Hexasodium 2,2'-[vinylenebis[(3-sulphonato-4,1-phenylene)imino[6-[(2-cyanoethyl)[2-(2-hydroxyethoxy)ethyl]amino]-1,3,5-triazine-4,2-diyl]imino]]bis[benzene-1,4-disulphonate]

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: inherent biodegradability

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From January the 4th to February the 1st, 1990

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Justification for type of information:

Justification for read across in detailed at section 13.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted on 12 May, 1981

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

sewage, domestic, non-adapted

Details on inoculum:

- Source of inoculum: domestic waste-water sewage plant (ARA Sissach/Swiitzerland).
- Preparation of inoculum for exposure: areated prior incubation (2.5-l cylindrical reactor flasks).
- Concentration of sludge: 0.2 g dry material per litre of the final test medium.

Duration of test (contact time):

28 d

Initial conc.:

40.6 mg/L

Based on:

test mat.

Initial conc.:

91.5 mg/L

Based on:

DOC

Parameter followed for biodegradation estimation:

DOC removal

Details on study design:

TEST CONDITIONS
- Composition of medium: prepared as described by OECD guideline 302B.
- Test temperature: 21 - 22.5 °C
- pH: ranged from 7.5 to 7.8.
- Aeration of dilution water: flasks areated with a flow rate of ca. 0.5 - 0.7 l/min, resulting in an oxygen concentration of 6.2 - 8.9 mg O2 per litre.
- Continuous darkness: yes.

TEST SYSTEM
- Test article concentration: 40.6 mg/l, corresponding to 91.5 mg DOC/l.
- Number of flasks: both test article and reference compounds were run in duplicate.
- Measuring equipment: Technicon Carbon Analyser (Autoanalyser II; Technicon Industrial Method No. 451-76W) equipped with an Automatic Data Processing System.
- Inoculum blank: an untreated inoculated control was run.

SAMPLING
- Sampling frequency: at day 0 (0 and 3 hours after treatment), 7, 14, 21 and 28 of the incubation period.
- Sampling method: two subsamples of 30 ml were taken per flask and analysed in duplicate for DOC.
- Sample preparation: filtered through a washed fluted filter paper. The first 5 ml of the filtrate were replaced into the reactor. The remaining 25 ml were used for DOC-analysis. Samples were analysed within 24 hours.
- Other: in case precipitated amounts of non-specified material were observed during incubation, it was scraped off into the solution on the respective sampling day. Starting with day 7, the test article precipitated and adhered to the wall of reaction vessel. Water evaporation losses were compensated by adding bidistilled water.

Reference substance:

aniline

Remarks:

100 mg/l, corresponding to a theoretical amount of 77.4 mg DOC/l.

Parameter:

% degradation (DOC removal)

Value:

10

Sampling time:

28 d

Results with reference substance:

Aniline showed a biodegradation of 96 % within 14 days.

Evaluation of % of biodegradation

Substance	Test set no.	% DOC-removal after
		3 h	7 d	14 d	21 d	28 d
Test item	1	2.0	1.0	-2.0	9.0	8.0
	2	12.0	6.0	6.0	13.0	11.0
	Mean	7.0	4.0	2.0	11.0	10.0
Aniline	3	7.0	96.0	96.0	98.0	100.0

Biodegradation of test article

Colture medium	Test set no.	Test article conc. [mg/l]	DOC [mg/l]
			0 h	3 h	7 d	14 d	21 d	28 d
Mineral nutrient solution, test article, inoculum	1A	40.6	82.0	86.0	88.0	89.0	75.0	82.0
	1B	40.6	93.0	86.0	88.0	89.0	85.0	82.0
	Mean	40.6	87.5	86.0	88.0	89.0	80.0	82.0
	2A	40.6	91.0	85.0	95.0	89.0	81.0	87.0
	2B	40.6	100.0	84.0	88.0	89.0	87.0	86.0
	Mean	40.6	95.5	84.5	91.5	89.0	84.0	86.5
Mineral nutrient solution, inoculum	4A	-	0.8	0.9	2.1	0.4	1.2	2.6
	4B	-	0.8	0.9	2.2	0.4	1.2	2.5
	Mean	-	0.8	0.9	2.2	0.4	1.2	2.6

Biodegradation of aniline

Culture medium	Test set N.	Aniline conc. [mg/l]	DOC [mg/l]
			0 h	3 h	7 d	14 d	21 d	28 d
Mineral nutrient solution, aniline, inoculum	3A	100	71.0	66.0	5.1	3.3	2.4	2.5
	3B	100	70.0	65.0	4.9	3.3	2.4	2.5
	Mean	100	70.5	65.5	5.0	3.3	2.4	2.5
Mineral nutrient solution, inoculum	4A	-	0.8	0.9	2.1	0.4	1.2	2.6
	4B	-	0.8	0.9	2.2	0.4	1.2	2.5
	Mean	-	0.8	0.9	2.2	0.4	1.2	2.6

Validity criteria fulfilled:

yes

Remarks:

degradation of aniline within 14 days was higher than 70 %

Interpretation of results:

not inherently biodegradable

Conclusions:

No significant degradation was recorded after 28 days (i.e. 10 %), thus the substance cannot be considered as inherently biodegradable.

