(8α,9R,8'''α,9'''R)-1,1'-[(2,3,5,6-tetrafluoro-1,4-phenylene)bis(methylene)]bis(6'-methoxycinchonan-1-ium-9-ol) dibromide

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:

Study Initiation Date: September 02, 2019 Experimental Starting Date: September 09, 2019 Experimental Completion Date: November 26, 2019 Study Completion Date: January 28, 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study according to OECD guideline 202 and under GLP

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Swiss GLP Monitoring Authorities, Date of decission 2018-01-10

Specific details on test material used for the study:

Appearance: Beige solid
Expiration Date: End of February 2021
Storage Conditions: < 30°C

Analytical monitoring:

yes

Details on sampling:

- Sampled concentrations: 4.6, 10, 22, 46, and 100 mg/L
- Sampling method: duplicate samples were taken from each test concentration and the control at
the start and at the end of the test (after 48 hours). For the 48-hour stability samples, the contents
of the respective replicates were combined prior to sampling.
- Sample storage conditions before analysis: all samples were frozen (at -20 ± 5 °C)
- Analysed samples: The concentrations of the test item CA5727A were measured in one of the duplicates for all taken samples.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test medium of the highest nominal concentration of 100 mg/L was prepared by mixing
100.16 mg of the test item into 1000 mL of test water using ultrasonic treatment for
15 minutes and intense stirring for 15 minutes at room temperature.
- Eluate:The highest test concentration of 100 mg/L was further diluted with test water to prepare the test
media with the lower test concentrations. The test medium was prepared just before the introduction of
the daphnids (i.e., start of the test).
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No remarkable observations were made concerning the appearance of the test media.
All test media were clear solutions throughout the entire test duration.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Daphnia magna Straus
- Source: Daphnia Collection of the University of Basel/Switzerland in 2015
- Age of parental stock: 3-4 Weeks. At the start of the test, the organisms used
in the test were 6 to 24 hours old and were not first brood progeny.
- Feeding during test: no


ACCLIMATION
- Acclimation period: no acclimatisation was necessary, since the bred of the daphnia
was under identical conditions to those of the test.
- Type and amount of food during the breeding: Algal suspension of the green algae
Desmodesmus subspicatus supplied by the Collection of Algal Cultures (SAG, Institute for
Plant Physiology, University of Göttingen, 37073 Göttingen / Germany) and cultivated at
IES Ltd Laboratories under standardized conditions or a mixture of this algal suspension and a
commercial fish diet (Tetra Min® Hauptfutter, supplied by TETRA-GmbH, 49324 Melle / Germany).
- Feeding frequency: three times a week
- Health during acclimation (any mortality observed): The daphnia were healthy, no mortality was observed.

Test type:

static

Water media type:

other: ISO Test water (Medium according to the guideline)

Limit test:

no

Total exposure duration:

48 h

Hardness:

2.5 mmol/L = 250 mg/L as CaCO3

Test temperature:

20-21 °C

pH:

7.8

Dissolved oxygen:

at the start of the test at all concentrations and the control: 8.4-8.7 mg/L oxygen
at the end of the test at all concentrations and the control: 8.3-8.7 mg/L oxygen

Salinity:

n.d.

Conductivity:

n.d.

Nominal and measured concentrations:

nominal concentrations (measured concentrations 0/48h): 4.6 (4.6/4.9), 10 (10/11), 22 (22/23), 46 (46/46) and
100 (103/99) mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Beakers
- Type (delete if not applicable): Open (The test vessels were Loosely coverd with glass sheets
to reduce the loss of water by evaporation and to avoid the entry of dust into the solutions.)
- Material, size, headspace, fill volume: glas, 100 mL, open, 50 mL
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): no, static test
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates):4
- No. of vessels per control (replicates):4
- No. of vessels per vehicle control (replicates):no
- Biomass loading rate: 1 Daphnia / 10 mL test media

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO Medium
Ingredients / Properties [mmol/L] / Concentration [mg/L]
CaCl2 × 2H2O / 2.0 / 294
MgSO4 × 7H2O / 0.5 / 123
NaHCO3 / 0.75 / 65
KCl / 0.075 / 5.8
- Alkalinity: 0.8 mmol/L
- Water Hardness: 2.5 mmol/L = 250 mg/L as CaCO3

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod:16-hour light to 8-hour dark cycle with a 30-minute transition period
- Light intensity:16 and 17 µE m-2 s-1

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobility of the daphnids and other non-lethal effects (e.g. abnormal behavior or appearance) after 24 and 48 hours of exposure.

