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Short-term toxicity to fish

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Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2020-01-17 to 2020-02-20
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:
2019-06-18
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch number of test material: From Sponsor. UW19417870
- Expiration date of the lot/batch: 2022-04-16
- Purity test date: 2019-12-12
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Dry, cool (room temperature)
- Stability under storage conditions: Stable
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The stock solution was prepared by adding 800.0 mg test item to 8 L test medium and stirring for 1 h using a magnetic stirrer at room temperature. This stock solution was used as single test item concentration in the test according to the following scheme. One replicate with seven fish was prepared for the test item treatment as well as the negative control (NC).


FORM AS APPLIED IN THE TEST (if different from that of starting material) liquid
Analytical monitoring:
yes
Details on sampling:
- Concentrations:
- Sampling method: TOC analysis
Vehicle:
yes
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
TEST ORGANISM
- Common name: Zebra Fish
- Source: in-house breeding of Hydrotox GmbH
- Age at study initiation: 2.5 months old juveniles
- Length at study initiation: total mean length of about 21.7 mm

ACCLIMATION
- Acclimation period: seven days
- Acclimation conditions (same as test or not): the fish were kept under test conditions in test medium at 23.0 – 24.1 °C.
- Type and amount of food during acclimation: workdays, they are fed with commercial fish food (TetraMin flake food, Tetra GmbH, Melle, Germany) and two times per week with freshly hatched Artemia salina larvae.
- Feeding frequency during acclimation: daily
- Health during acclimation (any mortality observed): no mortality observed
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Test temperature:
The temperature during the exposure was 23.1 – 24.5 °C in the control and 23.0 – 24.3 °C in
the test item treatment (required: 21 – 25 °C, constant within  1 °C).
pH:
The pH was 7.7 – 8.0 in the control and 8.1 – 8.9 in the test item treatment
Dissolved oxygen:
The dissolved oxygen concentration was 7.9 – 8.3 mg/L in the control and 7.9 – 8.3 mg/L in
the test item treatment (> 60 % of the air saturation).
Conductivity:
The conductivity was 678 – 702 μS/cm in the control and 671 – 715 μS/cm in the test item
treatment
Nominal and measured concentrations:
Nominal concentration 100 mg/mL
Details on test conditions:
TEST SYSTEM
The stock solution was prepared by adding 800.0 mg test item to 8 L test medium and stirring for
1 h using a magnetic stirrer at room temperature. This stock solution was used as single test item
concentration in the test according to the following scheme. One replicate with seven fish was
prepared for the test item treatment as well as the negative control (NC).
- Test vessel: Glass tanks 8 L
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The test was performed with ISO 7346 medium according to EN ISO 7346 (1997), prepared as following:
ISO 1 Calcium chloride dihydrate p.a. CaCl2 × 2 H2O 29.4 g/L
ISO 2 Magnesium sulfate heptahydrate p.a. MgSO4 × 7 H2O 12.33 g/L
ISO 3 Sodium hydrogene carbonate p.a. NaHCO3 6.3 g/L
ISO 4 Potassium chloride p.a. KCl 0.55 g/L

After addition of 300 mL ISO 1, ISO 2, ISO 3 and ISO 4, the medium was filled up to 30 L with ultrapure water, continuously aerated and the pH (7.8) as well as oxygen saturation (99.8 %) was measured.
- Total organic carbon: 99.8 – 100.3 %
OTHER TEST CONDITIONS
- Photoperiod: 14 h light:10 h dark
EFFECT PARAMETERS MEASURED (with observation intervals if applicable): The vitality of the fish was assessed after 24 h, 48 h, 72 h and 96 h exposure
TEST CONCENTRATIONS
A nominal concentration of 100 mg/L test item was used (limit test).


Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
LL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Key result
Duration:
96 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Details on results:
MORTALITY
No fish died until the end of the exposure. Thus, the mortality in the control as well in the test item treatment was 0%.

APPEARENCE AND BEHAVIOR
No Obvious effect on appearance and behavior of the fish in the test item treatment was observed. All test organisms in the control as well as in the test item treatment appeared healthy and lively throughout the exposure.
At the start of the test (0 h), the test item solution in the treatment A showed a white-turbit appearence. From the following day (24 h) on the test item solution in treatment A was clear and whitish deposit was observable on the bottom of the test vessel.

