2-ethyl-3-hydroxy-4-pyrone

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

5 September 2000 - 22 October 2000

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

analytics was done by TOC measurement (not substance specific method). No CoA included.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

1984

Deviations:

yes

Remarks:

There was no analytical test performed to determine the concentration of test substance directly. TOC values were determined and used to calculate the recovery of test substance.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

TOC values were determined and recalculated to test substance concentrations (via purity and % carbon content).

Details on sampling:

TOC-analyses were performed at t=0h and t=48h.
pH was measured at the beginning of the test and after 48h

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method:
Suspensions of the test substance were prepared in dilution water. For this purpose, the test substance was weighed into the glass vessels with the respective amount of water used for the preparation of the aqueous extracts. The test substance was introduced into the dilution water whilst shaking on a shaking machine (ca. 130 rpm) for 24h at room temperature. lt is assumed that during that time equilibration between the test substance and the water was achieved. Thereafter, the suspensions were filtered through filier paper which was previously rinsed thoroughly with ultrapure water (Seral, „Purelab Plus") in order to eliminate possible soluble impurities from the filier material. The filtered extracts were used directly for the test without any further dilution steps. Prior to using in the test the pH of the aqueous extracts was measured; pH was not adjusted. In the same way a Blank without any test substance was prepared.

As the test substance exhibits partial adsorptive properties the aqueous extracts of the test substance were transferred into the respective test vessels, and after approx. 1 hour, the aqueous extracts were replaced by a second pari of the aqueous extract. By that measure the surfaces of the glass walls were saturated with the test substance, and losses due to adsorption were avoided. After that measure the test animals were introduced into the test solutions and the incubation was started.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain/clone: STRAUS
- Source: laboratory bred (originally derived from Dr. Knie, LWA Düsseldorf)
- Age: not older than 24h.
- Feeding during test: No
- Food type: unicellular green alga Scenedesmus subspicatus according to SOP 5001 of Institut Fresenius Taunusstein.

Test type:

static

Water media type:

freshwater

Remarks:

closed system

Limit test:

no

Total exposure duration:

48 h

Test temperature:

19.4 - 20.0 °C

pH:

t= 0h: 7.11 - 7.37
t= 48h: 7.04 - 7.25

Dissolved oxygen:

t= 0h: 8.86 - 9.65 mgO2/L
t =48h: 8.59 - 9.01 mgO2/L

Nominal and measured concentrations:

Nominal concentrations: 0, 10, 20, 40, 80, 160 mg/L
Calculated concentrations (t=0): 0, 11.4, 20.4, 40.6, 76.6, 157.6 mg/L.
Calculated concentrations (t=48): 0, 11.3, 20.0, 39.5, 75.9, 152.3 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL-carboys made of glass.
- Type (delete if not applicable): closed with the glass-stoppers because the test substance contains volatile components.
- Fill volume: 100 mL
- Aeration: No, the oxygen content of the test solutions was sufficient for the whole incubation period.
- Renewal rate of test solution (frequency/flow rate): n.a.
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared from four separate stock solutions:
Solution 1: 0.22 g KCL (312TA387636 Merck p.a., screening-test and main test) were dissolved in 1L deionized water.
Solution 2: 2.52 g NaHCO3 (K20354929 424 Merck p.a., screening-test, 72008002130 Merck p.a., main test) were dissolved in 1L deionized water.
Solution 3: 11.76 g CaCL2 * 2 H2O (TA268482 718 Merck p.a., screening-test and main test.) were dissolved in 1L deionized water.
Solution 4: 4.93 g MgSO4 * 7 H2O (356006/1 796 Fluka p.a., screening-test and main test.) were dissolved in 1L deionized water.
250 mL of each of solutions 1 to 4 were given into deionized water, the final volume was made up to 10000 mL and the solution was stirred vigorously for about 30 min.
- Intervals of water quality measurement: pH, oxygen content and temperature in the test solutions were checked at the beginning and the end of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16h of light and 8h of darkness

EFFECT PARAMETERS MEASURED : After 24 and 48h the numbers of immobilised daphnia at each concentration of the test substance was recorded. The effect of the test substance to the animals was examined by the determination of the swimming behaviour of the daphnia. For this purpose the respective test vessel was gently agitated. Daphnia showing no swimming within 15 sec. were considered as immobile. Moving of antenna were not considered as swimming.

