Ethyl enantate

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:

From 2017-03-26 to 2017-04-03

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:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 1.5 mg/L
- Sampling method: The limit concentration and the control are analytically verified from all freshly prepared and aged test media.
- Sample storage conditions before analysis: All samples were stored at 6 ± 2 °C until the start of the analysis. Prepared samples were stored in the autosampler at room temperature until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Stock solution of 50 mg/L in demineralized water. After a stirring period of 24 hours ± 1 hour at room temperature with 600 rpm, an appropriate volume of this stock solution was pipetted into a test vessel filled with an appropriate volume of dilution water. The test media was mixed with an ultraturrax.
- Controls: 7
- Test concentration: 1.50 mg/L

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish
- Strain: Danio rerio
- Source: single brood, see study report for details on source.

ACCLIMATION
- Acclimation period: at least 12 days
- Acclimation conditions: same as test
- Type and amount of food during acclimation: Sera Vipan, 4 % of the fish body weight per feeding day.
- Feeding frequency during acclimation: 3 times per week, not fed 24 h before the test start

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

Total hardness of dilution water: 64 mg/L as CaCO3

Test temperature:

22.7-23°C

pH:

pH at the start of exposure: 7.61-7.72 (1.01 mg/L), 7.51-7.71 (control)
pH at the end of exposure: 7.51-7.71 (1.01 mg/L), 7.41-7.65 (control)

Dissolved oxygen:

Dissolved oxygen of dilution water: 9.97 mg/L
Dissolved oxygen at the start of exposure: 100% (1.01 mg/L), 100% (control)
Dissolved oxygen at the end of exposure: 90-92% (1.01 mg/L), 92-95% (control)

Nominal and measured concentrations:

Nominal: 1.5 mg/L
Measured: 1.01 mg/L (geometric mean)

Details on test conditions:

TEST SYSTEM
- Test vessel: 13 L glass aquaria
- Type (delete if not applicable): closed, covered with a floating lid
- Material, size, headspace, fill volume: Glass aquaria of 13 L, filled with approximately 8 L of dilution water
- Aeration: No aeration
- Renewal rate of test solution (frequency/flow rate): 24 h
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): One replicate
- No. of vessels per vehicle control (replicates): One replicate
- Biomass loading rate: 0.06 g fish per litre test solution

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Tap water of local origin
- Chlorine:filtered on activated charcoal and aerated for at least 24 h to remove chlorine.
- Alkalinity: 1.00 mmol/L
- Intervals of water quality measurement: quarterly

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light / 8 h dark
- Light intensity: 7 - 750 Lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Biological Parameters: observed for visible abnormalities after 2, 24, 48, 72 and 96 h. Measire size and weight of the control fish at the end of the exposure.
- Physical Properties: The pH-value, temperature and oxygen saturation were measured in all test vessels at the start of the exposure and every 24 hours and after renewal of the test media from fresh and old test media. Total hardness was determined at the start of the exposure out of the control. Water temperature was recorded once per hour during the test. Light intensity was measured at the start of the exposure.

TEST CONCENTRATIONS
- Justification for using less concentrations than requested by guideline: Taking into account the aquatic toxicity data of the test item the test was performed without a range-finding test. The threshold concentration (TC) is based on the toxicity data of the test item of an alga toxicity test (ErC50(0-72 h) = 0.440 mg/L). Considering the loss of the substance during the exposition, a nominal concentration of 1.50 mg/L was performed in the limit test.
- Test concentrations: 1.50 mg/L were tested as a threshold concentration in a limit test.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 1.01 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

1.01 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC100

Effect conc.:

> 1.01 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Biological data: No symptoms of toxicity were observed for both test and control group. No mortality occurred.
- Physicochemical data: The environmental conditions (pH-value, temperature, O2 saturation) were within the acceptable limits. The continuous measuring of water temperature was in the valid range of 23 ± 2 °C. The measured light intensity was in the given range of 7 - 750 lux and ranged from 106 to 112 lux (mean value 109 lux).

