[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:

From 2018-01-31 to 2018-05-09

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Principles of method if other than guideline:

Not relevant.

GLP compliance:

yes (incl. QA statement)

Remarks:

Signed on 2017-01-10

Specific details on test material used for the study:

None.

Analytical monitoring:

yes

Details on sampling:

Duplicate samples for analysis were taken from the control and the limit test loading rate at the start (t=0h), at t=24h (new and old solutions) and at the end of the test (t=48h). Concentration of Zirconium, a constituent of the test item, was checked in the control medium and the limit test loading rate. Zirconium analysis was not performed in compliance with the OECD GLP principles but in accordance with ISO 17025.

Vehicle:

yes

Details on test solutions:

The study was carried out using WAFs (Water Accommodated Fractions). The WAFs (for fresh media at t=0h and t=24h) were prepared under closed conditions and by slow-stirring.
The mixing vessels were 1-L cylindrical glass bottles sealed with screw caps and fitted with a drain port near the bottom for drawing off the WAFs. A magnetic stirring bar was placed in each mixing vessel and test water was added. The loading rates of the test item were weighed on weighing boats that afterwards were placed above each mixing vessel and rinsed with test water. The mixing vessels were then carefully filled with the remaining volume of test water to obtain 1.1 L of test medium in order to use a maximum volume and to minimise headspace. Then the mixing vessels were closed immediately. The mixing was initiated with the vortex in the centre extending maximally around 10% vessel depth from the top to the bottom of the vessel. After 24 +/- 2 hours of gentle stirring in the dark at room temperature, the WAFs were allowed to stand for at least 1 hour before use. The first 100 mL were discarded via the drain port. Then the WAFs were filtered and added into test vessels that were immediately sealed with screw caps after introduction of daphnids. No small bubble was observed in the test vessels. The test solution in test vessels was observed to be clear and colourless. The test was carried out without adjustment of the pH.

Test organisms (species):

Daphnia magna

Details on test organisms:

Species: Daphnia magna (Straus), clone 5
Sex: Female
Origin: 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.
Reason for selection: Characteristic and common representative of freshwater zooplankton which has been selected as an internationally accepted invertebrate species.
Validity of batch: Daphnids originated from a healthy stock, showing no signs of stress such as mortality, presence of males, ephippia or discoloured animals.
Age at test start: < 24 hours old
Breeding conditions: 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 three times a week. These culture conditions maintained the daphnids in the parthenogenetic reproductive stage.

Test type:

semi-static

Water media type:

freshwater

Remarks:

Reconstituted water (Elendt M4 medium), as prescribed by OECD Guideline 202

Limit test:

yes

Total exposure duration:

48 h

Remarks on exposure duration:

None.

Post exposure observation period:

None.

Hardness:

Approximately 250 mg.L-1 (as CaCO3)

Test temperature:

20.7 - 20.9 °C (average; 20.8°C)

pH:

In the range of 6 to 9,not varying by more than 1.5 units.

Dissolved oxygen:

Oxygen: ≥ 60% of the air-saturation value at the end of the test

Nominal and measured concentrations:

Limit test: 100 mg/L (loading rate)

Details on test conditions:

TEST SYSTEM
- Test vessel: All-glass test tubes of approximately 20 mL capacity sealed with screw caps. Each test vessel will be uniquely identified with study code, replicate number, date of experimentation and concentration.
- Aeration: No aeration of the test solutions occurred throughout the test.
- Renewal rate of test solution: A semi-static test was performed with renewal of test solutions after 24 h (because of the properties of the test item)
- No. of daphnids: 20 per treatment group (including controls), divided into 4 groups of 5 animals
- Loading: 5 daphnids per vessel each completely filled with test solution and without headspace.
- Introduction of Daphnids: Daphnids were introduced into the test vessels each completely filled with test solution and without headspace immediately after filling the test vessels with test solutions.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water as prescribed by the OECD Guideline 202

OTHER TEST CONDITIONS
- Photoperiod: 16 h light : 8 h dark

EFFECT PARAMETERS MEASURED:
- Immobility: Immobility and abnormal behaviour were determined by visual observation after 24 and 48 hours. Immobile animals were eliminated from the vessels as soon as they were discovered. Daphnids were considered to be immobile if they were not able to swim within 15 seconds after gentle agitation of test vessels.
- pH and dissolved O2: At start (t=0h), at t=24h (new and old solutions) and at the end of the test (t=48h) from all test concentrations and control.
- Temperature of Medium: Measured continuously in a temperature controlled vessel next to the test vessels, over the entire study period, beginning at the start of the test.

