5-amino-1-[2,6-dichloro-4-(trifluoromethyl)phenyl]-4-(ethylsulfinyl)-1H-pyrazole-3-carbonitrile

Bioaccumulation: aquatic / sediment

Link to relevant study record(s)

Reference

Endpoint:

bioaccumulation in aquatic species: fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 Jan - 14 Feb 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 305 B (Bioaccumulation: Semi-static Fish Test)

Version / remarks:

adopted in 1996

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Ministry of Health, Welfare and Sport, The Netherlands, 22 Apr 2002

Radiolabelling:

yes

Details on sampling:

- Sampling intervals/frequency for test organisms: At each sampling, two fish were collected from the control and four fish from each test concentration. During the uptake phase, fish were collected at test start and after 3, 6, 12, 24, 48, 72, 102, 126 and 144 h. During the depuration phase, fish were collected in 24 h intervals.
- Sampling intervals/frequency for test medium samples: 10 mL of the test solutions were sampled from each test aquarium. During the uptake phase, water samples were taken at test start and after 24, 48, 72, 102, 126 and 144 h. During the depuration phase, water samples were taken every 24 h.
- Sample storage conditions before analysis: Water samples were directly transferred to scintillation flasks for radioanalysis. Fish samples were stored at – 20 °C.
- Details on sampling and analysis of test organisms and test media samples: The whole fish were individually combusted in a Packard sample oxidizer for 2 min. The resulting 14CO2 was trapped in 6 mL of Carbosorb solution and eluted into a scintillation vial for radioanalysis. The 10 mL water samples were mixed with 10 mL of insta-gel XF (Packard) for analysis.

Vehicle:

yes

Details on preparation of test solutions, spiked fish food or sediment:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The stock solution for low concentration was prepared from one primary stock solution of 100 mg of test substance non-radiolabeled per liter of dilution water plus acetone 2%. 50 mL of this solution was transferred and made up to 500 mL with dilution water and then, 43 µL of [14C]-test substance was added to this solution, resulting in a solution with 10 mg of test substance/L and 51,141,328 dpm/mg of radioactivity. Then, 15 mL of this solution was transferred to the test aquarium, resulting in 0.01 mg of test substance/L and 32,537,000 dpm/mg. The average concentration of test solution during the assimilation phase confirmed by radioanalysis was 0.01 mg/L.
The stock solution for high concentration was prepared by transferring 43 µL of [14C]-test substance and 50 mg of active ingredient (non-radiolabeled test substance) in 500 mL of dilution water plus acetone 2 %, resulting in a solution with 100 mg of test substance/L and 5,114,132 dpm/mg of radioactivity. Then, 15 mL of this solution was transferred to the test aquarium, resulting in 0.10 mg of test substance/L and 3,771,460 dpm/mg. The average concentration during the uptake phase confirmed by radioanalysis was 0.11 mg/L.
- Controls: In the LC50 preliminary test, the concentration of 0.2% acetone did not show any effect in fish, so a solvent control in the bioconcentration test was not necessary.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium (stock solution and final test solution(s) at different concentrations and in control(s)): 0.004%

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Source: The fish were acquired from suppliers maintained and acclimated under controlled laboratorial conditions.
- Weight at study initiation (range): 0.321 - 0.933 g (uptake phase), 0.316 - 0.871 g (depuration phase)
- Description of housing/holding area: 250 L tanks
- Feeding during test
- Food type: standard commercial fish food TetraMin with 45% of protein, 1 - 2 h before test solution renewal

ACCLIMATION
- Acclimation period: 72 h
- Acclimation conditions (same as test or not): same as test
- Type and amount of food: standard commercial fish food TetraMin with 45% of protein

Route of exposure:

aqueous

Test type:

semi-static

Water / sediment media type:

natural water: freshwater

Total exposure / uptake duration:

6 d

Total depuration duration:

4 d

Test temperature:

23.4 - 24.1 °C

pH:

7.5 - 7.68

Dissolved oxygen:

7.0 - 7.18 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: aquarium
- Material, size, fill volume: stainless steel, 45 x 25 x 25.5 cm, 15 L
- Aeration: Only the dilution water was aerated.
- Renewal rate of test solution (frequency/flow rate): every 24 h
- No. of organisms per vessel: 80 fish per test aquarium, 40 fish per control aquarium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Water distributed by the Municipal Service of Water and Sewage Treatment was dechlorinated and aerated before use.
- Intervals of water quality measurement: Temperature, dissolved oxygen and pH were recorded at test start and after 24, 48, 72, 102, 126 and 144 h during the uptake phase and at the start and every 24 h during the depuration phase.

