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Long-term toxicity to fish

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Endpoint:
fish early-life stage toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
Data from the related substance imazalil base is used to cover this endpoint. The justification for read across is attached in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across source
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
0.299 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Remarks:
imazalil sulfate (recalculated from imazalil concentration)
Endpoint:
adult fish: sub(lethal) effects
Remarks:
prolonged toxicity test
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
OECD Guideline 204 (Fish, Prolonged Toxicity Test: 14-day Study)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: batch BEB191, synthesized in own laboratories (97.6% purity)
- Expiration date of the lot/batch: not indicated
- Purity test date: not indicated

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Because the solubility of the free base was too low to prepare the required stock solutions, a sufficient amount was transformed to the sulphate salt and received the batch number PFA011 (purity 99.5%)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.01, 0.03, 0.10, 0.30, 1 mg/l
- Sampling method: water samples were sampled daily from all aquaria during the test period
- Sample storage conditions before analysis: samples were frozen at -20°C until determination of the test substance concentration
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Imazalil from batch BEB191, synthesized in own laboratories and with a purity of 97.6%, was used. Because the solubitliy of the free base was too low to prepare the required stock solutions, a sufficient amount was transformed to the sulphate salt and received the batch number PFA011. The purity of the latter was 99.5%
- Controls: yes
Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: rainbow trout
- Strain: certified by the supplier to belong to the genus Salmo
- Source: collected from a local hatchery (Wijnant Meussen, Voeren)
- Feeding during test : yes
- Food type: commercial dry fish food
- Amount: 2% of the wet fish weight
- Frequency: daily

ACCLIMATION
- Health during acclimation (any mortality observed): Upon arrival at the test facilities, the fish were free from diseases and malformations. The mortality during the acclimation period recorded after a settling-in period of 48 hours was < 5%, so that the lot was accepted.
Test type:
flow-through
Water media type:
freshwater
Limit test:
no
Total exposure duration:
28 d
Hardness:
174.8 - 179.2 mg/l CaCO3
Test temperature:
12.0 - 15.4 °C
pH:
8.02 - 8.32
Dissolved oxygen:
8.6 - 9.8 mg/l O2
Salinity:
not applicable
Conductivity:
not applicable
Nominal and measured concentrations:
Nominal concentrations: 0.01, 0.03, 0.10, 0.30, 1.00 mg/l
Measured median concentrations: 0.006, 0.021, 0.067, 0.225, 0.759 mg/l
Details on test conditions:
TEST SYSTEM
- Test vessel: aquaria
- Size of vessel: 40l
- Material, size, headspace, fill volume: all-glass aquaria
- Aeration: The aquaria were fitted out with an aeration device and covered with a glass pane to prevent the fish from jumping out.
- Type of flow-through (e.g. peristaltic or proportional diluter): peristaltic pump
- Renewal rate of test solution (frequency/flow rate): 139 ml/min (tap water) and 1 ml/min (concentrated stock solution), leading to an aquaria renewal of 5/day
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Tap water, dechlorinated over activated charcoal, was used as dilution water
- Intervals of water quality measurement: three times weekly

OTHER TEST CONDITIONS
- Photoperiod: 16h light - 8h dark
- Light intensity: fluorescent lamp (Philips 58W/33)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : mortality

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2

Reference substance (positive control):
no
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
> 0.225 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
28 d
Dose descriptor:
LOEC
Effect conc.:
> 0.759 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
28 d
Dose descriptor:
LC50
Effect conc.:
0.668 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- Cumulative mortality at each concentration and for each recommended observation time if possible:
Day: 1 2 4 7 14 21 28
Control: 0 0 0 0 0 0 1/10
0.01 mg/l: 0 0 0 0 1/10 1/10 1/10
0.03 mg/l: 0 0 0 0 0 0 0
0.10 mg/l: 0 0 0 0 0 0 0
0.30 mg/l: 0 0 0 0 0 1/10 1/10
1.00 mg/l: 0 0 0 1/10 2/10 7/10 7/10

- Mortality in the controls:
Up to 27 days, none of the controls died. On day 28, mortality of the controls was 1/10.

- Behavioural observation of the fish:
Possible differences in behaviour from the control group were classified into 4 types: 1) remaining on the bottom with retarded movements, 2) hyper-ventilation, 3) loss of equilibrium, and 4) reduced eating capacity. No difference in behaviour could be observed between the control group and fish exposed to test substance concentrations of 0.01, 0.03 and 0.10 mg/l. Fish exposed to a test substance concentration of 0.30 mg/l swam somwhat slower than the control group but this altered bahaviour was not too distinct. After 5 days of exposure, all fish sbjected to an imazalil concentration of 1 mg/l moved very slowly. Their eating capacity was strongly reduced. Consequently, after an exposure period of 28 days, they were leaner than the control group, which can be derived from the weight results. This explains further the abnormal mortality in the 1 mg/l dosage group, because the general condition of the fish was rather bad. Therefore, at this concentration, the effect of imazalil can be regarded of second order, rather than having a direct impact on the survival.

- Incidents in the course of the test which might have influenced the results:
Up to 0.3 mg imazalil per litre, there was no difference in mortality in comparison with the control group. At a concentration of 1 mg imazalil per litre, 7 fish from the 10 died. However, this mortality might have been caused by other factors than the test substance. In fact, after 15 days of exposure the mortality was not higher than 20%. The high mortality within the following days might have been caused by a sudden rise of the temperature by 3°C. Although, the temperature was not that high to affect the fish, the sudden rise might have been harmful.
Reported statistics and error estimates:
Probit analysis, following the method of Finney, was applied to determine the LC50 after 1, 2, 4, 7, 14, 21 and 28 days of exposure. The homogeneity of the data was tested with chi-square statistics.
Validity criteria fulfilled:
yes
Conclusions:
A chronic study with the rainbow trout (Oncorhynchus mykiss) was performed with the test substance Imazalil according to OECD guideline 204. The NOEC with respect to mortality calculated on the basis of the measured concentrations was the NOEC > 0.225 mg imazalil / L. The results can be considered reliable without restrictions.

Description of key information

The study of Weytjens (1989), investigating the chronic toxicity of Imazalil in the rainbow trout (Oncorhynchus mykiss) according to OECD guideline 204, was considered as the key study for endpoint coverage. The NOEC with respect to mortality was 0.225 mg imazalil / L which corresponds to 0.2995 mg imazalil sulphate/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish
Effect concentration:
0.299 mg/L

Additional information