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EC number: 600-520-3 | CAS number: 1040874-53-0
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
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:
- 04 Apri l- 30 May 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 018
- Report date:
- 2018
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
- Version / remarks:
- July 17, 1992
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)
- Version / remarks:
- May 30, 2008
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 835.3110 (Ready Biodegradability)
- Version / remarks:
- January 1998
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- Reaction product of 4-aminophenol with 2-ethyl-6-methylbenzenamine, sodium polysulfide and sodium metabisulfite
- EC Number:
- 600-520-3
- Cas Number:
- 1040874-53-0
- Molecular formula:
- not applicable
- IUPAC Name:
- Reaction product of 4-aminophenol with 2-ethyl-6-methylbenzenamine, sodium polysulfide and sodium metabisulfite
- Test material form:
- solid: particulate/powder
- Details on test material:
- Test item: Blue TBR
Appearance: black bluish powder
CAS No: 1040874-53-0
1
- Specific details on test material used for the study:
- Expiration date: 31.05.2022
Study design
- Oxygen conditions:
- anaerobic
- Inoculum or test system:
- activated sludge, domestic, non-adapted
- Details on inoculum:
- Test System
Species:
Activated sludge, microorganisms from a domestic waste water treatment plant.
Origin:
The (controlled) activated sludge was supplied by the sewage plant for domestic sewage in Balatonfüred, Hungary, on 06 April 2018 (seven days before the main test). The prepared activated sludge was continuously aerated (2L/minute) at the test temperature of 22±2oC, for about 7 days.
Preparation of Activated Sludge Inoculum:
The activated sludge used for this study was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in isotonic saline solution with shaking and again centrifuged. This procedure was repeated twice. An aliquot of the final sludge suspension was weighed (5.218 g wet weight), dried and the ratio of wet sludge to dry weight (0.4927 g dry weight) determined. Based on this ratio, calculated amount of wet sludge (5 g dry weight that was equivalent to 59.953 g wet sludge) was suspended in mineral medium (Section 5.4; ad. 1000 mL) to yield a concentration equivalent to about 5 g per litre (on dry weight basis). The prepared activated sludge inoculum was aerated under test conditions (for 7 days) until use.
The pH of the activated sludge inoculum after preparation was 7.42, just before use the pH was: 7.39. A pH adjustment of activated sludge inoculum was not performed.
Pre-conditioning of Activated Sludge Inoculum:
Pre-conditioning consisted of aerating (2 L/minute) activated sludge (in mineral medium ) for 7 days (from April 06 to 13, 2018) at test temperature (the actual temperature: 20.0 – 20.8 oC). Before use the cell count of inoculum was checked as follows: the viability of the cultured sludge was determined by plating 0.1 mL of the different, 10-1, 10-2, 10-3 and 10-4 dilutions of cultures on nutrient agar plates. Plates were incubated at 37 °C for 24 hours. The viable cell number of the cultures was determined by these plating experiments by manual colony counting. The approximately cell count of aerated inoculum was ~10^8/L; therefore, before the test the inoculum was further diluted 50 000x with mineral medium to reach the necessary ~10^4 – 10^6 cells/L cell concentration. After preparation the sludge was filtered through cotton wool. Pre-conditioning (see above) improves the precision of the method. The inoculum was not pre-adapted to the test chemical. - Duration of test (contact time):
- 28 d
Initial test substance concentration
- Initial conc.:
- 4 mg/L
- Based on:
- other: was based on the results of the preliminary solubility and toxicity tests
Parameter followed for biodegradation estimation
- Parameter followed for biodegradation estimation:
- O2 consumption
- Details on study design:
- Environmental Conditions
The test was carried out in a controlled environment room (during the preparation, aeration and incubation of the mineral medium, preparation of test bottles (units), during the formulation, oxygen and pH measuring) at a temperature of 22 ± 2 ºC according to the guideline. The actual temperature range was 20.8 - 21.3 °C.
The test bottles were incubated in an incubator at 22 2 C, in the dark. During the incubation (28 days) of the test units the temperature range was 20.0-20.2 °C.
During the pre-conditioning of activated sludge inoculum the temperature was 20.0 – 20.8 oC.
