1-amino-4-hydroxy-2-(2-phenoxyethoxy)anthraquinone

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:

18 June 2021 to 23 June 2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)

Version / remarks:

2016

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OECD 23 (Guidance Document on Aqueous-phase Aquatic Toxicity Testing of Difficult Test Chemicals)

Version / remarks:

2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

All of the samples were directly analysed by HPLC

Details on sampling:

For determination of the test material concentration, samples were taken from the testing concentration and from the control at the start and end of each water renewal period of the experiment.

Vehicle:

yes

Remarks:

Reconstituted water (ISO Medium)

Details on test solutions:

TEST SOLUTION PREPARATION
As the test material is poorly soluble in deionized water as well in the test medium, for preparation of test solution the water-accommodated fraction (WAF) approach was taken, as described in OECD Guidance Document No. 23. A supersaturated solution (100 mg/L nominal loading) was prepared by suspending an amount of 0.0302 g test material in 302 mL test medium (ISO Medium) for the first renewal period and 0.0303 g test mateiral in 303 mL test medium (ISO Medium) for the second renewal period.
The test solutions were shaken rigorously (~ 350 rpm) for a period of 72 hours to achieve an equilibrated test item concentration for both of the renewal periods. The test solution was then filtrated through a membrane filter (0.45 μm; Thermo Nalgene® membrane) to separate the possible non-dissolved test material. After the formulation procedure the test animals were immediately introduced into the test solution at each water renewal period.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Straus
- Age at study initiation: <24 hours
- Method of breeding: bred at the test facility
- Sex: Female
- Animal health: Apparently healthy animals were used in this test with a known history
- Feeding during test: The Daphnia were fed with concentrated algal suspension of Raphidocelis subcapitata during the holding. Test animals were not fed during the exposure.
- Acclimatisation: Test animals were bred under similar (or the same) conditions as that used during the exposure period (holding water, temperature, background colour etc.), therefore additional acclimatisation before the test was not necessary. Brood daphnids were maintained in dilution water at the test temperature for at least 48 hours prior to the start of the test.

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

The measured hardness of the ISO Medium used in the test was determined to be 245.64 mg/L (as CaCO3).

Test temperature:

The test temperature was in the range of 21.2 – 21.3 °C measured in the test vessels during the test. The additionally measured ambient temperature within the climate chamber was between 20.7 – 21.6 °C.

pH:

7.11 - 8.23

Dissolved oxygen:

7.80 – 8.31 mg/L

Nominal and measured concentrations:

Nominal concentrations: 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 50 mL glass beaker
- Fill volume: 40 mL
- Renewal rate of test solution: Solutions were renewed every 24 hours
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST WATER (DILUTION WATER)
- Reconstituted water (ISO Medium) was used as dilution water in the experiment. Separate stock solutions of individual substances were first prepared in deionised water. The ISO Medium was prepared by adding 25 mL from each of four stock solutions up to one litre deionised water proportionally.
Concentration of the stock solutions:
CaCl2.2H2O - 11.76 g/L
MgSO4.7H2O - 4.93 g/L
NaHCO3 - 2.59 g/L
KCl - 0.23 g/L

The water temperature, dissolved oxygen concentration and pH were measured in each test group at the start (before test solutions had been distributed into the test vessels) and in each test vessel at the end of each water renewal period of the experiment. Additionally, the ambient temperature was measured continuously using a min/max thermometer in the climate chamber.


EFFECT PARAMETERS MEASURED
Daphnia were observed for immobility or mortality by visual observation after 24 and 48 hours of exposure. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile. In addition to immobility, any abnormal behaviour or appearance is reported.


RANGE-FINDING STUDY
- Test concentrations: Nominal concentrations of 0, 10, 50, 100 and 500 mg/L
- Results used to determine the conditions for the definitive study: yes
- Description of range-finding study: In the preliminary range-finding test ten (2 x 5) Daphnids in each test concentration and in the control were exposed for 48 hours in a static test. No immobilisation of the Daphnids was observed at any concentration.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: No toxic effects observed at saturation

Details on results:

ANALYTICAL RESULTS
The concentration of the test material was analysed at the start and at the end of each renewal period of the experiment. The test material was not detected in the untreated control group (i.e. no interfering component was detected in the control samples). The measured test material concentration was below limit of quantification (LOQ) over the test period of 48 hours. Therefore, the biological results are based on the nominal test material concentration and on the half of the LOQ (LOQ = 0.005 mg/L).

