Tetrasodium [2-({4-fluoro-6-[(2-{[4-fluoro-6-({5-(hydroxykappaO)-6-[(2-{[2-(hydroxy-kappaO)-5-sulfophenyl] diazenyl-kappaN1}-4,5-dimethoxyphenyl) diazenyl-kappaN2]-7-sulfo-2-naphthyl} amino)-1,3,5-triazin-2-yl] amino} propyl) amino]-1,3,5-triazin-2-yl} amino) benzene-1,4- disulfonato(6-)] cuprate(4-)

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experiment start date - 02 May 2006;
Experiment completion date - 31 May 2006;
Study completion date - 19 July 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Identity: FAT 40825/A
Batch number: CHU 297 / BOP 04/05
Purity: Organic part (Na-salt): approx. 83.7 %; All coloured components: approx. 80.54 %; Main component: approx. 59.1 %.
Appearance: Solid, black powder
Storage conditions: At room temperature at about 20 °C
Expiration date: December 31, 2010

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

The study was performed with aerobic activated sludge from a wastewater treatment plant (ARA Ergolz II, Füllinsdorf, Switzerland), treating predominantly domestic wastewater. The sludge was washed twice with tap water by centrifugation and the supernatant liquid phase was decanted. A homogenized aliquot of the final sludge suspension was weighed, thereafter dried and the ratio of wet to dry weight was calculated.
Based on this ratio, calculated amounts of wet sludge were suspended in test water to obtain a concentration equivalent to 4 g (±10 %) dry material per liter. During holding, the sludge was aerated at room temperature until use. Prior to use, the sludge was first thoroughly mixed and then diluted with test water to a concentration of 1 g per liter (dry weight basis). Based on the determined dry weight of this diluted activated sludge, defined amounts were added to test water to obtain a final concentration of 30 mg dry material per liter.

Duration of test (contact time):

28 d

Initial conc.:

101 mg/L

Based on:

test mat.

Initial conc.:

100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

Test concentrations:
The test item was weighed by means of an analytical balance and transferred to the test flasks with test water. No emulsifiers or solvents were used. The reference item sodium benzoate was tested simultaneously under the same conditions as the test item, and functioned as a procedure control. A stock solution containing 2.5 g sodium benzoate per liter test water was prepared by completely dissolving 250 mg sodium benzoate in 100 mL test water. From this stock solution, 10 mL aliquots were added to the corresponding test flasks containing test water.
Finally, with the exception of the abiotic control flask, activated sludge was added to each test flask. The final volume was 250 mL per test flask.

Conditions of the test:
Light conditions: Darkness
Test temperature: 22 °C, maintained with a built-in thermostat and checked once per week.
pH: Prior to test start, the pH was measured in each test flask before the addition of the activated sludge inoculum. At the end of incubation, the pH was measured again in each test flask.

Reference substance:

benzoic acid, sodium salt

Remarks:

Sodium benzoate

Parameter:

% degradation (O2 consumption)

Value:

-3

Sampling time:

28 d

Remarks on result:

other: based on ThODNH4

Parameter:

% degradation (O2 consumption)

Value:

-2

Sampling time:

28 d

Remarks on result:

other: based on ThODN03

Details on results:

The biochemical oxygen demand (BOD) of the test item FAT 40825/A in the test media was in the range of the inoculum controls throughout the entire study period of 28 days. Therefore, FAT 40825/A was not biodegradable under the test conditions within 28 days.
In the toxicity control, the course of biodegradation over the 28-day exposure period was similar to the two procedure controls, containing only the reference item. Within 14 days of exposure, biodegradation amounted to 60 % and 46 % based on the ThODNH4 and ThODN03, respectively. Thus, according to the test guidelines, the test item had no inhibitory effect on activated sludge microorganisms at the tested concentration of 100 mg/L, because biodegradation in the toxicity control was >25 % within 14 days. The pH measured in all flasks at the start of the test was in the range 7.3 - 7.4. At the end of exposure (Day 28), pH values in the range 7.4 - 7.8 were measured.

Parameter:

BOD5

Value:

0 mg O2/g test mat.

Parameter:

COD

Value:

0.85 mg O2/g test mat.

Results with reference substance:

28 d biodegradation: 99 % based on ThOD

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

FAT 40825/A was not biodegradable under the test conditions within 28 days.

