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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
Study period:
1999-02-10 to 1999-03-11
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guidelineopen allclose all
according to guideline
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Version / remarks:
July, 1992
according to guideline
EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)
Version / remarks:
December 1992
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
1-(3,5-Dihydroxyphenyl)-1-oxo-2-(N-1-(4-methoxyphenyl)isopropylamino)ethane Hydrobromide
EC Number:
Molecular formula:
C18-H21-N-O4 x HBr
1-(3,5-Dihydroxyphenyl)-1-oxo-2-(N-1-(4-methoxyphenyl)isopropylamino)ethane Hydrobromide
Test material form:
solid: particulate/powder

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, domestic (adaptation not specified)
Details on inoculum:
- Source of inoculum: The source of test organisms was activated sludge freshly obtained from a municipal sewage treatment plant: 'Waterschap de Maaskant', 's-Hertogenbosch, the Netherlands
- Storage conditions: The sludge was kept under continuous aeration until further treatment. The concentration of suspended solids was 3.0 g/L in the concentrated sludge (information obtained from the municipal sewage treatment plant). Before use, the sludge was allowed to settle for at least 30 minutes and the liquid decanted for use as inoculum at the amount of 10 ml/L of mineral medium.
- Concentration of sludge: 1% of final solution
- Water filtered: yes
- Type and size of filter used, if any: reverse osmosis, activated carbon and ion-exchange cartridges
Duration of test (contact time):
28 d
Initial test substance concentrationopen allclose all
Initial conc.:
12 mg/L
Based on:
Initial conc.:
1 g/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
- Composition of medium: grade salts were added to deionised water to give stock solutions. Stock solutions were diluted in deionised water.
- Test temperature: between 20.5 and 22°C
- pH: between 7.5 and 7.9
- pH adjusted: no
- Aeration of dilution water: Mineral components, deionised water and inoculum were added to each bottle. This mixture was aerated with CO2-free air overnight to purge the system of CO2.

- Culturing apparatus: 2 litre all-glass brown coloured bottles
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: A mixture of oxygen (21%) and nitrogen (79%) was led through a bottle, containing 0.5 -1 litre 0.0125 Ba(OH)2 solution to trap CO2 which might be present in small amounts. The CO2 free air was sparged through the scrubbing solutions at a constant rate.
- Measuring equipment: Three CO2-absorbers (bottles filled with 100 ml 0.0125 M Ba(OH)2), Titration

- Sampling frequency: Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until the 28th day. Each time the CO2-absorber nearest to the test bottle was removed for titration; each of the remaining two absorbers was moved one position in the direction of the test bottle. A new CO2-absorber was placed at the far end of the series. Phenolphthalein was used as pH indicator. On the 28th day, the pH of the test suspensions was measured and 1 ml of concentrated HCl was added to each bottle. The bottles were aerated overnight to drive off CO2 present in the test suspension. The final titration was made on day 29.
- Sampling method: The CO2 produced in each test bottle reacted with the barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl.

- Inoculum blank: containing only inoculum (2 bottles)
- Positive control: containing reference substance (ca. 40 mg/L sodium acetate), (1 bottle)
- Toxicity control: containing test substance, reference substance and inoculum (1 bottle)
Reference substance
Reference substance:
other: sodium acetate

Results and discussion

% Degradation
% degradation (CO2 evolution)
>= 0.2 - <= 1.6
Sampling time:
28 h
Details on results:
The difference of duplicate values for %-degradation of the test item was always less than 20%.

BOD5 / COD results

Results with reference substance:
The positive control substance was degraded at least 60% within 14 days.

Any other information on results incl. tables

In the modified Sturm test 42 ml of the 1 g test item/Lsolution was tested in 2 litres test medium, corresponding to 12 mg TOC.

The relative degradation values calculated from the measurements performed during the test period revealed no significant degradation of the test item.

In the toxicity control the test item was found to be not inhibitory.

Since all acceptability criteria prescribed by the protocol were met, this study was considered to be valid.

In conclusion, the test item was not readily biodegradable under the conditions of the modified Sturm test performed.

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
not readily biodegradable
The test item was not readily biodegradable under the conditions of the modified Sturm test performed.

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