Executive summary:

The inherent biodegradability test was performed according to OECD 302B “modified Zahn-Wellens test" by measuring the DOC concentration.

The substance showed a degradation of 10 % within 28 days under the following test conditions: t range from 21 to 22.5 °C, pH range from 7.5 to 7.8, darkness. In addition, it did not show toxicity to microorganisms. The validity of the test was fulfilled since it met all the criteria of the followed guideline.

The substance can not be seen has inherently biodegradable.

Conclusion

No significant degradation was recorded after 28 days (i.e. 10 %), thus the substance cannot be considered as inherently biodegradable.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From June the 15th to July the 13th, 1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Justification for type of information:

Justification for read across is detailed in section 13.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

Version / remarks:

adopted on 12 May, 1981

Qualifier:

according to guideline

Guideline:

EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)

Version / remarks:

adopted in September, 1984

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

sewage, domestic, non-adapted

Details on inoculum:

- Source of inoculum: Waterschap de Aa, Schijndel, the Netherlands.
- Water filtered: yes.
- Type and size of filter used: coarse filter paper, the first 200 ml were discarded.
- Preparation of inoculum for exposure: the filtrate was kept areated until inoculation.
- Temperature: 20 ± 2 °C

Duration of test (contact time):

28 d

Initial conc.:

0.615 other: ml O2/mg

Based on:

ThOD

Remarks:

with nitrogen eliminated as ammonia

Initial conc.:

3 mg/L

Based on:

test mat.

Remarks:

(low conc.)

Initial conc.:

15 mg/L

Based on:

test mat.

Remarks:

(high conc.)

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Test medium: air saturated Milli-Q water with nutrients, according to the followed guideline. All nutrients were added to Milli-Q water in such a way that no precipitation occurred.
- Test vessels: calibrated 300 ml Winckler oxygen bottles with glass stoppers.
- Temperature of different stock solutions: range from 19.6 to 21.0 °C.
- pH values of different stock solutions: 7.4
- Temperature of medium: 21.0 °C
- O2 concentration of test medium: 9.00 mg O2/l
- Continuous darkness: yes.

TEST SYSTEM
- Test substance concentration: because the maximum consumable oxygen was considered to be 3.4 mg O2/l medium, two trials of different test substance concentration were assessed; low at 3.0 mg/l and high at 15.0 mg/l.
- Inoculation: one drop filtrate of the secondary effluent per litre of final volume.
- Milli-Q water: first purified by reverse osmosis and thereafter passed over carbon and ionexchange cartridges.

CONTROL AND BLANK SYSTEM
- Inoculum blank: contained inoculum but neither test nor control substance.
- Positive control: 2.0 mg/l sodium acetate with inoculum.
- Inhibition control: 2.0 mg/l sodium acetate with inoculum and the low concentration (3.0 mg/l) of the test substance.
- Oxygen control: test medium without inoculum.

OTHER
- Expected oxygen demand: the ThOD of the test item was calculated from the bruto formula.
- Potential toxicity: the toxicity of the test substance for the bacteria in the inoculum was evaluated by comparison of the sum of the BOD's of the positive control and the low concentration of the test substance with the BOD recorded for the inhibition control.

MEASUREMENTS
- Sampling frequency: at the start of the experiment (day 0), and at day 5, 15 and 28 in duplicate.
- Measuring equipment: oxygen electrode; accuracy 0.5 %.
- Calibration: directly before each series of determinations.

Reference substance:

acetic acid, sodium salt

Remarks:

0.78 mgl O2/mg

Parameter:

% degradation (O2 consumption)

Remarks:

low concentration test item (3.0 mg/L)

Value:

35

Sampling time:

28 d

Parameter:

% degradation (O2 consumption)

Remarks:

high concentration test item (15.0 mg/L)

Value:

9

Sampling time:

28 d

Details on results:

Although substantial biodegradation (35 %) was recorded after 28 days of exposure, the test substance appears to be not readily biodegradable under the test conditions, as according to OECD 301D.

Since oxygen consumption of the inhibition control was in agreement with the total oxygen depletion expected on basis of the results for the positive control and the low concentration group, the test item appeared to be not toxic to the bacteria present in the inoculum.

Results with reference substance:

Positive control was degraded for more than 60 % within 15 days.

