3,3'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[N-cyclohexyl-2,4,6-trimethylbenzenesulphonamide]

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: well performed OECD study with GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

3,3'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[N-cyclohexyl-2,4,6-trimethylbenzenesulphonamide]

EC Number:

245-728-1

EC Name:

3,3'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[N-cyclohexyl-2,4,6-trimethylbenzenesulphonamide]

Cas Number:

23552-74-1

Molecular formula:

C44H52N4O6S2

IUPAC Name:

3,3'-[(9,10-dioxo-9,10-dihydroanthracene-1,4-diyl)diimino]bis(N-cyclohexyl-2,4,6-trimethylbenzenesulfonamide)

Sampling and analysis

Analytical monitoring:

no

Test solutions

Vehicle:

no

Details on test solutions:

Preparation of the The saturated solution (100 mg/L test item were weighed out) was
saturated solution prepared with dilution water (as specified below) one day prior to the start of the exposure (-24 hours).

Treatment The test item was dispersed in the dilution water by stirring with approximately 1100 rpm for 24 hours at room
temperature.
After completion of stirring, the test item dispersion was filtered with a membrane filter
(0.45 µm, RC, MACHEREY-NAGEL) to achieve the saturated solution.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Clone 5
- Source:
Origin: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu)
Breeder: DR.U.NOACK-LABORATORIEN, Käthe-Paulus-Str. 1, 31157 Sarstedt, Germany
- Age at study initiation (mean and range, SD): 2 to 24 h old daphnids from a healthy stock were used for the study
- Method of breeding:
Culture: In glass vessels (2 - 3 L capacity) with approximately 1.8 L culture medium, at 20 +- 2°C, in an incubator, 16 h illumination; light intensity
of max. 20 µE x m-2 x s-1
Culture medium: Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L
Cultur feeding: At least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and
Desmodesmus subspicatus, with an algae cell density of > 106 cells/mL
- Feeding during test: No feeding

ACCLIMATION
- Acclimation period: At least 2 h in dilution water.
- Acclimation conditions (same as test or not): Same as test conditions.
- Health during acclimation (any mortality observed): No mortality observed.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

265 mgCaCO3/L

Test temperature:

18 - 22°C, constant within ± 1°C
At start of exposure: 21.0

pH:

pH at the Start of the Exposure (0 hours)
(measured in one additional replicate of the limit concentration and the control)

Nominal loading
of the test item
[mg/L] pH-value

100 7.85
(Saturated solution)
Control 7.91


pH at the End of the Exposure (48 hours)
(measured in all replicates)

Nominal loading
of the test item
[mg/L] pH-values
Replicates
1 2 3 4
100 7.70 7.72 7.73 7.72
(Saturated solution)
Control 7.53 7.63 7.67 7.68

Dissolved oxygen:

O2-concentration at the Start of the Exposure (0 hours)
(measured in one additional replicate of the limit concentration and the control)

Nomonal loading
of the test item
[mg/L]
Dissolved
O2-concentration
[mg/L]
100 5.02
(Saturated solution)
Control 8.54



O2-concentration at the End of the Exposure (48 hours)
(measured in all replicates)

Nominaltest loading
of the test item
[mg/L] Dissolved O2 -concentration [mg/L]
Replicates
1 2 3 4
100 8.43 8.49 8.58 8.55
(Saturated solution
Control 8.36 8.27 8.34 8.29

Nominal and measured concentrations:

Nominal loading: 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers (4 cm ID x 7 cm H), 50 mL capacity, loosely covered with watch glasses
- Test volume: 20 mL
- Aeration: None
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO Test water, according to OECD 202, Annex 3
Component Concentration [mg/L]
KCl 5.76
NaHCO3 64.8
CaCl2 x 2 H2O 294
MgSO4 x 7 H2O 123

- Water Quality Parameters of the Dilution Water prior to the Start of the Exposure (0 h):
pH-value DissolvedO2-concentration[mg/L] Temperature[°C] Conductivity[µS/cm] Total hardness [mg CaCO3/L]
7.91 8.54 21.0 661 265

- Culture medium different from test medium: Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L
- Intervals of water quality measurement: test start (0h) and test end (48h)

OTHER TEST CONDITIONS
- Photoperiod: 16/8 h light/dark cycle
- Light intensity: Diffuse light, light intensity of max. 20 µE x m-2 x s-1

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Immobilisation, 24 h interval

TEST CONCENTRATIONS
-The effects of the saturated solution of the test item were tested in a limit test.

- Range finding study:
A non GLP preliminary range finding test under static conditions over a period of 48 hours was conducted at the test facility with a saturated solution of the test item with a nominal loading of 100 mg/L. In the range finding test, two replicates each of the saturated solution and the control, each with
ten daphnids, were tested. No significant effects were observed at the saturated solution. The biologically results are presented in the table below.


