1-imino-1H-isoindol-3-amine

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-08-07 to 2017-08-11

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Nominal concentrations: Control and 43.4, 52.1, 62.5, 75.0 and 90.0 mg/L test item nominal
- Sampling method: taken in 24 h intervals from fresh and aged test solutions. For each sampling also a retain sample was taken.
- Sample storage conditions before analysis: 10 mL samples were taken and stabilized with 50 µL acetic acid and analyzed immediately

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: stock solution: weighed necessary amounts of test item into a volumetric flask and added test medium up to the bench mark then homogenized by 2 minutes of ultrasonication

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Source: own stock breeding of the lab, originally purchased from the Karlsruhe Institute of Technology
- Method of breeding: fed with commercial fish flake food (TetraMin) and protein-enriched pellet food (Caviar) at least once per day
- Maintenance of the brood fish: continuously reared in the lab

FEEDING DURING TEST
- Frequency: none

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

250 mg CaCO3/L

Test temperature:

21.9 – 24.4 °C

pH:

7.20 – 8.78

Dissolved oxygen:

68 - 104 % of air saturation

Conductivity:

792 µS/cm

Nominal and measured concentrations:

Nominal: 43.4, 52.1, 62.5, 75.0 and 90.0 mg/L
Measured: 23.3, 28.4, 32.4, 42.1, 42.6 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 2 L glass beaker
- Fill volume: filled with 2 L of test solution
- Aeration: none
- Renewal of test solution: every 24 hours
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: max. loading of 0.41 g fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted test water consisting of analytical grade salts dissolved in purified water with a ratio of Ca:Mg and Na:K of 4:1 and 10:1, respectively

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours daily

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Biological Assessment: effects and mortality assessed at 0, 4, 24, 48, 72 and 96 hours after test start; all fish were euthanized, weighed and measured at the end of test.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 50, 70, 90 mg/L
- Results used to determine the conditions for the definitive study: Mortality [%]: 33% at 70 mg/L, 100% at 100 mg/L.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

25.7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

23.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Mortality: see section 'Any other information on results'
- Clinical signs: No sublethal effects were observed after 4 hours. After 24 hours many fish showed reduced activity and were swimming at the bottom of the test vessel in the two highest test concentrations. Additionally, after 24 hours, some fish showed increased respiration at 42.1 mg/L. After 48 hours, one fish was swimming at the bottom at 42.1 mg/L. After 72 hours, fish were swimming at the surface at 28.4 mg/L and 32.4 mg/L.
- Length and weight: The average weight of the test organisms was 0.0838 ± 0.0443 g; the average length was 19.1 ± 2.8 mm.
- Analytical Results: The measured content of test in fresh test solutions was between 73 % and 92 % of nominal and 26-38 % of nominal in aged test solutions

Reported statistics and error estimates:

The NOEC (mortality) was established based on the highest test concentration at which no mortality above the allowed control mortality was observed. The LC50-values after 4, 24, 48 and 72 h were determined by Logit analysis using linear max. likelihood regression.

Sublethal observations / clinical signs:

Biological results:

% mortality of fish in the main test:

Nominal test item conc. [mg/L]	Control	43.4	52.1	62.5	75.0	90.0
Measured test item conc (mg/L)	Control	23.3	28.4	32.4	42.1	42.6
Time [h]	Mortality [%]
4 h	0	0	0	0	0	0
24 h	0	0	0	0	0	14
48 h	0	0	0	14	57	100
72 h	0	0	29	71	100	100
96 h	0	14	100	100	100	100

 

pre-test

Determined concentrations of test item during the test:

