RM23

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

March 12, 1997 - April, 10 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Version / remarks:

17 July 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Version / remarks:

December 1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Waterschap de Maaskant', 's-Hertogenbosch, the Netherl ands
- Storage conditions: The sludge was kept under continuous aeration until further treatment.
- Preparation of inoculum for exposure: 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: 3.5 g/L (concentrated sludge)

Duration of test (contact time):

28 d

Initial conc.:

12 mg/L

Based on:

TOC

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium:
Stock solutions
A) 8.50 g KH2PO4
21.75 g K2HPO4
67.20 g Na2HPO4 x 12H2O
0.50 g NH4Cl
dissolved in 1 1 Milli-Q water, pH 7.4 ± 0.2

B) 22.50 g MgSO4 x 7H2O dissolved in 1 L Milli-Q water.

C) 36.40 g CaCl2 x 2H2O dissolved in 1 L Milli-Q water.

D) 0.25 g FeCl3 x 6H2O dissolved in 1 L Milli-Q water.

1 L mineral medium contains: 10 mL of solution (a), 1 mL of solutions (b) to (d) and Milli-Q water.

- Test temperature: 20 - 21°C
- pH: 7.5 - 7.8
- pH adjusted: no
- Aeration of dilution water: A mixture of mineral components, Milli-Q water (ca. 80% total volume) and inoculum (1% final volume) was aerated with CO2-free air overnight.

TEST SYSTEM
- Culturing apparatus: 2 litre all-glass brown coloured bottles
- Number of culture flasks/concentration: 2 bottles (test suspension, inoculum blank); 1 bottle (positive control, toxicity control)
- Method used to create aerobic conditions: Aeration was done with CO2-free air. To contain CO2-free air, a mixture of oxygen (21%) and nnitrogen (79%) was led through a bottle, containing 0.5 - 1 L 0.0125 M 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) were connected in series to the exit air line of each test bottle. The C02 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.


SAMPLING
- 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.
- Sampling method: 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.


CONTROL AND BLANK SYSTEM
- Inoculum blank: containing only inoculum
- Toxicity control:containing test substance, reference substance and inoculum

Reference substance:

acetic acid, sodium salt

Key result

Parameter:

% degradation (CO2 evolution)

Value:

< 20

Sampling time:

28 d

Details on results:

The relative degradation values calculated from the measurements performed during the test period revealed 11% degradation of the test item in test bottle B and no significant degradation, i.e. 7.0% in test bottle A (significant: >10%). In the toxicity control more than 25% degradation occurred in 14 days (based an ThCO2)· Therefore, the test substance was assumed to be not inhibitory.
The positive control substance was degraded 72% in 14 days. The total CO2 release in the blank reached a total value of 30 mg CO2 per 2 litres of medium. The difference of duplicate values for %-degradation of the test item was always less than 20. Thus, the acceptability criteria of the test were fulfilled.

Results with reference substance:

The positive control substance was degraded 72% in 14 days.

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

The test item was not readily biodegrdable under the conditions of the modified Sturm test according to OECD Guideline 301B.

Executive summary:

The test item was tested for its ready biodegradability in the carbon dioxide (CO2) evolution test (modified sturm test) at ca. 34 mg per 2 litres, corresponding to 12 mg TOC/L.

The study procedure was based a EEC directive 92/69, C.4-C, December 1992, and OECD guideline No. 301 B July 17, 1992.

The Theoretical CO2 production (ThCO2) of the test item was calculated to be 2.572 mg CO2/mg.

The relative degradation values calculated from the measurements performed during the test period revealed 11% degradation of the test item in test bottle B and no significant degradation in test bottle A (significant: >10%). 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 presently performed.

Description of key information

The test item was not readily biodegrdable under the conditions of the modified Sturm test according to OECD Guideline 301B.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Type of water:

freshwater