Diallyl hexahydrophthalate

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:

13th December 2016 - 6th September 2017

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

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Source: Osaka Soda (sponsor); lot/batch no.: 40201
- Expiration date of the lot/batch: 26 January 2019
- Purity test date: 99.2%

RADIOLABELLING INFORMATION (if applicable)
- Radiochemical purity: n/a
- Specific activity: n/a
- Locations of the label: n/a
- Expiration date of radiochemical substance: n/a

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Controlled room temperature (15 - 25 ºC, below 70 RH%), protected from light
- Stability under test conditions: Not specofied
- Solubility and stability of the test substance in the solvent/vehicle: Not specified
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: Not specified

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: A saturated test item solution (with a nominal loading rate of 100 mg/L) was prepared with direct addition of the test item, mixed into the test medium (ISO medium) using ultrasonic bath approximately 35 minutes). The test solutions were prepared by appropriate dilution of this stock solution just before introduction of the Daphnia (start of treatments)
- Preliminary purification step (if any): Not specified
- Final dilution of a dissolved solid, stock liquid or gel: n/a
- Final preparation of a solid: n/a

FORM AS APPLIED IN THE TEST (if different from that of starting material) : n/a

TYPE OF BIOCIDE/PESTICIDE FORMULATION (if applicable) : n/a

OTHER SPECIFICS: n/a

Analytical monitoring:

yes

Remarks:

Analytical measurements using HPLC-UV performed from the control and at the applied test concentration levels at the beginning and at the end of the renewal periods.

Details on sampling:

- Concentrations: control (0.0), 6.21, 12.10, 23.83, 47.04 and 90.00 mg/L
- Sampling method: Samples were taken from the test solutions and one sample from the control solution. Samples were properly diluted and analysed by HPLC-UV method.
- Sample storage conditions before analysis: Not specified

Vehicle:

no

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia
- Strain/clone: Daphnia magna
- Justification for species other than prescribed by test guideline: Standard species of the acute immobilisation test
- Source: István Szent University, Hungary
- Age of parental stock (mean and range, SD): at least 24 hours old at the beginning of the test
- Feeding during test: No
- Food type: n/a
- Amount: 20 animals in test group and control group, divided into 4 replicates (5 animals / replicate)
- Frequency: water renewal frequency was 24 hours

ACCLIMATION
- Acclimation period: Not required because water used was similar to the culture water
- Acclimation conditions (same as test or not): n/a
- Type and amount of food: n/a
- Feeding frequency: n/a
- Health during acclimation (any mortality observed): n/a

QUARANTINE (wild caught)
- Duration: n/a
- Health/mortality: n/a

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES: n/a

Test type:

semi-static

Water media type:

not specified

Total exposure duration:

48 h

Hardness:

246 mg/L (as CaCO3)

Test temperature:

20.2 - 20.6 ºC

pH:

7.47 - 8.06

Dissolved oxygen:

8.0 - 8.6 mg/L

Salinity:

not stated

Conductivity:

not stated

Nominal and measured concentrations:

Nominal (% sat.sol.) = control, 6.25, 12.5, 25, 50, 100
Measured (mg/L) = 0.0, 6.21, 12.10, 23.83, 47.04, 90.00

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beaker
- Type (delete if not applicable): open / closed Not specified
- Material, size, headspace, fill volume: at least 5 mL test solution/animal
- Aeration: not specified
- Type of flow-through (e.g. peristaltic or proportional diluter): not specified
- Renewal rate of test solution (frequency/flow rate): 24 hour
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): n/a
- Biomass loading rate: not specified

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water (ISO medium)
- Total organic carbon: not specified
- Particulate matter: not specified
- Metals: not specified
- Pesticides: not specified
- Chlorine: not specified
- Alkalinity: not specified
- Ca/mg ratio: not specified
- Conductivity: not specified
- Salinity: not specified
- Culture medium different from test medium: not specified
- Intervals of water quality measurement: Beginning and end of the renewal periods

OTHER TEST CONDITIONS
- Adjustment of pH: Not adjusted, did not vary by more than 1.5 units in any one test
- Photoperiod: 16 hour light and 8 hour dark cycle
- Light intensity: not specified

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobility or mortality determined by visual observation 24 and 48 hours after the start of the test.

VEHICLE CONTROL PERFORMED: no (no vehicle). An untreated control was performed (dilution water ISO-medium)

RANGE-FINDING STUDY: Yes.
- Test concentrations: Ten daphnids (divided into two replicates) in each test concentration and control were exposed for 48 hours under semi-static conditions.
- Results used to determine the conditions for the definitive study: Significant immobility detected at higher test item concentrations (10 and 100 % sat. solution - 1 and 10 immobilised animals observed respectivily).

