Registration Dossier

Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Currently viewing:

Administrative data

Link to relevant study record(s)

Referenceopen allclose all

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22 April 2013 - 21 June 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Test Item: SE7B Batch 2137-0 (CAS 1365345-64-7)
Analytical monitoring:
yes
Details on sampling:

- Concentrations: 0 and 10 g/L loading rate WAFs
- Sampling method: Analysis of the WAFs was carried out by Total Organic Carbon (TOC) analysis. Water samples were taken from the control and the 10 g/L loading rate WAF test group
- Sample storage conditions before analysis: Duplicate samples were taken and stored at approximately -20 °C for further analysis if necessary.
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
Due to the low aqueous solubility and complex nature of the test item, for the purposes of the test, the test item was prepared as a Water Accommodated Fraction (WAF).

Reconstituted water used for both the range-finding and definitive tests. The reconstituted water was made up as follows:

Stock Solutions
a) CaCl2.2H2O 11.76 g/L
b) MgSO4.7H2O 4.93 g/L
c) NaHCO3 2.59 g/L
d) KCl 0.23 g/L


Preparation:
An aliquot (25 mL) of each of solutions a-d was added to each liter (final volume) of deionized water with a conductivity of <5 μS cm-1. The reconstituted water had a pH of 7.8 ± 0.2 adjusted (if necessary) with NaOH or HCl and was aerated until the dissolved oxygen concentration was approximately air-saturation value.
The reconstituted water had an approximate theoretical total hardness of 250 mg/L as CaCO3.
Test organisms (species):
Daphnia magna
Details on test organisms:

The test was carried out using 1st instar Daphnia magna derived from in-house laboratory cultures.
Adult Daphnia were maintained in 150 mL glass beakers containing Elendt M7 medium in a temperature controlled room at approximately 21 °C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition 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, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing. The diet and diluent water are considered not to contain any contaminant that would affect the integrity or outcome of the study.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
The reconstituted water had an approximate theoretical total hardness of 250 mg/L as CaCO3.
Test temperature:
Temperature was maintained at 21 °C to 24 ºC throughout the test.
pH:
There were no treatment related differences for pH.
Dissolved oxygen:
There were no treatment related differences for dissolved oxygen concentration.
Salinity:
Not applicable as the study is a freshwater study
Nominal and measured concentrations:
Range finding test: nominal concentrations of 1, 10 and 100 mg/L.
Details on test conditions:
TEST SYSTEM
In the initial test 250 mL glass jars containing approximately 200 mL of test preparation were used. At the start of the test 5 daphnids were placed in each test and control vessel at random, in the test preparations. Four replicate test and control vessels were prepared. The test vessels were then covered to reduce evaporation and maintained in a temperature controlled room at 20 °C to 22 °C with a photoperiod of 16 hours light (603 to 612 lux) and 8 hours darkness with 20 minute dawn and dusk transition periods. The daphnids were not individually identified, received no food during exposure and the test vessels were not aerated.

The control group was maintained under identical conditions but not exposed to the test item.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
The test preparations were not renewed during the exposure period. Any immobilization or adverse reactions to exposure were recorded at 24 and 48 hours after the start of exposure. The criterion of effect used was that Daphnia were considered to be immobilized if they were unable to swim for approximately 15 seconds after gentle agitation.

TEST CONCENTRATIONS
- Justification for using less concentrations than requested by guideline: 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 immobilization or adverse reactions to exposure were observed.
- Range finding study:
- Test concentrations:1, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study:
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 immobilization or adverse reactions to exposure were observed.

Additional test (required for other world wide notifications):
At the request of the Sponsor, an additional "Limit test" was conducted at a single loading rate of 10 g/L to confirm that no immobilization or adverse reactions to exposure were observed at this higher nominal loading rate.

An amount of test item (20 g) was added to the surface of 2 liters of reconstituted water to give the 10 g/L loading rate. After the addition of the test item, the reconstituted 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 after 23 hours and the mixture allowed to stand for 1 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 WAF 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 10 g/L loading rate WAF.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 10 g/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
10 g/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
Range finding test:
No immobilization was observed at 1.0, 10 and 100 mg/L loading rate WAF.

