[3aR-(3aα,5aβ,9aα,9bβ)]decahydro-3a,6,6,9a-tetramethylnaphth[2,1-b]furan-2(1H)-one

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:

2018-07-22 to 2019-08-02

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 on July 17, 1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

300 mL of test samples from each group was drawn at 0 and 24 h of every renewal. The representative samples were divided into two equal portions (150 mL). One portion (150 mL) was sent for test concentration analysis and the second portion (150 mL) was stored at -20 ± 5 ºC temperature till the study completion.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
A quantity of 45 mg sclareolide was dissolved in RO water and made up to 5000 mL with RO water to obtain the test concentration of 0.009 mg/mL for each stock A, B and C.
Volumes of 1111.2 mL from the stock C for (G2), 1666.7 mL from the stock C for (G3), 777.7 mL and 1666.8 mL from the stock B and C (G4) , respectively, 3666.7 mL from the stock B (G5), 5000 mL from the stock A and 555.6 mL from the stock B (G6), respectively were taken and diluted to 10 L with RO water in fish tank to obtain the nominal test concentration of 1.0 (G2), 1.5 (G3), 2.2 (G4), 3.3(G5) and 5.0 (G6) mg sclareolide/L, respectively.

The test solutions were thoroughly mixed to achieve uniform distribution of the test item within the aquaria. The test medium in the aquaria was changed every 24 h and replaced with the freshly prepared test item solution. At each renewal, approximately 100% volume of the tank was renewed.

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: Rainbow trout
- Source: Department of Fisheries, Government of Jammu and Kashmir, India
- Length at study initiation: Fish length maintained between 4.0 and 5.0 cm.

ACCLIMATION
- Acclimation period: 7 days
- Acclimation conditions: 14.6 - 15.2°C water temperature, 7.25 - 7.65 pH, 88.3 - 96.3% dissolved oxygen and total hardness of 84.0 mg/L as CaCO3.
- Fish length: maintained between 4.0 and 5.0 cm
- Health during acclimation (any mortality observed): No signs of physical or behavioural abnormalities or mortalities of fish.

FEEDING DURING TEST
- Food type: Trout starter feed
- Frequency: daily
- Feed was withheld 24 h before the exposure to sclareolide and throughout the exposure period.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

N/A

Hardness:

78.4 - 89.6 mg/L as CaCO3

Test temperature:

14.0 - 15.2 °C

pH:

6.65 - 7.31

Dissolved oxygen:

84.0 - 96.8%

Salinity:

N/A

Conductivity:

not reported

Nominal and measured concentrations:

A preliminary range finding study was conducted with the test concentrations of 0.0 (control), 0.01, 0.1, 1.0, 5.0, and 9.0 mg sclareolide/L for a period of 96 h with ten fish in each group to determine the concentrations to be selected for the main study. The cumulative percent mortalities at 96 h observed were 0, 0, 0, 0, 100, and 100% at 0.0 (control), 0.01, 0.1, 1.0, 5.0, and 9.0 mg sclareolide/L, respectively.

Based on the result of preliminary range finding study, the test item concentrations of control (0.0), 1.0, 1.5, 2.2, 3.3, and 5.0 mg sclareolide/L along with the control was selected with geometric factor of 1.5 for main study.

Main study:
Nominal concentrations: 0 (control), 1, 1.5, 2.2, 3.3, and 5.0 mg /L
Measured concentrations: 0 (control), 0.9, 1.4, 2.0, 3.1 and 4.9 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: All glass aquaria
- Vessel parameters: The volume of water used was 10 liters in a glass aquarium of 15 liters capacity for control and treated groups.
- Renewal rate of test solution (frequency/flow rate): Test solutions were changed at every 24 h with freshly prepared solutions. At each renewal 100% of volume of the tank was renewed. During renewal, fish were transferred by netting to new test vessels. The test solutions were not aerated during the exposure.
- No. of organisms per vessel: 10 fish per test concentration. A maximum loading of 1.0 g fish per liter of water was used.

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

EFFECT PARAMETERS MEASURED: Behavioural responses, clinical signs and mortality.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

1.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

2.3 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.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: mortality and behaviour

Duration:

96 h

Dose descriptor:

LOEC

Effect conc.:

1.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: mortality and behaviour

Details on results:

MORTALITY
- The cumulative percent mortality at 96 h observed was 0, 40, 60, 100, and 100 at 1.0, 1.5, 2.2, 3.3, and 5.0 mg/L, respectively. No mortality was observed in fish in the control group.

BEHAVIOURAL ABNORMALITIES
- Loss of equilibrium at 5.0 mg/L
- Lethargy at 1.5, 2.2, 3.3 mg/L.
- No behavioural symptoms in control and at 1 mg/L

Reported statistics and error estimates:

The 24 and 96 h LC50 with 95% confidence limits were calculated following the probit analysis method (Finney, 1971) using in-house developed validated software. NOEC and LOEC were determined and reported.

Validity criteria fulfilled:

yes

Conclusions:

The No Observed Effect Concentration (NOEC) and the Lowest Observed Effect Concentration (LOEC) of test item was 1.0 and 1.5 mg/L, respectively. The 24 and 96 h LC50 of sclareolide was 2.3 and 1.8mg/L, respectively.

Executive summary:

A GLP compliant fish acute toxicity test was conducted on sclareolide according to OECD 203 guideline. Based on results of the preliminary range finding study, 6 rainbow trout’s (Oncorhynchus mykiss) were exposed to the test item under semi-static (being renewed every 24 h) conditions at the dose concentrations of 0 (control), 1.0, 1.5, 2.2, 3.3, and 5.0 mg/L for a period of 96 h. The stability test of sclareolide in test media was performed and was stable up to 24 h (>80% of nominal concentration). Test media was analyzed for active ingredient at 0 and 24 h, during the main study. The active ingredient concentration of sclareolide in test media was within acceptable limit (>80% of nominal concentration). The measured active ingredient concentrations at 24 h interval was 0, 0.9, 1.4, 2.0, 3.1 and 4.9 mg/L. The test conditions passed all the validity criteria. The highest concentration of thetest itemcausing no mortality or behavioral symptom, and the lowest concentration tested causing 100 percent mortality, within the 96 h test period was 1.0 and 3.3 mg/L, respectively. The NOEC and the LOEC of test item was 1.0 and 1.5 mg/L, respectively. The 24 and 96 h LC₅₀ of sclareolidewas 2.3 and 1.8mg/L,respectively.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Remarks:

96-hr

Effect concentration:

1.8 mg/L