3-[[3-[[(2-cyanoethyl)amino]methyl]-3,5,5-trimethylcyclohexyl]amino]propiononitrile

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:

2020-03-10 to 2020-10-08 (Exposure period 2020-03-16 to 2020-03-20)

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP study with analytical test item concentration verification

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

GLP compliance:

yes

Specific details on test material used for the study:

DETAILS OF TEST MATERIAL
- Origin of test item: Mr. D. Zenker, RB/TE, Z470
- lot/batch number of test material: 18315014
- Purity: 97.0 corr. area-%
- Physical state: liquid
- Color: colorless, clear
- Density: 0.9927 g/mL (BASF, see IUCLID Ch. 4.4)
- Vapor pressure: p/hPa = 2.2E-06 (20°C); 4.2E-06 (25°C); 8.3E-05 (50°C) (BASF, see IUCLID Ch. 4.6)
- Water solubility: 32 g/L ± 1 g/L at 20°C ± 0.5 °C (BASF, see IUCLID Ch. 4.8)

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage conditions: ambient (room temperature)
- Storage stability of test material: The stability of the test substance under storage conditions over the test period was guaranteed by the sponsor, and the sponsor holds this responsibility.
- Stability and homogeneity of the test material in the vehicle/solvent under test conditions (e.g. in the exposure medium) and during storage: The stability of the test substance as a solution in test water and under testing conditions was determined by concentration control analysis. The test solutions were visibly homogeneous.

OTHER SPECIFICS
- adjustment of pH: no

Analytical monitoring:

yes

Details on sampling:

- For the first interval, samples were collected within 30 min before insertion of the test organisms (0 h) from all test vessels and after 24 hours from all test vessels with aged test solution. For the second interval, samples were collected from fresh test solutions within 30 min of test solution renewal at 72 h and after 96 hours from all test vessels with aged test solution.
- The samples for analysis were collected with a glass pipette inserted in the middle of the test vessel. The required sample volume (exactly 10 mL, stabilized with 10 μL formic acid) was deposited into glass ampoules.
- Since the measured concentrations of the test substance in the test solutions were within ±20% of the nominal concentrations, the effect concentration can be expressed relative to the nominal concentration for the evaluation of the test substance.

Vehicle:

no

Details on test solutions:

- The test substance was homogenized by shaking the container before use then approx. 1.2 g of the test substance was weighed and the defined volume of test water (10 L) was filled into the test vessel. An aliquot from the test water in the test vessel was taken and mixed with the test substance using a high shear mixer and the mixture was poured into the test vessel and the whole test batch was homogenized again with an ultra turrax.
- Test solution was checked for complete dissolution of the test substance.
- Nominal concentration: 0 mg/L (control), 120 mg/L

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebrafish
- Length: 2.0 cm (range 1.8 cm – 2.1 cm)
- Weight: 0.07 g (range 0.05 g – 0.08 g)
- Age of the animals: approx. 3 months
- Feeding during test: no

HOUSING CONDITIONS
- Light conditions: 16 hours light, 8 hours dark
- pH-value: Generally, 7.5 – 8.5
- TOC: Generally, < 2 mg/L
- Oxygen content: > 80% saturation
- Temperature: 21 - 25°C

- Withdrawal of feed: No feeding from approximately 24 hours before start of exposure
- Mortality during the last week before start of exposure: 0%

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Test temperature:

- Control and Treatment: 22°C,

pH:

- Control: 8.0 - 8.2,
- Treatment: 8.4 - 8.5

Dissolved oxygen:

- Control: 8.6 - 8.8 mg/L,
- Treatment: 8.5 - 8.8

Nominal and measured concentrations:

Nominal: 0 (control), 100 mg/L;
Measured: <5.0 mg/L (control), 97.3 mg/L

Details on test conditions:

