Reaction products of alkenyl dihydrofuran-2,5-dione and polyisoalkenyl dihydro-2,5-furandione with alkylene glycol

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:

2014-03-18 to 2014-04-24

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:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Principles of method if other than guideline:

Studiy complies with latest guideline in that environmental measurements were only taken from one replicate vessel of the control and test group. This deviates from the original study plan.

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature in the dark

Analytical monitoring:

yes

Details on sampling:

Range-finding study
Stability - A sample of each loading rate WAF was taken for analysis at 0 and 48 hours. All samples were stored at approximately -20°C prior to analysis. Only concentrations within the range to be used for the definitive test were analysed.

Definitive test
Concentration and stability - Samples taken at 0 and 48 hours.
Water samples taken from control and 100 mg/L loading rate WAF test group at 0 (from bulk test preparation) and 48 hours (replicates R1 - R4 pooled) for quantitative analysis. All samples were stored at approximately -20°C prior to analysis.
Duplicate samples were taken and stored at approximately -20°C for further analysis if necessary.

Details on test solutions:

Validation of mixing period
Pre-study investigational work was performed to determine if stirring for a prolonged period produced significantly higher levels of dissolved test item in the WAF.
A WAF of nominal loading rate of 100 mg/L was prepared, in duplicate, in deionised reverse osmosis water. One loading rate was stirred for a period of 23 hours and the other for 95 hours. After a 1-hour standing period, the mixtures were removed by siphon and samples taken for TOC.

Range-finder
Nominal amounts of test item (5, 50 and 500 mg) were added separately to the surface of 5 litres of test water to give the 1.0, 10 and 100 mg/L loading rates respectively. After the addition of the test item the water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped aafter 23 hours and the mixtures allowed to stand for 1 hour. Microscopic observations made on the WAF indicated that a significant amount of dispersed test material was present in the water column and hence it was justified to remove the WAF by filtering through a glass wool plug (2-4 cm in length). A wide bore glass tube covered at one end with Nescofilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. A length of Tygon tubing was inserted into the glass tube and pushed through the Nescofilm seal. A glass wool plug was inserted into the opposite end of the tubing and the WAF removed by mid-depth siphoning (the first 75 - 100 mL discarded) to give the 1.0, 10 and 100 mg/L loading rate WAF. Microscopic observations were performed on the WAF after filtering and showed no dispersed test item present.
Definitive test
A limit test at a single loading rate of 100 mg/L was conducted to confirm the range-finder results.
The test concentration was prepared in the same manner as for the range finder.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Source: in-house laboratory cultures
- Age at study initiation (mean and range, SD): 1st instar, < 24 hours
- Method of breeding: Culture conditions ensured that reproduction was by parthenogenesis.
- Feeding during test no
- Food type: mixture of algal suspension (Desmodesmus subspicatus) and Tetramin® flake food suspension
- Frequency: Cultures fed daily prior to the test

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

none

Hardness:

250 mg/L as CaCO3

Test temperature:

21°C

pH:

7.8-8.1

Dissolved oxygen:

8.6-8.9 mg O2/L

Salinity:

not applicable

Nominal and measured concentrations:

nominal: 0.1, 10 and 100 mg/L loading rate WAFs

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL glass jars, covered to reduce evaporation
- fill volume: 200 mL
- Aeration: no
- Renewal rate of test solution (frequency): 24 hours
- No. of organisms per vessel: 5 (range-finder 10)
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water (ISO medium) used for both the range-finding and definitive tests.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours light/8 hours darkness
- Light intensity: 556-794 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : immobility 24 & 48 hours

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0.1, 10 or 100 mg/L loading rate WAFs
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: loading rate WAF

Duration:

48 h

Dose descriptor:

NOELR

Effect conc.:

100

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: loading rate WAF

Details on results:

Rnage-finder
No immobilisation was observed at 1.0, 10 and 100 mg/L loading rate WAF.
Based on this information a single loading rate of four replicates,of 100 mg/L was seleceted for the definitive test. This experimental design conforms to a "Limit Test" to confirm no immobilisation or adverse reactions to exposure are observed.
Chemical analysis of the 100 mg/L loading rate WAF test preparations at 0 and 48 hours showed that measured concentrations of 0.58 and 0.32 mg/L were obtained respectively.

Definitive test
Chemical analysis of the 100 mg/L loading rate at 0 and 48 hours showed measured test concentrations of 0.62 and 0.49 mg/L respectively.
The dissolved test item may have been one or several components of the test item. Given that toxicity cannot be attributed to a single component or mixture of components but to the test item as a whole, the results were based on nominal loading rates only.

