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Short-term toxicity to aquatic invertebrates

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Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Principles of method if other than guideline:
The 48 h acute toxicity of water accomodated fractions and dispersions of the test substance to the marine copepod Acartia tonsa were tested under static conditions. After 24 and 48 h of exposure, the number of living and dead organisms were counted. Dissolved oxygen concentrations, pH, temperature and salinity were measured at various time points during the tests.

GLP compliance:
no
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
- In a first series of studies, Water Accommodated Fraction (WAF) methodology was used. The test medium was natural seawater.

- The other studies were conducted using dispersions produced in Tween 80.

WATER ACCOMODATED FRACTIONS
Water accomodated fractions were prepared by adding the test substance (generally 2,000 mg) to 2,000 mL of dilution water to achieve the desired loading rates. The media were then stirred on a magnetic stirrer for ca. 16 h and allowed to settle for 4 h to allow phase separation before use. From these media, the sublayers were tapped off under the meniscus and used for the test. The first 200 mL were discarded. The control media were prepared similarly. For the acute tests, about 10 mL of each WAF and the control media were placed in each of the ten 20 mL glass vials (10 test vials, 10 controls). One test animal was transferred to each test vessel. The test vessels were then covered. Dilution natural seawater was used as a control medium.

DISPERSIONS WITH TWEEN 80
Weighed quantities of test substance (generally 2,000 mg) were added to 2,000 mL of water. A quantity of dispersant Tween 80 was added (generally 200 mg). The media were stirred until a homogeneous dispersion was achieved.
Test organisms (species):
other aquatic crustacea: Acartia tonsa
Details on test organisms:
- Conditions: Cultured under standard conditions according to the relevant standard operating procedures
- Age: 6-8 d old at start of test
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20.1 - 21.3
pH:
7.5 - 8.1
Dissolved oxygen:
6.2 - 9.9 mg/L
Salinity:
29.1 - 29.3
Nominal and measured concentrations:
CRUDE PALM OIL
WAF
- 0 and 1,000 mg/L
- 0, 320, 560, 1,000, 1,800 and 3,200 mg/L
Dispersion
- 0, 100, 180, 320, 560, 1,000 mg/L

REFINED PALM OIL
WAF
- 0 and 1,000 mg/L
Dispersion
- 0, 100, 180, 320, 560, 1,000 mg/L
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
other: LC50/LL50
Effect conc.:
> 3 200 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Crude palm oil, water accomodated fraction
Key result
Duration:
48 h
Dose descriptor:
other: LC50/LL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Crude palm oil, water accomodated fraction
Key result
Duration:
48 h
Dose descriptor:
other: LC50/LL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Crude palm oil, dispersion in Tween 80
Key result
Duration:
48 h
Dose descriptor:
other: LC50/LL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Refined palm oil, water accomodated fraction
Key result
Duration:
48 h
Dose descriptor:
other: LC50/LL50
Effect conc.:
ca. 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: Refined palm oil, dispersion in Tween 80

 Test substance  Dosing method  Survival (out of 10 animals) at different test concentration (mg/L)  Dispersing agent 48 h LC50/LL50 (mg/L) 
     0 100  180  320  560  1000  1800  3200     
 Crude palm oil WAF   9 > 1000 
  WAF   9 10  > 3200 
  Dispersion   5  10  10  9 Tween 80  > 1000 
 Refined palm oil  WAF  9 - 10  > 1000 
   Dispersion  10 10  10  7*  Tween 80  ca. 1000 

* Physical hindering

Validity criteria fulfilled:
not specified
Conclusions:
Based on the results obtained, the 48 h acute toxicity (LC50/LL50) of crude palm oil to Acartia tonsa was considered to be >3,200 mg/L for WAFs and >1,000 mg/L for dispersions. The 48 h acute toxicity (LC50/LL50) of refined palm oil to Acartia tonsa were considered to be >1,000 mg/L for WAFs and ca. 1,000 mg/L for dispersions.

The 48 h acute toxicity (LC50/LL50) of water accomodated fractions or Tween 80 dispersions of refined palm oil to Acartia tonsa were >1,000 mg/L and ca. 1,000 mg/L respectively.
Executive summary:

