Registration Dossier

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
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study conducted under GLP

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Details on test material:
- Name of test material (as cited in study report): Methyl propyl ketone (MPK)
- Analytical purity: 99.7%
- Lot/batch No.: 7054784
- Storage condition of test material: room temperature in a dark ventilated cabinet

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
Approximately 4 L of activated sludge was collected from the Wareham, Massachusetts Wastewater Treatment Plant which treats primary domestic sewage. Upon arrival at the lab, the untreated sludge was centrifuged at 1000 rpm for 10 minutes. The supernatant was kept and the solids discarded. The resultant supernatant was filtered through coarse filter paper. Following filtration, the first 200 mL of supernatant was discarded. The next 500 mL of supernatant was retained and aerated. The inoculum was used the same day as it was prepared.
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
2.5 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
The test solutions were prepared by first adding approximately 4 L of aerated water by siphon to each of three 9-L bottles, one for the test substance, one for the sodium benzoate and one for the inoculum control blank. Each 9-L bottle then received 6 ml of each of the four nutrient solutions. The test substance bottle next received 960 uL of the inoculum and 18.75 uL of the 2.5 mg/L test substance solution, the sodium benzoate bottle received 960 uL of the inoculum and 3.0 ml of the 10 mg/ml sodium benzoate stock solution and the inoculum blank bottle received 960 uL of the inoculum. Each of the three bottles was then brought to a final volume of 6 L with the aerated water by siphon.

Immediately after preparation, each solution was siphoned (bubble-free) from the 9-L bottles into the respective series of 18 BOD bottles (for a total of 54 bottles for the three solution types). The bottles were slightly overfilled to eliminate all air bubbles. Three bottles from each set were labeled for each interval (i.e., days 7, 14, 21 and 28) and two bottles from each set were labeled for day 0. Once filled, the bottles were closed, wrapped in foil and labeled. With the exception of the BOD bottles to be used for day 0 analysis, all bottles were then placed in the environmental chamber. Four additional BOD bottles were prepared from each set of solutions to provide additional samples for analysis if necessary. Oxygen depletion in the test substance and sodium benzoate solutions was calculated at each sampling interval.
Reference substance
Reference substance:
benzoic acid, sodium salt

Results and discussion

% Degradation
% degradation (O2 consumption)
Sampling time:
28 d

BOD5 / COD results

Results with reference substance:
Degradation in the sodium benzoate test solutions was 72% on day 28 which indicated that the inoculum contained a viable microbial population
and that the test procedures used during the study were adequate.

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
readily biodegradable
Executive summary:

Ready biodegradability of methyl propyl ketone (MPK) was evaulated following the OECD Guideline 301 D (Closed Bottle Test). MPK achieved 70% biodegradation within the 28 days and can be classified as readily biodegradable under the conditions of this test.