VEHICLE CONTROL PERFORMED: no, not needed


RANGE-FINDING STUDY:
- Test concentrations: 1.0 / 10 / 100 mg/L
- Results used to determine the conditions for the definitive study:
48-hour immobilization: 1.0 mg/L / 0%; 10 mg/L / 20%; 100 mg/L / 100%

Reference substance (positive control):

yes

Remarks:

Potassium dichromate / Separate reference studies under GLP twice a year.

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: no abnormalities in the control and up and including the highest test concentration
100 mg/L.
- Mortality of control: no
- Other adverse effects control: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: 0.19-3.0 mg/L
- Dose-response test: Yes
- ECx: 24-hour EC50: 1.8 mg/L
48-hour EC50: 1.4 mg/L

Reported statistics and error estimates:

No statistics needed.
The 24-hour and 48-hour NOEC were determined directly from the raw data. The
24-hour and 48-hour EC50 of the test item could not be calculated, because none of the responses
exceeded 50%. The EC50 was, therefore, also determined directly from the raw data.

Analytical results

Sampling Day/ Sample Age	Nominal Concentra-tion of Test Item	Measured Concentration  of Test Item	Sample Preparation Factor	Determined Concentration of Test Item	% of Nominal Concentration
[day/hours]	[mg/L]	[µg/L]		[mg/L]	[%]
0/0	Control	<LOD	42	n.a.	n.a.
	4.6	3.62	1260	4.56	99
	10	3.70	2772	10.3	103
	22	3.28	6552	21.5	98
	46	3.60	12852	46.3	101
	100	4.04	25452	103	103
2/48	Control	<LOD	42	n.a.	n.a.
	4.6	3.85	1260	4.85	105
	10	3.88	2772	10.7	107
	22	3.43	6552	22.5	102
	46	3.60	12852	46.3	101
	100	3.89	25452	99.0	99

LOD      = 0.0497 µg test item/L

n.a.        = not applicable

The tabulated values of the samples represent rounded results obtained by calculation using the exact Raw Data.

Biological Results: Effect of CA5727A on the Mobility of Daphnia magna

 

Nominal Test Item Concentration	No. of Daphnids Tested	Immobilized Daphnids after 24 Hours	Immobilized Daphnids after 24 Hours	Immobilized Daphnids after 48 Hours	Immobilized Daphnids after 48 Hours
[mg/L]		No.	[%]	No.	[%]
Control	20	0	0	0	0
4.6	20	0	0	0	0
10	20	0	0	0	0
22	20	0	0	0	0
46	20	0	0	0	0
100	20	0(3F)	0	0	0

F: number of mobile test animals with slower swimming activity compared to the control animals

 

 

Validity criteria fulfilled:

yes

Conclusions:

The substance had not adverse effects on aquatic freshwater invertebrates at a test concentration of 100 mg/L over a test period of 48 hours.

Executive summary:

The acute toxicity of the test item to freshwater invertebrates (Daphnia magna) was determined in a 48‑hour static test according to the OECD Guideline for Testing of Chemicals, No. 202 (2004) and the Commission Regulation (EC) No. 440/2008, Part C.2.

The nominal test item concentrations tested were 4.6, 10, 22, 46 and 100 mg/L. A control (test water without test item) was tested in parallel. The measured concentrations of the test item in the test mediawith the concentrations from 4.6 to 100 mg/L were between 98 and 103% of the nominal values at the start of the test and between 99 and 107 % of nominal at the end of the test. The reported biological results were based on the nominal concentrations of the test item, since the correct dosage and the stability of the test item were confirmed.

The biological test results (based on nominal concentrations) were as follows:

48-hour EC₅₀ >100 mg/L

48-hour NOEC = 100 mg/L

 

Description of key information

48-hour EC50 >100 mg/L (Daphnia magna, OECD TG 202, GLP)