OTHER TEST CONDITIONS
- Photoperiod: 16 h light

RANGE-FINDING / PRELIMINARY STUDY
- In order to find the high and low concentrations for the bioconcentration test, a 96-hour toxicity test was carried out in semi-static system according to OECD Methodologies, in order to determine the acute toxicity to Danio rerio.
- Results used to determine the conditions for the definitive study: The 96 h LC50 value of the test substance was 1.06 mg/L.

Type:

BCF

Value:

8.92 dimensionless

Basis:

whole body w.w.

Calculation basis:

steady state

Remarks on result:

other: Conc.in environment / dose:0.01 mg/L

Type:

BCF

Value:

10.24 dimensionless

Basis:

whole body w.w.

Calculation basis:

steady state

Remarks on result:

other: Conc.in environment / dose:0.11 mg/L

Elimination:

yes

Parameter:

other: DT50 for 0.01 mg/L

Depuration time (DT):

31.06 h

Elimination:

yes

Parameter:

other: DT50 for 0.11 mg/L

Depuration time (DT):

35.46 h

Elimination:

yes

Parameter:

other: DT90 for 0.01 mg/L

Depuration time (DT):

103.19 h

Elimination:

yes

Parameter:

other: DT90 for 0.11 mg/L

Depuration time (DT):

117.8 h

Details on kinetic parameters:

- Uptake rate constant (k1): 0.1991 ± 0.0557 (low concentration), 0.2002 ± 0.0768 (high concentration)
- Depuration (loss) rate constant (k2): 0.0223 ± 0.0013 (low concentration), 0.0195 ± 0.0045 (high concentration)
- Computation/data analysis: The parameters were determined by BIOCINE software.

- Mortality of control: 4.5% during the test period

- Test substance accumulation in whole fish after six days of exposure: 0.089 and 1.09 mg/kg for low and high concentrations, respectively.

- Elimination by the end of the depuration period: 88.76% and 83.49% elimination in fish kept at low and high concentrations, respectively.

Table 1: Validity criteria for OECD 305

Criterion from the guideline	Outcome	Validity criterion fulfilled
The temperature variation is less than ± 2 °C	Temperature ranged from 23.4 to 24.1 °C	Yes
The concentration of dissolved oxygen does not fall below 60% saturation	Oxygen ranged from 6.99 to 7.18 mg/L ( ~83% - 85% saturation)	Yes
The concentration of the test substance in the chambers is maintained within ± 20% of the mean of the measured values during the uptake phase	Concentrations ranged from 86% - 143% for the low concentration, and from 66% - 144% for the high concentration of the respective mean measured value	No
The mortality or other adverse effects/disease in both control and treated fish is less than 10% at the end of the test; where the test is extended over several weeks or months, death or other adverse effects in both sets of fish should be less than 5% per month and not exceed 30% in all.	Fish mortality was 4.5%	Yes

Validity criteria fulfilled:

no

Remarks:

See Table 1 in 'any other information on results incl. tables'

Description of key information

BCF = 8.92 - 10.24 for Danio rerio (OECD 305 B)

Additional information

The bioaccumulation characteristics of the substance were investigated in one study according to OECD 305 B and GLP. In a semi-static system, zebrafish (Danio rerio) were exposed to 0.01 mg/L (low concentration) and 0.11 mg/L (high concentration) for 6 days. The exposure period was followed by a depuration phase of 4 days. The depuration half-life (DT50) of the substance was 31 h and 35.5 h for the low and high concentration, respectively. The BCF was determined to be 8.92 for the low and 10.24 for the high concentration.