tetrakis[({bis[4-(dimethylamino)phenyl] and 4-(methylamino)phenyl} or {4-(dimethylamino)phenyl and bis[4-(methylamino)phenyl]} or {tris[4-(dimethylamino)phenyl]}methylium)]C12-(branched and/or linear)-alkyl-(4-sulfonatophenoxy)benzenesulfonate and oxybis[C12-(branched and/or linear)-alkyl-benzenesulfonate].

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 2008/06/10 to 2008/06/13

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The reliability is rated 2 because the study followed a standard guideline of reference (NF EN ISO 8692, 2005), which describes a procedure designed to evaluate this endpoint. The results were reviewed for reliability and assessed as valid. However the study was not conducted under GLP conditions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Deviations:

no

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

bis(bis[4-(dimethylamino)phenyl][4-(methylamino)phenyl]methylium) bis(tris[4-(dimethylamino)phenyl]methylium) 3-(dodecan-3-yl)-4-(2-dodecyl-4-sulfonatophenoxy)benzene-1-sulfonate 3-dodecyl-4-(4-sulfonatophenoxy)benzene-1-sulfonate

EC Number:

700-759-4

Molecular formula:

not applicable

IUPAC Name:

bis(bis[4-(dimethylamino)phenyl][4-(methylamino)phenyl]methylium) bis(tris[4-(dimethylamino)phenyl]methylium) 3-(dodecan-3-yl)-4-(2-dodecyl-4-sulfonatophenoxy)benzene-1-sulfonate 3-dodecyl-4-(4-sulfonatophenoxy)benzene-1-sulfonate

Details on test material:

- Name of test material (as cited in study report): Sepisol Fast Violet 881239
- Lot/batch No.: 601204
- Substance type: organic salt - UVCB
- Storage condition of test material: at ambient temperature

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES
- Water solubility (under test conditions): 10 mg/L with a correspondig pH of 2.8

Sampling and analysis

Analytical monitoring:

no

Test solutions

Vehicle:

no

Test organisms

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Age of inoculum (at test initiation): 3 days

Study design

Test type:

not specified

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Test temperature:

22°C +/2

pH:

T0: From 8.1 to 8.2
T72: From 8.2 to 9.3

Nominal and measured concentrations:

Nominal concentration:
0 - 1.0 - 1.5 - 2.0 - 2.9 - 4.0 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer
- Material, size, headspace, fill volume: Erlenmeyer with 60 mL of the tested solution
- Initial cells density: 10 000 cells/mL
- No. of vessels per concentration : 3
- No. of vessels per control (replicates): 6

OTHER TEST CONDITIONS
- Light intensity and quality:4500 lux
- stirring: rev/250 min-1

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Mallassez counting chamber


TEST CONCENTRATIONS
- the maximum tested concentration was 4.0 mg/L.

Reference substance (positive control):

yes

Remarks:

K2Cr2O7

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 1.96-2.13

Effect concentrations 2

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

2.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 2.27-2.42

Effect concentrations 3

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

2.7 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI : 2.65-2.74

Effect concentrations 4

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

1.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Results with reference substance (positive control):

- Results with reference substance valid: Yes
- EC50 (72h): 1.21 mg/L (belongs to the recommended range of values: 0.92 - 1.46 mg/L)
- Other:K2CrO7

Reported statistics and error estimates:

CE x % calculated using the Hill equation (macro Regtox_ev6.6.2xls)
NOEC calculated with the T-test of Bonferroni (Toxcalc software)

Any other information on results incl. tables

Results at t = 24h, 48h and 72h:

 

 

t = 72 h
Concentration (mg/L)	Cells Concentration x 10^6
	1	2	3	4	5	6	Average Cells number	Cells Inhibition
0	3.59	3.78	3.69	4.16	3.72	4.03	3.83
1.0	4.34	4.08	3.81				4.08	-7%
1.5	4.13	3.96	3.56				3.88	-1%
2.0	1.93	1.46	1.60				1.66	57%
2.9	0.09	0.12	0.12				0.11	97%
4.0	0.01	0.01	0.01				0.01	100%

