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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:
25 September 2007 (start) - 23 October 2007 (end)
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was performed according to OECD and EC guidelines and according to GLP.

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 301 C (Ready Biodegradability: Modified MITI Test (I))
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Reference substance name:
EC Number:
EC Name:
Molecular formula:
5,11,17,23,29,35,41,47-octa-tert-butyl-49,50,51,52,53,54,55,56-octahydroxy-2λ⁶,8λ⁶,14λ⁶,20λ⁶,26λ⁶,32λ⁶,38λ⁶,44λ⁶-octathianonacyclo[³,⁷.1⁹,¹³.1¹⁵,¹⁹.1²¹,²⁵.1²⁷,³¹.1³³,³⁷.1³⁹,⁴³]hexapentaconta-1(49),3,5,7(56),9,11,13(55),15,17,19(54),21(53),22,24,27,29,31(52),33,35,37(51),39(50),40,42,45,47-tetracosaene-2,2,8,8,14,14,20,20,26,26,32,32,38,38,44,44-hexadecone; 5,11,17,23-tetra-tert-butyl-25,26,27,28-tetrahydroxy-2λ⁶,8λ⁶,14λ⁶,20λ⁶-tetrathiapentacyclo[³,⁷.1⁹,¹³.1¹⁵,¹⁹]octacosa-1(25),3,5,7(28),9,11,13(27),15(26),16,18,21,23-dodecaen-2,2,8,8,14,14,20,20-octone
Constituent 2
Reference substance name:
MIxture of TC4A-SO2 and TC8A-SO2
MIxture of TC4A-SO2 and TC8A-SO2
Details on test material:
- Name of test material (as cited in study report): mixture of TC4A-SO2 and TC8A-SO2
- Substance type: multi-constituent
- Physical state: solid
- Stability under test conditions: Stable under storage conditions. There was no difference between the IR spectrum taken at the start of the study and after completion of the study.
- Storage condition of test material: in the archives of the room where the test substance is handled.
- Other: The IR spectrum of the test substance was shown to be similar to that supplied by the study sponsor, confirming the identity.

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, industrial (adaptation not specified)
Details on inoculum:
Standard activated sludge, purchased from Chemicals Evaluation and Research Institute Japan, Chemical Biotesting Center, Tokyo Japan.

Preparation inoculum:
After ceasing aeration of the sludge mixture for 30 minutes, container was left still for the gravity sedimentation of sludge particles. 300 ml supernatant was removed and 300 ml synthetic sewage was added to the mixture and the container was aerated again. The pH was kept at 7.0 +/- 1.0 .

Synthetic sewage: glucose, peptone and potassium phosphate monobasic were dissolved in dechlorinated water to obtain 0.1% (w/v) of each component and pH was adjusted with 0.1 M sodium hydroxide to 7.0 +/- 1.0.

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:
test mat. analysis
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:

Basal culture medium: In accordance with "Test methods of industrial liquid waste" (JIS K 0102-1998-21). In purified water adjusted to pH 7.2.

The concentration of suspended solid of the cultivated activated sludge was 3800 mg/L. The cultivated activated sludge was diluted with a standard medium so as to make the sludge concentration of 900 mg/L.

Vessel 1 (water degradation test group): 300 ml of water and 30 mg of test substance (100 mg/l)
Vessel 2 (inoculum blank): 290 ml of basal culture medium
Vessel 3 (reference group): 290 ml of basal culture medium and 30 mg of aniline
Vessels 4, 5 and 6 (biodegradation test group): 290 ml of basal culture medium and 30 mg of test substance

10 ml of the diluted cultivated activated sludge was inoculated to the vessels 2, 3 and 4, 5, 6 : volume total in test vessel: 300 mL. Suspended solid concentration: 30 mg/L.


Temperature: 25 +/- 1 deg C
Incubation time: 28 days
Light conditions: Light-shielded (except during the additions and the observations)


Closed system oxygen consumption measuring apparatus
Absorbent for carbon dioxide: Soda Lime
The test solutions were stirred by a magnetic stirer.
It was confirmed that there was no leakage in the closed system before the BOD measurement.


As the test substance is a mixture of 2 components, the amount of residual test substance was calculated not only as a sum of 2 components but also for each component (TC4A-SO2 and TC8A-SO2).

The percentage biodegradation by BOD as well as by HPLC were calculated.

Reference substance
Reference substance:

Results and discussion

Preliminary study:
Not performed.
Test performance:
Appearance test solution (sludge + test substance) after incubation: Not dissolved. No growth of sludge. No coloration.
The pH of the test solution after incubation was 7.93 (water+test substance) and 7.02, 7.08 and 7.10 in the solutions containing sludge + test substance.
% Degradationopen allclose all
% degradation (O2 consumption)
Sampling time:
28 d
Remarks on result:
other: BOD
% degradation (test mat. analysis)
Sampling time:
28 d
Details on results:
HPLC: -2% based on both components. -2% and 0% based on the individual components (TC4A-SO2 and TC8A-SO2, resp.)

BOD5 / COD results

Results with reference substance:
The percentage biodegradation of the reference substance after 14 days was 76% , which is higher than 60%.

Any other information on results incl. tables

Since the mean degree of degradation by BOD and by HPLC (sum of 2 components) was -8% and -2%, respectively, the test substance was considered to be persistent.

Analytical results confirmed that the composition of the test substance was not changed.

The residual percentage (concentration analysed relative to the concentration formulated) was 98 -100% for all test solutions containing test substance.

The difference between the maximum value and the minimum value for degradation of the test substance at the end of the test was 1% for the degradation determined from the BOD and 0-2% for the degradation determined by HPLC, both of which were less than 20%.

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
under test conditions no biodegradation observed