Reported statistics and error estimates:
As no effect on the test organisms was observed, no statistical data evaluation was performed.
Sublethal observations / clinical signs:



























































Nominal test item loading rate [mg/L]Sample Time [h]Mean Measured TOC conc. [mg/L]Mean measure TOC conc., NC-Corrected [mg/L]Recovery rate (C-content test item: 66.12%) [%]Geometric mean recovery rate [%]
NC00.82------
NC480.68------
NC960.29------
100067.1666.34100.3100.0
1004866.6765.9999.8100.0
1009666.3866.0899.9100.0
Validity criteria fulfilled:
yes
Remarks:
-Mortality of the test organisms in the control until the end of the test ≤ 1 fish -DOC in the control and the test item treatment throughout the test ≥ 60 % of the air saturation -Analytical determination of test item concentration
Conclusions:
An acute toxicity study (OECD 203) limit test on the effects of N,N-bis[3-(dimethylamino)propyl]-N',N'-dimethylpropane-1,3-diamine on zebra fish (Danio rerio) was conducted.
The measured TOC concentrations in the test item treatment were 99.8-100.3% of the nominal concentrations.
No fish died until the end of the exposure. Thus, the mortality in the control and in the test item as well treatment was 0%. No obvious effect on appearance and behavior of the fist in the test item treatment was observed. All test organism in the control as well as in the test item treatment appeared healthy and lively throughout the exposure.
The NOEC was reported to be >= 100 mg/L and the LOEC > 100 mg/L.
The validity of the test according to OECD Test Guideline 203 (18 June 2019) was determined by the fulfillment of the validity criteria.

Description of key information

The key study for the acute fish toxicity of the substance was based upon the results of an acute toxicity study (limit test)on the effects of N,N-bis[3-(dimethylamino)propyl]-N',N'-dimethylpropane-1,3-diamine on zebra fish (Danio rerio). The test was conducted in accordance with OECD 203 (18 June 2019).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish
Effect concentration:
100 mg/L

Additional information

An acute toxicity study (OECD 203) limit test on the effects of N,N-bis[3-(dimethylamino)propyl]-N',N'-dimethylpropane-1,3-diamine on zebra fish (Danio rerio) was conducted.


The measured TOC concentrations in the test item treatment were 99.8-100.3% of the nominal concentrations.


No fish died until the end of the exposure. Thus, the mortality in the control and in the test item as well treatment was 0%. No obvious effect on appearance and behavior of the fish in the test item treatment was observed. All test organism in the control as well as in the test item treatment appeared healthy and lively throughout the exposure. Thus, the LC50 (96h) is > 100 mg/L.


The NOEC was reported to be >= 100 mg/L and the LOEC > 100 mg/L.


The validity of the test according to OECD Test Guideline 203 (18 June 2019) was determined by the fulfillment of the validity criteria.


An acute toxicity study on the effects of a N;N;N';N'';N''-Pentamethyldipropylentriamin on zebra fish (Brachdanio rerio HAM. and BUCH.) serves as supporting study for the short-term toxicity to fish endpoint, based upon read-across from a supporting substance (structural analogue or surrogate). Based on nominal concentrations, the 96-h LC50 was reported as ca. 100 mg/L. Mortality occurred first at the concentration of 46.4 mg/L (nominal). Other substance-related effects were the symptoms of apathy, tumbling, and swimming near the water surface. The NOEC was reported as 21.5 mg/L (nominal). Based on the arithmetic mean values of the analytically detected concentrations after 1+96 hours, the LC50 was reported at about 92.5 mg/L. The pH adjustment of a parallel group with the highest nominal concentration (464 mg/L) did not significantly reduce mortality and toxic signs.


A further supporting study on the acute fish toxicity of the substance was conducted as part of a fish bioaccumulation study, which evaluated the effects of the substance on himedaka (Oryzias latipes); this study resulted in a 48-h TLm (median lethal dose) of 430 mg/L. The 96-h test on zebra fish is considered to be more reliable test; the LC50 value is lower than the TLm value reported from the 48-h test on himedaka and is used as the key value for the LC50.