Disodium 2,2'-[(9,10-dihydro-9,10-dioxo-1,4-anthrylene)diimino]bis[3-bromo-5-butyltoluene-4-sulphonate]

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

Study period:

28 August 2018 to 30 August 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Frequency of measurement: At the start of exposure, before and after the renewal, and at the end of exposure.
- Sample for measurement: Another solution sampled separately from the preparation container (at the preparation). The mixed solution taken out with equal volume from the middle layer of the test solution in the test vessels in each test level (before the renewal and at the end of exposure).
- Volume of sample: Approximately 10 mL (all test levels).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- The test sample (0.150 g) and dilution water (1350 mL) were mixed to prepare the nominal concentration of 100 mg/L, and they were stirred with a magnetic stirrer for 30 minutes. Then, it was filtered with a glass fibre filter (GB-140, 0.4 µm pore size, Toyo Roshi) by suction. The filtrate was stirred for 30 minutes at about 20 °C to recover dissolved oxygen concentration decreased at the suction filtration, and then used as the stock solution. Required volumes of the stock solution and dilution water processing similar to control were mixed and stirred in a preparation container to prepare the test solution which was divided into each test vessel.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Strain/clone: Clone A
- Source: The University of Sheffield, UK
- Feeding during test: no

ACCLIMATION
- Young daphnids produced by parents that were cultured in this test facility were used. The parents to obtain young daphnids were bred in the same quality of water (dechlorinated tap water), water temperature (20 ± 1 °C) and photoperiod (16-hour light/8-hour dark) as used in the study. The parent animals used for the test were same lot and their age and survival rate were 21-day old and 95 %, respectively. Chlorella vulgaris of 0.1- 0.2 mgC (Organic carbon content)/day per Daphnia was fed to the parents once a day.
- Selection of young daphnia : Less than 24-hour-old daphnids were selected

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test temperature:

20.1 - 20.2 °C

pH:

8.0 - 8.1

Dissolved oxygen:

7.9 - 8.8 mg/L

Nominal and measured concentrations:

Nominal: 100, 45.5, 20.7, 9.39, 4.27 and 1.94 % solution
Geometric mean measured: 97.3, 42.9, 19.0, 8.72, 3.80, 1.59 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL glass beaker with a transparent plastic lid
- Fill volume: 100 mL/vessel
- Aeration: no
- Renewal rate of test solution: after 24 hours
- 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: Dechlorinated tap water, aerated sufficiently and temperature-controlled, was used. It was confirmed that the concentration of residual chlorine in dilution water was below 0.02 mg/L at the use. The result of chemical characteristics of dilution water met the standard that was provided with the standard operating procedure of the test facility.
- Intervals of water quality measurement: Appearance of the test solution was recorded at the start of exposure and before the renewal (at 24 hours). Dissolved oxygen concentration, pH and temperature were recorded at the start of exposure, before and after the renewal, and at the end of exposure.

OTHER TEST CONDITIONS
- Photoperiod: Room light, 16-hour light/8-hour dark
- Adjustment of pH: no

EFFECT PARAMETERS MEASURED:
- Observations about immobility and symptoms of the test organisms were conducted at 24 and 48 hours after exposure. Daphnids were considered immobile when they were not able to swim within 15 seconds after gentle agitation of the test vessel.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

11.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limits: 9.41-13.9 mg/L

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

72.9 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limits: 56.6-106 mg/L

Details on results:

IMMOBILITY
- The minimum concentration causing 100 % immobility at 48 hours was 42.9 mg/L. Maximum concentration causing no immobility at 48 hours was 3.80 mg/L.
- Immobility in the control during exposure was 0 %, which met the criterion for the validity of the test (i.e. not more than 10 %).

OBSERVED ABNORMAL RESPONSE
- Observed abnormal responses during exposure were immobilisation (including lethargic) and reduced activity.
- In the control, no abnormal response (discolour of body, trapping at the surface of the water and so on), which met the criterion for the validity of the study (i.e. not more than 10 %), was observed during exposure.

OBSERVATION AND MEASUREMENT OF TEST SOLUTION
- Appearance of test solution: The test solutions in the exposure levels were depended on concentration blue and clear at the start and before renewal. That in the control was colourless and clear at the start and before renewal.
- Condition of test solutions: The measured values of dissolved oxygen concentration, pH and temperature of the test solutions during exposure were 7.9-8.8 mg/L, 8.0-8.1 and 20.1-20.2 °C, respectively. The measured values of dissolved oxygen concentration met the criterion for the validity of the test (more than 3 mg/L at the end of exposure).
- Concentration of test material in test solution: The measured concentrations of the test material in the test solutions at the start of exposure and after the renewal (at the preparation) were 1.57-97.5 mg/L, and those before the renewal and at the end of exposure were 1.51-107 mg/L which were 96.4-109 % of the concentrations at the preparation.

