6-chloro-2-(6-chloro-4-methyl-3-oxobenzo[b]thien-2(3H)-ylidene)-4-methylbenzo[b]thiophene-3(2H)-one

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

According to OECD 202 and GLP

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

The saturated solution (2.00 mg/L test item were weighed out) was prepared with dilution water two days prior to the start of the exposure (0 h).
The test item was dispersed in the dilution water by stirring with approximately 1100 rpm for 48 h 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. Due to the test item
properties centrifugation was not possible. The effects of the saturated solution of the test item were tested in a limit test.

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.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

265 mgCaCO3/L

Test temperature:

18 - 22°C, constant within ± 1°C

pH:

pH at the Start of the Exposure (0 h)
(measured in one additional replicate of the saturated solution and the control)

Nominal loading
of the test item
[mg/L] pH-value
2.00 (Saturated solution) 7.58
Control 7.81



pH at the End of the Exposure (48 h)
(measured in all replicates of the saturated solution and the control)

Nominal loading
of the test item
[mg/L] pH-values
Replicates
1 2 3 4
2.00 (Saturated solution) 7.80 7.80 7.81 7.82
Control 7.65 7.62 7.62 7.61

Dissolved oxygen:

O2-concentration at the Start of the Exposure (0 h)
(measured in one additional replicate of the saturated solution and the control)

Nominal loading
of the test item
[mg/L] DissolvedO2-concentration[mg/L]
2.00 (Saturated solution) 4.74
Control 8.82


O2-concentration at the End of the Exposure (48 h)
(measured in all replicates of the saturated solution and the control)

Nominal loading
of the test item
[mg/L] DissolvedO2-concentration[mg/L]
Replicates
1 2 3 4
2.00 (Saturated solution) 8.21 8.29 8.25 8.35
Control 8.13 8.26 8.18 8.23

Nominal and measured concentrations:

Limit concentration: saturated solution (nominal loading level 2.00 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
- 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]
7.81 8.82 21.5 658 265

- 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
- Test concentration: limit concentration (saturated solution, nominal loading 2.00 mg/L)

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

48 h

Dose descriptor:

NOELR

Effect conc.:

2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Duration:

48 h

Dose descriptor:

LOELR

Effect conc.:

> 2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Duration:

48 h

Dose descriptor:

EL100

Effect conc.:

> 2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

> 2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Duration:

48 h

Dose descriptor:

EL10

Effect conc.:

> 2 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 h. The EC50-value with 95 % confidence interval (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 1.68 - 2.28 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.

The percentage immobility, determined in the saturated solution with a nominal loading of 2.00 mg/L of the test item Pigment Red 181 and the control after 24 and 48 h under static conditions, is presented in the table below. There was no effect observed at the saturated solution and the control.

 

Immobilisation Rates of the definitive Testafter 24 and 48 hours of Exposure

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

Pigment Red181 Nominal loading of the test item   [mg/L]	IMMOBILISATION [%]
	24 h					48 h
	Replicates					Replicates
	1	2	3	4	MV	1	2	3	4	MV
2.00 (Saturated solution)	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

MV   = mean value

 

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

 

Validity criteria fulfilled:

yes

Conclusions:

At the saturated solution with a nominal loading of 2.00 mg/L of the test item Pigment Red 181, 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 2.00 mg/L of the test item Pigment Red 181 were determined according to OECD 202 (2004).The limit test was conducted under static conditionsover a period of 48 hours..

The test item is a red powder and sparingly soluble in water (< 0.02 mg/L at 20 °C). Therefore, a saturated solution was prepared by stirring with approximately 1100 rpm for 48 h at room temperature followed by filtration (membrane filter: 0.45 µm, RC,Macherey-Nagel) to achieve the saturated solution. Twenty daphnids were exposed to the saturated solution and the control.

No analytical monitoring was performed during the test period because the test item has a very low solubility in water

and organic solvents. Therefore, an analytical standard method with a sufficient sensitivity could not be established. Exposures are expressed in terms of the original concentration of the test item Pigment Red181in 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.

 

 

At the saturated solution* with a nominal loading of 2.00 mg/L of the test item Pigment Red 181, 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]	PigmentRed 181
		Nominal loading level of the test item [mg/L]
EL10	24	> 2.00
	48	> 2.00
EL50	24	> 2.00
	48	> 2.00
EL100	24	> 2.00
	48	> 2.00
NOEL	48	2.00
LOEL	48	> 2.00

*)  Saturated solution: The maximum dissolved concentration of the test item that can be achieved under the test conditions in the test medium, acc.

to OECD Series, No. 23 (2000).

Description of key information

One study according OECD Guideline 202 is available. At the saturated solution with a nominal loading of 2.00 mg/L of the test item Pigment Red 181, no effects were observed on Daphnia magna.