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Short-term toxicity to fish

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Endpoint:
short-term toxicity to fish
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: read-across from a guideline study
Justification for type of information:
The read-across justification is presented in the Endpoint summary Aquatic toxicity, because for fish, Daphnia, and algae the same source substance is used, and the reasoning is included in one document. The accompanying files are also attached there.
Reason / purpose:
read-across source
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
0.805 mg/L
Basis for effect:
mortality
Remarks on result:
other: read-across from Terpinolene
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
November 2012 - April 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
This information is used for read-across to the Terpene hydrocarbon alcohols.
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
yes
Remarks:
Control temperature was slightly lower than 23+/-2°C at initial time point
Qualifier:
according to
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Principles of method if other than guideline:
not applicable
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
PHYSICO-CHEMICAL PROPERTIES
Vapour pressure: 101 Pa (20°C)
Water solubility (column elution method): 4.6 mg/L
Water solubility (slow-stirring method): 7.0 mg/L
Analytical monitoring:
yes
Details on sampling:
Concentrations: single samples for analysis were taken from the control and all test concentrations.

Sampling method: samples were taken at the start (t = 0 h) and every 24 h thereafter in old and new solutions and the control.

Storage: samples were analysed directly after sampling.

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
Pre-experiments (non-GLP) were performed to assess stability of test substance under test conditions:
Stock solutions (x4) were prepared in closed conditions and minimum headspace to avoid loss of test substance through volatilisation from water. A water volume of 10 L was placed in vessel. An excess of substance (5-7 g) was added to the surface of water. The stock solution was prepared by slow-stirring and maintaining the vortex in the centre extending maximally ca. 10% of vessel depth. After 24 ± 2 hours of gentle stirring, the saturated aqueous phase was taken out of the drain port. The first 100 mL were discarded and samples were taken from the following stock solution and chemically analyzed. No evidence of undissolved particle is reported. The test sample appears soluble and stable when mixed with test water at the target concentrations.

Method of preparation for final test: based on the results of a range-finding test, solutions used in the definitive test were prepared by direct addition of the required amounts of test solution to test water.

Controls: test water without test substance but treated in the same way as the test substance solutions.
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
TEST ORGANISM
Common name: Zebrafish
Source: Elevage de la Grande Rivière - B. et C. Romano - La Fond Garel – 69490 SAINT FORGEUX
Length at study initiation: 2.53 cm (range: 2.11 to 2.80 cm) for range-finding test, 2.66 cm (range: 2.34 to 2.93 cm) for final test
Feeding during test: no feeding from 24 h prior to the test and during the total test period.

ACCLIMATION
Acclimation period: at least 12 days
Acclimation conditions: same as test
Type and amount of food: pelleted fish food
Feeding frequency: daily

VALIDITY OF BATCH OF FISH
In the batch of fish which was used for the test, mortality during the seven days prior to the start of the test was less than 5 %.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
not applicable
Hardness:
250 mg/L (CaCO3)
Test temperature:
17.0-21.8°C (measured, whole test period)
pH:
7.48-7.98 (measured, whole test period)
Dissolved oxygen:
7.60-9.37 mg/L (measured, whole test period)
Salinity:
not applicable
Nominal and measured concentrations:
Nominal concentrations: 0.60, 0.80, 1.10, 1.50, 2.00 mg/L
Measured concentrations: 0.48, 0.65, 0.93, 1.12, 1.48 mg/L
Details on test conditions:
TEST SYSTEM
Test vessel: glass aquaria (3.5 L) filled with minimum headspace (approximately 1 cm) and closed by placing a glass plate on the aquarium and sealed with vaseline.
Aeration: no
Renewal rate of test solution (frequency): daily
No. of organisms per vessel: 7
No. of vessels per concentration (replicates): single
No. of vessels per blank control (replicates): single
Biomass loading rate: 1.0 g/L

TEST MEDIUM / WATER PARAMETERS
Source/preparation of dilution water: reconstituted water, as prescribed by the OECD Guideline 203
Conductivity: < 10 µS/cm
Culture medium is not different from test medium.
Measurements: pH, nitrate and nitrite concentration, ammonia concentration, total water hardness and dissolved oxygen were measured once a week. Temperature was recorded daily.