TEST CONCENTRATIONS
Limit test: 100 mg/L (loading rate)

RANGE-FINDING STUDY
The range-finding test was carried out using WAFs (Water Accommodated Fractions) of the test item over a range of nominal loading rate of 1.0, 3.2, 10.0, 32.0 and 100.0 mg.L-1 and to a control. The WAFs (for fresh media at t=0h and t=24h) were prepared in the dark under closed conditions and by slow-stirring. No effect was observed.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

After 24 and 48 hours of exposure, no immobilisation of the test animals was observed in the control and the loading rate of 100 mg.L-1, confirming the observations of the range-finding test.
Based on these results, the 24 and 48-hour EL50 values were therefore > 100 mg.L-1.

Results with reference substance (positive control):

On March 8, 2018 (KD18-001; most recent test), the 24h-EC50 was 1.31 mg.L-1. Hence, the sensitivity of the clone of Daphnia magna was in agreement with OECD 202 (1) (expected 24h-EC50: 0.6 mg.L-1 to 2.1 mg.L-1) at this time.

Reported statistics and error estimates:

Not relevant, limit test.

Biological results

Acute immobilisation of daphnids after 24 and 48 hours in the final test.

Nominal concentration*(mg test item.L^-1)	Replicate	Number of daphnids exposed	Response at 24h		Response at 48h
			Number	Total %	Number	Total %
Control	1 2 3 4	5 5 5 5	0 0 0 0	0	0 0 0 0	0
100	1 2 3 4	5 5 5 5	0 0 0 0	0	0 0 0 0	0

* WAF prepared at the given loading rate.

Analytical results

Concentration of Zirconium (one of the main constituents of the test item) was checked in the control and the loading rate of 100 mg.L^-1at start (t=0h), at t=24h (new and old solutions) and at the end of the test (t=48h).

Analytical results indicated that very few Zirconium was found throughout the test in the WAFs at 100 mg.L^-1, which was unexpected given the analytical results obtained during the range-finding test, where Zr levels were much higher at the tested loading rates and after 48 hours. It was assumed that Zirconium from the test item sample was oxidised or eventually formed a precipitate since the first opening of the test item flask, and therefore was not easy to detect in this form with the analytical method.

In any case, it was considered that the concentration of the test item at 100mg.L^-1did not vary throughout the study duration as it was renewed at t=24h. Moreover, despite the analytical results, the study was considered as valid given the results fromthe range-finding test which are consistent with the final test outcome, in agreement with the Study Monitor.Therefore, and since the test item was a multi-constituent substance, the results were based onnominalloading rates.

Concentrations of Zirconium (mg.L^-1) in test water -Final test

 

Nominal concentration (mg test item.L^-1)*	Start (t=0h)	t=24old	t=24fresh	End (t=48h)
Control	< 0.01	< 0.01	< 0.01	< 0.01
Control	< 0.01	< 0.01	< 0.01	< 0.01
100	0.04	0.04	0.01	< 0.01
100	0.02	< 0.01	< 0.01	0.01

* WAF prepared at the given loading rate.

Water quality parameters and environmental conditions throughout the test:

pH and oxygen concentrations remained within the limits prescribed by the study plan (pH: 6.0-9.0, not varying by more than 1.5 units; oxygen:≥ 60% of the air-saturation valueat the end of the test), suggesting oxygen concentrations were not influenced by the test item; pH values were however lower in presence of test item, suggesting a slight test item effect on the pH of the test medium.

Furthermore, the temperature of the test medium was situated between 20.7 and 20.9°C throughout the test (average value: 20.8°C), and complied with the requirements as laid down in the study plan (20°C ± 2°C, constant within 1°C).

pH-values during the final test.

		Nominal concentration(mg test item.L^-1)*
		Control	100
Start t=0h		8.1	6.77
t=24h	Old	8.13	6.79
	Fresh	8.2	6.68
End t=48h		8.38	6.8

.

  

Dissolved oxygen concentrations (mg.L^-1) during the final test

 

		Nominal concentration(mg test item.L^-1)*
		Control	100
Start t=0h		8.33	8.27
t=24h	Old	7.96	7.79
	Fresh	8.30	8.18
End t=48h		8.34	8.13

* WAF prepared at the given loading rate.

All these dissolved-oxygen concentrations correspond to values > 60% of the air-saturation value.

Validity criteria of the study

Controls: In the control, no daphnids became immobilised nor trapped at the surface of the water nor showed signs of stress.

