2-{3-[(4-amino-2-methylpyrimidin-5-yl)methyl]-4-methyl-1,3-thiazol-3-ium-5-yl}ethyl hydrogen phosphate

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:

12 - 25 Apr 2017

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:

adopted 17 Jul 1992

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

GLP compliance:

yes (incl. QA statement)

Remarks:

Hess. Ministerium für Umwelt, Energie, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany (18 Nov 2014)

Analytical monitoring:

yes

Remarks:

HPLC-UV

Details on sampling:

- Concentrations: Control and 100 mg/L (limit concentration) at test start (0 h) and at test end (96 h).
- Sampling method: One replicate was given to an analytical laboratory for analysis. The second replicate was used as backup.
- Sample storage conditions before analysis: Deep frozen at ≤ - 20 °C. The stability of the test item during freezing was demonstrated in a separate non-GLP study.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A stock solution with 100 mg/L test item in reconstituted water was prepared and stirred for 30 min with a magnetic stirrer.
- Controls: Reconstituted water (untreated test medium)
- Evidence of undissolved material: The test medium was a slightly yellowish preparation and stayed unchanged throughout the study.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebrafish
- Strain: Tübinger Strain
- Source: Max-Planck-Institute, Tübingen, Germany
- Length at study initiation: 2.0 ± 1.0 cm
- Maintenance of the brood fish: Before the start of exposure, the fish were kept in a glass aquarium for at least 12 d in an air-conditioned laboratory at a water temperature of 21 - 25 °C and a 12 h light and 12 h dark regime to ensure similar conditions as in the experiment. Aerated reconstituted water was used as holding medium. The fish were fed daily with Tetra Min diet (Tetra GmbH, Melle, Germany) and frozen artemia. The fish were last fed 24 h before test start.

FEEDING DURING TEST
No feeding during the test.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

223 mg CaCO3/L (reconstituted water)

Test temperature:

21.5 - 22.2 °C

pH:

0 h: 7.91 (control), 7.40 (treatment)
96 h: 7.48 (control), 7.54 (treatment)

Dissolved oxygen:

0 h: 95.6% O2 (control), 96.4% O2 (treatment)
96 h: 88.1% O2 (control), 87.9% O2 (treatment)

Conductivity:

676 µS/cm (reconstituted water)

Nominal and measured concentrations:

Control and 100 mg/L (nominal)
< LOD and 97.88 mg/L (0 h)/ 98.29 mg/L (96 h)

Details on test conditions:

TEST SYSTEM
- Test vessel: All-glass aquariums containing about 10 L test solution.
- Type: Open vessels, covered to allow air exchange.
- Aeration: No aeration during exposure
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: A maximum stocking density of 1 g fish per Liter medium was not exceeded.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water was prepared by dissolving the medium components in fully demineralized water (Milli-Q Water Purification System).
- Acid capacity: 0.8 mmol/L
- Ca/Mg ratio: 4:1
- Na/K ratio: 10:1
- Culture medium different from test medium: Culture medium same as test medium.
- Intervals of water quality measurement: 24 h

OTHER TEST CONDITIONS
- Adjustment of pH: The pH of the test solution was not adjusted.
- Photoperiod: 12 h light and 12 h dark

EFFECT PARAMETERS MEASURED:
- Mortality: 3 h after start of exposure and then daily.
- Clinical symptoms: 3 h after start of exposure and then daily (e.g. equilibrium, swimming behaviour, respiration, pigmentation, etc.).
- Length and Body Weight: At the end of exposure

TEST CONCENTRATIONS
- Range finding study: Yes
- Results used to determine the conditions for the definitive study: Yes. In a static pre-test no toxic effects were observed at 100 mg/L test item in open vessels. In a separate study the stability of the test item in reconstituted water for fish was confirmed over a period of 96 h in open vessels and after deep-freezing.

Reference substance (positive control):

not required

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: No clinical symptoms were observed.
- Observations on body length and weight: Length: 2.6 - 2.7 cm (control), 2.6 - 2.8 cm (treatment); Body Weight: 0.3 - 0.5 g (control), 0.4 - 0.6 g (treatment)
- Mortality of control: The mortality at the end of the study in the control did not exceed the 10% limit.

Sublethal observations / clinical signs:

VALIDITY CRITERIA

 The test fulfilled the validity criteria of the OECD guideline 201 (Table 1).

 

Table 1: Validity criteria of OECD 201.

Criterion from the guideline	Study results	Validity criterion fulfilled
The mortality in the control(s) should not exceed 10% (or one fish if less than ten are used) at the end of the test	Mortality in the control was below 10% at the end of the study.	Yes

 

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Description of key information

LC50 (96 h) > 100 mg/L (nominal, OECD 203, D. rerio)

Additional information

There is one study available, in which the short-term toxicity of 2-{3-[(4-amino-2-methylpyrimidin-5-yl)methyl]-4-methyl-1,3-thiazol-3-ium-5-yl}ethyl hydrogen phosphate (CAS 10023-48-0) to fish was assessed according to OECD guideline 203 and GLP.

In a static limit test, Danio rerio was exposed to a limit concentration of 100 mg/L test item in open test vessels for 96 h. Test item concentrations were analytically verified at the beginning (0 h) and end of the test (96 h) by HPLC-UV.

The measured test item concentrations were 97.9% and 98.3% of the nominal concentration at 0 h and 96 h, respectively, indicating that the substance was stable in reconstituted water for the duration of the test. Therefore, the actual concentrations were within ± 20% of the nominal concentration and the LC50 value was based on the nominal concentration.

After 96 h, neither acute toxicity nor clinical symptoms were observed at the nominal concentration of 100 mg/L. The reported LC50 (96 h) is > 100 mg/L (nominal).