Tall oil, reaction products with ethanolamine

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:

06 January 2015 to 20 February 2015

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

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

RANGE-FINDING TEST
- A sample of each loading rate WAF was taken for chemical analysis at 0 and 48 hours in order to determine the tability of the test item under test conditions.
- The 0-hour samples were stored frozen prior to analysis.
- Only concentrations within the range to be used for the definitive test were analysed.

DEFINITIVE TEST
- Water samples were taken from the control and the 100 mg/L loading rate WAF test goup at 0 hours (taken from bulk test preparation) and 48 hours (replicates R1 to R4 pooled) for quantitative analysis.
- The 0-hour samples were stored frozen prior to analysis.
- Duplicate samples were taken and stored frozen for further analysis if necessary.

Vehicle:

no

Details on test solutions:

TEST WATER
- Reconstituted water (ISO medium) used for both the range-finding and definitive test is defined in Appendix 4 (attached).

PROCEDURE
- Due to the low 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).

VALIDATION OF MIXING PERIOD
- Preliminary work (see Appendix 5, attached) was carried out to determine whether stirring for a prolonged period produced significantly higher measured test concentrations in the WAF.

RANGE_FINDING TEST
- The loading rate to be used in the definitive test was determined by a preliminary range-finding test.
- Nominal amounts of test item (5.0, 50 and 500 mg) were each separately added to the surface of 5 L of test water to give 1.0, 10 and 100 mg/L loading rates respectively.
- After addition of the test item, the test water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface.
- Stirring was stopped after 23 hours and the mixtures were allowed to stand for one hour.
- 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.
- Microscopic inspection of the WAFs showed no micro-dispersions or undissolved test item to be present.
- The aqueous phase or WAF was removed by mid-depth siphoning (the first approximate 75-100 mL discarded) to give 1.0, 10 and 100 mg/L loading rate WAFs.

DEFINITIVE TEST
- Based on the results of the range-finding test a "limit test" was conducted at a single loading rate of 100 mg/L to confirm that no immobilisation or adverse reactions to exposure wwere observed.
- A nominal amount of test item (500 mg) were each separately added to the surface of 5 L of test water to give the 100 mg/L loading rate.
- After addition of the test item, the test water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface.
- Stirring was stopped after 23 hours and the mixtures were allowed to stand for one hour.
- 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.
- Microscopic inspection of the WAFs showed no micro-dispersions or undissolved test item to be present.
- The aqueous phase or WAF was removed by mid-depth siphoning (the first approximate 75-100 mL discarded) to give the 100 mg/L loading rate WAF.

Test organisms (species):

Daphnia magna

Details on test organisms:

- The test was carried out using 1st instar Daphnia magna dervied from in-house laboratory cultures.
- Adult Daphnia were maintained in 150 mL glass beakers containing Elendt M7 medium (see Appendix 2, attached) in a temperature controlled room at approximately 20°C.
- The lighting cycle was controlled to give 16 hours light and 8 hours darkness with 20 minute dawn and dusk periods.
- Each culture was fed daily with a mixture of algal suspension (Desmodesmus subspicatus) and Tetramin flake food suspension.
- Culture conditions ensured that reproduction was by parthenogenesis.
- Gravid adults were isolated the day before initiation of the test so that young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing.
- The diet and dilutent water were considered not to contain any contaminant that would affect the integrity or outcome of the study.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

None

Hardness:

250 mg/L total theoretical hardness as CaCO3

Test temperature:

21 °C

pH:

7.5 to 8.2

Dissolved oxygen:

8.7 to 9.0 mg O2/L

Salinity:

Not applicab;e

Nominal and measured concentrations:

RANGE-FINDING TEST
- Nominal loading rates of 1.0, 10 and 100 mg/L.

DEFINITIVE TES
- Nominal loading rate of 100 mg/L.

Details on test conditions:

RANGE FINDING TEST
- Daphnids (10) were placed in each test and control vessel and maintained at a temperature controlled room at approximately 21 °C with a photoperiod of 16 hours light and 8 hours darkness for a period of 48 hours with 20 minute dawn and dusk transition periods.
- Each 250 mL test and control vessel contained 200 mL of test media and was covered to reduce evaporation.
- After 24 and 48 hours, the number of immobilised Daphnia magna were recorded.
- The control group was maintained under identical conditions but not exposed to the test item.

DEFINITIVE TEST
- As in the range-finding test, 250 mL glass jars containing approximately 200 mL of test preparation were used.
- Daphnids (5) were placed in each test and control vessel at random.
- Four replicate test and control vessels were prepared.
- Test vessels were covered to reduce evaporation and maintained in a temperature controlled room at approximately 21 °C with a photoperiod of 16 hours light (523 to 561 lux) and 8 hours darkness with 20 minute dawn and dusk transition periods.
- Daphnids were not individually identified, received no food during exposure, and the test vessels were not aerated.

