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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:
key study
Study period:
12 November 1997 - 14 November 1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted in accordance with EC, OECD, and US EPA test guidelines, and in complaince with GLP.
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OTS 797.1300 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: Nominal loadings - 0 (control), 1.0, 2.2, 4.6, 10, 22, 46, and 100 mg/L.
- Sampling method: Duplicate pooled medium samples were taken at 0 and 48 hours; sampling methodology not specified.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Method followed advice given in ECETOC 1996, Monograph No. 26 as the test material has poor water solubility.
Test material was added by accurate pipette to 2 L glass vessels of Elendt M4 medium and stirred for at least 18 hours. Once stirred the solutions were left to stand for one hour prior to removing the water accommodated fraction. Approximately 800 mL of the solution was removed from a mid-water position.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): In preparation of the water accommodated fraction, when it was left to stand (without stirring), globules of test material were seen at the bottom of the vessels and a film was seen on the surface of the water. In the exposure vessels, expired solutions were noted to be cloudy after 24 hours.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain: Straus
- Source: Cultured in-house from a strain originating from the Institute National de Recherche Chimique Applique (IRChA), France
- Age at study initiation (mean and range, SD): Less than 24 hours old
- Method of breeding: The Daphnia used were bred under conditions which ensures that cultures reproduce by parthenogenesis

ACCLIMATION
- Acclimation period: Not applicable - the Daphnia used were all less than 24 hours old. Stock culture conditions are provided below.
- Type and amount of food: A suspension of the unicellular green alga, Chlorella vulgaris
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
19.3°C
pH:
7.7 - 7.8
Dissolved oxygen:
8.1 - 9.0 mgO2/L
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal loadings: 0 (control), 1.0, 2.2, 4.6, 10, 22, 46, and 100 mg/L.
Measured concentrations for diethylene glycol dibenzoate (expressed as % loading rate)
1.0 mg/L: 54.2 (Fresh prepared), 8.7 (expired / 48 hours old)
2.2 mg/L: 17.9, 2.7
4.6 mg/L: 76.1, 46.9
10 mg/L: 37.1, 20.1
22 mg/L: 60.0, 60.9
46 mg/L: 9.3, 6.0
100 mg/L: 17.8, 17.7
Measured concentrations for diethylene glycol monobenzoate (expressed as % loading rate in terms of diethylene glycol dibenzoate)
1.0 mg/L: 8.5 (Fresh prepared), 1.9 (expired / 48 hours old)
2.2 mg/L: 2.6, 0.0
4.6 mg/L: 11.8, 26.4
10 mg/L: 9.4, 14.3
22 mg/L: 9.9, 20.0
46 mg/L: 8.2, 9.6
100 mg/L: 8.9, 8.7
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type : closed (loosely covered to minimise evaporative losses)
- Material, size, headspace, fill volume: 250 mL glass jars, filled with 200 mL test medium
- Renewal rate of test solution (frequency/flow rate): None (static test)
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 40 mL test solution per organism.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4 medium, prepared in house using analytical reagents and deionised water

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light, 8 hours dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : immobilisation

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
Reference substance (positive control):
no
Duration:
24 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
6.7 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% confidence limits : 4.4 - 10.0 mg/L
Duration:
48 h
Dose descriptor:
NOELR
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 10% immobilisation was caused at 1.0 mg/L; this level of immobilisation is not considered significant.
Details on results:
- Mortality of control: Two Daphnia in the control level were immobilised after 48 hours. As this corresponded to the no-effect level of 10%, the study results were still considered valid.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Precipitation was observed, as noted previously. The analytical method used to confirm the concentration of test material in solution determined concentrations of diethylene glycol monobenzoate DEGMB) as well as diethylene glycol dibenzoate (DEGDB); the concentration of DEGDB was found to be lower after 48 hours than at the start of the test in most cases, whereas the concentration of DEGMB had generally increased. This observation suggests that the monobenzoate was a degradation product of the dibenzoate.
Validity criteria fulfilled:
yes
Conclusions:
The 48 hour EL50 (immobilisation value for DEGDB with Daphnia Magna) was determined to be 6.7 mg/L. The "No-observed effect loading rate" (NOELR) was 1.0 mg/L.
Executive summary:

An acute Daphnia Immobilisation study was performed to determine the acute toxicity to Daphnia Magna of the test material diethylene glycol dibenzoate (DEGDB). The study was conducted according to EC, OECD, and US EPA test guidelines, and in compliance with GLP.

The 48 hour EL50 (immobilisation value for DEGDB with Daphnia Magna) was determined to be 6.7 mg/L. The "No-observed effect loading rate" (NOELR) was 1.0 mg/L.