Immobilisation data
There was no immobilisation in 20 daphnids exposed to 100 mg/L loading rate WAF for a period of 48 hours.

The No Observed Effect Loading Rate after 24 and 48 hours was 100 mg/L loading rate WAF.

It was considered unneccessary and unrealistic to test at loading rates in excess of 100 mg/L

Test item solubility
Observations on the test media were carried out during the mixing and testing of the WAF.
At the start of the mixing period the 100 mg/L loading rate was observed to be a clear colourless water column with test item globules at the surface. After 23 hours stirring and a 1-hour standing period the 100 mg/L loading rate was observed to remain as at the start of stirring. Microscopic examination of the WAF showed that microparticles of the test item were present in the water column and therefore it was considered justifiable to remove the WAF by filtering. Microscopic examination after filtering showed the undissolved test item had been removed. During the test the 100 mg/L loading rate was observed to be a clear, colourless solution.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Mortality:
- EC50/LC50 (48h) = 0.77 mg/L (95% CI = 0.73 - 0.82 mg/L)
- Other: NOEC (48h) = 0.56 mg/L

Validation of mixing period

Preliminary investigation work in dicated that there was no significant increase in the amount of dissolved test item when the preparation period was extended for longer than 24 hours.

Validity criteria fulfilled:

yes

Conclusions:

The acute toxicity of the test item to the freshwater invertebrate Daphnia magna was assessed in accordance with OECD Guideline 202 and gave a 48-Hour EL50 value of greater than 100 mg/L loading rate WAF. The No Observed Effect Loading Rate was 100 mg/L loading rate WAF.

Executive summary:

Introduction

A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method followed was designed to becompatible with the OECD Guidelines for Testing of Chemicals (April 2004) No 202, "Daphnia sp. Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.

Methods

Due to the low aqueous solubility and complex nature of the test item, for the purposes of the test, the test medium was prepared as a Water Accommodated Fraction (WAF)

Following a preliminary range-finding test, twenty daphnids (4 replicates of 5 animals) were exposed to a WAF of the test item at a single nominal loading rate of 100 mg/L for 48 hours at a temperature of approximately 21 °C under static test conditions. The numbers of immobilized Daphnia and any adverse reactions were recorded after 24 and 48 hours.

Results

Chemical analysis of the 100 mg/L loading rate at 0 and 48 hours showed measured test concentrations of 0.62 and 0.49 mg/L respectively. The dissolved test item may have been one or several components of the test item. Given that toxicity cannot be attributed to a single component or mixture of components but to the test item as a whole, the results were based on nominal loading rates only.

Exposure of Daphnia magna to the test item gave EL50 values of greater than 100 mg/L loading rate WAF. The No Observed Effect Loading Rate was 100 mg/L loading rate WAF.

It was considered unneccessary and unrealistic to test at loading rates in excess of 100 mg/L

Description of key information

The acute toxicity of the test material to Daphnia magna was assessed in accordance with OECD Guideline 202.  Exposure of Daphnia magna to the test item gave EL50 values of greater than 100 mg/L loading rate WAF. The No Observed Effect Loading Rate was 100 mg/L loading rate WAF.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

The key study was performed on the reference substance.

A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method followed was designed to becompatible with the OECD Guidelines for Testing of Chemicals (April 2004) No 202, "Daphnia sp. Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.

Due to the low aqueous solubility and complex nature of the test item, for the purposes of the test, the test medium was prepared as a Water Accommodated Fraction (WAF)

Following a preliminary range-finding test, twenty daphnids (4 replicates of 5 animals) were exposed to a WAF of the test item at a single nominal loading rate of 100 mg/L for 48 hours at a temperature of approximately 21 °C under static test conditions. The numbers of immobilized Daphnia and any adverse reactions were recorded after 24 and 48 hours.

Chemical analysis of the 100 mg/L loading rate at 0 and 48 hours showed measured test concentrations of 0.62 and 0.49 mg/L respectively. The dissolved test item may have been one or several components of the test item. Given that toxicity cannot be attributed to a single component or mixture of components but to the test item as a whole, the results were based on nominal loading rates only.

Exposure of Daphnia magna to the test item gave EL50 values of greater than 100 mg/L loading rate WAF. The No Observed Effect Loading Rate was 100 mg/L loading rate WAF.

It was considered unneccessary and unrealistic to test at loading rates in excess of 100 mg/L.