A study was conducted to evaluate the acute toxicity of water accommodated fractions (WAFs) or dispersions (in Tween 80) of the constituent glycerides, C16 -18, C18 -unsatd. (crude and refined palm oil) to Acartia tonsa under static conditions. Acartia tonsa was exposed to crude and refined palm oil at concentrations up to either 3,200 (crude oil) or 1,000 (refined oil) mg/L for 48 h. Based on the results obtained, the 48 h acute toxicity (LC50/LL50) of crude palm oil to Acartia tonsa was considered to be >3,200 mg/L (WAFs) and >1,000 mg/L (dispersions). The 48 h acute toxicity (LC50/LL50) of refined palm oil to Acartia tonsa were considered to be >1,000 mg/L (WAFs) and ca. 1,000 mg/L (dispersions) (Bowmer, 1998).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
March 2002 to April 2002
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: ISO 14669, ISO 10253 and GESAMP protocol for preparation of poorly soluble complex mixtures
Deviations:
no
GLP compliance:
not specified
Analytical monitoring:
yes
Details on sampling:
Samples were taken for carbon analysis at 0, 24 and 48 h.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water accommodated fraction (WAF) methodology
- Preparation: WAFs were prepared by adding test substance to the test medium (sterilised natural seawater) to give loading rates of 10, 100 and 1,000 mg/L. In order to mix the test substance, which was solid at room temperature, the sample was microwaved on full power for approximately 2 min until it became liquid. The contents of the WAF vessels were then stirred using a magnetic stirrer, at 300 rpm, for 24 h, in a controlled environment room set at 20 ± 2°C and then left to settle for a minimum of 2 h (range 2 – 4 h), to allow the undissolved material to separate out. The aqueous phases - the WAFs – were then drawn off via the tap for use in the tests. Control media was sterilised seawater.

Test organisms (species):
other aquatic crustacea: Acartia tonsa
Details on test organisms:
TEST ORGANISMS
- Age: 12-14 d old.
- Source: Obtained from Guernsey Sea Farms, Vale, Guernsey

ENVIRONMENTAL CONDITIONS
- Temperature: 20-22°C
- Light: 16 h light/8 h dark cycle
- Diet: Fed ad libitum with mixed marine algae
Test type:
static
Water media type:
saltwater
Limit test:
yes
Total exposure duration:
48 h
Test temperature:
20.5-20.7°C
pH:
8.2 - 8.1
Dissolved oxygen:
6.1 - 6.2 mg/L
Salinity:
32-33%
Nominal and measured concentrations:
Nominal: 0 and 1,000 mg/L
Details on test conditions:
-All media and glassware were autoclaved at 121°C for 15 min before use.
-A preliminary test was made to decide stirring speed. A speed of 300 rpm was selected.
-2 vessels where used during the test, containing 2 L of media and known volume of test item.
-The tests were conducted in 150 mL glass crystal dishes containing 100 mL of either control or test solution. A. tonsa were added using modified glass Pasteur pipettes. A. tonsa were considered to be immobilised if they failed to respond to touching with the end of a sealed glass pipette. At the end, a few drops of formalin were added. Immobilisation, carbon concentration and growth were estimated.
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
% immobilised after exposure:
24 h: 3%
48 h: 3%
Results with reference substance (positive control):
% immobilised after exposure:
24 h: 22%
48 h: 53%
Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the 48 h EL50 of the constituent was determined to be >1,000 mg/L.
Executive summary:

A study was conducted to evaluate the acute toxicity of the constituent glycerides C16 -18, C18 -unsatd (palm oil) to Acartia tonsa under static conditions. The test method was based on guidelines issued by ISO 14669, ISO 10253 and GESAMP. Acartia tonsa were exposed to water accomodated fractions prepared from the constituent at concentrations of 0 and 1,000 mg/L and immobility was determined at 0, 24 and 48 h. 3,5 -dichlorophenol was used as a reference substance. Under the test conditions, the 48 h EL50 of the constituent was determined to be >1000 mg/L (Worden and Sherren, 2002b).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
March 2002 to April 2002
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: ISO 14669, ISO 10253 and GESAMP guidelines
Deviations:
not specified
GLP compliance:
not specified
Analytical monitoring:
yes
Details on sampling:
Samples were taken for carbon analysis at 0, 24 and 48 h.
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficultly soluble test substances)
- Method: Water accommodated fraction (WAF) methodology
- Preparation: WAFs were prepared by adding test substance to the test medium (sterilised natural seawater) to give loading rates of 10, 100 and 1,000 mg/L. The contents of the WAF vessels were then stirred using a magnetic stirrer, at 300 rpm, for 24 h, in a controlled environment room set at 20 ± 2°C and then left to settle for a minimum of 2 h (range 2 – 4 h), to allow the undissolved material to separate out. The aqueous phases - the WAFs – were then drawn off via the tap for use in the tests. Control media was sterilised seawater.
Test organisms (species):
other aquatic crustacea: Acartia tonsa
Details on test organisms:
TEST ORGANISMS
- Age: 12-14 d old.
- Source: Obtained from Guernsey Sea Farms, Vale, Guernsey

ENVIRONMENTAL CONDITIONS
- Temperature: 20-22°C
- Light: 16 h light/8 h dark cycle
- Diet: Fed ad libitum with mixed marine algae
Test type:
static
Water media type:
saltwater
Limit test:
yes
Total exposure duration:
48 h
Test temperature:
20-22°C
pH:
8.5-8.2
Dissolved oxygen:
6.1-6.2 mg/L
Salinity:
32-33%
Nominal and measured concentrations:
Nominal: 0 and 1,000 mg/L
Details on test conditions:
- All media and glassware were autoclaved at 121°C for 15 min before use.
- A preliminary test was made to decide stirring speed. A speed of 300 rpm was selected.
- Two vessels where used during the test, containing 2 L of media and known volume of test item. The test was conducted on 150 mL glass crystallising dishes containing 100 mL of either control or test solution.
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
% immobilised after exposure:
24 h: 0%
48 h: 3%
Results with reference substance (positive control):
% immobilised after exposure:
24 h: 22%
48 h: 53%
Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the 48 h EL50 of the constituent was determined to be >1,000 mg/L.