Immobilisation Rates in the non GLP Preliminary Range Finding Test
(n = 20, divided into 2 replicates with 10 daphnids each)

Nominal loading
of the test item

[mg/L] IMMOBILISATION [%]
24 hours 48 hours
Replicates Replicates
1 2 MV 1 2 MV
100 0 0 0 0 0 0
(Saturated solution)
Control 0 0 0 0 0 0


MV = mean value


The exposure concentrations were monitored at the start (0 hours) and at the end (48 hours) of the exposure. The results of the analytical
monitoring during the range finding test are presented in the table below. The measured values were below the system quantification limit of
0.4 mg test item/L.



Measured Concentrations of the Test Item during the non GLP Preliminary Range Finding Test


Sampling date 2015-02-10 2015-02-12
0 hours 48 hours
Start of the exposure interval End of the exposure interval


Nominal loading Polysynthren Blue RLS
of the test item
[mg/L]
Meas. conc. Meas. conc.
[mg/L] [mg/L]

100 < SysQL < SysQL
(Saturated solution) < SysQL < SysQL
Control


Meas. conc. = measured concentration, mean value of 2 injections
SysQL = system quantification limit (0.4 mg/L of the test item)

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

24 h

Dose descriptor:

other: EL10,50,100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Effect concentrations 2

Duration:

48 h

Dose descriptor:

other: EL10,50,100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Effect concentrations 3

Duration:

48 h

Dose descriptor:

other: NOEL

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Effect concentrations 4

Duration:

48 h

Dose descriptor:

other: LOEL

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Details on results:

See below

Results with reference substance (positive control):

The percentage immobility for the reference item was determined after 24 hours. The EC50-value with 95 % confidence limits (CI) was calculated by sigmoidal dose-response regression. The EC50-value for the most recent of the monthly performed reference tests was:
EC50: 1.96 mg/L (CI 1.78 - 2.13 mg/L)
The EC50-value of the reference item potassium dichromate after 24 h is within the prescribed concentration range of 0.6 - 2.4 mg/L of quality criteria according to AQS P 9/2 (02/2000) for daphnids clone 5 cultured in Elendt M4 medium. The EC50-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD-Guideline 202.

Any other information on results incl. tables

Biological Data

The percentage immobility, determined in the saturated solution with a nominal loading of 100 mg/L of the test item and the control after 24 and 48 h under static conditions, is presented inthe table below. There was no effect observed in the saturated solution and the control.

 

Table5:   Immobilisation Rates of the definitive Test after 24 and 48 hours of Exposure

             (n = 20, divided into 4 replicates with 5 daphnids each)

Test item Nominal loading of the test item   [mg/L]	IMMOBILISATION [%]
	24 hours					48 hours
	Replicates					Replicates
	1	2	3	4	MV	1	2	3	4	MV
100 (Saturated solution)	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

 

All replicates of the saturated solution of the test item and the control were colourless and visually clear throughout the exposure period.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

In the saturated solution with a nominal loading of 100 mg/L of the test item, no effects were observed on Daphnia magna.

Executive summary:

In the acute immobilisation test with Daphnia magna (STRAUS), the effects of the saturated solution with a nominal loading of 100 mg/L of the test item were determined according to OECD 202 (2004).The limit test was conducted under static conditions over a period of 48 hours .

The test item is a blue powder and sparingly soluble in water (approximately 12 mg/L at 20 °C). Therefore, a saturated solution (100 mg/L) was prepared by stirring with approximately 1100 rpm for 24 hours at room temperature followed by filtration with a membrane filter (0.45 µm, RC,Macherey-Nagel) to remove undissolved test item to achieve the saturated solution.The saturated solution was colourless and visually clear throughout the exposure phase of 48 hours.Twenty daphnids were exposed to the saturated solution and the control.

No analytical monitoring was performed during the test period in agreement with the sponsor,because a measured concentration below the system quantification limit of the analytical method (0.4 mg/L of the test item) was determined during the non GLP preliminary range finding test in the saturated solution. This was also expected for the definitive study. Exposures are expressed in terms of the original concentration of the test item in water at the start of the mixing period (loading rate). The EL-values are presented in the table below. 

Water quality parameters (i.e. pH-value and dissolved oxygen concentration), measured at the start (0 h) and at the end of the exposure (48 h), were within the acceptable limits. The validity criteria of the test guidelines were fulfilled.

 

 

In the saturated solution with a nominal loading of 100 mg/L of the test item ,

no effects were observed on Daphnia magna.

 

 

 

 EL₁₀-, EL₅₀-EL₁₀₀-Values, NOEL and LOEL

 (based on the nominal loading levels of the test item)

Endpoints	Test duration [h]	Test item
		Nominal loading level of the test item [mg/L]
EL10 / 50 / 100	24	> 100
	48	> 100
NOEL	48	100
LOEL	48	> 100