RANGE-FINDING STUDY
- Test concentrations: 0, 1, 10, and 100 mg/L
- Results used to determine the conditions for the definitive study: yes, immobilisation results were used. No immobilisation was observed at 0, 1, and 10 mg/L after 48 hours. 100 mg/L caused immobilisation in 7/10 Daphnia after 48 hours.

Reference substance (positive control):

yes

Remarks:

potassium dichromate (tested once a month)

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

27 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Remarks on result:

other: SD%: 11%; 95% Confidence level: 34%

Details on results:

Under the conditions used for the test, immobilisation of the daphnia was observed at nominal concentrations ≥20 mg/L.
Within TOC-analyses performed with the test solutions at t =0 and 48 h, it could be shown, that the organic matter of the test substance remained constant in the aqueous phase. The recoveries of the test substance on the basis of the nominal concentrations were observed to be in the range of >90%.

Validity of the test:
• In the control without any test substance no dead animals were observed.
• In the test solutions the oxygen concentration did not drop below a value of 60% of the saturation value (5 mg O2/L).

Results with reference substance (positive control):

- Results with reference substance are valid.
- 48-h EC50: 0.9 mg/L in most recent test

Reported statistics and error estimates:

EC0 (48h): 7.5 mg/L (SD: 20%, 95% confidence level: 63%)
EC50 (48h): 27 mg/L (SD: 11%, 95% confidence level: 34%)
EC100 (48h): 98 mg/L (SD: 14%, 95% confidence level: 43%)

Main test results

Concentration of the test substance (mg/L)	Ratio immobilised Daphnia [%]
	24h	48h
Control	0 [0/20]	0 [0/20]
10	0 [0/20]	0 [0/20]
20	0 [0/20]	30 [6/20]
40	60 [12/20]	95 [19/20]
80	85 [17/20]	100 [20/20]
160	90 [18/20]	100 [20/20]

Numbers in brackets give the absolute numbers of immobilised animals at each concentration.

 

Calculated test substance recoveries in the main test, based on measured TOC-values*.

Nominal Concentration [mg/L]	Mean calculated Amount of Test Substance [mg/L] at T0	Mean calculated Amount of Test Substance [mg/L] at t48h	% Recovery at t0	% Recovery at t48h
0	0.0	0.0	0	0
10	11.4	11.3	113.7	112.8
20	20.4	20.0	101.8	100.2
40	40.6	39.5	101.6	98.7
80	76.6	75.9	95.8	94.9
160	157.6	152.3	98.5	95.2

*The measured TOC-values and the known carbon content (59.39%) including the % purity of the test substance (99%) and the chemical formula of the test substance, the concentrations and % recoveries on basis of the nominal concentrations were calculated.

Validity criteria fulfilled:

yes

Remarks:

see details on results

Conclusions:

The 48h EC50 Daphnia magna is 27 mg/L (based on the nominal concentrations, TOC confirmed levels > 80%).

Executive summary:

An acute toxicity study was performed with Daphnia magna, in accordance with OECD Guideline for Testing of Chemicals No. 202 (including GLP, Kl. 1). Groups of twenty daphnids (less than 24 hours old), were exposed for 48 hours to 0, 10, 20, 40, 80, and 160 mg/L concentrations of the substance in closed systems. The mobility of the daphnids was observed at 24 and 48 hours after study initiation. TOC-analyses were performed with the test solutions at t =0 hours and t = 48h. TOC analysis showed >90% of the nominal concentration. In view of the stability of the substance 48h EC50 value is 27 mg/L based on nominal concentrations. In addition, in the fish test the substance was stable over 24 hours supporting the TOC results.

Description of key information

An acute toxicity study was performed with Daphnia magna, in accordance with OECD Guideline for Testing of Chemicals No. 202 (including GLP, Kl. 1). Groups of twenty daphnids (less than 24 hours old), were exposed for 48 hours to 0, 10, 20, 40, 80, and 160 mg/L concentrations of the substance in closed systems. The mobility of the daphnids was observed at 24 and 48 hours after study initiation. TOC-analyses were performed with the test solutions at t =0 hours and t = 48h. TOC analysis showed >90% of the nominal concentration. In view of the stability of the substance 48h EC50 value is 27 mg/L based on nominal concentrations. In addition, in the fish test the substance was stable over 24 hours supporting the TOC results.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

27 mg/L