Executive summary:

A study was conducted to evaluate the acute toxicity of the constituent glycerides C16 -18, C18 -unsatd. (soybean oil) to Acartia tonsa under static conditions. The test method was based on the guidelines ISO 14669, ISO 10253 and GESAMP. Acartia tonsa were exposed to water accomodated fractions prepared from the constituent at concentrations of 0 and 1,000 mg/L and immobility was determined at 0, 24 and 48 h. 3,5 -dichlorophenol was used as a reference substance. Under the test conditions, the 48 h EL50 of the constituent was determined to be >1,000 mg/L (Worden and Sherren, 2002a).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
January 12 to 17, 2004
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Reason / purpose for cross-reference:
reference to other study
Qualifier:
according to guideline
Guideline:
other: ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea)
Deviations:
no
GLP compliance:
not specified
Analytical monitoring:
not specified
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water accommodated fraction (WAF) methodology
- Preparation: WAFs were prepared by adding test substance to the test medium (sterilised natural seawater) to give loading rates of 10, 100 and 1000 mg/L. In order to mix the test substance, which was solid at room temperature, the sample was microwaved on full power for approximately 2 min until it became liquid. The contents of the WAF vessels were then stirred using a magnetic stirrer, at 300 rpm, for 24 h, in a controlled environment room set at 20 ± 2°C and then left to settle for a minimum of 2 h (range 2 – 4 h), to allow the undissolved material to separate out. The aqueous phases (the WAFs) were then drawn off via the tap for use in the tests. Control media was sterilised seawater.
Test organisms (species):
other: Acartia tonsa
Details on test organisms:
TEST ORGANISM
- Source: Guernsey Sea Farms, Vale, Guernsey
- Age at study initiation (mean and range, SD): 9-12 d
- Feeding
- Food type: Mixed marine algae supplied with the organisms
- Amount, frequency: Ad libitum


ACCLIMATION
- Acclimation period: 1 d
- Acclimation conditions (same as test or not): Temperature: 18-20°C
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20±2°C
pH:
8.1-8.6
Dissolved oxygen:
6.4-8.3 mg/L
Salinity:
33-34%
Nominal and measured concentrations:
Nominal concentration: 0, 10, 100 & 1,000 mg/L
- 1,000 mg/L was equivalent to 4 mg total carbon/L
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: Glass crystallising dishes, 150 mL
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): Not renewed
- No. of organisms per vessel: 32 (control), 35 (control, filtered), 31-33 (test material) and 33 (positive control)
- No. of vessels per concentration (replicates): Three
- No. of vessels per control (replicates): Three
- No. of vessels per vehicle control (replicates): Three


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Natural seawater, collected off the beach at Kinmel Bay to the west of Rhyl, Flintshire, N. Wales
- Total organic carbon: 28 & 29 mg/L for filtered and unfiltered controls; 25, 27 & 31 mg/L for test substance at 10, 100 & 1,000 mg/L, respectively


OTHER TEST CONDITIONS
- Photoperiod: 16 h light/8 h dark cycle
- Temperature: 20±2°C
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol; 1 mg/L
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
>= 100 - < 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
dissolved
Basis for effect:
mobility
Details on results:
Observation of immobilised organisms under a low power, binocular microscope did not reveal any adverse signs and is therefore concluded that the effects seen were due to toxicity.
Results with reference substance (positive control):
% immobilisation: 21 (after 24 h); 76 (after 48 h)

Preliminary study: The results from the study indicated that the test material was toxic to Acartia tonsa at a loading rate of 1000 mg/L.

Table 1: Immobilisation ofA. tonsaexposed to WAF of coconut oil and 1 mg/L solution of reference compound 3,5-DCP

Test material Loading rate (mg/L) Number of A. tonsa exposed Cumulative Number (N) and Percentage (P) of A. tonsa immobilised after exposure period of:
24 h 48 h
N P N P
Control   32 0 0 0 0
Control (filtered) 35 0 0 2 6
 Coconut oil 10 31 0 0 3 10
  100 32 0 0 0 0
1000 33 2 6 17 52
  3,5-DCP  1 33 21  25  76 
Validity criteria fulfilled:
yes
Conclusions:
Based on the results, the 48 h EL50 of the constituent was determined to be between 100 and 1,000 mg/L.
Executive summary:

