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Biodegradation in water: screening tests

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biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From April 12th 2018 to June 20th 2018.
1 (reliable without restriction)
according to guideline
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Version / remarks:
according to EU method C.4-C
GLP compliance:
yes (incl. QA statement)
The certificate is included in the Study Report attached.
Specific details on test material used for the study:
Batch number: N6A3I
Expiry date: 04 August 2022
Oxygen conditions:
Inoculum or test system:
sewage, domestic (adaptation not specified)
Activated sludge from a biologic sewage treatment plant was used as inoculum. the chosen plant treats mostly domestic sewage.
Details on inoculum:
- Source of inoculum/activated sludge: The sludge was taken from the activation basin of the sewage treatment plant, In den Seewiesen, 67480 Edenkoben, Germany. Date of collection: 18 May 2018, batch n°: 20180518.

- Pretreatment: The sludge was filtrated, washed with test medium (2X) and re-suspended in test medium. It was then aerated until use. The dry matter was determined to contain 4520 mg of suspended solids.
- Concentration of sludge: 25.0 mg/L
Duration of test (contact time):
28 d
Initial conc.:
ca. 30 mg/L
Based on:
test mat.
Initial conc.:
ca. 20 mg/L
Based on:
Parameter followed for biodegradation estimation:
CO2 evolution
Analyses of the emitted CO2 were made by IC measurement using the carbon analyser TOC multi N/C 2100S, Analytik Jena. Each sample was measured in duplicate or triplicate, respectively (depending of the variation between the measured values).
Details on study design:

- Composition of medium:
10 mL of Solution A (8.5 g of Potassium dihydrogen phosphate + 21.75 g of Dipotassium hydrogen phosphate + 33.4 g of Disodiumhydrogen phosphate dihydrate + 0.5 g of Ammonium chloride in 1000 mL H2O demin.)
1 mL of Solution B (27.5 g of Calcium chloride in 1000mL H2O demin.)
1 mL of Solution C (22.5 g of Magnesium sulphate heptahydrate in 1000mL H2O demin.)
1 mL of Solution D (0.25 g of Iron (III) chloride hexahydrate + 0.4 g of Di-sodium-ethylene diaminetetraacetate dihydrate in 1000mL H2O demin.)
in 1000mL H2O demin.

- Test temperature: 20.4 - 23.0 °C
- pH: 7.6-7.7
- pH adjusted: no

- Culturing apparatus: The test vessels were aerated with purified (by activated charcoal), CO2-scrubbed, moistened air. The scrubbing of carbon dioxide was achieved by bubbling the purified air through a flask containing 1.5 M NaOH. To control the absence of CO2, the air was then led througha flask containing a solution of Ba(OH)2 before reaching the test vessels.

- Number of culture flasks/concentration: 2 flasks of 1500 mL (30 mg/L of the test substance)

- Measuring equipment: IC measurement using the carbon analyser TOC multi N/C 2100S, Analytik Jena.

- Details of trap for CO2 and volatile organics if used: The emitted CO2 was trapped in 0.25 M NaOH. Two scrubbers containing 100 mL each were connected in series to the test vessels. The initial IC value of the 0.25M NaOH was separately determined in each flask.

- Sampling frequency: 9 samples were taken on day 0, 2, 6, 8, 10, 14, 17, 23 and 29.
- Sampling method: The sample volume was 1 mL; The resulting chane in the volume of the front flask was considered in the calculation of emitted CO2.. On day 28, 5 mL HCl 2M was added to each test flask in order to drive off dissolved CO2. On day 29, samples were from both scrubber flasks were taken.

- Blank controls: 2 flasks of 1500 mL containing mineral medium and inoculum
- Positive control flasks: 2 flasks of 1500 mL containing Aniline, mineral medium and inoculum
- Abiotic control: 1 flask of 1500 mL containing test item, mineral medium and HgCl2
- Toxicity control: 1 flask of 1500 mL containing test item, Aniline, mineral medium and inoculum.

Reference substance:
Key result
% degradation (inorg. C analysis)
>= 86.9 - <= 88.8
Sampling time:
29 d
Validity criteria fulfilled:
Interpretation of results:
readily biodegradable
The test item 5-methyl-1H-benzotriazole is considered as "readily biodegradable". The degree of biodegradation reached 87% after 28 days. The 10-day-window began on day 9 at its end, 82% degradation were reached surpassing the pass level of 60% given in the OECD guideline. Abiotic degradation was not observed.

Description of key information

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information