Reaction mass of 3-[(diphenoxyphosphoryl)oxy]phenyl triphenyl 1,3-phenylene bis(phosphate) and tetraphenyl 1,3-phenylene bis(phosphate)

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:

STUDY INITIATION DATE:November 27, 2018 STUDY COMPLETION DATE: March 11, 2019

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)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Sampling method:
Test water samples were collected from one test chamber of each treatment and control group
nine, eight, four and two days prior to the start of exposure to confirm concentrations after conditioning
the diluter system for one, two, six and eight days, respectively. Test water samples also were collected
from one replicate test chamber in each treatment and control group at the beginning of the test and at
48 hours (± 1 hour) to measure concentrations of the test substance. Sampling alternated between the
replicate test chambers in each group at each sampling interval. The samples (10.0 mL) were collected
from mid-depth, placed in glass vials containing 10.0 mL of acetonitrile and processed immediately for
analysis. Stock solution samples also were collected at the beginning of the test.

- Sample storage conditions before analysis:
The samples (approximately 2 mL) were placed in glass vials and stored under refrigerated conditions until analysis.The analytical method consisted of diluting samples with acetonitrile upon collection. The samples were further diluted into the calibration curve range with 50 : 50 (v/v) acetonitrile : freshwater, as necessary

Vehicle:

yes

Remarks:

Dimethylforamide (DMF)

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A 50-mL primary stock solution was prepared by mixing a calculated amount of test substance into HPLC-grade dimethylformamide (DMF) at a nominal concentration of 1500 μg/mL. The primary stock solution was brought to volume and inverted to mix. The stock solution appeared clear and colorless with no visible precipitates. Four secondary stock solutions (20 mL each) were prepared DMF at nominal concentrations of 95, 190, 375 and 750 μg/mL by proportional dilution of the primary stock.
- Controls: The negative control received dilution water only. The solvent control was prepared by delivering HPLC-grade DMF to the mixing chamber for the solvent control at the same rate as the test substance stock solutions.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Dimethylformamide
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): The concentration of DMF in the solvent control and all treatment groups was 20 μL/L.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): no.
- Other relevant information:
During the course of the test, the appearance of the test solutions at these nominal concentrations was observed in the test chambers, as well as in the diluter mixing chambers where test substance stocks and dilution water were combined prior to delivery to the test chambers. The test solutions in the mixing chambers and test chambers
appeared clear and colorless during the test, with no evidence of precipitation observed in any control or treatment solution.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: cladoceran, Daphnia magna,
- Age at study initiation (mean and range, SD): < 24-hour old neonates
- Source: cultures maintained by Eurofins EAG Agroscience, LLC in Easton, Maryland
- Age of parental stock (mean and range, SD): Adult daphnids
- Feeding during test : not fed during the test
- Food type: yeast, cereal grass media and trout chow (YCT), supplemented with a vitamin stock solution and a suspension of the freshwater green alga, Raphidocelis subcapitata.
- Frequency: Daphnids in the cultures were fed daily. The adults were fed during the 24-hour period prior to test initiation, but neonates were not fed during the test.

ACCLIMATION
- Acclimation period: 2-week period
- Acclimation conditions (same as test or not): same as test. water temperatures in the cultures ranged from 19.3 to 20.4ºC, the pH of the water ranged from 8.1 to 8.4, and the dissolved oxygen concentrations were >= 7.3 mg/L (>= 80% of saturation).
- Type and amount of food:
yeast, cereal grass media and trout chow (YCT), supplemented with a vitamin stock solution and a suspension of the freshwater green alga, Raphidocelis subcapitata.
- Feeding frequency: The adults were fed during the 24-hour period prior to test initiation, but neonates were not fed during the test.
- Health during acclimation (any mortality observed): The four adult daphnids used to supply neonates for the test were held for at least 24 days prior to collection of the juveniles for testing and had each produced at least five previous broods. Adult daphnids in the culture had produced an average of at least three young per adult per day over the 7-day period prior to the test. The adults showed no signs of disease or stress, no ephippia were produced during the holding period, and mortality in the culture stock was < 5% in the two-day period prior to test initiation