Executive summary:

The test item FAT 40825/A was investigated for its ready biodegradability in a manometric respirometry test over 28 days according to EU Commission Directive 92/69 EEC, C.4-D (1992) and OECD Guideline for Testing of Chemicals No. 301 F (1992). The biochemical oxygen demand (BOD) of the test item FAT 40825/A in the test media was in the range of the inoculum controls throughout the entire 28-day exposure period. Consequently, FAT 40825/A was not biodegradable under the test conditions within 28 days. No degradation of the test item occurred in the abiotic control under the test conditions within 28 days. In the toxicity control, containing both FAT 40825/A and the reference item sodium benzoate, no inhibitory effect on the biodegradation of the reference item was determined. Thus, FAT 40825/A had no inhibitory effect on the activity of activated sludge microorganisms at the tested concentration of 100 mg/L. In the procedure controls, the reference item sodium benzoate was degraded by an average of 92 % by Exposure Day 14, and reached complete biodegradation (99 %) by the end of the test (Exposure Day 28), thus confirming suitability of the activated sludge.

Reference 2

Endpoint:

biodegradation in water: inherent biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experiment start date - 10 April 2006;
Experiment completion date - 09 May 2006;
Study completion date - 08 September 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 302 B (Inherent biodegradability: Zahn-Wellens/EMPA Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.9 (Biodegradation: Zahn-Wellens Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Identity: FAT 40825/A
Batch number: CHU 297 / BOP 04/05
Purity: Organic part (Na-salt): approx. 83.7 %; All coloured components: approx. 80.54 %; Main component: approx. 59.1 %.
Appearance: Solid, black powder
Storage conditions: At room temperature at about 20 °C
Expiration date: December 31, 2010

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

The study was performed with aerobic activated sludge from a wastewater treatment plant (ARA Ergolz II, Füllinsdorf, Switzerland), treating predominantly domestic wastewater. The sludge was washed three times with tap water by centrifugation and the supernatant liquid phase was decanted. A homogenized aliquot of the final sludge suspension was weighed, thereafter dried and the ratio of wet to dry weight was calculated. Based on this ratio, calculated amounts of wet sludge were suspended in test water to obtain a concentration equivalent to 4 g (± 10 %) dry material per liter. During holding, the sludge was aerated at room temperature until use. Before use, this diluted activated sludge was mixed, the dry weight was determined again, and defined amounts were added to test water to obtain a final concentration of 300 mg dry material per liter. The ratio between inoculum and test item (based on the mean DOC measured) was 3 : 1.

Duration of test (contact time):

28 d

Initial conc.:

331.6 mg/L

Based on:

test mat.

Initial conc.:

329.1 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

Test concentrations:
The test item was directly weighed into the test flasks, and about 900 ml of test water was added. No emulsifiers or solvents were used. The reference item diethylene glycol was tested simultaneously under the same conditions as the test item, and functioned as a procedure control. A stock solution containing 8.8 g diethylene glycol per liter test water was prepared, by completely dissolving 2.2 g diethylene glycol in 250 mL test water. From this, 25 mL aliquots were added to the corresponding test flasks. To each flask, activated sludge was added. Finally, the flasks were made up to a volume of 1000 mL with test water.

Test conditions:
Test vessels: 2000-mL Erlenmeyer flasks, cleaned with alcoholic hydrochloric acid, rinsed with deionized water and dried. The final volume was 1000 mL per flask. Each flask was loosely covered with aluminium foil to allow the exchange of air between the flask and the surrounding atmosphere. The test media were continuously stirred by magnetic stirrers.
Test temperature: 21 - 24 °C. The inoculated flasks were incubated in a temperature-controlled room. The temperature was checked on each sampling date in one of the inoculum blanks. Additionally, the room temperature was continuously recorded.
pH: Prior to test start (after the addition of activated sludge inoculum) and before each sampling, the pH was measured in each test flask. Adjustment of pH was not necessary since the pH was at each measurement within the guideline recommended range of 7.0-8.0.
Light conditions: The test flasks were incubated in the dark.
Oxygen concentration: 7.0 - 9.0 mg O2/L. The oxygen concentration was measured at the start of the test and before each sampling.

SAMPLING:
Samples were taken on Day 0 (0 hours and 3 hours ± 30 minutes after the addition of the test and/or reference item) and on Day 2, 7, 9, 14, 21, and 28 of the incubation period. Prior to sampling, water evaporation losses were compensated by adding purified water up to the mark prior to sampling. Deposits on the test vessels were scraped off and resuspended in the test vessel. Per sampling interval, one sample of about 10 mL was taken and analyzed for DOC.

