Reaction mass of 6-Chloro-2-{4[ethyl(2-hydroxyethyl)amino]phenyl}-1-methylbenzo[cd]indolium hydroxide and 6-chloro-2-[4-[ethyl(2-hydroxyethyl)amino]phenyl]-1-methylbenz[cd]indolium acetate

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:

Test start date: 24 April, 1990; Test end date: 10 August 1990; Study completion date: 16 November, 1990

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

GLP compliance:

yes

Specific details on test material used for the study:

Identity: FAT 31'064/F
Batch: EN 158496.82/HEW 133/6
Aggregate State: Powder
Appearance in the Test Medium: Violet suspension (determined by RCC)
Solubility: Aqueous: >50 g/L at 20 °C
Stability: Stable for at least 2 hours in aqueous medium
Expiration Date: November, 1994
Storage: At room temperature, in the dark
Safety Precautions: Routine hygienic procedures were sufficient to assure personnel health and safety.

Analytical monitoring:

yes

Details on sampling:

The concentration of the test article at the beginning and at the end of Test set 2 was analysed for the control and for the 0.098, 6.25 and 100 mg/L treated test medium. For this purpose, duplicate 10 mL samples were taken from the test vessels at hour 0 and at hour 48. At the same time, samples of 10 mL and 100 ml were taken from a test vessel containing 100 mg/L but without daphnids (stability test). All samples were taken without stirring as well as the samples of the stability test which showed a precipitation after 48 hours of exposure on the bottom of the test vessel. All samples were stored at -20 °C pending analysis.

Vehicle:

no

Details on test solutions:

For both test sets FAT 31064/F was dissolved in test medium at a concentration of 100 mg/100 mL (stock suspension) by means of ultrasound source. Based on pretest results, series of sequential dilutions with the test medium were prepared to obtain the final test concentrations of the test article which were in the range 100 to 0.098 mg/l (nominal).

Untreated control samples were also tested.
A separate test with the reference compound potassium dichromate was carried out.

Test organisms (species):

Daphnia magna

Details on test organisms:

The tests were run with the species Daphnia magna bred in the Laboratories of RCC under standardized conditions. For the tests, animals with an age of less than 24 hours were used.
The species daphnia magna was selected as an aquatic test organism since the animal is regarded to be a sensitive animal for acute aquatic toxicity testing. The quality of the animals used will be checked by determining the EC 50 value for potassium dichromate twice per year. The method of application and the test system are recognized by the OECD guideline.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

Study terminated after 48 hrs

Test temperature:

20 - 20.5

pH:

8- 9.1

Dissolved oxygen:

8 - 8.5

Nominal and measured concentrations:

0.098, 0.391, 1.563, 6.25, 25, and 100 mg/L

Details on test conditions:

The daphnids were exposed to the test article in 50 mL beakers containing 20 mL test medium with 10 daphnids per beaker. During exposure, the animals were kept at 20 - 20.5 °C for Test set 1 and 2 with 16 hours of illumination (about 500- 2000 Lux). All tests were run in duplicate. The test medium used was reconstituted water prepared according to the EEC Directive.

Reference substance (positive control):

yes

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

4.571 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

EC50 calculated by the Logit-model:

After 24 hours of exposure, the EC50 value was found to be >100 mg/l. The values of EC0 after 48 hours (0.016 mg/l), EC50 after 48 hours (2.751 mg/l) and EC100 after 48 hours (466.196 mg/l) were determined by linear regression of values used for the Logit estimation.
When the EC50 was calculated by the Logit-model, values of EC 50 after 48 hours of 4.571 mg/l were obtained with the following confidence limits:
90 % confidence limit (48 hours): 2.206 - 10.846 mg/l
95 % confidence limit (48 hours): 1.894 - 13.555 mg/l
99 % confidence limit (48 hours): 1.354 - 23.754 mg/l
The dose response curve was found to be:
Logit (48 hours) = -0.55 + 0.360 x log (dose).
The 95 %-confidence limits for the slopes were:
48 hours : 0.347 - 0.373.

EC50 determined by linear regression:

The values of EC 0 after 24 and 48 hours (0.804 mg/1 and 0.623 mg/1), EC 50 after 24 and 48 hours (1.608 mg/1 and 1.247 mg/1) and EC 100 after 24 and 48 hours (3.215 mg/1 and 2.494 mg/1) were determined by linear regression of values used for logit estimation. When the EC 50 was calculated by the Logit-model, values of EC 50 after 24 and 48 hours of 1.625 mg/1 and 1.21 mg/1 were obtained with the following confidence limits:
90 % confidence limit (24 hours): 1.441 - 1.867 mg/l
90 % confidence lmit (48 hours): 0.989 - 1.405 mg/l
95 % confidence lmit (24 hours): 1.407 - 1.931 mg/l
95 % confidence lmit (48 hours): 0.937 - 1.444 mg/l

99 % confidence limit (24 hours): 1.336 - 2.092 mg/l
99 % confidence limit (48 hours): 0.821 - 1.526 mg/l
The dose response curve was found to be:
Logit (24 hours) = -2.60 + 5.358 x log (dose)
Logit (48 hours) = -1.02 + 5.358 x log (dose)

The 95 %-confidence limits for the slopes were:
24 hours : 3.114 - 7.601
48 hours : 3.114 - 7.601

Results with reference substance (positive control):

The EC50 values determined for potassium dichromate after 24 and 48 hours were 1.61 and 1.25 mg/l, respectively.