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

140 mg/L as CaCO3

Test temperature:

20.1 – 20.4°C

pH:

8.1 – 8.2

Dissolved oxygen:

≥ 79% ASV; no aeration

Salinity:

Alkalinity 182 mg/L as CaCO3

Conductivity:

338 μS/cm

Nominal and measured concentrations:

Nominal Fyrolflex RDP Nominal n=1 Component Mean Measured n=1 Component Mean Measured Fyrolflex RDP
Negative Control Negative Control < LOQ < LOQ
Solvent Control Solvent Control < LOQ < LOQ
1.9 μg/L 1.3 μg/L 0.94 μg/L 1.3 μg/L
3.8 μg/L 2.7 μg/L 0.90 μg/L 1.3 μg/L
7.5 μg/L 5.3 μg/L 2.5 μg/L 3.6 μg/L
15 μg/L 11 μg/L 5.6 μg/L 7.9 μg/L
30 μg/L 21 μg/L 14 μg/L 20 μg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Test chambers were 25-L Teflon®-lined stainless steel aquaria filled with approximately 22 L of test water. The depth of the test water in a representative chamber was 28.8 cm and was maintained by a standpipe within the test chamber. Each test chamber contained two test compartments constructed from a glass beaker approximately 6.5 cm in diameter and 12 cm in height, with two nylon mesh-covered holes on opposite sides of the container to allow for the flow of water through the test compartment. The depth of the test water in a representative compartment was 8.8 cm.
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 22 L
- Volume of solution: filled with approximately 22 L of test water
- Aeration: No
- Type of flow-through (e.g. peristaltic or proportional diluter): proportional diluter
- Renewal rate of test solution (frequency/flow rate): The proportion of the test water that was delivered to each replicate test chamber was checked to ensure that flow rates varied by no more than ± 10% of the mean flow rate for the two replicates.During the exposure period, the stock solutions were pumped into the diluter mixing chambers assigned to the treatment groups at a target rate of 3.10 μL/minute and were mixed with dilution water in the mixing chambers, delivered at a target rate of 155 mL/minute to achieve the desired nominal test concentrations.
- No. of organisms per vessel: five daphnids in each test compartment
- No. of vessels per concentration (replicates): two replicate test chambers in each treatment group.
- No. of vessels per control (replicates):
two replicate test chambers in each control.
- No. of vessels per vehicle control (replicates): two replicate
- Biomass loading rate:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The water used for organism holding and testing was freshwater obtained from a well approximately 40 meters deep located on the Eurofins site in Easton, Maryland.
- Total organic carbon: <1
- Particulate matter: See Appendix 4 in the attached report
- Metals: See Appendix 4 in the attached report
- Pesticides: See Appendix 4 in the attached report
- Chlorine: See Appendix 4 in the attached report
- Alkalinity: 162 – 180mg/L as CaCO3
- Ca/mg ratio: 136 mg/L as CaCO3
- Conductivity: 305 – 339 μS/cm
- Salinity:
- Culture medium different from test medium:
- Intervals of water quality measurement: approximate four-week period immediately preceding the test

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours of light and 8 hours of darkness. A 30-minute transition period of low light intensity was provided when lights
went on and off to avoid sudden changes in light intensity.
- Light intensity: Light intensity at the beginning of the test was 570 lux at the surface of the water of one representative test chamber

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Dissolved oxygen and pH were measured in each replicate test chamber of each treatment and control group at the beginning of the test, at approximately 24 hours during the test, and at the end of the test.