Initial test:
No immobilization was observed at 100 mg/L loading rate WAF.

At the request of the Sponsor, a single loading rate of four replicates, of 10 g/L, using a stirring period of 23 hours followed by a 1-Hour standing period, was selected for the definitive test. This experimental design conforms to a "Limit test" to confirm that no immobilization or adverse reactions to exposure were observed.

Definitive test:
There was no immobilization in 20 daphnids exposed to a 10 g/L loading rate WAF for a period of 48 hours. Inspection of the immobilization data gave the following results:

Time (h) EL 50 (g/L loading rate WAF)
24 > 10
48 > 10
Results with reference substance (positive control):
The results from the positive control with potassium dichromate were within the normal range for this reference item.

Based on this information, a single loading rate of four replicates, of 100 mg/L, using a stirring period of 23 hours followed by a 1-Hour standing period, was selected for the initial test. This experimental design conforms to a "Limit test" to confirm that no immobilization or adverse reactions to exposure were observed.

Positive control:

Analysis of the immobilization data by the trimmed Spearman-Karber method (Hamilton et al 19771) at 24 and 48 hours based on the nominal test concentrations gave the following results. The results from the positive control with potassium dichromate were within the normal range for this reference item , as can be seen below:

 Time point (hours)

 EC50 (mg/L)

 95% confidence limits

 NOEC

 LOEC

 24

0.91 - 1.2 

0.56 

1.0 

 48

0.71 

0.65 - 0.76 

0.32 

0.56 

 

Validity criteria fulfilled:
yes
Conclusions:
The acute toxicity of the test item to the freshwater invertebrate Daphnia magna has been investigated and gave a 48-Hour EL*50 value of greater than 10 g/L loading rate WAF. The No Observed Effect Loading rate was 10 g/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 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 aqueous solubility and complex nature of the test item, for the purposes of the test, the test item was prepared as a Water Accommodated Fraction (WAF).

Following a preliminary range-finding test and an initial test, at the request of the Sponsor, twenty daphnids (4 replicates of 5 animals) were exposed to a Water Accommodated Fraction (WAF) of the test item at a single nominal loading rate of 10 g/L for 48 hours at a temperature of 21 °C to 24 °C under static test conditions. The number of immobilized Daphnia and any adverse reactions to exposure were recorded after 24 and 48 hours.

Results:

Exposure of Daphnia magna to the test item gave EL*50 values of greater than 10 g/L loading rate WAF. The No Observed Effect Loading Rate was 10 g/L loading rate WAF.

Total Organic Carbon (TOC) analysis of the test preparations at 0 hours showed a measured concentration of 1.69 mg C/L. Analysis of the old media at 48 hours showed a measured concentration of less than the control.

The dissolved test item may have been one or several components of the test item. Given that the toxicity of the test item 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.

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
disregarded due to major methodological deficiencies
Study period:
26 July-19 Aug
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
significant methodological deficiencies
Qualifier:
according to
Guideline:
other: ISO 14669 (1999) Water Quality - Determination of acute lethal toxicity to marine copepods (Copepoda; Crustacea)
Version / remarks:
ISO 14669 (1999) Water Quality -
Determination of acute lethal toxicity to
marine copepods (Copepoda; Crustacea)
Deviations:
yes
Remarks:
See note below
Principles of method if other than guideline:
The test was conducted in accordance with the study plan except for the following deviation:

The control mortality of 12% exceeds the guideline figure of 10%. This was deemed acceptable as it is
only slightly higher and the reference chemical mortalities appeared normal.
The test met all other relevant validity criteria.
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Test material: LubriGreen BioSynthetic SE6
Behaviour in seawater: Poorly Soluble
Preparation method WAF’s