Temperature was measured continuously using min/max thermometer (in controlled environment room) or built-in thermometer (in incubator) and recorded at least once a day.
The oxygen concentration of test water (mineral medium) was in the range of 8-9 mg/L. It was measured at the start of the test and found to be 8.24 mg/L.
The pH was checked prior study start and found to be 7.41; further pH adjustment was considered as not necessary. The test conditions were measured with suitable instruments and documented in the raw data.
Equipment
Large glass tank (volume: ~30 L) and large glass bottles (volume: 5 L),
Narrow necked, Winkler bottles with glass stoppers,
Funnels and coarse filter papers,
Oxygen and pH meter with appropriate O2 and pH electrode,
Aeration system, Moisture analyzer,
Temperature controlled (in the range of 22 ± 2 °C with a temperature deviation of ±1 °C) environment room (and/or incubator) with thermometer with exclusion of light,
Balance, Centrifuge, Ultrasonic bath, Freezer,
Spectrophotometer (for nitrate, nitrite and COD determination).
Test Units
Type and Size: Winkler bottles (300 mL, coded) with special neck and glass stoppers.
Identification: Each test bottle was uniquely identified with study number, test group, days of measurement and replicate number.
Preliminary Experiments
The pre-experiments on solubility of the test item, and the 14-day toxicity test were conducted non-GLP, and are excluded from the Statement of Compliance in the final report. The raw data of these tests will be archived under the study code of present study.
Preliminary Toxicity Test
The test item solubility, behavior, and toxicity were tested in a 14-day preliminary experiment. The test design was the same as described at the main experiment. In the preliminary experiment the test item was investigated at the concentration of 4 mg/L. Unequivocal toxic effect of the test item was not found at this investigated concentration (in the toxicity control group 30.3 % (higher than 25 %) degradation occurred within 14 days).
Reference substance
- Reference substance:
- benzoic acid, sodium salt
Results and discussion
- Preliminary study:
- The test item solubility, behavior, and toxicity were tested in a 14-day preliminary experiment. The test design was the same as described at the main experiment. In the preliminary experiment the test item was investigated at the concentration of 4 mg/L. Unequivocal toxic effect of the test item was not found at this investigated concentration (in the toxicity control group 30.3 % (higher than 25 %) degradation occurred within 14 days).
- Test performance:
- The chosen test item concentration of 4.0 mg/L investigated in the main test was based on the results of the preliminary solubility and toxicity tests. The COD of 1.23 mg O2/ mg test item of Blue TBR was determined at the start of the main experiment.
Under the test conditions ready biodegradation of this test item was not noticed. The percentage biodegradation of Blue TBR reached a mean of 18.7 % after 28 days based on its COD. Based on the dissolved oxygen depletion, the resulting biodegradation values reached a plateau on about the 14th day of the experiment. From this day the slight changes were considered as being within the biological variability range of the applied test system.
The concurrently conducted analytical determination of possible nitrite and nitrate development showed slight changes in nitrite concentrations in all measured 28-day samples; however the measured dissolved oxygen concentrations in the inoculum control, test item and toxicity control bottles did not correspond to the consumed oxygen of ammonium oxidation processes. Likely technical effects (turbidity and/or discoloration) also influenced the nitrite concentration determinations. Therefore the biodegradability value of the test item was calculated based on its COD; any correction, based on the measured nitrite and/or
nitrate content was not performed.
The reference item Sodium benzoate was sufficiently degraded to a mean of 71.4 % after 14 days, and to a mean of 73.6 % after 28 days of incubation based on ThODNH3, thus confirming the suitability of the used activated sludge inoculum.
In the toxicity control containing both, the test item and the reference item, a mean of 43.1 % biodegradation was noted within 14 days and 46.1 % biodegradation after 28 days of incubation. Thus, the test item can be assumed to not inhibit the activated sludge microorganisms (higher than 25 % degradation occurred within 14 days).