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

ca. 38.19 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

ca. 24.03 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

ca. 47.04 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

ca. 12.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

ca. 23.83 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: Not specified
- Observations on body length and weight: Not specified
- Other biological observations: Not specified
- Mortality of control: 0
- Other adverse effects control: Not specified
- Abnormal responses: Not specified
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Not specified
- Effect concentrations exceeding solubility of substance in test medium: Not specified

Results with reference substance (positive control):

- Results with reference substance valid? Yes - no immobilisation observed
- Relevant effect levels: not specified
- Limit test: not specified
- Dose-response test: no immobilisation observed
- ECx: not specified
- Other: n/a

Reported statistics and error estimates:

Dunnett's Test used - for measured concentration of 12.10 mg/L, Dunnett's test showed that there was no statistical difference between that concentration and the results of the copntrol.

For test concentrations 23.83, 47.04 and 90.00 mg/L, Dunnett's test showed that there was a statistically significant difference in the results compared to the control.

Validity criteria fulfilled:

yes

Conclusions:

Acute toxicity of MDAC on Daphnia magna was assessed in an acute immobilisation test, over an exposure period of 48 hours in a semi-static test system. The percentage of Daphnia being no longer capable of swimming at the end of the test or being dead was recorded for each test concentration. The nominal concentrations of test item used in the main experiment were: 6.25, 12.5, 25, 50 and 100 % saturated solution. The test concentrations were analytically determined at the start and at the end of the renewal periods. The corresponding measured geometric mean test item concentrations were: 6.21; 12.10; 23.83; 47.04 and 90.00 mg/L.

The immobility or mortality of the Daphnia was determined by visual observation 24 and 48 hours after the start of the test. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile. The number of immobilised animals and the percentage of immobility were determined at 24 and 48 hours. All validity criteria were within acceptable limits and therefore the study can be considered as valid.

Under the conditions of this Daphnia magna acute immobilisation study the observed endpoints for the effect of MDAC were the followings:
- The 24h EC50 value: 38.19 mg/L (measured) (95 % conf. limits: 35.60 – 40.98 mg/L)
- The 48h EC50 value: 24.03 mg/L (measured) (95 % conf. limits: 22.25 – 25.96 mg/L)
- The 48h EC100 value: 47.04 mg/L (measured)
- The 48h No-Observed Effect Concentration (NOEC): 12.10 mg/L (measured)
- The 48h Lowest Observed Effect Concentration (LOEC): 23.83 mg/L (measured)

Description of key information

Acute toxicity of MDAC on Daphnia magna was assessed in an acute immobilisation test, over an exposure period of 48 hours in a semi-static test system. The percentage of Daphnia being no longer capable of swimming at the end of the test or being dead was recorded for each test concentration. Because significant immobility was observed at the highest examined concentration level during the preliminary range-finding test, five test concentrations in a geometric series (factor 2.0) and one control group were tested in the definitive test under semi-static conditions.

The nominal concentrations of test item used in the main experiment were: 6.25, 12.5, 25, 50 and 100 % saturated solution. The test concentrations were analytically determined at the start and at the end of the renewal periods. The corresponding measured geometric mean test item concentrations were: 6.21; 12.10; 23.83; 47.04 and 90.00 mg/L. Twenty animals (young Daphnia magna, less than 24hr old) were divided into four groups (replicates) within glass beakers of five animals each at the test concentrations and for the control as well. The animals were not fed during the test.

The choice of the test concentration was made on the basis of the results of a preliminary range-finding test, which consisted of ten daphnids (divided into two replicates) in each test concentration and control being exposed for 48 hours under semi-static conditions. At concentrations of 10% sat.solution, 1 daphnia was immobile. At 100% sat.solution, 100% of all daphnia were immobile.

The immobility or mortality of the Daphnia was determined by visual observation 24 and 48 hours after the start of the test. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile. The number of immobilised animals and the percentage of immobility were determined at 24 and 48 hours.

All validity criteria were within acceptable limits and therefore the study can be considered as valid.

Under the conditions of this Daphnia magna acute immobilisation study the observed endpoints for the effect of MDAC were the followings:

- The 24h EC50 value: 38.19 mg/L (measured) (95 % conf. limits: 35.60 – 40.98 mg/L)

- The 48h EC50 value: 24.03 mg/L (measured) (95 % conf. limits: 22.25 – 25.96 mg/L)

- The 48h EC100 value: 47.04 mg/L (measured)

- The 48h No-Observed Effect Concentration (NOEC): 12.10 mg/L (measured)

- The 48h Lowest Observed Effect Concentration (LOEC): 23.83 mg/L (measured)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

24.03 mg/L