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Diss Factsheets

Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 2011-07-01 to 2011-08-11
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline-conform study performed under GLP without deviations

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 301 C (Ready Biodegradability: Modified MITI Test (I))
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: "Method for Testing the Biodegradability of Chemical Substances by Microorganisms" (March 31, 2011, N0.0331-7, Pharmaceutical and Food Safety Bureau, Ministry of Health, Labour and Welfare)
Deviations:
no
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Details on test material:
Identifier: C-C10
Description: Blackish blue powder
Lot/batch No.: MB-1
Stability under test conditions: The stability of the test item was confirmed by comparing the IR spectrum of the test item after the completion of the experiment with that before the start of the experiment
Storage condition of test material: The test sample was stored in a dark storage place at room temperature.

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge: mixture of samples from at least 10 different locations in Japan (surface water, surface soil from rivers, lakes and inland sea, sludge from sewage treatment plants)
- Laboratory culture: as described in guideline
- Method of cultivation: as described in guideline
- Storage conditions: as described in guideline
- Storage length: not reported
- Preparation of inoculum for exposure: synthetic sewage was produced from glucose, peptone and potassium dihydrogenphosphate dissolved in purified water and the pH of the solution was adjusted to 7.0 ± 1; activated sludge was fed with the synthetic sewage and sludge was used 19.5 hours after being fed
- Pretreatment: no pretreatment
- Concentration of sludge: 2820 mg/L suspended solids in final sludge

- Water filtered: no
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimationopen allclose all
Parameter followed for biodegradation estimation:
O2 consumption
Remarks:
: During the cultivation period, the change in BOD of the test solutions was measured by a closed system oxygen consumption measuring apparatus.
Parameter followed for biodegradation estimation:
DOC removal
Remarks:
: After the end of the cultivation, dissolved organic carbon (DOC) was determined.
Parameter followed for biodegradation estimation:
test mat. analysis
Remarks:
: After the end of the cultivation, the test item in the test solutions were determined.
Details on study design:
TEST CONDITIONS
- Composition of medium: 5 L of basal culture medium was prepared by adding purified water to each 3 mL aliquots of solutions a), b), c) and d) as described in the guideline; a total of 5L culture medium was prepared; test material (at 100 mg/L) or reference material (aniline at 100 mg/L) and sludge (at 30 mg suspended solids/L) was added
- Additional substrate: none
- Solubilising agent (type and concentration if used): not applicable
- Test temperature: 25 ± 1 °C
- pH: water + test item: 6.6 - 7.0, sludge + test item: 6.8 - 7.3
- pH adjusted: pH adjustment of culture medium and activated sludge
- Aeration of dilution water: no aeration during test
- Suspended solids concentration: 30 mg/L
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: glass vessels
- Number of culture flasks/concentration: 3
- Method used to create aerobic conditions: not reported
- Measuring equipment: closed system oxygen consumption measuring apparatus with data sampler OM7000A (Ohkura Electric Co. Ltd), temperature controlled bath
- Test performed in closed vessels: yes
- Test performed in open system: not applicable
- Details of trap for CO2: glass vessel containing soda lime

SAMPLING
- Sampling frequency: The oxygen consumption was measured continuously during the incubation period
- Sampling method: Continuous sampling of oxygen; continuous absorption of CO2 in trap
- Sterility check if applicable: one abiotic control was included
- Sample storage before analysis: not applicable

CONTROL AND BLANK SYSTEM
- Inoculum blank: 1 vessel
- Abiotic sterile control: 1 vessel
- Toxicity control: not performed
Reference substance
Reference substance:
aniline

Results and discussion

% Degradationopen allclose all
Parameter:
% degradation (O2 consumption)
Value:
4
Sampling time:
28 d
Parameter:
% degradation (DOC removal)
Value:
2
Sampling time:
28 d
Parameter:
% degradation (test mat. analysis)
Value:
2
Sampling time:
28 d
Details on results:
The average percentage biodegradation by BOD, that by DOC and that of the test item wae 4%, 2% and 2%, respectively. In addition, thae was no signiscant change of the test item in peak shape on the HPLC chromatograms. From the above-mentioned results, it was considered that the test item was not biodegraded under the test conditions of this study.

BOD5 / COD results

Results with reference substance:
The reference substance aniline showed 58 % degradation after 7 days, 78 % degradation after 14 days, 81% degradation after 21 days and 83% degradation after 28 days (based on BOD).

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
under test conditions no biodegradation observed
Conclusions:
The substance is not biodegradable under the conditions of a modified MITI test.
Executive summary:

The ready biodegradability of the test substance C-C10 was studied under GLP in a modified MITI test in accordance with OECD TG 301-C (dated from 1992). Activated sludge was obtained by mixing samples from 10 locations in Japan (samples from surface water, river sediment, lakes and inland sea plus sludge from sewage treatment plants). Inoculum was used in the test 19.5 hours after the unit had been fed.

Culture medium prepared in accordance with the guideline was used. The following test media (all at final volume of 300 mL) were tested:

1) water and test substance (100 mg/L) in one abiotic control;

2) activated sludge (30 mg/L suspended solids) and test substance (100 mg/L) in three testing vessels;

3) activated sludge (30 mg/L suspended solids) and reference substance aniline (100 mg/L) in one positive control;

4) activated sludge (30 mg/L suspended solids) as one inoculum control.

The pH value of the 3 test vessels was between 6.8 and 7.3, that of the abiotic control between 6.6 and 7.0. All test media were stirred and inoculated during a period of 28 days at 25 ± 1 °C in the dark in closed vessels connected to an oxygen consumption measuring apparatus. The biodegradation was monitored by continuously measuring the biological oxygen demand. In addition, the dissolved organic carbon was measured and the concentration of the test material was determined by HPLC at the end of the test.

The test material C-C10 was not biodegradable under the conditions of the test since the percentage of biodegradation after 28 days was only 4% based on BOD, 2% based on DOC and 2% based on HPLC analysis.

The reference substance aniline was degraded by 58% after 7 days and 78% after 14 days showing the suitability of the inoculum.

All validity criteria were fulfilled because the difference between extremes of replicates was less than 20%, the BOD of the control blank after 28 days was less than 60 mg/L and the biodegradation was above the required percentages after 7 and 14 days.

In conclusion, the substance C-C10 is not biodegradable under the conditions of a modified MITI test.