EC50
- The 24-hour and 48-hour EC50 values of the test material for Daphnia magna were 72.9 mg/L (95 % confidence limits: 56.6-106 mg/L) and 11.4 mg/L (95 % confidence limits: 9.41-13.9 mg/L), respectively.

Results with reference substance (positive control):

- A 48-hour acute immobilisation test of a reference substance with the test organisms was periodically conducted.
- Period of study: 9 April 2018 to 11 April 2018
- 48-hour EC50: 0.14 mg/L
- This value was within the stipulated range (mean ± 2 S.D.: 0.10-0.35 mg/L, n= 103) to background data in this test facility.

Reported statistics and error estimates:

- The 24-hour EC50 and 48-hour EC50 values were calculated by Probit analysis, and also the 95 % confidence limits and the slope of the concentration-response curve were calculated.
- The EC50 was estimated using computer program (running on Microsoft software "Excel") developed by our test facility.
- The results of this study were estimated based on the geometric mean of the measured concentrations as the test concentration

Table 1: Immobility

Measured Concentration (mg/L)	Vessel	24 Hours		48 Hours
		No. of Immobilised Daphnids/ Total Daphnids	Immobility (%)	No. of Immobilised Daphnids/ Total Daphnids	Immobility (%)
Control	A	0/5	0	0/5	0
	B	0/5		0/5
	C	0/5		0/5
	D	0/5		0/5
1.59	A	0/5	0	0/5	0
	B	0/5		0/5
	C	0/5		0/5
	D	0/5		0/5
3.80	A	0/5	0	0/5	0
	B	0/5		0/5
	C	0/5		0/5
	D	0/5		0/5
8.72	A	0/5	0	2/5	25
	B	0/5		1/5
	C	0/5		1/5
	D	0/5		1/5
19.0	A	0/5	0	5/5	90
	B	0/5		5/5
	C	0/5		4/5
	D	0/5		4/5
42.9	A	3/5	25	5/5	100
	B	2/5		5/5
	C	0/5		5/5
	D	0/5		5/5
97.3	A	4/5	65	5/5	100
	B	3/5		5/5
	C	4/5		5/5
	D	2/5		5/5

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of this study, the 48-hour EC50 was 11.4 mg/L.

Executive summary:

The acute toxicity of the test material to aquatic invertebrates was investigated in accordance with the standardised guideline OECD 202, under GLP conditions.

Daphnia magna were exposed to six test concentrations of the test material of 100, 45.5, 20.7, 9.39, 4.27, 1.94 % as the stock solution content (a geometric series with a factor of 2.2) [97.3, 42.9, 19.0, 8.72, 3.80, 1.59 mg/L as measured concentrations] and a control. A semi-static regime (renewal at 24 hours) was followed for a 48 hour exposure period.

The 24-hour and 48-hour EC50 values of the test material for Daphnia magna were 72.9 mg/L (95 % confidence limits: 56.6-106 mg/L) and 11.4 mg/L (95 % confidence limits: 9.41-13.9 mg/L), respectively.

The measured concentrations of the test material in the test solution were within the range of ± 20 % of that at the start of exposure. The environmental conditions were within the suitable range. Therefore, it is concluded that this study complied with the applied test guideline.

Under the conditions of this study, the 48-hour EC50 was 11.4 mg/L.

Description of key information

Under the conditions of the study, the 48-hour EC50 was 11.4 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

11.4 mg/L

Additional information

The acute toxicity of the test material to aquatic invertebrates was investigated in accordance with the standardised guideline OECD 202, under GLP conditions.

The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).

Daphnia magna were exposed to six test concentrations of the test material of 100, 45.5, 20.7, 9.39, 4.27, 1.94 % as the stock solution content (a geometric series with a factor of 2.2) [97.3, 42.9, 19.0, 8.72, 3.80, 1.59 mg/L as measured concentrations] and a control. A semi-static regime (renewal at 24 hours) was followed for a 48 hour exposure period.

The 24-hour and 48-hour EC50 values of the test material for Daphnia magna were 72.9 mg/L (95 % confidence limits: 56.6-106 mg/L) and 11.4 mg/L (95 % confidence limits: 9.41-13.9 mg/L), respectively.

The measured concentrations of the test material in the test solution were within the range of ± 20 % of that at the start of exposure. The environmental conditions were within the suitable range. Therefore, it is concluded that this study complied with the applied test guideline.

Under the conditions of the study, the 48-hour EC50 was 11.4 mg/L.