VEHICLE CONTROL PERFORMED: yes

RANGE-FINDING STUDY
- Test concentrations: Nominal test concentrations were selected in consultation with the Sponsor based on the study range finding test: "Claude, M. 2014. Fyrolflex RDP: A Flow-Through Life-Cycle Toxicity Test with the Cladoceran (Daphnia magna). Wildlife International, Ltd. Project Number 238A-131A. 24 October 2014", and the reported water solubility (8.91 μg a.i./L for the n=1 component and 10738 μg a.i./L for the TPP component ).
- Results used to determine the conditions for the definitive study: were taken from the study: Claude, M. 2014. Fyrolflex RDP: A Flow-Through Life-Cycle Toxicity Test with the Cladoceran (Daphnia magna). Wildlife International, Ltd. Project Number 238A-131A. 24 October 2014.

Reference substance (positive control):

yes

Remarks:

Dimethylforamide (DMF)

Key result

Duration:

48 h

Dose descriptor:

EC50

Remarks:

based on measured n=1 component

Effect conc.:

> 14 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Remarks:

n=1 component

Basis for effect:

mortality

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 20 µg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat. (total fraction)

Basis for effect:

mortality

Details on results:

- Behavioural abnormalities: no
- Observations on body length and weight: no
- Other biological observations: No signs of toxicity were observed among the surviving daphnids any control or treatment group at test termination.
- Mortality of control: All daphnids in the negative and solvent control groups appeared normal throughout the test.
- Other adverse effects control: No signs of toxicity were observed among the surviving daphnids any control or treatment group at test termination.
- Immobilisation of control: no immobile daphnids or overt signs of toxicity observed.
- Abnormal responses: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values:
- Effect concentrations exceeding solubility of substance in test medium:no immobile daphnids or overt signs of toxicity observed above solubility level (14 μg/L (calculated to > 20 μg/L Fyrolflex RDP from the measured n=1 component)).

Please see table 9 attached below and on page 31 in the attached report

Validity criteria fulfilled:

yes

Conclusions:

The cladoceran, Daphnia magna, was exposed for 48 hours under flow-through conditions to five mean measured concentrations of the n=1 component of Fyrolflex RDP ranging from 0.90 to 14 μg/L, which calculate to 1.3 to 20 μg/L Fyrolflex RDP from the measured n=1 component. Based on the mean measured concentrations of the n=1 component of Fyrolflex RDP, the 48-hour EC50 value was > 14 μg/L (calculated to > 20 μg/L Fyrolflex RDP from the measured n=1 component), the highest concentration tested. The no-immobility concentration was 14 μg/L (calculated to 20 μg/L Fyrolflex RDP from the measured n=1 component), the 100% immobility concentration was > 14 μg/L (calculated to > 20 μg/L Fyrolflex RDP from the measured n=1 component) and the NOEC was 14 μg/L (calculated to 20 μg/L Fyrolflex RDP from the measured n=1 component). Even though a solvent used for increasing substance solubility no toxicity was observed under the experimental conditions.

Executive summary:

The cladoceran, Daphnia magna, was exposed for 48 hours under flow-through conditions to five mean measured concentrations of the n=1 component of Fyrolflex RDP ranging from 0.90 to 14 μg/L, which calculate to 1.3 to 20 μg/L Fyrolflex RDP from the measured n=1 component. The 48-hour EC50 value was > 14 μg/L (calculated to > 20 μg/L Fyrolflex RDP from the measured n=1 component), the highest concentration tested.

The EC50 value is above water solubility value (8.9 μg/L) of the substance. Even though a solvent used for increasing substance solubility no toxicity was observed under the experimental conditions.

Description of key information

In a key study according to OECD guideline 202 under GLP, the acute toxicity (48h-EC50) of Fyrolflex RDP towards Daphnia magna was found to be > 20 μg/L. The EC50 value is above water solubility value (8.9 μg/L) of the substance. No toxicity was obsered under the experimental conditions. Even though a solvent used for increasing substance solubility no toxicity was observed under the experimental conditions.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

20 µg/L