A study was conducted to evaluate the acute toxicity of the constituent "Glycerides C8 -18, C18 -unsatd (coconut oil) to Acartia tonsa under static conditions. The method followed the guideline ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea). Acartia tonsa were exposed to water accommodated fractions prepared from the constituent at concentrations ranging from 10-1,000 mg/L for 48 h and immobility was determined at 24 and 48 h. Based on the results, the 48 h EL50 of the constituent was determined to be between 100 and 1,000 mg/L (Worden and Sherren, 2004b).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea)
Deviations:
no
GLP compliance:
not specified
Analytical monitoring:
not specified
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water accommodated fraction (WAF) methodology
- Preparation: WAFs were prepared by adding test substance to the test medium (sterilised natural seawater) to give loading rates of 10, 100 and 1000 mg/L. In order to mix the test substance, which was solid at room temperature, the sample was microwaved on full power for approximately 2 min until it became liquid. The contents of the WAF vessels were then stirred using a magnetic stirrer, at 300 rpm, for 24 h, in a controlled environment room set at 20 ± 2°C and then left to settle for a minimum of 2 h (range 2 – 4 h), to allow the undissolved material to separate out. The aqueous phases - the WAFs – were then drawn off via the tap for use in the tests. Control media was sterilised seawater.
Test organisms (species):
other: Acartia tonsa
Details on test organisms:
TEST ORGANISM
- Source: Guernsey Sea Farms, Vale, Guernsey
- Age at study initiation (mean and range, SD): 13-15 d
- Food type: Mixed marine algae supplied with the organisms
- Amount, frequency: Ad libitum
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20 ± 2°C

pH:
7.7-8.3 (at time 0), 8.1-8.2 (after 48 h)

Dissolved oxygen:
8.6-8.9 mg/L (at time 0), 7.6-8.9 (after 48 h)

8.6-8.9 mg/L (at time 0), 7.6-8.9 (after 48 h)

Salinity:
32-33%
Nominal and measured concentrations:
Nominal concentration: 10, 100 and 1,000 mg/L


Details on test conditions:
TEST SYSTEM
- Material, size, headspace, fill volume: Glass crystallising dishes, 150 mL
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): Not renewed
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): Three
- No. of vessels per control (replicates): Three
- No. of vessels per vehicle control (replicates): Three


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Natural seawater, collected off the beach at Kinmel Bay to the west of Rhyl, Flintshire, N. Wales
- Total organic carbon (mean of three replicates at time 0): 28.1, 29.1 and 33.2 mg/L for 10, 100 and 1000 mg/L loading rate, respectively


OTHER TEST CONDITIONS
- Photoperiod: 16 h light/8 h dark cycle
- Temperature: 20 ± 2°C
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol; 1 mg/L
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
The WAF prepared from palm kernel oil was found to be non-toxic to A. tonsa, with 48 h EL50 values exceeding 1,000 mg/L, the highest loading rate tested.
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Other: The percentage of A. tonsa that were immobilised when exposed to 1 mg/L 3,5-DCP was 58% after 48 h.

Table 1: Immobilisation of A. tonsa exposed to WAF of palm kernel oil and 1 mg/L solution of reference compound 3,5-DCP

Test material Loading rate (mg/L) Number of A. tonsa exposed Cumulative Number (N) and Percentage (P) of A. tonsa immobilised after exposure period of:
24 h 48 h
N P N P
Control   35 1 3 4 11
Palm kernel oil 10 35 0 0 6 17
  100 34 1 3 3 9
  1000 32 2 6 5 16
3,5-DCP 1 33 4 12 19 58
Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the 48 h EL50 of the test substance was determined to be >1,000 mg/L.


Executive summary:

A study was conducted to evaluate the acute toxicity of the constituent C8 -18, C18 -unsatd. (palm kernel oil) to Acartia tonsa under static conditions. The method followed the guideline ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea). Acartia tonsa were exposed to water accomodated fractions prepared from the constituent at concentrations ranging from 10 to 1,000 mg/L for 48 h and immobility was determined at 24 and 48 h. Under the test conditions, the 48 h EL50 of the constituent was determined to be >1,000 mg/L (Worden and Sherren, 2004d).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea)
Deviations:
no
GLP compliance:
not specified
Analytical monitoring:
not specified
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficultly soluble test substances)
- Method: Water accommodated fraction (WAF) methodology
- Preparation: WAFs were prepared by adding test substance to the test medium (sterilised natural seawater) to give loading rates of 10, 100 and 1000 mg/L. In order to mix the test substance, which was solid at room temperature, the sample was microwaved on full power for approximately 2 min until it became liquid. The contents of the WAF vessels were then stirred using a magnetic stirrer, at 300 rpm, for 24 h, in a controlled environment room set at 20 ± 2°C and then left to settle for a minimum of 2 h (range 2 – 4 h), to allow the undissolved material to separate out. The aqueous phases - the WAFs – were then drawn off via the tap for use in the tests. Control media was sterilised seawater.
Test organisms (species):
other: Acartia tonsa
Details on test organisms:
TEST ORGANISM
- Source: Guernsey Sea Farms, Vale, Guernsey
- Age at study initiation (mean and range, SD): 13-15 d
- Food type: Mixed marine algae supplied with the organisms
- Amount, frequency: Ad libitum
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20 ± 2°C
pH:
7.7-8.3 (at time 0), 8.1-8.2 (after 48 h)
Dissolved oxygen:
8.6-8.9 mg/L (at time 0), 7.6-8.9 (after 48 h)
Salinity:
32-33%