OTHER TEST CONDITIONS
Photoperiod: 16 h light and 8 h dark

EFFECT PARAMETERS MEASURED
Mortality was observed at 3, 6, 24, 48, 72 and 96 h following the start of exposure.

TEST CONCENTRATIONS
Spacing factor for test concentrations: 1.35

Range finding study
Test concentrations: 0.6-1.0-2.0 mg/L
Results used to determine the conditions for the definitive study: mortality rate of 3.3% was observed at 1.0 mg/L and mortality rate of 100% was observed at 2.0 mg/L.

Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
0.805 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: 95% CL: 0.670 - 0.928 mg/L
Details on results:
- Behavioural abnormalities: sublethal effects (lack of balance/swimming on the back) were observed at 0.93-1.12 and 1.48 mg/L measured test concentrations
- Mortality of control: no
- Effect concentrations exceeding solubility of substance in test medium: no
- LC0 = 0.48 mg/L
- LC100 = 1.12 mg/L
Results with reference substance (positive control):
- Results with reference substance were valid.
- LC50: 179 mg/L
Reported statistics and error estimates:
The software ToxRat® Professional was used for the determination of the effective concentrations. The LC50 value was determined by Probit analysis using linear max. likelihood regression.
Validity criteria fulfilled:
yes
Conclusions:
The 96-h LC50 value for fish (D. rerio) is 0.805 mg/L, based on nominal concentrations.
Executive summary:

A fish acute toxicity study was performed according to OECD TG 203 and in compliance with GLP criteria. Following a preliminary range-finding test, groups of seven zebrafish (D. rerio) were exposed to Terpinolene at nominal test concentrations of 0.6, 0.8, 1.1, 1.5 and 2.0 mg/L, in aquaria closed over the study period with a minimum of headspace. The total test period was 96 hours and test solutions were renewed on a daily basis. The mortality of the fish was determined in a semi-static 96-h test by visual observation after 24, 48, 72 and 96 h. Exposure concentrations were measured at the start of the test and every 24 h thereafter in old and new solutions. Concentrations of the test item were not satisfactorily maintained within ± 20 % of the initial concentrations throughout the test. Thus, the evaluation of the effects on fish was based on the geometric mean of the measured concentrations: 0.48, 0.65, 0.93, 1.12 and 1.48 mg/L. Mortalities were recorded at 3, 6, 24, 48, 72 and 96 h.

Sub-lethal effects of lack of balance /swimming on the back were observed in the groups exposed to 0.93, 1.12 and 1.48 mg/L of test substance. Based on mean measured concentrations, the 96-h LC50 of Terpinolene to zebrafish was 0.805 mg/L

Description of key information

The Terpene hydrocarbon alcohols' fish toxicity is based on read-across from the close structural analogue Terpinolene.

Terpinolene and its acute fish toxicity

An acute toxicity study was performed according to OECD TG 203 and in compliance with GLP criteria. Following a preliminary range-finding test, groups of seven zebrafish (D. rerio) were exposed to Terpinolene at nominal test concentrations of 0.6, 0.8, 1.1, 1.5 and 2.0 mg/L, in aquaria closed over the study period with a minimum of headspace. The total test period was 96 hours and test solutions were renewed on a daily basis. The mortality of the fish was determined in a semi-static 96-h test by visual observation after 24, 48, 72 and 96 h. Exposure concentrations were measured at the start of the test and every 24 h thereafter in old and new solutions. Concentrations of the test item were not satisfactorily maintained within ± 20 % of the initial concentrations throughout the test. Thus, the evaluation of the effects on fish was based on the geometric mean of the measured concentrations: 0.48, 0.65, 0.93, 1.12 and 1.48 mg/L. Mortalities were recorded at 3, 6, 24, 48, 72 and 96 h. Sub-lethal effects of lack of balance /swimming on the back were observed in the groups exposed to 0.93, 1.12 and 1.48 mg/L of test substance. Based on mean measured concentrations, the 96-h LC50 of Terpinolene to zebrafish was 0.805 mg/L.  

Key value for chemical safety assessment

LC50 for freshwater fish:
0.805 mg/L

Additional information