Test item nominal	Sampling	Test item		Geometric mean	Test item concentration measured
[mg/L]		[mg/L]	% of nominal	[%]	[mg/L]
0	0 h fresh	n.d.	-	-
	24 h aged	n.d.	-
	24 h fresh	n.d.	-
	48 h fresh	n.d.	-
	72 h fresh	n.d.	-
	96 h aged	n.d.	-
43.4	0 h fresh	31.6	73	53.7	23.3
	24 h aged	16.7	38
	24 h fresh	36.9	85
	48 h fresh	36.1	83
	72 h fresh	34.5	79
	96 h aged	14.9	34
52.1	0 h fresh	47.8	92	54.6	28.4
	24 h aged	18.8	36
	24 h fresh	41.3	79
	48 h fresh	45.6	88
	72 h fresh	42.5	82
	96 h aged	17.8	34
62.5	0 h fresh	56.9	91	51.8	32.4
	24 h aged	21.1	34
	24 h fresh	48.9	78
	48 h fresh	55.6	89
	72 h fresh	48.9	78
	96 h aged	18.5	30
75.0	0 h fresh	67.3	90	56.1	42.1
	24 h aged	27.9	37
	24 h fresh	59.7	80
	48 h fresh	64.0	85
90.0	0 h	82.1	91	47.3	42.6
	24 h aged	23.4	26
	24 h fresh	73.3	81

Validity criteria fulfilled:

yes

Conclusions:

The LC50 (96 h) of the test item was determined to be 25.7 mg/L (geometric mean of measured concentrations). The corresponding NOEC (mortality) (96 h) was 23.3 mg/L (geometric mean of measured concentrations). No sublethal effects were observed in the control and at 23.3 mg/L, 28.4 mg/L and 32.4 mg/L (geometric mean of measured concentrations) after 96 h.

Executive summary:

The effects of the test item on the mortality of the Zebrafish Danio rerio were determined under worst-case exposure conditions according to OECD 203. The test item was evaluated in a semi-static dose response test with daily water renewal, with five test item (43.4, 52.1, 62.5, 75.0 and 90.0 mg/L) nominal concentrations and one control with untreated test medium. The test concentrations and control contained seven fish. The test duration was 96 hours. Assessments on sublethal effects and mortality were conducted after 0, 4, 24, 48, 72 and 96 hours.

The measured content of test item in fresh test solutions was between 73 % and 92 % of nominal concentration. In the aged solutions, 26-38 % of the nominal concentration was measured. The toxicological endpoints were evaluated using geometric mean of the measured test item concentrations.

The LC50 (96 h) of the test item was determined to be 25.7 mg/L (geometric mean of measured concentrations). The corresponding NOEC (mortality) (96 h) was 23.3 mg/L (geometric mean of measured concentrations). No sublethal effects were observed in the control and at 23.3 mg/L, 28.4 mg/L and 32.4 mg/L  (geometric mean of measured concentrations) after 96 h.

Description of key information

For this endpoint, one short-term toxicity to fish study is available which has been performed according to OECD 203. The toxicity towards Danio rerio was assessed in a semi-static dose response test with daily water renewal and 5 test item concentrations plus control. Fish were exposed for 96h.

The LC50 (96 h) of the test item was determined to be 25.7 mg/L (geometric mean, measured concentration). The corresponding NOEC (mortality) (96 h) was 23.3 mg/L (geometric mean, measured concentration).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

25.7 mg/L

Additional information

For this endpoint only one test is available. The effects of the test item on the mortality of the Zebrafish Danio rerio were determined under worst-case exposure conditions according to OECD 203.

The test item was evaluated in a semi-static dose response test with daily water renewal, with five test item (43.4, 52.1, 62.5, 75.0 and 90.0 mg/L) nominal concentrations and one control with untreated test medium. The test concentrations and control contained seven fish. The test duration was 96 hours. Assessments on sublethal effects and mortality were conducted after 0, 4, 24, 48, 72 and 96 hours.

The LC50 (96 h) of the test item was determined to be 25.7 mg/L (geometric mean, measured concentration). The corresponding NOEC (mortality) (96 h) was 23.3 mg/L (geometric mean, measured concentration). No sublethal effects were observed in the control and at 23.3 mg/L, 28.4 mg/L and 32.4 mg/L (geometric mean, measured concentration) after 96h.