Oxygen depletion at different points in time

Test medium	Oxygen depletion [mg BOD/l] after
	5 d	15 d	28 d
Positive control	0.93	1.05	0.96
Low concentration test item (3.0 mg/l)	0.05	0.02	0.64
High concentration test item (15.0 mg/l)	0.11	0.03	0.81
Inhibition control	1. 23	1.16	1.66

Percentage biodegradation at different points in time

Test medium	% biodegradation after
	5 d	15 d	28 d
Positive control	60	67	62
Low concentration test item (3.0 mg/l)	3	1	35
High concentration test item (15.0 mg/l)	1	0	9

O₂ determinations

Serie	Content	Stock solution		Flask no.	mg O2/l after
		pH	T (°C)		0 d	5 d	15 d	28 d
Oxygen control	Medium	7.4	21	1A	9.00	8.95	8.81	8.70
				1B	8.97	9.02	8.80	8.78
				Mean	8.99	8.99	8.81	8.74
Inoculum blank	Medium, inoculum, positive control	7.4	20	2A	9.19	8.82	8.83	8.43
				2B	9.12	8.98	8.68	8.60
				Mean	9.16	8.90	8.76	8.52
Positive control	Medium, inoculum, positive control	7.4	20.8	3A	9.10	7.90	7.56	7.56
				3B	8.99	7.82	7.63	7.33
				Mean	9.05	7.86	7.60	7.45
Test substance (3.0 mg/l)	Medium, inoculum, test substance (low conc.)	7.4	19.6	4A	9.22	9.02	8.78	8.55
				4B	9.15	8.74	8.74	7.26
				Mean	9.19	8.88	8.76	7.91
Test substance (15.0 mg/l)	Medium, inoculum, test substance (high conc.)	7.4	20.1	5A	9.17	8.70	8.66	8.00
				5B	9.13	8.87	8.78	7.40
				Mean	9.15	8.79	8.72	7.70
Inhibition control	Medium, inoculum, test substance (3.0 mg/l), positive control	7.4	19.6	6A	9.29	7.70	7.66	6.15
				6B	9.19	7.82	7.70	7.74
				Mean	9.24	7.76	7.68	6.95

Validity criteria fulfilled:

yes

Remarks:

control substance biodegradation > 60 % within 28 days; oxygen depletion in the control not exceeded 0.3 and 0.4 mg O2/l after 5 and 28 days; oxygen depletion in inoculum blank not exceeded 0.5 mg O2/l after 5 days and 0.6 mg O2/l after 15 and 28 days.

Interpretation of results:

not readily biodegradable

Conclusions:

The test item appears to be not readily biodegradable under the test conditions.

Executive summary:

Test item was tested for its ready biodegradability, according to OECD 301D (closed bottle test) at concentrations of 3.0 and 15.0 mg/l.

Under the test conditions (T of 20 ± 2 °C, pH of 7.4, darkness), the low concentration of test item (3.0 mg/l) and the high concentration test item (15.0 mg/l) showed a biodegradation of 35 % and 9 %, respectively. The positive control showed a degradation higher than 60 % within 15 days.

In addition, the test item appeared to be not toxic to the microbial activity present in the medium. The test met all the validity criteria of the followed guideline.

Although substantial biodegradation was reached after 28 days of exposure, the test item appears to be not readily biodegradable under the test conditions.

Conclusion

The test item appears to be not readily biodegradable under the test conditions.

Description of key information

The substance has to be considered neither readily, nor inherently biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Type of water:

freshwater

Additional information

There is no information about the biodegradability potential of Optical Brightener 380, therefore, the available data on the structural analogous Similar Substance 01 have been taken into consideration. The read across approach can be considered reliable and appropriate to investigate the property (details for the approach are included into the IUCLID section 13).

Two experiments for the assessment of the biodegradability of the test substance in water were performed, in order to assess both the readily and inherently biodegradability potential.

The ready biodegradability test was performed according to OECD 301D "closed bottle test". The degradation of the substance was determined to be lower than 60 % within 28 days under the following test conditions: temperature of 20 ± 2 °C, pH of 7.4, darkness. Therefore, it cannot be seen as readily biodegradable. No toxicity against microorganisms was detected. The validity of the test was fulfilled since it met all the criteria of the followed guideline.

The inherent biodegradability test was performed according to OECD 302B “modified Zahn-Wellens test". The substance showed a degradation of 10 % within 28 days under the following test conditions: temperature range from 21 to 22.5 °C, pH range from 7.5 to 7.8, darkness. Therefore, it cannot be seen as inherently biodegradable. The substance did not show toxicity to microorganisms. The validity of the test was fulfilled since it met all the criteria of the followed guideline.

In conclusion, the test substance can be considered as neither readily nor inherently biodegradable.