- Test groups: 0 (control), 120 mg/L as nominal concentrations
- Test replicates: One aquarium with 7 fish per test group
- Test temperature: 22°C
- Test water: non chlorinated charcoal-filtered drinking water mixed with deionized water, hardness approx. 100 mg/L as CaCO3, pH generally 7.5 - 8.5
- Test vessels: approx. 10 Liter stainless steel aquaria (29 x 21 x 22 cm)
- Test volume: 10 L
- Loading: 7 Fish/ test vessel, max. 0.8 g fish/L (laoding = 0.049 g fish/L)
- Feeding: None
- Photoperiod: 16h light, 8h dark
- Light intensity: approx. 115 - 287 lux
- Oxygen content: >60% ASV (air saturation value)
- Aeration: None
- Adjustment of the pH-value: None
- Renewal of test solution: 24-hour test solution renewal

Reference substance (positive control):

not specified

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 120 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: test concentration analytically verified

Details on results:

No mortality and no additional adverse effects or abnormal behavior were observed in any of the test treatments during the 96 h exposure period.

Sublethal observations / clinical signs:

Table: Analytically determined concentrations of the test substance during exposure

Nominal Concentration [mg/L]				Measured concentration as % of nominal concentration[a]
0				<LOQ				<LOQ						<LOQ						<LOQ
120				108				106						112						105
[a] mean values of two measurements, see analytical report for detailed results

 

Table: Mortality

Nominal concentration [mg/L]	Initial number of fish	Cumulative mortality (numer of dead animals)
		1 hour	6 hours	24 hours	48 hours	72 hours	96 hours
0	7	0	0	0	0	0	0
120	7	0	0	0	0	0	0

 

Table: Effect concentrations (mg/L) w based on nominal concentrations

Effect concentration (mg/L)

Observation time (hours after start of exposure) 1 hour 6 hours 24 hours 48 hours 72 hours 96 hours

LC50 (95% confidence limits)

>120 -

>120 -

>120 -

>120 -

>120 -

>120 -

NOEC

120

120

120

120

120

120

LOEC

>120

>120

>120

>120

>120

>120

- insufficient data for calculation

 

Table: Test temperature

Nominal concentration [mg/L]			Instantaneous temperature in °C during exposure
			Day 0					Day 1										Day 2							Day 3							Day 4
			fresh					fresh							- aged			fresh				- aged			fresh				aged			aged
0			22					22							22			22				22			22				22			22
120			22					22							22			22				22			22				22			22

 

Table: pH

Nominal concentration [mg/L]		pH values during each day of exposure
		Day 0					Day 1										Day 2							Day 3							Day 4
		fresh					fresh							aged			fresh				aged			fresh				aged			aged
0		8.0					8.1							8.2			8.2				8.1			8.2				8.2			8.2
120		8.4					8.4							8.4			8.5				8.4			8.5				8.5			8.5

 

Table: Dissolved oxygen

Nominal concentration [mg/L]		Dissolved oxygen during each day of exposure, mg/L
		Day 0					Day 1										Day 2							Day 3							Day 4
		fresh					fresh							aged			fresh				aged			fresh				aged			aged
0		8.7					8.7							8.6			8.8				8.6			8.8				8.6			8.6
120		8.6					8.7							8.5			8.8				8.6			8.8				8.6			8.5

 

All the following validity criteria according to the guidelines of this study (OECD 203) were met and the experiment is considered valid.
Exposure criteria
- The pH did not vary by more than one unit during the course of the test
- The concentrations of dissolved oxygen were maintained above 60% of the maximum saturation at the test temperature of 22°C (i.e. >5.3 mg/L).
- measured concentrations of the test substance in the test solutions were within ±20% of the nominal concentrations

Control performance criteria
- At least one negative control group was included in the test design.
- ≤ 10 % mortality in the control group

Validity criteria fulfilled:

yes

Description of key information

With high probability acutely not harmful for fish

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

> 120 mg/L

Additional information

The short-term toxicity of modified Isophoronediamine (MIPDA, CAS 93940 -97 -7) to fish was assessed under semi-static conditions in an OECD 203 study using Danio rerio as test organism (BASF, 2020, Report no. 17F0241/13E077).

Based on the mortality of the fish the following effect value was determined: 96-h LC50 > 120 mg/L (based on nominal concentrations, verified by analytical measurement).The pH of the test solutions were not adjusted and all validity criteria according to the OECD guideline were fullfilled.

With high probability the substance is concluded to be acutely not harmful to fish.