Sublethal observations / clinical signs:

Stability test (non-GLP):

Exposure time (h)	measured conc (mg/L)	% of nominal concentration
0	0.529	88
24	0.379	63

Observations in the Test Vessels:

Geometric mean measured concentration [mg/L]	Effect	Number of fish effected after different exposure periods [hours]
		2	24	48	72	96
1.01	Normal behaviour	7/7	7/7	7/7	7/7	7/7
Control	Normal behaviour	7/7	7/7	7/7	7/7	7/7

Cumulative Mortality [%] in the Test Vessels:

Geometric mean measured concentration [mg/L]	Cumulative Mortality [%] after different exposure periods [hours]
	2	24	48	72	96
1.01	0	0	0	0	0
Control	0	0	0	0	0

Analytical data:

Nominal concentration (mg/L)	Measured concentration (mg/L)								geometric mean
	0h, fresh	24h, aged	24h, fresh	48h, aged	48h, fresh	72h, aged	72h, fresh	95h, aged
1.5	1.43	0.804	1.22	0.693	1.34	0.735	1.33	0.859	1.01

Validity criteria fulfilled:

yes

Conclusions:

The threshold concentration of the test item (1,5 mg/L nominal, 1.01 mg/L geometric mean) was found to have no effect on the vitality of zebrafish. In conclusion, the LC0 corresponds to the geometric mean measured concentration 1.01 mg/L. The LC50 and the LC100 correspond to the geometric mean measured concentration > 1.01 mg/L. All effect levels are given based on the geometric mean measured concentration of the test item.

Executive summary:

The acute toxicity of the test item to fish (zebrafish) was determined according to the OECD Guideline for Testing of Chemicals No. 203 (1992) and EU Council Regulation No. 440/2008/C.1 (2008).

A semi-static limit test was performed with a threshold concentration of 1.50 mg/L. The test had a duration of 96 hours. 7 test organisms were exposed each for the test concentration and the control. The water quality parameters pH-value, temperature and oxygen saturation, measured after 0, 24, 48, 72 and 96 h, were determined to be within the acceptable limits.

The determination of the test concentration was carried out via SPME-GC-MS with a polydimethylsiloxane (PDMS) fiber. Evaluation was carried out from all freshly prepared media after 0, 24, 48 and 72 h and from the corresponding 24 h aged test media after 24, 48, 72 and 96 h. The measured concentrations in freshly prepared media were in the range of 82 to 96 % of the nominal values. In corresponding 24 h aged test media, the measured concentrations were in the range of 46 to 57 % of the nominal values. Therefore, all effect values were based on the geometric mean measured concentration of the test item.

In conclusion, the LC0 corresponds to the geometric mean measured concentration 1.01 mg/L. The LC50 and the LC100 correspond to the geometric mean measured concentration > 1.01 mg/L.

Description of key information

For this endpoint, there is one acute fish test available performed according to OECD 203, limit test and semi-static test set-up. Fish were exposed for 96h. The LC0 corresponds to the geometric mean measured concentration 1.01 mg/L. The LC50 and the LC100 correspond to the geometric mean measured concentration > 1.01 mg/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

1.01 mg/L

Additional information

For this endpoint only one test is available. In this acute toxicity test with fish (zebrafish), the effects of a threshold concentration of 1.50 mg/L of the test item were determined according to OECD 203 (1992).

The test had a duration of 96 hours. 7 test organisms were exposed each for the test concentration and the control. The water quality parameters pH-value, temperature and oxygen saturation, measured after 0, 24, 48, 72 and 96 h, were determined to be within the acceptable limits.

The determination of the test concentration was carried out via SPME-GC-MS with a polydimethylsiloxane (PDMS) fiber. Evaluation was carried out from all freshly prepared media after 0, 24, 48 and 72 h and from the corresponding 24 h aged test media after 24, 48, 72 and 96 h. The measured concentrations in freshly prepared media were in the range of 82 to 96 % of the nominal values. In corresponding 24 h aged test media, the measured concentrations were in the range of 46 to 57 % of the nominal values. Therefore, all effect values were based on the geometric mean measured concentration of the test item.

In conclusion, the LC0 corresponds to the geometric mean measured concentration 1.01 mg/L. The LC50 and the LC100 correspond to the geometric mean measured concentration > 1.01 mg/L.

As key value for LC50 the lowest concentration from the range was selected: 1.01 mg/L.