Reported statistics and error estimates:

Test evaluations were done according to the guideline 202 of the OECD by visual assessment of the mobility of the daphnids after 24 and 48 hours. The EC50 after 48 hours was estimated by using the Logit-model. EC0, EC50 and EC100 values were determined by linear regression of values used for the Logit estimation.

Test set 1: Influence of FAT 31'064/F on the immobilization of Daphnia magna

Concentration of test article (mg/L)	No of animals	No of responses		% of responses
		24 h	48 h	24 h	48 h
0	10	0	0	0	0
0	10	0	0	0	0
0.098	10	0	0	0	0
0.098	10	0	0	0	0
0.391	10	0	6	0	60
0.391	10	1	4	10	40
1.563	10	0	5	0	50
1.563	10	3	3	30	30
6.25	10	0	4	0	40
6.25	10	0	9	0	90
25	10	0	6	0	60
25	10	0	5	0	50
100	10	3	10	30	100
100	10	3	10	30	100

Test set 2: Influence of FAT 31'064/F on the immobilization of Daphnia magna

Concentration of test article (mg/L)	No of animals	No of responses		% of responses
		24 h	48 h	24 h	48 h
0	10	0	0	0	0
0	10	0	1	0	10
0.098	10	0	1	0	10
0.098	10	1	1	10	10
0.391	10	1	2	10	20
0.391	10	0	3	0	30
1.563	10	0	10	0	100
1.563	10	2	10	20	100
6.25	10	2	5	20	50
6.25	10	2	3	20	30
25	10	1	2	10	20
25	10	4	4	40	40
100	10	4	10	30	100
100	10	3	10	30	100

Validity criteria fulfilled:

yes

Conclusions:

FAT 31064/F is considered to be moderately toxic to daphnids when exposed for 48 hours in a static system as recommended by the OECD Guideline.
The following results were obtained:
EC 0 (48 hours):0.016 mg/L
EC 50 (24 hours): >100 mg/L
EC 50 (48 hours): 4.571 mg/L
EC 100 (48 hours): 466.196 mg/L

Executive summary:

FAT 31064/F was tested for its acute toxicity to Daphnia magna in a 48 hours static test. This study was performed in compliance with the OECD Guideline 202, with the OECD Principles of Good Laboratory Practice and the Procedure and Principles of Good Laboratory Practice in Switzerland as well as with the EEC Directive 84/449. In this study, following concentration were tested: 0 (blank), 0.098, 0.391, 1.563, 6.25, 25 and 100 mg/L.. The EC 50 after 24 hours of exposure was found to be >100 mg/L. The EC0 value after 48 hours was determined to be 0.016 mg/L and the EC100 value was found to be 466.196 mg/L after 48 hours of exposure. The EC50 value was estimated to be (Logit-model) 4.571 mg/L. The 95 %-confidence limits for the EC50 value was 1.894 to 13.555 mg/L. The immobilization observed with the control animals after 48 hours was 0 % in Test set 1 and 5 % in Test set 2. The tests were validated by testing potassium dichromate. The EC50 values determined for potassium dichromate after 24 and 48 hours were 1.61 and 1.25 mg/L, respectively. In conclusion, FAT 31064/F is considered to be moderately toxic to daphnids when exposed for 48 hours in a static system as recommended by the OECD Guideline. The following results were obtained:

EC0 (48 hours):0.016 mg/L

EC50 (24 hours): >100 mg/L

EC50 (48 hours): 4.571 mg/L

EC100 (48 hours): 466.196 mg/L

Description of key information

FAT 31064/F was tested for its acute toxicity to Daphnia magna in a 48 hours static test. This study was performed in compliance with the OECD Guideline 202, with the OECD Principles of Good Laboratory Practice and the Procedure and Principles of Good Laboratory Practice in Switzerland as well as with the EEC Directive 84/449.In this study, following concentration were tested: 0 (blank), 0.098, 0.391, 1.563, 6.25, 25 and 100 mg/L.. The EC50 after 24 hours of exposure was found to be >100 mg/L. The EC0 value after 48 hours was determined to be 0.016 mg/L and the EC100 value was found to be 466.196 mg/L after 48 hours of exposure. The EC50 value was estimated to be (Logit-model) 4.571 mg/L. The 95 %-confidence limits for the EC50 value was 1.894 to 13.555 mg/L. The immobilization observed with the control animals after 48 hours was 0 % in Test set 1 and 5 % in Test set 2. The tests were validated by testing potassium dichromate. The EC50 values determined for potassium dichromate after 24 and 48 hours were 1.61 and 1.25 mg/L, respectively. In conclusion, FAT 31064/F is considered to be moderately toxic to daphnids when exposed for 48 hours in a static system as recommended by the OECD Guideline. The following results were obtained:

EC0 (48 hours):0.016 mg/L

EC50 (24 hours): >100 mg/L

EC50 (48 hours): 4.571 mg/L

EC100 (48 hours): 466.196 mg/L

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

4.571 mg/L