[No public or meaningful name is available]

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:

2020-12-21 - 2021-01-28

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Also conducted according to GLP, and designed according to OECD GD 23.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

April 13, 2004

Qualifier:

according to guideline

Guideline:

ISO 6341 (Water quality - Determination of the Inhibition of the Mobility of Daphnia magna Straus (Cladocera, Crustacea))

Version / remarks:

December 2012

Qualifier:

according to guideline

Guideline:

other: OECD Series on testing and assessment, No.23 “Guidance document on aqueous phase aquatic toxicity testing of difficult test chemicals”,

Version / remarks:

February 2019

GLP compliance:

yes (incl. QA statement)

Remarks:

2018-11-15

Analytical monitoring:

yes

Details on sampling:

Samples for analysis were taken from the control and all test solutions at the start (t=0 h), at t=24 h (new and old solutions) and at the end of the test (t=48 h).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)

- Method: Test item weighed and added to vessel to obtain the stock solution. The solution was mixed for 23 hours and then left undisturbed for at least 1 hour before use. Samples from the stock solution were collected and analyzed to verify the concentration. The stock solution was then diluted with test water to obtain the test concentrations (dilutions based on measured concentration in the stock solution).
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): N/A
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): N/A
- Test concentration separation factor: 1.35
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): Not observed, solutions were clear and colorless. The solutions were also tested for the Tyndall effect (none observed).

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: clone 5
- Justification for species other than prescribed by test guideline: N/A
- Age at study initiation (mean and range, SD): < 24 hours
- Method of breeding: Daphnids were cultured in the Laboratoires des Pyrénées et des Landes under similar temperature and light conditions as used in the test. The cultivation of the parental daphnids was performed in all-glass vessel containing test water. Cultures were maintained at a density of 1 adult daphnid per 25 mL of culture medium. Daphnids were fed at least three times a week with a suspension of algal cells (Pseudokirchneriella subcapitata) up to 0.1-0.2 mg C.Daphnia.-1day.-1. The water was changed twice a week. These culture conditions maintained the daphnids in the parthenogenetic reproductive stage.
- Source: LIEBE - CNRS UMR 7146 - UFR SciFA - Université de Lorraine Campus Bridoux - Bât. IBISE, 8, rue du Général Delestraint - 57070 METZ, bred in the Laboratoires des Pyrénées et des Landes
- Feeding during test: No

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

ca. 250 mg.L-1

Test temperature:

ca. 20 °C

pH:

Start: 7.86-7.88
End: 7.67-7.85

Dissolved oxygen:

Start: 8.73-8.78 mg.L-1
End: 8.55-8.73 mg.L-1

Nominal and measured concentrations:

Nominal: 0, 3.0, 4.1, 5.5, 7.4, and 10.0 mg.L-1

Details on test conditions:

TEST SYSTEM
- Test vessel: 60-mL glass flasks sealed with assembled screw cap with hole and PTFE/silicone septum
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 60 mL, no headspace, glass
- Volume of solution: to the rim
- Aeration: None
- Renewal rate of test solution (frequency/flow rate): Renewed after 24h
- 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: Reconstituted water (Elendt M4 medium), as prescribed in the OECD Guideline 202

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16L:8D

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Immobility and abnormal behavior were determined by visual observation after 24 and 48 hours. Daphnids were considered to be immobile if they were not able to swim within 15 seconds after gentle agitation of test vessels.

RANGE-FINDING STUDY
- Test concentrations: 0, 0.32, 1.00, 3.20, 10.00, 12.00 mg.L-1
- Results used to determine the conditions for the definitive study: 0% immobilization for 0 to 3.20 mg.L-1; 100% immobilizatio for 10 and 12 mg.L-1

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

3.856 mg/L

95% CI:

3.488 - 4.22

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mobility

Results with reference substance (positive control):

24h-EC50 1.365 mg.L-1.
The sensitivity of Daphnia magna was in agreement with the OECD 202 recommendations (24h-EC50 between 0.6 mg.L-1 and 2.1 mg.L-1).

Table 1. Acute immobilization of daphnids after 24 and 48 hours in the final test

Analytically confirmed nominal concentrations (mg test item.L-1)	Replicate	Number of daphnids exposed	Immobilization at 24h		Immobilization at 48h
			Number	Total %	Number	Total %
0 (control)	1	5	0	0	0	0
	2	5	0		0
	3	5	0		0
	4	5	0		0
3	1	5	2	15	2	20
	2	5	0		0
	3	5	1		2
	4	5	0		0
4.1	1	5	5	35	5	45
	2	5	0		0
	3	5	1		2
	4	5	1		2
5.5	1	5	1	10	5	100
	2	5	1		5
	3	5	0		5
	4	5	0		5
7.4	1	5	5	90	5	100
	2	5	5		5
	3	5	4		5
	4	5	4		5
10	1	5	5	100	5	100
	2	5	5		5
	3	5	5		5
	4	5	5		5

Two daphnids at 3 mg.L-1 [Rep. 2] were weakened at t=48 h. Mobile daphnids at 4.1 mg.L-1 were weaker at the end of the test than those from lower concentrations.