BIOLOGICAL RESULTS
No immobilisation was observed either in the control group or in the test material treated group. In addition to immobility, no abnormal behaviour or appearance of test animals was detected. All biological results are based on the nominal test material concentration and on the analytical determined and calculated test material concentration.

Results with reference substance (positive control):

For the evaluation of the quality of the Daphnia clone and the experimental conditions, Potassium dichromate is tested at least twice a year to demonstrate satisfactory test conditions. The most recent 24h EC50 value was determined to be 0.94 mg/L (95% conf. limits: not determined).

Reported statistics and error estimates:

A limit test was performed and no toxic effects were observed, therefore statistical analysis was not necessary. The NOEC and LOEC values of the test item were determined directly from the raw data.

Immobilisation of test animals

Test Group	Replicate	No. of treated animals	No. of immobilised animals
			24 h	48 h
Control	1	5	0	0
	2	5	0	0
	3	5	0	0
	4	5	0	0
Saturated test concentration (0.0025 mg/L measured)	1	5	0	0
	2	5	0	0
	3	5	0	0
	4	5	0	0

 

Concentration of test material in the test solutions for both the 1st and 2nd renewal period

Nominal concentration, mg/L	Measured concentrations, mg/L
	Start	End
100	<LOQ	<LOQ
	<LOQ	<LOQ
	<LOQ	<LOQ
	<LOQ	<LOQ
Mean:	-	-
RSD:	-	-

 

 

Validity of the study

Immobilisation was 0.0 % (< 10 %) in the control group and none of the daphnids showed signs of disease or stress. The dissolved oxygen concentration during the test in control and test vessels was in the range of 7.80 – 8.31 mg/L (more than 3 mg/L in all cases). All validity criteria were within acceptable limits and therefore the study is considered as valid.

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of this study, no effects were observed at saturation and hence the 48-h NOEC was determined to be 0.0025 mg/L (nominal 100 mg/L) and the 48-h LOEC was determined to be > 0.0025 mg/L (nominal > 100 mg/L).

Executive summary:

The acute toxicity of the test material to Daphnia magna was assessed according to OECD Test Guideline 202 and EU Method C.2. and in compliance with GLP in a 48 hour semi-static test. During the study, twenty Daphnia (divided into 4 replicates) were tested each, exposed to the saturated test concentration and in the control. ISO Medium was used as test medium. The immobilisation of the test animals was observed 24 and 48 hours after test start. Environmental conditions were recorded at the start and at the end of each renewal period of the experiment. All measured values remained within the acceptable ranges.
No immobilisation was observed either in the control group or in the test material treated group. In addition to immobility, no abnormal behaviour or appearance of test animals was detected. A limit test was performed and no toxic effects were observed, therefore NOEC and LOEC values of the test material were determined directly from the raw data.

Under the conditions of this study, no effects were observed at saturation and hence the 48-h NOEC was determined to be 0.0025 mg/L (nominal 100 mg/L) and the 48-h LOEC was determined to be > 0.0025 mg/L (nominal > 100 mg/L).

Description of key information

Under the conditions of this study, no effects were observed at saturation and hence the 48-h NOEC was determined to be 0.0025 mg/L (nominal 100 mg/L) and the 48-h LOEC was determined to be > 0.0025 mg/L (nominal > 100 mg/L).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Effect concentration:

> 100 mg/L

Additional information

The acute toxicity of the test material to Daphnia magna was assessed according to OECD Test Guideline 202 and EU Method C.2. and in compliance with GLP in a 48 hour semi-static test. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).

Durinbg ths study, twenty Daphnia (divided into 4 replicates) were tested each, exposed to the saturated test concentration and in the control. ISO Medium was used as test medium. The immobilisation of the test animals was observed 24 and 48 hours after test start. Environmental conditions were recorded at the start and at the end of each renewal period of the experiment. All measured values remained within the acceptable ranges.
No immobilisation was observed either in the control group or in the test material treated group. In addition to immobility, no abnormal behaviour or appearance of test animals was detected. A limit test was performed and no toxic effects were observed, therefore NOEC and LOEC values of the test material were determined directly from the raw data.

Under the conditions of this study, no effects were observed at saturation and hence the 48-h NOEC was determined to be 0.0025 mg/L (nominal 100 mg/L) and the 48-h LOEC was determined to be > 0.0025 mg/L (nominal > 100 mg/L).