Nominal and measured concentrations:
Nominal concentration: 10, 100 and 1,000 mg/L


Details on test conditions:
TEST SYSTEM
- Material, size, headspace, fill volume: Glass crystallising dishes, 150 mL
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): Not renewed
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): Three
- No. of vessels per control (replicates): Three
- No. of vessels per vehicle control (replicates): Three


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Natural seawater, collected off the beach at Kinmel Bay to the west of Rhyl, Flintshire, N. Wales
- Total organic carbon (mean of three replicates at time 0): 28.4, 29.7 and 33.9 mg/L for 10, 100 and 1000 mg/L loading rate, respectively


OTHER TEST CONDITIONS
- Photoperiod: 16 h light/8 h dark cycle
- Temperature: 20 ± 2°C
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol; 1 mg/L
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
The WAF prepared from palm kernel olein was found to be non-toxic to A. tonsa, with 48 h EL50 values exceeding 1,000 mg/L, the highest loading rate tested.
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Other: The percentage of A. tonsa that were immobilised when exposed to 1 mg/L 3,5-DCP was 58% after 48 h.

Table 1: Immobilisation of A. tonsa exposed to WAF of palm kernel olein and 1 mg/L solution of reference compound 3,5-DCP

Test material Loading rate (mg/L) Number of A. tonsa exposed Cumulative Number (N) and Percentage (P) of A. tonsa immobilised after exposure period of:
24 h 48 h
N P N P
Control   35 1 3 4 11
Palm kernel olein 10 31 1 3 7 23
  100 34 1 3 2 6
  1000 35 0 0 2 6
3,5-DCP 1 33 4 12 19 58
Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the 48 h EL50 of the constituent was determined to be >1,000 mg/L.


Executive summary:

A study was conducted to evaluate the acute toxicity of the constituent C8 -18, C18 -unsatd. (palm kernel olein) to Acartia tonsa under static conditions. The method followed the guideline ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea).

Acartia tonsa were exposed to water accomodated fractions prepared from the constituent at concentrations ranging from 10 to 1,000 mg/L for 48 h and immobility was determined at 24 and 48 h. Under the test conditions, the 48 h EL50 of the constituent was determined to be >1,000 mg/L (Worden and Sherren, 2004c).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
other: ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea)
Deviations:
no
GLP compliance:
not specified
Analytical monitoring:
not specified
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water accommodated fraction (WAF) methodology
- Preparation: WAFs were prepared by adding test material to the test medium (sterilised natural seawater) to give loading rates of 10, 100 and 1,000 mg/L. In order to mix the test substance (solid at room temperature), the sample was microwaved on full power for approximately 2 min until it became liquid. The contents of the WAF vessels were then stirred using a magnetic stirrer, at 300 rpm, for 24 h, in a controlled environment room set at 20 ± 2°C and then left to settle for a minimum of 2 h (range 2 – 4 h), to allow the undissolved material to separate out. The aqueous phases - the WAFs – were then drawn off via the tap for use in the tests. Control media was sterilised seawater.
Test organisms (species):
other: Acartia tonsa
Details on test organisms:
TEST ORGANISM
- Source: Guernsey Sea Farms, Vale, Guernsey
- Age at study initiation (mean and range, SD): 13-15 d
- Food type: Mixed marine algae supplied with the organisms
- Amount, frequency: Ad libitum
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20 ± 2°C
pH:
7.7-8.3 (at time 0), 8.1-8.2 (after 48 h)
Dissolved oxygen:
8.6-8.9 mg/L (at time 0), 7.6-8.9 (after 48 h)
Salinity:
32-33%
Nominal and measured concentrations:
Nominal concentrations: 0, 10, 100 & 1,000 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: Glass crystallising dishes, 150 mL
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): Not renewed
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): Three
- No. of vessels per control (replicates): Three
- No. of vessels per vehicle control (replicates): Three


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Natural seawater, collected off the beach at Kinmel Bay to the west of Rhyl, Flintshire, N. Wales
- Total organic carbon (mean of three replicates at time 0): 28.3, 28.6 and 29.2 mg/L for 10, 100 and 1000 mg/L loading rate, respectively


OTHER TEST CONDITIONS
- Photoperiod: 16 h light/8 h dark cycle
- Temperature: 20 ± 2°C
Reference substance (positive control):
yes
Remarks:
3,5-dichlorophenol; 1 mg/L
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Results with reference substance (positive control):
- Results with reference substance valid: yes
- Other: The percentage of A. tonsa that were immobilised when exposed to 1 mg/L 3,5-DCP was 58% after 48 h.

