2-ethyl-3-hydroxy-4-pyrone

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:

31 October 2016 - 10 January 2017

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)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0 and 100 mg/L
- Sampling method: Water samples were taken from the control and 100 mg/L test vessel at 0 and 72 hours from fresh media and at 24 and 96 hours from old media for quantitative analysis. The samples were stored frozen prior to analysis.
Duplicate samples and samples at 24 (fresh media), 48 (old and fresh media) and 72 hours (old media) were taken and stored frozen for further analysis if necessary.
- Sample storage conditions before analysis: Frozen prior to analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A nominal amount of test item (2500 mg) was dissolved in test water and the volume adjusted to 1 liter to give the 2500 mg/L stock solution. An aliquot (800 mL) of the stock solution was taken and the volume adjusted to 20 liters in order to produce the 100 mg/L test concentration. The stock solution was inverted several times and the test concentration was stirred with a flat bladed mixer for approximately 1 minute to ensure adequate mixing and homogeneity.

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rainbow trout
- Source: Brow Well Fisheries Limited, Hebden, near Skipton, Yorkshire, UK
- Age at study initiation: juvenile
- Length: mean standard length of 5.7 cm (sd = 0.20) at the end of the definitive test.
- Weight: mean weight of 1.44 g (sd = 0.25) at the end of the definitive test.
- Maintenance of the brood fish: Fish were maintained in a glass fiber tank with a "single pass" water renewal system.

ACCLIMATION
- Acclimation period: From 5 December 2016 to 12 December 2016.
- Type and amount of food during acclimation: commercial trout pellets
- Health during acclimation (any mortality observed): no mortalities in the 7 days prior to the start of the test

FEEDING DURING TEST: No

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

approximately 140 mg/L as CaCO3

Test temperature:

14 - 15 °C

pH:

7.6 - 8.2

Dissolved oxygen:

9.3 - 11.1 mg O2/L

Conductivity:

average 392.342 μS/cm at 20°C

Details on test conditions:

TEST SYSTEM
- Test vessel: 25 to 30 liter glass exposure vessels
- Type: The test vessels were then covered to reduce evaporation.
- Fill volume: 20 liters
- Aeration: The test vessels were aerated via narrow bore glass tubes
- Renewal rate of test solution: A semi-static test regime was employed in the test involving a daily renewal of the test preparations to prevent the build up of nitrogenous waste products.
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.50 g bodyweight/liter (static volume)

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Laboratory tap water was dechlorinated by passage through an activated carbon filter (Purite Series 500) and partly softened (Elga Nimbus 1248D Duplex Water Softener)
- Total organic carbon: 1.060 mg/L average (Typical Water Quality Characteristics, WATER SUPPLY ZONE ZDB13 ALVASTON, 01/01/2014 – 31/12/2014)
- Pesticides: 0.012 mg/L average (Typical Water Quality Characteristics, WATER SUPPLY ZONE ZDB13 ALVASTON, 01/01/2014 – 31/12/2014)
- Chlorine: 0.333 mg/L average before de chlorination by passage through an activated carbon filter (Typical Water Quality Characteristics, WATER SUPPLY ZONE ZDB13 ALVASTON, 01/01/2014 – 31/12/2014)
- Culture medium different from test medium: No

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods

EFFECT PARAMETERS MEASURED : Any mortalities and sub-lethal effects of exposure were recorded at 1, 3, 6, 24, 48, 72 and 96 hours after the start of exposure. The criteria of death were taken to be the absence of both respiratory movement and response to physical stimulation.

TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 10 and 100 mg/L.
- Results used to determine the conditions for the definitive study: yes, no mortality was observed in the preliminary test. After 72h exposure to 100 mg/L increased pigmentation was observed in all animals (n=3).

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 85 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

Analysis of the freshly prepared test preparations at 24, 48 and 72 hours showed measured test concentrations to range from 95 to 102 mg/L.
Analysis of the old (or expired) test preparations at 48, 72 and 96 hours showed measured test concentrations to range from 95 to 101 mg/L.
The 0-Hour fresh test preparation and corresponding 24-Hour old (or expired) test solution gave measured concentrations of 50 and 49 mg/L respectively, indicating a potential error in the test solution preparation at 0 hours.
Given the lower measured test concentrations of the 0-Hour fresh test preparation and corresponding 24-Hour old (or expired) test solution it was considered justifiable to base the results on the geometric mean measured test concentration to give a "worst case" analysis of the data. The geometric mean measured concentration was determined for 24-Hour intervals, 0 to 24, 24 to 48, 48 to 72 and 72 to 96 Hours and the arithmetic average calculated of these values.

Other biological observations: A sub-lethal effect of exposure was observed at the test concentration of 85 mg/L. Increased pigmentation was recorded in all seven fish at both 72 and 96-Hour observations.

None of the control fish died or showed signs of stress during the test.
The oxygen concentration at the end of the test was greater than or equal to 60% of ASV (6.1 mg O2/L at 15 °C) in the control and test vessels.

Reported statistics and error estimates:

No statistics applied as no mortality was observed.

Sublethal observations / clinical signs:

Cumulative Mortality Data

Geometric Mean Measured Concentration (mg/L)	Cumulative Mortality (Initial Population = 7)							% Mortality
	1 Hour	3 Hours	6 Hours	24 Hours	48 Hours	72 Hours	96 Hours	96 Hours
Control	0	0	0	0	0	0	0	0
85	0	0	0	0	0	0	0	0

Sub-lethal Effects

Geometric Mean Measured Concentration (mg/L)	Sub-lethal Effects	Time (Hours)
		1	3	6	24	48	72	96
Control	No abnormalities detected	7	7	7	7	7	7	7
85	No abnormalities detected	7	7	7	7	7	0	0
	Increased pigmentation	0	0	0	0	0	7	7

Validity criteria fulfilled:

yes

Remarks:

none of the control fish died or showed signs of stress during the test. Oxygen concentration at the end of the test was greater than or equal to 60% of ASV in control and test substance vessels.

Conclusions:

The 96-h LC50 value is > 85 mg/L in rainbow trout (Oncorhynchus mykiss), based on geometric mean measured concentrations.

Executive summary:

Fish acute toxicity was examined in a limit study in accordance with OECD TG 203 and GLP. In this study seven rainbow trouts (Oncorhynchus mykiss) were exposed to nominal concentrations of 100 mg/L under semi static conditions for 96 hours. The concentrations were analytically verified and geomean measured concentrations were considered worst-case 85% of nominal throughout the test. No mortality was observed during the study, however increased pigmentation was observed at the test concentration of 85 mg/L after 72 and 96 hours for all exposed fish. Based on these data the 96-h LC50 was determined >85 mg/L (based on geomean measured concentrations).

Description of key information

Fish acute toxicity was examined in a limit study in accordance with OECD TG 203 and GLP. In this study seven rainbow trouts (Oncorhynchus mykiss) were exposed to nominal concentrations of 100 mg/L under semi static conditions for 96 hours. The concentrations were analytically verified and geomean measured concentrations were considered worst-case 85% of nominal throughout the test. No mortality was observed during the study, however increased pigmentation was observed at the test concentration of 85 mg/L after 72 and 96 hours for all exposed fish. Based on these data the 96-h LC50 was determined >85 mg/L (based on geomean measured concentrations).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

85 mg/L

Additional information

The 96h LC50 value is > 85 mg/L.