Dissolved [0₂]                        Dissolved oxygen concentration at the end of the test was ≥ 60% of the air-saturation value in controls and test vessels. 

Thus the validity criteria have been fulfilled in the present study.

Validity criteria fulfilled:

yes

Conclusions:

Under the experimental conditions and based on nominal loading rates, the 48-hour EL50 was determined to be higher than 100 mg.L-1.

Executive summary:

A study was performed to assess the acute toxicity of the test item SOCOGEL PART I toDaphnia magna. The followed method was designed to be compliant with OECD Guideline for Testing of Chemicals No. 202, “Daphnia sp., Acute Immobilisation Test”, referenced as Method C.2 of Commission Regulation No. 440/2008 (amended byCommission Regulation (EU) 2016/266) and with the “Guidance document on aquatic toxicity testing of difficult substances and mixtures” (OECD No. 23).

A limit test was performed following the results of a range-finding test. Twenty daphnids (four replicates, five daphnids per replicate) were exposed to Water Accommodated Fractions (WAFs) of the test item at a nominal loading rate of 100 mg test item.L^-1and to a control.The immobilitisation of the daphnids was determined in a closed semi-static 48 -hour test by visual observation after 24 and 48 hours. Concentration of Zirconium (one of the main constituents of the test item) was checked in the control and the loading rate of 100 mg.L^-1at start (t=0h), at t=24h (new and old solutions) and at the end of the test (t=48h).

Analytical results indicated that very few Zirconium was found throughout the test in the WAFs at 100 mg.L^-1. However,it was considered that the concentration of the test item at 100mg.L^-1did not vary throughout the study duration as it was renewed at t=24h.Therefore, and since the test item was a multi-constituent substance, the results were based on nominal loading rates. After 24 and 48 hours of exposure, no immobilisation of the test animals was observed in the control and the loading rate of 100 mg.L^-1.

The toxic effect of test item SOCOGEL PART I to the freshwater invertebrate Daphnia magna was investigated in a closed semi-static limit test using Water Accommodated Fractions. Under the experimental conditions and based on nominal loading rates,the 48 -hour EL₅₀was determined to be higher than 100 mg.L^-1. Moreover, the highest nominal loading rate without observed effects value was determined to be at least 100 mg.L^-1

Description of key information

The toxic effect of test item SOCOGEL PART I to the freshwater invertebrate Daphnia magna was investigated in a closed semi-static limit test using Water Accommodated Fractions. Under the experimental conditions and based on nominal loading rates,the 48 -hour EL₅₀was determined to be higher than 100 mg.L^-1. Moreover, the highest nominal loading rate without observed effects value was determined to be at least 100 mg.L^-1.

Additional information

A study was performed to assess the acute toxicity of the test itemSOCOGEL PART I to Daphnia magna. The followed method was designed to be compliant with OECD Guideline for Testing of Chemicals No. 202, “Daphniasp., Acute Immobilisation Test”, referenced as Method C.2 of Commission Regulation No. 440/2008 (amended byCommission Regulation (EU) 2016/266)and with the “Guidance document on aquatic toxicity testing of difficult substances and mixtures” (OECD No. 23).

A limit test was performed following the results of a range-finding test. Twenty daphnids (four replicates, five daphnids per replicate) were exposed toWater Accommodated Fractions (WAFs) of the test item at a nominal loading rate of 100 mg test item.L^-1and to a control.The immobilitisation of the daphnids was determined in a closed semi-static 48 -hour test by visual observation after 24 and 48 hours.Concentration of Zirconium (one of the main constituents of the test item) was checked in the control and the loading rate of 100 mg.L^-1at start (t=0h), at t=24h (new and old solutions) and at the end of the test (t=48h).

Analytical results indicated that very few Zirconium was found throughout the test in the WAFs at 100 mg.L^-1. However,it was considered that the concentration of the test item at 100mg.L^-1did not vary throughout the study duration as it was renewed at t=24h.Therefore, and since the test item was a multi-constituent substance, the results were based onnominalloading rates.After 24 and 48 hours of exposure, no immobilisation of the test animals was observed in the control and the loading rate of 100 mg.L^-1.

The toxic effect of test item SOCOGEL PART Ito the freshwater invertebrate Daphnia magna was investigated in a closed semi-static limit test using Water Accommodated Fractions. Under the experimental conditions andbased on nominal loading rates,the 48 -hour EL₅₀was determined to be higher than 100 mg.L^-1. Moreover, the highest nominal loading rate without observed effects value was determined to be at least 100 mg.L^-1