Reference substance:

diethylene glycol

Parameter:

% degradation (DOC removal)

Value:

6

Sampling time:

28 d

Details on results:

In the test flasks, containing the test item and activated sludge (inoculum), the mean concentration of DOC (dissolved organic carbon) varied between 94 and 101 mg/L over the 28-day exposure period, and was not significantly different from the initial mean DOC concentration of 101 mg/L, measured on Day 0 (starting value after 3 hours of exposure). Expressed as percentage DOC-removal, mean values in the range from 1 to 7 % were noted. Therefore, FAT 40825/A was not biodegradable under the test conditions within 28 days. No DOC removal was observed during the first three hours of exposure, which is an indication that the test item did not adsorb on the activated sludge (inoculum). No elimination of the test item occurred in the abiotic control under the test conditions after 28 days of exposure.

Results with reference substance:

In the procedure controls, containing the reference item diethylene glycol and activated sludge (inoculum), the reference item was completely degraded within the first seven days of exposure, thus confirming suitability of the activated sludge.

In the toxicity control, containing the test item (corresponding to 49 % of total DOC), the reference item (corresponding to 51 % of total DOC) and activated sludge (inoculum), the initial DOC concentration of 202 mg/L measured on Day 0 (after three hours) rapidly decreased by 47 % within the first 14 days of exposure. Thus, the test item was not inhibitory to activated sludge at the tested concentration of about 330 mg/L, because degradation was >35 % within 14 days of incubation (based on the test guidelines).

During the test, the pH was in the range 7.0 - 7.6.

During the test, the oxygen concentration was in the range 7.0 - 9.0 mg/L.

During the test, the temperature was in the range 21 - 24 °C.

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

FAT 40825/A was not biodegradable under the test conditions within 28 days.

Executive summary:

The test item FAT 40825/A was investigated for its potential (inherent) ultimate biodegradability in a Zahn-Wellens / EMPA test over 28 days, based on the OECD Guideline for Testing of Chemicals, No. 302 B (1992) and the EU Commission Directive 88/302/EEC, Part C.9 (1988). In the test flasks, containing the test item and activated sludge (inoculum), the mean concentration of DOC (dissolved organic carbon) varied between 94 and 101 mg/L over the 28-day exposure period, and was not significantly different from the initial mean DOC concentration of 101 mg/L, measured on Day 0 (starting value after 3 hours of exposure). Expressed as percentage DOC-removal, mean values in the range from 1 to 7 % were noted. FAT 40825/A was not biodegradable under the test conditions within 28 days. No DOC removal was observed during the first three hours of exposure, which is an indication that the test item did not adsorb on the activated sludge (inoculum). The reference item diethylene glycol ultimately and completely degraded within the first seven days of exposure, thus confirming suitability of the activated sludge. In the abiotic control, containing the test item and poisoned mineral medium, no degradation was noted after 28 days of exposure (based on DOC measurements). In the toxicity control, containing the test item, the reference item diethylene glycol and activated sludge (inoculum), the initial DOC decreased by 47 % within 14 days of exposure. Thus, FAT 40825/A was not inhibitory to activated sludge at the tested concentration of about 330 mg/L, because degradation was >35 % within 14 days (based on the test guidelines).

Description of key information

FAT 40825/A was not biodegradable under the test conditions within 28 days.

Key value for chemical safety assessment

Biodegradation in water:

not biodegradable

Type of water:

freshwater

Additional information

Test reports for ready biodegradation and inherent biodegradation tests are available. The results are summarized as below:

The test item FAT 40825/A was investigated for its ready biodegradability in a manometric respirometry test over 28 days according to EU Commission Directive 92/69 EEC, C.4-D (1992) and OECD Guideline for Testing of Chemicals No. 301 F (1992). The biochemical oxygen demand (BOD) of the test item FAT 40825/A in the test media was in the range of the inoculum controls throughout the entire 28-day exposure period. Consequently, FAT 40825/A was not readily biodegradable under the test conditions within 28 days. The test item FAT 40825/A was investigated for its potential (inherent) ultimate biodegradability in a Zahn-Wellens / EMPA test over 28 days, based on the OECD Guideline for Testing of Chemicals, No. 302 B (1992) and the EU Commission Directive 88/302/EEC, Part C.9 (1988). In the test flasks, containing the test item and activated sludge (inoculum), the mean concentration of DOC (dissolved organic carbon) varied between 94 and 101 mg/L over the 28-day exposure period, and was not significantly different from the initial mean DOC concentration of 101 mg/L, measured on Day 0 (starting value after 3 hours of exposure). Expressed as percentage DOC-removal, mean values in the range from 1 to 7 % were noted. FAT 40825/A was not biodegradable under the test conditions within 28 days.