Spiro[piperidine-1,1'-pyrrolidinium] bifluoride

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

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

2008

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

adopted April 13, 2004

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Stability of test concentration/s during exposure: Examined by chemical analysis (DOC) in the freshly prepared media on 2017-12-12 and 2017-12-13 and in the media after 24 hours of exposure on 2017-12-13 and 2017-12-14 according to the semi-static test conditions.

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

PRE-TREATMENT OF TEST ITEM AND PREPARATION OF TEST ITEM CONCENTRATIONS
A stock solution was prepared to give the desired series of test concentrations. 100.3 mg of the test item were added to 1 litre of dilution water on 2017-12-12 and 101.0 mg of the test item were added to 1 litre of dilution water on 2017-12-13 and stirred for 1 h on a magnetic stirrer. The pH was measured to be 6.2 on 2017-12-12 and 6.1 on 2017-12-13.
To produce the different test item concentrations appropriate amounts of the stock solution were diluted with dilution water to a volume of 500 mL. 50 mL of the prepared solutions were taken per replicate and 0.5 mL of dilution water containing 5 daphnids was given to all replicates resulting in the final nominal concentrations. For each test item concentration and the control 4 replicates were prepared.

Test organisms (species):

Daphnia magna

Details on test organisms:

Name: Daphnia magna STRAUS, parthenogenetic females
Source: Strain of Bundesgesundheitsamt Berlin
Maintenance and Acclimatisation: A population of parthenogenetic females of synchronized age structure has been maintained for more than 20 years in the test facility under constant temperature conditions (20 +/- 2 °C) at a 16 : 8 hour light-dark photoperiod (light intensity: < 20 μE x m-2 x s-1). The culture water (so-called 'M4 medium') was partly renewed once a week. The Daphnia were exclusively fed unicellular green algae (Desmodesmus subspicatus) 'ad libitum'. Mortalities of parent Daphnia during the culture period were recorded daily. The neonates were separated from their parent Daphnia by filtration prior to the acute test.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

The hardness of the dilution water used was 14.6 °dH (= 261 mg/L CaCO3).

Test temperature:

18 - 22 °C with a maximum temperature fluctuation of +/- 1 °C in each individual test.

pH:

6.2 - 8.1

Dissolved oxygen:

8.5 - 8.9 mg/L (94 - 101 %)

Nominal and measured concentrations:

nominal: 6.25, 12.5, 25, 50 and 100 mg/L

Details on test conditions:

EXPOSURE CONDITIONS
Test vessels: 100 mL glass beakers covered with watch glasses holding 5 neonates in 50 mL of test medium.
Experimental design: 5 test concentrations plus 1 control; 5 neonates per vessel, 4 replicates per concentration/control; no feeding during the exposure period; semi-static system.
Method of initiation: neonates were placed in prepared media.
Photoperiod: 16 h light : 8 h dark
Temperature of incubation unit: 19.4 to 19.8 °C
Aeration: none
Test item concentration/s: 6.25, 12.5, 25, 50 and 100 mg/L
Method of administration: stock solution (100 mg/L)
Medium renewal: daily
Duration of exposure: 48 hours
Criteria of effects: The criterion of adverse effects used in this study was the item-induced alteration of the normal mobility behaviour and the loss of locomotory actions of the neonates, observed at 24 and 48 hours.

Reference substance (positive control):

no

Duration:

24 h

Dose descriptor:

EC0

Effect conc.:

50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limit: not determined due to mathematical reasons or inappropriate data

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limit: not determined due to mathematical reasons or inappropriate data

Duration:

24 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limit: not determined due to mathematical reasons or inappropriate data

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

12.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limit: 29.5 - 46.8 mg/L

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

37 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limit: 29.5 - 46.8 mg/L

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95 % confidence limit: 29.5 - 46.8 mg/L

Details on results:

The results are expressed in terms of nominal concentrations. The substance has not a definite or unique structure and consists of serveral components. The content of the test item during the exposure period was verified by DOC determination.

Reported statistics and error estimates:

The EC 50 was calculated by probit analysis using the statistics programme ToxRatPro Version 2.10 (released 2010-09-10).

Analysis (DOC)

Nominal concentration	0 h	24 h	0 h	24 h
[mg/L]	2017 -12 -12	2017 -12 -13	2017 -12 -13	2017 -12 -14
control	--	--	--	< 2
6.25	4.1803	4.3940	3.8539	3.7476
12.5	7.8568	7.5472	7.2924	7.1740
25	15.064	13.294	12.729	12.422
50	29.869	27.079	25.468	25.002
100	57.324	53.113	51.046	50.392

Results of the probit analysis

Results of the probit analysis: Selected effective concentrations (ECx) of the test item and their 95%- confidence limits (according to Fieller`s theorem).

Parameter	EC10	EC20	EC50
Value [mg/L]	16.531	21.799	37.006
lower 95%-cl	10.377	15.248	29.498
upper 95%-cl	21.730	27.551	46.833

Slope function after Litchfield and Wilcoxon: 1.875

Validity criteria fulfilled:

yes

Remarks:

The immobilisation and other abnormalities in the controls did not exceed 10 % by the end of the test. The dissolved oxygen concentration remained above 3 mg/L throughout the exposure period.

Conclusions:

The acute toxicity of RFR 6634 to Daphnia magna under semi-static conditions was determined to be EC50 = 37 mg/L after 48 hours.

Executive summary:

A study was performed to assess the acute toxicity of RFR 6634 to Daphnia magna STRAUS under semi-static conditions.

The study was conducted in accordance with the Council Regulation (EC) No 440/2008, Method C.2 ‘Acute toxicity for Daphnia’ (2008) which is equivalent to OECD Guideline for Testing of Chemicals No. 202 'Daphnia sp., Acute Immobilisation Test' (adopted April 13, 2004).

The Daphnia were exposed to a range of concentrations, nominally 6.25, 12.5, 25, 50 and 100 mg/L of RFR 6634 dissolved in dilution water. Auxiliary used to prepare the test media was a magnetic stirrer.

Observations were made on the swimming ability and the immobilisation rate, respectively, after 24 and 48 hours of exposure. The following values were determined:

Time	EC50
[h]	[mg/L]
24	> 100
48	37.0

The results are expressed in terms of nominal concentrations. The substance has not a definite or unique structure and consists of serveral components. The content of the test item during the exposure period was verified by DOC determination.

The hardness of the dilution water used was 14.6 °dH (= 261 mg/L CaCO3).

Description of key information

A study was performed to assess the acute toxicity of RFR 6634 to Daphnia magna STRAUS under semi-static conditions.

The study was conducted in accordance with the Council Regulation (EC) No 440/2008, Method C.2 ‘Acute toxicity for Daphnia’ (2008) which is equivalent to OECD Guideline for Testing of Chemicals No. 202 'Daphnia sp., Acute Immobilisation Test' (adopted April 13, 2004).

The Daphnia were exposed to a range of concentrations, nominally 6.25, 12.5, 25, 50 and 100 mg/L of RFR 6634 dissolved in dilution water. Auxiliary used to prepare the test media was a magnetic stirrer.

Observations were made on the swimming ability and the immobilisation rate, respectively, after 24 and 48 hours of exposure. The following values were determined:

Time	EC50
[h]	[mg/L]
24	> 100
48	37.0

The results are expressed in terms of nominal concentrations. The substance has not a definite or unique structure and consists of serveral components. The content of the test item during the exposure period was verified by DOC determination.

The hardness of the dilution water used was 14.6 °dH (= 261 mg/L CaCO3).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

37 mg/L