% Degradation
- Key result
- Parameter:
- % degradation (O2 consumption)
- Value:
- 18.7
- Sampling time:
- 28 d
- Details on results:
- Under the test conditions the percentage biodegradation of the test item reached a mean of 18.7 % after 28 days based on its COD. Based on the dissolved oxygen depletion, the resulting biodegradation values reached a plateau on about the 14th day of the experiment. From this day on, the slight subsequent changes were considered as being within the biological variability range of the applied test system. The test item can be considered to be not readily biodegradable.
BOD5 / COD results
BOD5 / COD
- Key result
- Parameter:
- COD
- Value:
- 1.23 g O2/g test mat.
- Results with reference substance:
- Biodegradation of the Reference Item
The reference item Sodium benzoate was sufficiently degraded to a mean of 71.4% after 14 days, and to a mean of 73.6 % after 28 days of incubation, based on ThODNH3, thus confirming the suitability of the used activated sludge inoculum.
Biodegradation of the Toxicity Control
In the toxicity control containing both, the test item and the reference item, a mean of 43.1 % biodegradation was noted within 14 days and 46.1 % biodegradation after 28 days of incubation. Thus, the test item can be assumed to not inhibit the activated sludge microorganisms (higher than 25 % degradation occurred within 14 days).
Any other information on results incl. tables
Dissolved Oxygen Concentrations at Different Time Intervals during the Exposure Period of 28 Days
Treatment |
Concentration |
Bottle |
mg O2/L after n days of exposure |
||||
[mg/L] |
No. |
0 |
7 |
14 |
21 |
28 |
|
Test item |
|
1a |
8.18 |
6.72 |
6.30 |
6.09 |
5.97 |
4.0 |
1b |
8.16 |
6.77 |
6.43 |
6.18 |
6.09 |
|
|
mean |
8.17 |
6.75 |
6.37 |
6.14 |
6.03 |
|
Reference item |
|
2a |
8.28 |
4.37 |
3.79 |
3.51 |
3.35 |
3.0 |
2b |
8.26 |
4.10 |
3.54 |
3.46 |
3.39 |
|
|
mean |
8.27 |
4.24 |
3.67 |
3.49 |
3.37 |
|
Inoculum control |
– |
3a |
8.23 |
7.34 |
7.27 |
7.21 |
7.16 |
3b |
8.22 |
7.22 |
7.11 |
6.94 |
6.85 |
||
mean |
8.23 |
7.28 |
7.19 |
7.08 |
7.01 |
||
Toxicity control |
Test item: 4.0 |
4a |
8.18 |
4.23 |
2.87 |
2.28 |
2.21 |
4b |
8.15 |
3.37 |
2.66 |
2.42 |
2.34 |
||
mean |
8.17 |
3.80 |
2.77 |
2.35 |
2.28 |
Oxygen Depletion at Different Time Intervals during the Exposure Period of 28 Days
Treatment |
Concentration |
Bottle |
mg O2/L after n days of exposure |
|||
[mg/L] |
No. |
7 |
14 |
21 |
28 |
|
Test item |
4.0 |
1a |
0.51 |
0.84 |
0.94 |
0.99 |
1b |
0.44 |
0.69 |
0.83 |
0.85 |
||
Reference item |
3.0 |
2a |
2.97 |
3.46 |
3.62 |
3.71 |
2b |
3.22 |
3.69 |
3.65 |
3.65 |
||
Toxicity control |
Test item: 4.0 |
4a |
3.01 |
4.28 |
4.75 |
4.75 |
4b |
3.84 |
4.46 |
4.58 |
4.59 |
oxygen depletion : (mt0- mtx) - (mbo- mbx), where:
mt0: oxygen concentration (mg/L) of test group on day 0 (1a, 2a, 4a and 1b, 2b, 4b from Table 2)
mtx: oxygen concentration (mg/L) of test group on day x (1a, 2a, 4a and 1b, 2b, 4b from Table 2)
mb0: oxygen concentration (mg/L) of inoculum blank on day 0 (mean of 3a and 3b from Table 2)
mbx: oxygen concentration (mg/L) of inoculum blank on day x (mean of 3a and 3b from Table 2)
BOD at Different Time Intervals during the Exposure Period of 28 Days
Treatment |
Concentration |
Bottle |
BOD after n days of exposure |
|||
[mg/L] |
No. |
7 |
14 |
21 |
28 |
|
Test item |
4.0 |
1a |
0.13 |
0.21 |
0.24 |
0.25 |
1b |
0.11 |
0.17 |
0.21 |
0.21 |
||
Reference item |
3.0 |
2a |
0.99 |
1.15 |
1.21 |
1.24 |
2b |
1.07 |
1.23 |
1.22 |
1.22 |
||
Toxicity control |
Test item: 4.0 |
4a |
0.43 |
0.61 |
0.68 |
0.68 |
4b |
0.55 |
0.64 |
0.65 |