Reference substance (positive control):

yes

Remarks:

potassium dichromate conducted between 23 February 2015 and 25 February 2015 (see Appendix 3, attached)

Duration:

24 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

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

Duration:

24 h

Dose descriptor:

NOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

VALIDATION OF MIXING PERIOD
- Preliminary investigational work (see Appendix 5, attached) indicated that there was no increase in the amount of dissolved test item when the preparation period was extended for longer than 24 hours.
- Therefore, for the purpose of testing, the WAF was prepared using a stirring period of 23 hours followed by a 1-hour settlement period.

RANGE-FINDING TEST
- Cumulative immobilisation data from exposure of Daphnia magna during the range-finding test are shown in Table 1 (attached).
- No immobilisation was observed at 1.0, 10 and 100 mg/L loading ragte WAF.
- Based on this information, a single loading rate of 100 mg/L (four replicates) was selected for the definitive test.
- Chemical analysis of the 100% v/v saturated solution at 0 and 48 hours showed measured test concentrations of less than 0.0072 mg/L, which was determined to be the limit of quantification (LOQ) for the analytical method employed.
- This result does not infer that no test item was in solution. However, any dissolved test item was present at a concentration of less than the LOQ.

DEFINITIVE TEST - CHEMICAL ANALYSIS OF TEST LOADING RATES
- Chemical analysis of the test preparations at 0 and 48 hours showed measured test concentrations of less than 0.0072 mg/L, which was determined to be the limit of quantification (LOQ) for the analytical method employed.
- This result does not infer that no test item was in solution. However, any dissolved test item was present at a concentration of less than the LOQ.
- 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.

DEFINITIVE TEST - IMMOBILISATION DATA
- Cumulative immobilisation data from exposure of Daphnia magna during the definitive test are shown in Table 2 (attached).
- There was no immobilisation in 20 daphnids exposed to a 100 mg/L loading rate WAF for a period of 48 hours.
- It was considered unnecessary and unrealistic to test at loading rates in excess of 100 mg/L.

DEFINITIVE TEST - VALIDATION CRITERIA
- The test was considered to be valid given that none of the control daphnids showed immobilisation or other signs of disease or stress and that the oxygen concentration at the end of the test was ≥ 3 mg/L in the control and test vessels.

DEFINITIVE TEST - WATER QUALITY CRITERIA
- The test was considered to be valid given that none of the control daphnids showed immobilisation or other signs of disease or stress and that the oxygen concentration at the end of the test was ≥ 3 mg/L in the control and test vessels.

DEFINITIVE TEST - VORTEX DEPTH MEASUREMENTS
- The vortex depth recorded at the start and end of the mixing period was observed to be a dimple at the water surface on each occasion.

DEFINITIVE TEST - OBSERVATIONS ON TEST ITEM SOLUBILITY
- 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 floating at the surface.
- After 23 hours stirring and a 1 hour standing period, the 100 mg/L loading rate was observed to be as at the start of stirring.
- Microscopic inspection of the WAF showed no micro-dispersions or undissolved test item to be present.
- After siphoning and for the duration of the test, the 100 mg/L loading rate was observed to be a clear colourless solution.

Results with reference substance (positive control):

- Results from the positive control with potassium dichromate were within the normal range for this reference item (see Appendix 3, attached).

Validity criteria fulfilled:

yes

Conclusions:

The acute toxicity of the test item to the freshwater invertebrate Daphnia magna has been investigated 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:

GUIDELINE

A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method was designed to be compatible 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 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 Water Accomodated Fraction (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 number of immobilised Daphnia and any adverse reactions to exposure were recorded after 24 and 48 hours.

RESULTS

Chemical analysis of the test preparations at 0 and 48 hours showed measured test concentrations of less than the limit of quantification (LOQ) of the analytical method employed were obtained, which was determined to be 0.0072 mg/L. This does not infer that no test item was in solution, just that any dissolved test item was at a concentration of less than the LOQ.

Given that the toxicity cannot be attributed to a single component or a 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 unnecessary and unrealistic to test at loading rates in excess of 100 mg/L loading rate WAF.

CONCLUSION

The acute toxicity of the test item to the freshwater invertebrate Daphnia magna has been investigated 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.

Description of key information

The acute toxicity of the test item to the freshwater invertebrate Daphnia magna, investigated using a method compatible with OECD 202 and EU Method C.2, 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.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

100 mg/L

Additional information

GUIDELINE

A study was performed to assess the acute toxicity of the test item to Daphnia magna. The method was designed to be compatible 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 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 Water Accomodated Fraction (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 number of immobilised Daphnia and any adverse reactions to exposure were recorded after 24 and 48 hours.

RESULTS

Chemical analysis of the test preparations at 0 and 48 hours showed measured test concentrations of less than the limit of quantification (LOQ) of the analytical method employed were obtained, which was determined to be 0.0072 mg/L. This does not infer that no test item was in solution, just that any dissolved test item was at a concentration of less than the LOQ.

Given that the toxicity cannot be attributed to a single component or a 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 unnecessary and unrealistic to test at loading rates in excess of 100 mg/L loading rate WAF.

CONCLUSION

The acute toxicity of the test item to the freshwater invertebrate Daphnia magna has been investigated 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.