ethyl 5,5-diphenyl-2-isoxazoline-3-carboxylate

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:

14 May - 16 May 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

adopted in 1992

Qualifier:

according to guideline

Guideline:

EPA OPP 72-2 (Aquatic Invertebrate Acute Toxicity Test)

Version / remarks:

adopted in 1982

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Limit test. Each triplicate test chamber was sampled for analysis of Ethyl 5,5-diphenyl-2-isoxazoline-3-carboxylate, on Day -1, at 0 hour (prior to the introduction of the test organisms to the test chambers), and at 48 hours.
- Sampling method: 20 mL samples were taken at mid-depth for each time point
- Sample storage conditions before analysis: samples were immediately analyzed

Vehicle:

yes

Details on test solutions:

- Method: A primary stock solution of the test item was prepared in acetone. 500 mg of test substance was weighed out and placed into a 50 mL volumetric flask. The flask was then filled to the line with acetone for a final concentration of 10 mg/mL. Test solutions were prepared from the primary stock solution and delivered to triplicate test chambers via a proportional diluter. Approximately fifteen minutes before each diluter cycle, 0.25 mL of the primary stock solution was injected with a syringe pump into approximately 2.5 L of dilution water in the diluter mix box. This solution was allowed to mix via a magnetic stirrer using a Teflon stir bar until the diluter cycled.
The diluter cycled approximately 48 cycles per day, at intervals of approximately 30 minutes between cycles. The diluter delivered approximately 375 mL of test solution per replicate chamber per cycle, or approximately 1.3 volume additions per cycle and approximately 62 volume additions per day.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Source: Parental stock was purchased September 14, 1993 from Aquatic Research Organisms, Hampton, NH. The supplier stated the original source to be the EPA laboratories in Cincinnati, OH. Daphnids were cultured in the Environmental Biology Laboratory at the AgrEvo Research Center since September 14, 1993.
- Date of purchase: 1993-09-14
- Age at study initiation: < 24 h
- Method of culture: Daphnids were cultured in 1 liter Pyrex beakers containing 800 mL of hard synthetic freshwater (hardness: 160 to 180 mg/L as CaC03). Culture beakers were maintained in a temperature controlled environmental chamber.
- Feeding during test: none

CULTURE
- Culture conditions (same as test or not): same as test
- Type and amount of food: Pseudokirchneriella subcapitata twice daily; TetraFin fish food suspension every Monday, Wednesday and Friday
- Health during acclimation (any mortality observed): no data

Test type:

flow-through

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

160 - 180 mg/L CaCO3

Test temperature:

19.9 - 20.2 °C

pH:

8.2 - 8.3

Dissolved oxygen:

9.6 - 9.9 ppm
106 - 109%

Nominal and measured concentrations:

nominal: 1.0 mg/L
mean measured: 0.51 mg/L (maximum solubility)

Details on test conditions:

TEST SYSTEM
- Test vessel: beakers
- Type (delete if not applicable): open, but standing in covered overflow tanks
- Material, size, fill volume: Pyrex glass; diameter 6.6 cm, hight 8.8 cm; fill volume: 300 mL
- Aeration: none
- Type of flow-through (e.g. peristaltic or proportional diluter): proportional diluter
- Renewal rate of test solution (frequency/flow rate): The diluter cycled approximately 48 cycles per day, at intervals of approximately 30 minutes between cycles. The diluter delivered approximately 375 ml of test solution per replicate chamber per cycle, or approximately 1.3 volume additions per cycle and approximately 62 volume additions per day.
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3
- No. of vessels per vehicle control (replicates): 3

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: A synthetic hard fresh water was prepared from deionized well water
- Total organic carbon: < 1.0 mg/L
- Particulate matter: < 9.0 mg/L
- Metals (mg/L): Ba 0.23, Ca 16.8, Fe 0.47, Mg 3.84, Mn 0.043, K 2.57, Na 5.96, Zn 0.045, Cu 0.0066; 17 other metals were shown to be below detection limit
- Pesticides: all 34 pesticides analyzed were below detection limit
- Chloride: 8.2 mg/L
- Alkalinity: 106 mg/L
- Conductivity: 500 µmhos/cm
- Culture medium different from test medium: no, except for addition of SeO2 and Vitamin B12 to the test medium
- Intervals of water quality measurement: Temperature, dissolved oxygen, pH and conductivity were measured at test initiation and at 48 h

OTHER TEST CONDITIONS
- Photoperiod: 16:8, light:dark
- Light intensity: 700 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Observations for death (i.e. lack of response to gentle prodding), and for abnormal appearance and behavior, were performed at 3, 6, 24, and 48 hours (± 1 hour).

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: first range finding test: 0.06, 0.1, 0.2, 0.3, 0.5 mg/L nominal; second range finding test 0.13, 0.22, 0.36, 0.60, 1.0 mg/L nominal
- Results used to determine the conditions for the definitive study: No mortality or sublethal effects were observed in the control or any treatment level during the two studies

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 0.51 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

0.51 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Mortality of control: One random mortality was observed in one replicate of the control treatment. No mortality was observed in the solvent control
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: A precipitate was observed in the diluter mix box and on the walls of the toxicant "E" chambers during the study. 0.51 mg/L mean measured concentration represents the maximum solubility of the test substance in the dilution water.

Reported statistics and error estimates:

This test was designed to show that Ethyl 5,5-diphenyl-2-isoxazoline-3-carboxylate is not toxic to Daphnia magna at a concentration of 1.0 mg/L (nominal). Therefore no calculation of EC50 values were performed.

Table 1: Validity criteria for OECD 202.

Criterion from the guideline	Outcome	Validity criterion fulfilled
In the control, including the control containing the solubilising agent, not more than 10% of the daphnids should have been immobilized.	One random mortality was observed in one replicate of the control treatment. No mortality was observed in the solvent control	yes
The dissolved oxygen concentration at the end of the test should be ≥ 3 mg/L in control and test vessels.	9.6 - 9.9 mg/L	yes

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Description of key information

EC50 (96 h): > 0.51 mg/L (mean measured, Daphnia magna, OECD 202)

No acute toxic effects on aquatic invertebrates were observed up to the limit of water solubility.

Additional information

One short term limit test using a flow-through system with Daphnia magna according to OECD 202 and GLP is available. The maximum solubility of the test substance was shown to be 0.51 mg/L under the present test conditions. For this concentration, no acute toxic effects on Daphnia magna were observed.