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

Toxicity to aquatic plants other than algae

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic plants other than algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08 Feb - 15 Feb 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

other: EPA OPP 123-2 (Aquatic Plants Field Study, Tier III), adopted in 1982

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all replicate test concentrations were analyzed at test start and after 96 h
- Sampling method: Approximately 50 to 60 mL was sampled from each freshly prepared test solution on Day 0 and Day 4. Approximately 30 mL of old test solutions (composite of replicate chambers within a treatment) was taken on Day 2 and Day 7.
- Sample storage conditions before analysis: not reported

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Acetone
- A primary stock solution was prepared by weighing 200.4 mg of the test substance, transferring it to a 10 mL volumetric flask and bringing it to volume with acetone for a final concentration of 20 mg/mL. The 2.0 mg/L treatment concentration was prepared by the addition of 0.4 ml of the primary stock (20 mg/mL) into a sterile pre-measured 4 L Erlenmeyer flask filled with 20X-AAP medium. The 2.0 mg/L treatment concentration was allowed to stir for one hour in EC-3 at test temperature. A white, flaky precipitate was observed following the stirring procedure. A solvent control was prepared by the addition of 0.4 mL acetone in 4 L of sterile 20X-AAP media for a final solvent loading factor of 0.1 mL/L. The control treatment contained only 20X-AAP medium in a 4 L Erlenmeyer flask.

Test organisms (species):

Lemna gibba

Details on test organisms:

TEST ORGANISM
- Common name: duckweed
- Source (laboratory, culture collection): Climate Stress Laboratory, USDA/ARS Beltsville Agricultural Center, Beltsville, MD, USA
- Age of inoculum (at test initiation): Cultures were maintained since 01 Apr 1998
- Method of cultivation: aseptic in 20X-AAP nutrient medium under continuous light

ACCLIMATION
- Culturing media and conditions (same as test or not): light intensity during 12 days acclimation was slightly different to main test: acclimation 4400 - 5400 lux, main test 4200 - 5800

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

7 d

Hardness:

No details in report given, standard 20X-AAP medium was used.

Test temperature:

Freshly prepared solutions: 24.7 - 25.0 °C
Old solutions from treatment vessels: 23.2 - 24.5 °C

pH:

Freshly prepared solutions: 6.8 - 7.4
Old solutions from treatment vessels: 8.4 - 8.5

Dissolved oxygen:

Freshly prepared solutions: 6.8 - 7.4 ppm, 82 - 88% saturation
Old solutions from treatment vessels: 8.3 - 10.1 ppm, 96 - 118% saturation

Nominal and measured concentrations:

Nominal: 2 mg/mL
Measured in freshly prepared treatment solutions: 1.478 mg/mL (mean value from two measurements, maximum solubility)
Measured in 2 day-old solutions: not detectable

Details on test conditions:

TEST SYSTEM
- Incubation chamber used: yes
- Test vessel: crystallizing dishes
- Type (delete if not applicable): closed
- Material, size, fill volume: crystallizing dish, 1200 mL, 600 mL
- Type of cover:glass sheet
- Renewal rate of test solution (frequency/flow rate): renewal every 2 days
- No. of plants per vessel: 5
- No. of fronds per plant: 3
- No. of fronds per vessel: 15
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes, 20X-AAP medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: filter sterilized 20X-AAP medium prepared from deionized well water
- Total organic carbon: < 1.0 mg/L
- Particulate matter: < 9.0 mg/L
- Metals (mg/L): Ba 0.21, Ca 34.2, Mg 3.20, Mn 0.027, K 2.24, Na 5.79; 20 other metals were below quantification limit
- Pesticides: all 34 pesticides analyzed were below quantification limit
- Chloride: 8.2 mg/L
- Alkalinity: 0.0 mg/L

- Conductivity: 20 µmhos/cm
- Culture medium different from test medium: no
- Intervals of water quality measurement: Discrete measurements of temperature, dissolved oxygen, pH, and specific conductivity were obtained on a representative sample of freshly prepared treatment solutions at test initiation (Day 0) and at renewal periods on Day 2 and Day 4. Discrete measurements were also performed on composite samples of replicates within treatments on Days 2, 4, and 7 (test termination).

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: Growth medium was adjusted to pH of 7.5 ± 0.1 with 0.1 N sodium hydroxide
- Photoperiod: continuous light
- Light intensity and quality: 4200 - 5800 lux, warm white fluorescent tubes

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of frond number: Frond counts and abnormal appearance were visually evaluated with the aid of an illuminated light box on Days 2, 4, and 7
- Determination of biomass: area under the curve calculations


RANGE-FINDING STUDY
- Test concentration: 1.0 mg/L
- Results used to determine the conditions for the definitive study: Average frond growth following 7 days of exposure for the solvent control and 1.0 mg/L treatment was 229 and 226, respectively.

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

>= 1.478 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Duration:

7 d

Dose descriptor:

LOEC

Effect conc.:

> 1.478 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: The 2.0 mg/L treatment concentration was allowed to stir for one hour in EC-3 at test temperature. A white, flaky precipitate was observed following the stirring procedure. The concentration of 1.478 mg/L at test start represents the maximum solubility. No test substance was detectable in the old test media after 2 days.
- Deviation from specification: The discrete temperature of test solutions at test termination (Day 7) deviated from specification (25.0 ± 1 °C) at 23.1 °C for all treatments. The low temperature was believed to be a result of pooling test solutions. The continuous temperature was verified to be within the specified range of 25.0 ±1 °C.

Mean number of fronds on Day 7 in the control, solvent control, and 1.478 mg/L treatment were 243, 244, and 240, respectively. The mean number of fronds in the pooled control chambers increased from 15 at study initiation to 244 at study termination representing an increase of 16 times. No phytotoxic effects were observed in the 1.478 mg/L treatment concentration.

Inhibition of biomass (area under the curve) and specific growth rate relative to the pooled controls were both 1%.

Validity criteria fulfilled:

not specified

Remarks:

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

Description of key information

NOEC (7 d): ≥ 1.478 mg/L (initial measured, Lemna gibba, EPA OPP 123-2)

No acute toxic effects on Duckweed growth were observed at the maximum solubility of the test substance. However, the test substance hydrolyse completely during the course of the study.

Additional information

A growth inhibition limit test with Duckweed (Lemna gibba G3) according to US-EPA OPP 123-2 (Aquatic Plants Field Study, Tier III) and GLP is available. In this semi-static 7-day exposure system (medium renawal: every 2 days), the test substance revealed no effects on Duckweed growth at its maximum solubility. However, due to hydrolysis during the study period at test termination, the test substance was not detectable in the test solution any more.