 

t = 72 h
Concentration (mg/L)	Growth Rate
	1	2	3	4	5	6	Average Cells number	Growth rate Inhibition
0	1.96	1.98	1.97	2.01	1.97	2.00	1.98
1.0	2.02	2.00	1.98				2.00	-1%
1.5	2.01	1.99	1.96				1.99	0%
2.0	1.75	1.66	1.69				1.70	14%
2.9	0.72	0.82	0.83				0.79	60%
4.0	0.00	0.00	0.00				0.00	100%

 

t = 48 h
Concentration (mg/L)	Cells Concentration x 10^6
	1	2	3	4	5	6	Average Cells number	Cells Inhibition
0	0.50	0.51	0.47	0.47	0.48	0.46	0.48
1.0	0.45	.049	0.44				0.46	5%
1.5	0.42	0.43	0.30				0.38	20%
2.0	0.13	0.15	0.18				0.16	68%
2.9	0.039	0.035	0.033				0.04	93%
4.0	0.02	0.01	0.01				0.02	97%

 

t = 48 h
Concentration (mg/L)	Growth Rate
	1	2	3	4	5	6	Average Cells number	Growth rate Inhibition
0	1.96	1.96	1.93	1.93	1.93	1.91	1.94
1.0	1.90	1.94	1.89				1.91	1%
1.5	1.87	1.88	1.71				1.82	6%
2.0	1.29	1.36	1.45				1.37	29%
2.9	0.68	0.63	0.59				0.63	67%
4.0	0.35	0.05	0.17				0.19	90%

 

 

  

t = 24 h
Concentration (mg/L)	Cells Concentration x 10^6
	1	2	3	4	5	6	Average Cells number	Cells Inhibition
0	0.075	0.070	0.063	0.077	0.080	0.075	0.07
1.0	0.064	0.068	0.061				0.064	12%
1.5	0.056	0.057	0.057				0.057	23%
2.0	0.045	0.031	0.027				0.034	53%
2.9	0.022	0.026	0.027				0.025	66%
4.0	0.006	0.009	0.011				0.009	88%

t = 24 h
Concentration (mg/L)	Growth Rate
	1	2	3	4	5	6	Average Cells number	Growth rate Inhibition
0	2.01	1.94	1.84	2.04	2.08	2.02	1.99
1.0	1.85	1.91	1.80				1.86	7%
1.5	1.72	1.75	1.74				1.73	13%
2.0	1.50	1.12	0.99				1.20	40%
2.9	0.77	0.97	0.98				0.91	54%
4.0	-0.52	-0.08	0.08				0.18	109%

 

 

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the experimental conditions:
ECr 10 - 72h = 2.0 mg/L (1.95-2.13)
ECr 20 - 72h = 2.2 mg/L (2.18-2.34)
ECr 50 - 72h = 2.7 mg/L (2.65-2.74)
NOEC - 72 h = 1.5 mg/L

Executive summary:

The acute algal toxicity test for the substance Sepisol Fast Violet 881239 was carried out to determine the ecotoxicological profile of the test substance against an unicellular green algae. As test organism Pseudokirchnerella subcapitata was used and the NF EN ISO 8692 (2005) standard method was followed.

 

The parameters which were to be determined were the effective concentrations (EC) and the no observed effect concentration 72h (NOEC-72h) values with regard to the growth rate (ErC).

The following nominal concentrations were tested: 0 - 1.0 - 1.5 - 2.0 - 2.9 - 4.0 mg/L. The maximum tested concentration was 4.0 mg/L.

A stock solution was prepared by diluting 10 mg of test material in 1 liter of pure water.

Each concentration and the controls were inoculated with approximately 10 000 algae per mL.

The test was carried out over 72 hours at a temperature of 22°C +/- 2 and constantly stirred (250 rev.min^-1). Each vessel concentration was tested 3 times. 6 controls were run with the same experimental conditions. Algal concentration was counted using a Mallassez counting chamber. Once a day the extinction of an aliquot of the test-vessels (incubation) was measured.

Validation experiments and the precultivation ensured that the algal were of appropriate vialibilty and susceptibility.

The results of this study showed, that the substance Sepisol Fast Violet 2B inhibit cells and reduce the growth rate of the test organism at the tested concentrations.  Under experimental conditions, the following parameters were determined:

ECr 10 - 72h = 2.0 mg/L (1.95-2.13)

ECr 20 - 72h = 2.2 mg/L (2.18-2.34)

ECr 50 - 72h = 2.7 mg/L (2.65-2.74)

NOEC - 72 h = 1.5 mg/L