 

Table 2. Concentrations of the test item (mg.L-1) in test water - Final test

Nominal concentration (mg.L-1)	Measured concentration (mg.L-1)				Geometric mean analytically confirmed concentrations
	Start (0h)	24h (old)	24h (new)	48h	mg.L-1	%Nominal
Control	<LOD	<LOD	<LOD	<LOD	N/A	N/A
3.0	3.23	2.91	2.94	3.11	3.04	101
4.1	4.08	4.22	4.13	4.28	4.18	102
5.5	5.18	5.43	5.63	5.58	5.45	99
7.4	7.39	6.70	7.25	8.32	7.39	100
10.0	10.18	8.78	-	-	9.45	95

No analysis were performed past 24h for the 10.0 mg.L-1 test concentration as all daphnids were immobilized after 24h.

Validity criteria fulfilled:

yes

Remarks:

In the control, no daphnids became immobilised nor trapped at the surface of the water nor showed signs of stress. Dissolved oxygen concentration at the end of the test was ≥ 3 mg.L-1 in controls and test vessels.

Conclusions:

The toxicity of the test item to the freshwater invertebrate Daphnia magna was investigated in a closed semi-static test (OECD TG 202, OECD GD 23, GLP). Under experimental conditions and based upon analytically confirmed nominal concentrations, the 48-hour EC50 was calculated to be 3.856 mg test item.L-1.

Executive summary:

The toxicity of the test item to Daphnia magna was investigated in a study conducted according to the OECD TG 202 (2004) and in compliance with GLP. The design further included the recommendations of the OECD GD 23 (testing difficult substances).

Following a preliminary range-finding study, twenty daphnids (four replicates, five daphnids per replicate) were exposed to an aqueous solution of the test item over 48 hours at the required nominal test concentrations 3.0, 4.1, 5.5, 7.4 and 10.0 mg.L-1, and to a control. The immobilization of the daphnids was determined in a closed semi-static 48-hour test by visual observation after 24 and 48 hours. Samples taken from the control and all test concentrations were analyzed at the start (t=0 h), at t=24 h (new and old solutions) and at the end of the test (t=48 h) in order to determine if concentrations of the test item were maintained.

The test item levels were found to be stable throughout the test (within ± 20% of the initial and nominal concentrations throughout the test). Thus, the evaluation of the effects on daphnids was based on analytically confirmed nominal concentrations. After 48 hours of exposure, immobilization rates were 20% at 3.0 mg.L-1, 45% at 4.1 mg.L-1 and 100% at 5.5, 7.4 and 10.0 mg.L-1.

Under the experimental conditions of this study, the 48-hour EC50 was calculated to be 3.856 mg.L-1, based upon analytically confirmed nominal concentrations.

Description of key information

The toxicity of the test item to the freshwater invertebrate Daphnia magna was investigated in a closed semi-static test (OECD TG 202, OECD GD 23, GLP). Under experimental conditions and based upon analytically confirmed nominal concentrations, the 48-hour EC50 was calculated to be 3.856 mg test item.L-1.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Effect concentration:

3.856 mg/L

Additional information

The toxicity of the test item to Daphnia magna was investigated in a study conducted according to the OECD TG 202 (2004) and in compliance with GLP. The design further included the recommendations of the OECD GD 23 (testing difficult substances).

Following a preliminary range-finding study, twenty daphnids (four replicates, five daphnids per replicate) were exposed to an aqueous solution of the test item over 48 hours at the required nominal test concentrations 3.0, 4.1, 5.5, 7.4 and 10.0 mg.L-1, and to a control. The immobilization of the daphnids was determined in a closed semi-static 48-hour test by visual observation after 24 and 48 hours. Samples taken from the control and all test concentrations were analyzed at the start (t=0 h), at t=24 h (new and old solutions) and at the end of the test (t=48 h) in order to determine if concentrations of the test item were maintained.

The test item levels were found to be stable throughout the test (within ± 20% of the initial and nominal concentrations throughout the test). Thus, the evaluation of the effects on daphnids was based on analytically confirmed nominal concentrations. After 48 hours of exposure, immobilization rates were 20% at 3.0 mg.L-1, 45% at 4.1 mg.L-1 and 100% at 5.5, 7.4 and 10.0 mg.L-1.

Under the experimental conditions of this study, the 48-hour EC50 was calculated to be 3.856 mg.L-1, based upon analytically confirmed nominal concentrations.