Table 1: Immobilisation of A. tonsa exposed to WAF of palm kernel stearin and 1 mg/L solution of reference compound 3,5-DCP

Test material Loading rate (mg/L) Number of A. tonsa exposed Cumulative Number (N) and Percentage (P) of A. tonsa immobilised after exposure period of:
24 h 48 h
N P N P
Control   35 1 3 4 11
Palm kernel stearin 10 33 2 6 3 9
  100 36 1 3 5 14
  1000 39 5 13 10 26
3,5-DCP 1 33 4 12 19 58
Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the 48 h EL50 of the constituent was determined to be > 1,000 mg/L.
Executive summary:

A study was conducted to evaluate the acute toxicity of the constituent palm kernel stearin to Acartia tonsa under static conditions. The procedures followed ISO 14669:1999 "Water quality" Part 5: Biological methods Section 5.24: Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea). Acartia tonsa were exposed to water accommodated fractions prepared from the constituent at concentrations ranging from 10 to 1,000 mg/L for 48 h and immobility was determined at 24 and 48 h. Under the test conditions, the 48 h EL50 of the constituent was determined to be > 1,000 mg/L (Worden and Sherren, 2004a).

 

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23 March 2010 to 21 July 2010
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Series on testing and assessment No. 23: Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Concentrations:
- Sampling method: The concentrations of the test substance were analysed in the duplicate test media samples from both sampling times (0 and 48 h). From the control samples only one of the duplicate samples was analysed from each of both sampling times.
- Sample storage conditions before analysis: All samples were stored in a freezer (≤ - 10°C), protected from light until analysis was performed

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 19.9, 23, 10.3, 9.4 and 4.3 mg of the test substance were added directly to 1990, 5000, 4900, 9900 and 10000 mL test water, respectively, and carefully stirred for about 24 h at room temperature in the dark to dissolve as much of the test substance as possible. After cessation of mixing and a following period of settling of about 1 h and 45 min to allow phase separation, the aqueous phases, i.e. the water accommodated fractions, were drawn off carefully and used as the test media of the nominal test concentrations.
- Controls: Yes
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Elendt "M4" reconstituted water
- Evidence of undissolved material (e.g. precipitate, surface film, etc): The appearance of the test substance in the test media was observed at the start of the test and after 24 and 48 h test duration in the only test concentration. No remarkable observations were made.


Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain: Clone 5
- Source: In house laboratory culture (i.e. IBACON testing laboratory)
- Age at study initiation (mean and range, SD): 6.25 to 21.5 h old
- Method of breeding: The cultivation of the parental Daphnia was performed in reconstituted water of a similar quality with regards to pH, the constituent salts and total hardness as the test water used in the test. The test organisms were not first brood progeny.
- Food type: Green algae (Desmodesmus subspicatus) freshly grown in the lab
-Frequency: Daily

ACCLIMATION
- Was not necessary, since the test was performed in the same medium as the culturing
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
250 mg/L as CaCO3
Test temperature:
20 - 22°C
pH:
7.7- 7.9
Dissolved oxygen:
8.0 - 9.0 mg/L
Nominal and measured concentrations:
Nominal: 0, 0.43, 0.95, 2.1, 4.6 and 10 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass beakers
- Type (delete if not applicable): Closed with lids
- Material, size, headspace, fill volume: Glass beakers of 100 mL volume containing approximately 60 mL of test medium.
- Type of flow-through (e.g. peristaltic or proportional diluter):
- Renewal rate of test solution (frequency/flow rate):
- No. of organisms per vessel: 20 Daphnia per test concentration and control, divided into 4 groups of 5 animals, each group in 50 mL test medium



TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt "M4" reconstituted water
- Alkalinity: 0.9 mmol/L
- Culture medium different from test medium: No


OTHER TEST CONDITIONS
- Adjustment of pH: Yes
- Photoperiod: 16 h light: 8 h dark
- Light intensity: 940 - 1150 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobility of the Daphnia was determined by visual observation after 24 and 48 h. Those animals not able to swim within 15 sec after gentle agitation of the test beaker were considered to be immobile (even if they could still move their antennae).


TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 1, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: Yes
A dose-related effect of the test material was observed in an additional range-finding test for the water accommodated fractions of nominal 10 and 1.0 mg/L. Nominal test concentrations 10, 4.6, 2.1, 0.95 and 0.43 mg/L were tested in the main experiment.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
0.95 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
2.1 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
5.17 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Key result
Duration:
24 h
Dose descriptor:
NOEC
Effect conc.:
0.95 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Key result
Duration:
24 h
Dose descriptor:
LOEC
Effect conc.:
2.1 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Details on results:
- Behavioural abnormalities: No
- Mortality of control: No
- After 48 h of exposure no immobilisation of the test animals was observed in the control and up to the nominal test substance concentration of 0.95 mg/L. At 2.1 mg/L, one daphnid was immobile after 48 h of exposure. 8 immobile daphnids were observed at nominal 4.6 mg/L. At the highest concentration of 10 mg/L, 18 daphnids were immobile.