Form: Liquid
Colour: Yellow/Amber
Density: 0.8906g/cm3 @ 20°C
Viscosity: Slight
pH: TSW=8.12, (1000 mg.l-1 stock)
Aqueous solubility: Insoluble / Floating and or settled particles at 1000 mg.l-1 in sea water after 20 hours stirring
Analytical monitoring:
not specified
Test organisms (species):
other aquatic crustacea: Acartia tonsa (Marine copepod)
Details on test organisms:
The Initial culture, CCAP 1077/5 was received from Dunstaffnage Marine Laboratory in 1995.
The test organisms were cultured in accordance with SOP 501 to provide age standardised cohorts.
Organisms used for testing were between 17 and 25 days old at test commencement; stage 5 in their life
cycle. The culture system was maintained with flowing 0.45 µm filtered ultra violet treated sea water,
supplied by pump from Scapa Flow in Orkney. The cultures were maintained on a mixed algal diet
comprising of between 2 and 4 species.
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
Temperature range 20 ± 2 °C 20.7 – 21.0
pH:
pH at 0h of dilution water: 8.05
pH at 0h of the test material stocks: 8.11 – 8.18
Dissolved oxygen:
Dissolved O2 range: 94 – 98%
Salinity:
Salinity at 0h of dilution water: 36%
Nominal and measured concentrations:
Nominal concentrations, Definitive Test: 1000, 1800, 3200, 5600, 10000 mg/L
Details on test conditions:
Test methods were consistent with ISO 14669 (1999) Water Quality: Determination of acute lethal toxicity
to marine adult copepods (Copepoda; Crustacea).
ISO 5667-16 (1998) Water Quality Sampling – Guidance on biotesting samples.
The method estimates the effect of chemicals on the mortality/immobility on adult copepods over a period
of 48h at a temperature of 20±2 °C.

Tests were conducted in 100 ml capacity borosilicate glass crystallising dishes, each containing 50 ml of
test medium covered with soda glass watch covers.
Rangefinding tests were conducted at 1000, 100, 10 and 1 mg.l-1, plus four control vessels. Definitive test
concentrations depend on the magnitude of response evident in the Rangefinding test and employ five
concentrations, plus four control vessels.
In the Rangefinding tests, five animals were exposed per concentration and in definitive tests ten animals
were exposed per replicate (20 per treatment). Test animals were transferred to test vessels using glass
pipettes of three to five millimetres in diameter (internal) and with fire polished rims.
Measurement of water quality (dissolved oxygen, pH and temperature) are carried out in one replicate at
each concentration at 0h and 48h. Observation of mortalities are carried out at 24h and 48h. An animal
is regarded as immobile if after gentle stimulation no movement is visible after 10 seconds.
A salinity measurement is carried out in the control medium at 0h.
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
> 10 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Key result
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
> 10 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
LubriGreen BioSynthetic SE6 exhibited a 48h LC50 value of greater than 10000mg.l-1 (Water
Accommodated Fractions (WAFs)) to the marine copepod Acartia tonsa in the aqueous phase.
The result is based on nominal concentrations and was calculated by Linear Interpolation within the
Toxcalc suite of statistical analysis.
It was not possible to calculate the 95% confidence limits due to no effect at the top test concentration.
There were no interferences in this test.
Reported statistics and error estimates:
The result is based on nominal concentrations and was calculated by Linear Interpolation within the Toxcalc suite of statistical analysis.
Validity criteria fulfilled:
yes
Conclusions:
LubriGreen BioSynthetic SE6 exhibited a 48h LC50 value of greater than 10000mg.l-1 (Water Accommodated Fractions (WAFs)) to the marine copepod Acartia tonsa in the aqueous phase.
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read-Across Justification is attached below.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 10 g/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
10 g/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Conclusions:
Read-Across is claimed between BT4 (target) and SE7B and SE6B (Sources), according to the justification attached to the target record, and based on structural and physical/chemical similarities.

