:
28 d
Details on results:
% ready biodegradable reached within 28 days.
Results with reference substance:
Satisfactory See table Below.

Calculation of Biodegredation

The biodegradation was calculated as the ratio of the biochemical oxygen demand (BOD) to the theoretical oxygen demand (ThOD). The ThOD of the pure active ingredient is 1.9O2/g test substance. The BOD of the test substance and sodium acetate were calculated from the oxygen concentrations in the bottles with and without test substance and the bottles with and without sodium acetate respectively.

Validity criteria fulfilled:
Interpretation of results:
readily biodegradable
Conducted to critical guidelines GLP no analysis certifiate. Reliable with restrictions.
Executive summary:

The test item is biodegraded in the closed bottle test (OECD Guideline 301 D) performed in conformity with the OECD principles of Good Laboratory Practices (GLP). The biodegradation percentage at day 28 is 79 and therefore the test item should be classified as readily biodegradable.

Description of key information

% Degradation of test substance: 79 after 28 d (Biochemical oxygen demand). Substance is considered to be readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

A study was conducted to determine ready biodegradability of the test substance in a closed bottle test according to OECD Guideline 301 D, in compliance with GLP. The test substance at 2 mg/L was incubated with non-adapted activated sludge and placed in closed bottles in the dark for 28 d. Degradation was assessed by determination of the dissolved oxygen concentration (DOC) on Days 5, 15 and 28. Control solutions containing the reference substance, sodium acetate (6.7 mg/L), together with inoculum was used for validation purposes. The test substance biodegraded 8, 68 and 79% on Day 5, 15 and 28, respectively, and fulfilled the 10 d window criteria. The theoretical oxygen demand (ThOD) of the test substance is 1.9 mg/mg. The validity of the test was demonstrated by an endogenous respiration of 0.6 mg/L on Day 28. Furthermore, the differences of the replicate values on Day 28 were lower than 20%. The biodegradation percentage of the reference compound, sodium acetate, on Day 15 was 85%. Finally, the validity of the study was shown by oxygen concentrations >0.5 mg/L in all bottles during the test period. Under the study conditions, the test substance was considered to be readily biodegradable (Ginkel, 1990).