0.66 |
BOD = = mg O2/mg T.i and/or R.i.where:T .i. =test itemR.i.=reference item i.control=inoculum control
Percentage Biodegradation at Different Time Intervals during the Exposure Period of 28 Days
Treatment |
Concentration |
Bottle |
Percent of biodegradation after n days of exposure |
|||
[mg/L] |
No. |
7 |
14 |
21 |
28 |
|
Test item |
|
1a |
10.5 |
17.2 |
19.1 |
20.1 |
4.0 |
1b |
9.0 |
14.1 |
16.9 |
17.3 |
|
|
mean |
9.8 |
15.7 |
18.0 |
18.7 |
|
Reference item |
|
2a |
59.3 |
69.1 |
72.4 |
74.2 |
3.0 |
2b |
64.3 |
73.7 |
73.0 |
73.0 |
|
|
mean |
61.8 |
71.4 |
72.7 |
73.6 |
|
Toxicity control |
Test item: 4.0 |
4a |
29.6 |
42.2 |
46.9 |
46.9 |
4b |
37.8 |
43.9 |
45.2 |
45.3 |
||
mean |
33.7 |
43.1 |
46.0 |
46.1 |
Applicant's summary and conclusion
- Validity criteria fulfilled:
- yes
- Interpretation of results:
- not readily biodegradable
- Conclusions:
- The test item was considered to be not readily biodegradable (18.7 % biodegradation on day 28).
- Executive summary:
The purpose of this study was to determine the ready biodegradability of the test item. The test item was exposed to activated sludge from the aeration tank of a domestic waste water treatment plant. The biodegradation was followed by oxygen uptake of the microorganisms during exposure. As a reference item sodium benzoate (at a concentration of 3.0 mg/L) was tested simultaneously under the same conditions as the test item, and functioned as a procedure control (reference control). Additionally inoculum (containing the filtered inoculum only) and toxicity (containing both the test item and reference item) controls were examined. The chosen test item concentration of 4.0 mg/L investigated in the main test was based on the results of the preliminary solubility and toxicity tests. The chemical oxygen demand (COD) of 1.23 mg O2/ mg test item was determined at the start of the main experiment.
Under the test conditions ready biodegradation of this test item was not noticed. The percentage biodegradation of test item reached a mean of 18.7 % after 28 days based on its COD. Based on the dissolved oxygen depletion, the resulting biodegradation values reached a plateau on about the 14th day of the experiment. From this day the slight changes were considered as being within the biological variability range of the applied test system. The concurrently conducted analytical determination of possible nitrite and nitrate development showed slight changes in nitrite concentrations in all measured 28-day samples; however the measured dissolved oxygen concentrations in the inoculum control, test item and toxicity control bottles did not correspond to the consumed oxygen of ammonium oxidation processes. Likely technical effects (turbidity and/or discoloration) also influenced the nitrite concentration determinations. Therefore the biodegradability value of the test item was calculated based on its COD; any correction, based on the measured nitrite and/or nitrate content was not performed.
The reference item Sodium benzoate was sufficiently degraded to a mean of 71.4 % after 14 days, and to a mean of 73.6 % after 28 days of incubation, based on ThODNH3, thus confirming the suitability of the used activated sludge inoculum.
In the toxicity control containing both, the test item and the reference item, a mean of 43.1 % biodegradation was noted within 14 days and 46.1 % biodegradation after 28 days of incubation. Thus, the test item can be assumed to not inhibit the activated sludge microorganisms (higher than 25 % degradation occurred within 14 days).
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