Therefore, the 48 h NOEC was determined to be 0.95 mg/L. The 48 h LOEC was determined to be 2.1 mg/L. The 48 h EC50 value was calculated to be 5.17 mg/L.
Results with reference substance (positive control):
- Results with reference substance valid: Yes
- 48 h EC50: 1.37 mg/L indicating that the sensitivity of the Daphnia was consistent with the level proposed by the OECD 202 guideline
- 48 h NOEC: 0.62 mg/L
- 48 h LOEC: 1.4 mg/L
- Other: Test was not conducted parallely with this study but prior to this study.

Analytical Results:

Determination of the Test substance

Based on the results of TOC measurements the concentration of the test substance was determined using a calibration curve.

Calibration Range

1 to 50 mg Carbon/L in pure water in the case of TC-calibration and
1 to 25 mg Carbon/L in pure water in the case of IC-calibration

Linearity of Response

Correlation of peak area of different standard solutions with their corresponding concentrations, using a linear regression

Regression Coefficient

At least 0.9999 in the case of TC
at least 0.9985 in the case of IC

Typical Calibration Curve

y = 4.3873 * x + 0.5934 in the case of TC

y = 4.4984 * x – 1.8233 in the case of IC

Limit of Detection

LODTC: 2.26 mg Carbon/L
LODIC: 2.45 mg Carbon/L

Limit of Quantification

3 mg Carbon/L
125 % (n = 5, RSD 6 %)

Mean Recovery in the Fortified Samples

118 % (n = 14, RSD 8 %)

 Mean recovery in the test samples Since the test substance is not well water soluble, all results were below LOQ
Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the nominal 48 h EC50 of the test substance was determined to be 5.17 mg/L and the nominal 48 h NOEC and LOEC were determined to be 0.95 and 2.1 mg/L respectively.
Executive summary:

A study was conducted to determine the acute toxicity of the test substance Oils, vegetable, deodorizer distillates to Daphnia magna in a static 48 h immobilisation test according to OECD Guideline 202 and EU method C.2. Twenty Daphnia magna per dose were exposed to water accommodated fraction (WAF) of the nominal test concentrations of 0, 0.43, 0.95, 2.1, 4.6 and 10 mg/L for 48 h under static conditions. Immobility of the Daphnia was determined by visual observation after 24 and 48 h. Analytical verification of the test concentrations at the start and the end of the test via the TOC-method revealed results below LOQ. Therefore, all concentrations were represented as the nominal concentration of the test substance. After 48 h of exposure, no immobilisation of the test animals was observed in the control and up to 0.95 mg/L. At 2.1 mg/L, one daphnid was immobile after 48 h of exposure. 8 immobile daphnids were observed at 4.6 mg/L and at the highest concentration of 10 mg/L, 18 daphnids were immobile. Therefore, under the test conditions the 48 h NOEC and LOEC were determined to be 0.95 and 2.1 mg/L respectively. The 48 h EC50 was determined to be 5.17 mg/L (Kley and Deierling, 2010c).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Study period:
23 March 2010 to 26 March 2010
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
read-across
Justification for type of information:
Refer to the section 13 for details on the read across justification. The acute fish study with the read across substance is used as a supporting study.
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Series on testing and assessment No. 23: Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Concentrations:
- Sampling method: The concentrations of the test substance were analysed in the duplicate test media samples from both sampling times (0 and 48 h). From the control samples only one of the duplicate samples was analysed from each of both sampling times.
- Sample storage conditions before analysis: All samples were stored in a freezer (≤ - 10°C), protected from light until analysis was performed

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficultly soluble test substances)
200.8 mg of test item were added directly to 200 mL of test water and carefully stirred for about 24 h at room temperature in the dark to dissolve as much of the test substance as possible. After cessation of stirring and a following period of settling to allow phase separation, the aqueous phase, i.e. the water accomodated fraction, was drawn off carefully and used as the test medium.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain: Clone 5
- Source: In house laboratory culture (i.e. IBACON testing laboratory)
- Age at study initiation (mean and range, SD): 6.25 to 21.5 h old
- Method of breeding: The cultivation of the parental Daphnia was performed in reconstituted water of a similar quality with regards to pH, the constituent salts and total hardness as the test water used in the test. The test organisms were not first brood progeny
- Food type: Green algae (Desmodesmus subspicatus) freshly grown in the lab
-Frequency: Daily

ACCLIMATION
- Was not necessary, since the test was performed in the same medium as the culturing
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
250 mg/L as CaCO3
Test temperature:
21°C
pH:
7.7- 7.8
Dissolved oxygen:
8.1 - 8.8 mg/L
Nominal and measured concentrations:
Nominal: 0 and 100 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass beakers
- Type (delete if not applicable): Closed with lids
- Material, size, headspace, fill volume: Glass beakers of 100 mL volume containing approximately 60 mL of test medium.
- No. of organisms per vessel: 20 Daphnia per control and test concentration, divided into 4 groups of 5 animals, each group in 50 mL test medium



TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt "M4" reconstituted water
- Alkalinity: 0.9 mmol/L
- Culture medium different from test medium: No


OTHER TEST CONDITIONS
- Adjustment of pH: Yes
- Photoperiod: 16 h light: 8 h dark
- Light intensity: 940 - 1150 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobility of the Daphnia was determined by visual observation after 24 and 48 h. Those animals not able to swim within 15 sec after gentle agitation of the test beaker were considered to be immobile (even if they could still move their antennae).


TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 1, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: Yes
Since a dose-related effect of the test material was observed in an additional range-finding test for the water accommodated fractions of nominal 10 and 1.0 mg/L. Nominal test concentrations 10, 4.6, 2.1, 0.95 and 0.43 mg/L were tested in the main experiment.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
other: water accommodated fraction
Basis for effect:
mobility
Details on results:
- Behavioural abnormalities: No
- Mortality of control: No
- Other adverse effects control: None

The concentration of test substance recorded by total organic carbon analysis was below the limit of quantification in the control and test vessels. The LOQ was equivalent to 3 mg/L. Test concentrations were therefore based on nominal values.

Validity criteria fulfilled:
yes
Conclusions:
Under the test conditions, the nominal 48 h EC50 of the read-across substance was determined to be >100 mg/L. The nominal 48 h NOEC was determined to be 100 mg/L.
Executive summary:

A study was conducted to determine the acute toxicity of the read-across substance Soybean oil, deodorizer distillates to Daphnia magna in a static 48 h immobilisation test (OECD Guideline 202 and EU method C.2). Twenty Daphnia magna were exposed to WAFs of the nominal test concentrations of 0 and 100 mg/L for 48 h. Immobility of the Daphnia was determined by visual observation after 24 and 48 h. Analytical verification of the test concentrations at the start and the end of the test via the TOC- method revealed results below LOQ.Therefore, all concentrations were represented as the nominal concentration of the test substance. Under the test conditions, the nominal 48 h NOEC of the read-across substance was determined to be 100 mg/L and the nominal 48 h EC 50 was >100 mg/L (Kley and Deierling, 2010d).

Description of key information

The 48 h EC50 and NOEC of the test substance were determined to be 5.17 and 0.95 mg/L (nominal) respectively.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
5.17 mg/L

Additional information

Taking into account the variable composition, the acute aquatic toxicity potential of the test substance 'oils, vegetable, deodorizer distillates' to freshwater invertebrates has been evaluated on the basis of below studies:

A study was conducted to determine the acute toxicity of the test substance 'oils, vegetable, deodorizer distillates’ to Daphnia magna in a static 48 h immobilisation test according to OECD 201 and EU Method C.1. In this study, twenty Daphnia magna per concentration were exposed to water accommodated fraction (WAF) of the nominal test concentrations of 0, 0.43, 0.95, 2.1, 4.6 and 10 mg/L for 48 h under static conditions. Immobility of the Daphnia was determined by visual observation after 24 and 48 h. Analytical verification of the test concentrations at the start and the end of the test via the TOC- method revealed results below LOQ. Therefore, all concentrations were represented the nominal concentration of the test substance. After 48 h of exposure, no immobilisation of the test animals was observed in the control and up to and including 0.95 mg/L. At 2.1 mg/L, one daphnid was immobile after 48 h of exposure. 8 immobile daphnids were observed at 4.6 mg/L. At the highest concentration of nominal 10 mg/L, 18 daphnids were immobile. Therefore, under the test conditions the 48 h NOEC and 48 h LOEC were determined to be 0.95 and 2.1 mg/L respectively and the 48 h EC50 value was calculated to be 5.17 mg/L (Kley and Deierling, 2010c) .

Another guideline compliant study was conducted to determine the acute toxicity potential of the read-across substance 'soybean oil, deodorizer distillates' using the same OECD guideline as above. Daphnia magna were exposed to WAFs of the nominal test concentrations of 0 and 100 mg/L for 48 h. Immobility of the Daphnia was determined by visual observation after 24 and 48 h. Analytical verification of the test concentrations at the start and the end of the test via the TOC- method revealed results below LOQ. Therefore, all concentrations were represented as the nominal concentration of the test substance. Under the test conditions and due to the absence of toxicity, the 48 h NOEC of the test substance was established at 100 mg/L and the 48 h EC50 was >100 mg/L (nominal) (Kley and Deierling, 2010d).

Further, several acute toxicity studies were conducted on the saltwater copepod Acartia tonsa using water accommodated fractions of glyceride constituent representative substances i.e., ‘glycerides, C8-18 and C18-unsatd.’ in the form of coconut oil, palm kernel stearin, palm kernel olein and palm kernel oil (Worden and Sherren, 2004a, b, c and d). The lowest loading rate which caused immobilisation in 50% of the exposed animals (48 h EL50) was above 100 mg/L, suggesting low acute toxicity. This is in line with results obtained with ‘glycerides, C16 -18 and C18-unsatd’ (Worden and Sherren, 2002a and b; Bowmer et al.,1998).

Overall based on the above studies, ‘oils, vegetable, deodorizer distillates’ is considered to have low acute toxicity potential in daphnia with the 48 h EC50 calculated at 5.17 mg/L.