Analogue diesters (SE7B, SE6B) and BT4 contain the same functional groups, i.e the ester group adjacent to the ethylhexane side chain, and the ester group at the opposite end of the molecules. The carbon range in the main backbone of the molecules is all the same (C18) though the acetate moiety is attached at slightly different positions (C12 for BT4, C9/10 for the analogue diesters). The analogue diesters have the additional alkane chain attached to the acetate cap. The alkane chains themselves are not typically considered to be functional groups, per se, as they are relatively inactive biologically. Thus the parent molecules BT4 and the analogue diesters are similar enough to allow for read across in that there are no differences with respect to functional groups, and their only real difference is number of, and length of, saturated hydrocarbon chains.

The Water Solubility of SE7B was measured as 0.778 mg/L @ 30 °C, whereas BT4 is found to be not stable in water but forms micelle structures resulting in suspension.
SE6B is characterised as poorly soluble in seawater (Floating and or settled particles at 1000 mg/L in sea water after 20 hours stirring)

Hydrolysis of BT4 would yield acetic acid plus 12-hydroxystearic acid (C18), versus either lauric acid (C12) or the coconut oil fatty acid mixture (C8-18) for SE6B and SE7B, respectively.
These substances are not expected to persist in the environment, consistent with the hazard assessment presented in the OECD SIDS (2009) for the category “Aliphatic Acids Category” where aliphatic fatty acids with a carbon chain length in the range of C8 – C22 were judged to be readily biodegradable.

The acute toxicity of the test item SE7B to the freshwater invertebrate Daphnia magna has been investigated and gave a 48-Hour EL*50 value of greater than 10 g/L loading rate WAF. The No Observed Effect Loading rate was 10 g/L loading rate WAF.
The test item SE6 exhibited a 48h LC50 value of greater than 10000mg/L (Water Accommodated Fractions (WAFs)) to the marine copepod Acartia tonsa in the aqueous phase.
Considering the structural similarity and low soubility of BT4, these results are also considered relevant for the read-across target BT4.

Description of key information

Key value for chemical safety assessment

Additional information

Read-Across is claimed between BT4 (target) and SE7B and SE6B (Sources), according to the justification attached to the target record, and based on structural and physical/chemical similarities.

Analogue diesters (SE7B, SE6B) and BT4 contain the same functional groups, i.e the ester group adjacent to the ethylhexane side chain, and the ester group at the opposite end of the molecules. The carbon range in the main backbone of the molecules is all the same (C18) though the acetate moiety is attached at slightly different positions (C12 for BT4, C9/10 for the analogue diesters). The analogue diesters have the additional alkane chain attached to the acetate cap. The alkane chains themselves are not typically considered to be functional groups, per se, as they are relatively inactive biologically. Thus the parent molecules BT4 and the analogue diesters are similar enough to allow for read across in that there are no differences with respect to functional groups, and their only real difference is number of, and length of, saturated hydrocarbon chains.

The Water Solubility of SE7B was measured as 0.778 mg/L @ 30 °C, whereas BT4 is found to be not stable in water but forms micelle structures resulting in suspension.

SE6B is characterised as poorly soluble in seawater (Floating and or settled particles at 1000 mg/L in sea water after 20 hours stirring)

Hydrolysis of BT4 would yield acetic acid plus 12-hydroxystearic acid (C18), versus either lauric acid (C12) or the coconut oil fatty acid mixture (C8-18) for SE6B and SE7B, respectively.

These substances are not expected to persist in the environment, consistent with the hazard assessment presented in the OECD SIDS (2009) for the category “Aliphatic Acids Category” where aliphatic fatty acids with a carbon chain length in the range of C8 – C22 were judged to be readily biodegradable.

The acute toxicity of the test item SE7B to the freshwater invertebrate Daphnia magna has been investigated and gave a 48-Hour EL*50 value of greater than 10 g/L loading rate WAF. The No Observed Effect Loading rate was 10 g/L loading rate WAF.

The test item SE6 exhibited a 48h LC50 value of greater than 10000mg/L (Water Accommodated Fractions (WAFs)) to the marine copepod Acartia tonsa in the aqueous phase.

Considering the structural similarity and low soubility of BT4, these results are also considered relevant for the read-across target BT4.