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
11 October 1997 - 13 October 1997.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study undertaken in GLP accredited laboratory.
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OTS 797.1300 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: nominal loadings - 0 (control), 1.0, 2.2, 4.6, 10, 22, 46 and 100 mg/L
- Sampling method: Duplicate pooled medium samples were taken at 0 and 48 hours; sampling methodology not specified.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The method of preparation was selected following advice given in ECETOC 1996 Monograph No 26 as the test material has poor water solubility.
Test material was added by accurate pipette to 2 L glass vessels of Elendt M4 medium and stirred for at least 18 hours. Once stirred the solutions were left to stand for one hour prior to removing the water accommodated fraction. Approximately 800 mL of the solution was removed from a mid-water position.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): In preparation of the water accommodated fraction, when it was left to stand (without stirring), globules of test material were seen at the bottom of the vessels and a film was seen on the surface of the water. In the exposure vessels, expired solutions were noted to be cloudy after 24 hours.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain: Straus
- Source: Cultured in-house from a strain originating from the Institute National de Recherche Chimique Applique (IRChA), France
- Age at study initiation (mean and range, SD): Less than 24 hours old
- Method of breeding: The Daphnia used were bred under conditions which ensures that cultures reproduce by parthenogenesis

ACCLIMATION
- Acclimation period: Not applicable - the Daphnia used were all less than 24 hours old. Stock culture conditions are provided below.
- Type and amount of food: A suspension of the unicellular green alga, Chlorella vulgaris
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
19 to 21°C
pH:
7.9 to 8.1
Dissolved oxygen:
7.0 to 8.0 mgO2 / L
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal loadings(fresh) ; 0 (control), 1.0, 2.2, 4.6, 10.0, 22, 46 and 100 mg/L
Measured concentrations of dipropylene glycol dibenzoate (expressed as % of loading)
1.0 mg/L: 33.9 (Fresh prepared), 18.1 (expired / 48 hours old)
2.2 mg/L: 40.7, 27.3
4.6 mg/L: 30.8, 26.8
10 mg/L: 49.4, 26.6
22 mg/L: 40.3, 27.5
46 mg/L: 13.8, 9.2
100 mg/L: 19.3, 7.1
Measured concentrations for dipropylene glycol monobenzoate (expressed as % loading rate in terms of dipropylene glycol dibenzoate)
1.0 mg/L: 3.6 (Fresh prepared), 9.0 (expired / 48 hours old)
2.2 mg/L: 5.0, 13.7
4.6 mg/L: 5.2, 16.0
10 mg/L: 4.8, 18.2
22 mg/L: 5.8, 11.3
46 mg/L: 7.6, 8.3
100 mg/L: 13.9, 6.9
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type : closed (loosely covered to minimise evaporative losses)
- Material, size, headspace, fill volume: 250 mL glass jars, filled with 200 mL test medium
- Renewal rate of test solution (frequency/flow rate): None (static test)
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 40 mL test solution per organism.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4 medium, prepared in house using analytical reagents and deionised water

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light / 8 hours dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobilisation

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
Reference substance (positive control):
no
Duration:
24 h
Dose descriptor:
EL50
Effect conc.:
43.2 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
19.3 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% confidence limits : 13.1 - 28.5 mg/L
Duration:
48 h
Dose descriptor:
NOELR
Effect conc.:
2.2 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 5% immobilisation was caused at 2.2 mg/L; this level of immobilisation is not considered significant.
Details on results:
- Mortality of control: Two Daphnia in the control level were immobilised after 48 hours. As this corresponded to the no-effect level of 10%, the study results were still considered valid.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Precipitation was observed, as noted previously. The analytical method used to confirm the concentration of test material in solution determined concentrations of dipropylene glycol monobenzoate DPGMB) as well as dipropylene glycol dibenzoate (DPGDB); the concentration of DPGDB was found to be lower after 48 hours than at the start of the test in all cases, whereas the concentration of DPGMB had generally increased. This observation suggests that the monobenzoate was a degradation product of the dibenzoate.
Validity criteria fulfilled:
yes
Conclusions:
The 48 hour EL50 (immobilisation value for DPGDB with Daphnia magna) was determined to be 19.3 mg/L. The "no-observed effect loading rate" (NOELR) was 2.2 mg/L.
Executive summary:

An acute Daphnia Immobilisation study was performed to determine the actue toxicity to Daphnia of the test material dipropylene glycol dibenzoate (DPGDB). The study was conducted according to EC, OECD, and US EPA test guidelines, and in compliance with GLP.

The 48 hour EL50 (immobilisation value for DPGDB with Daphnia Magna) was determined to be 19.3 mg/L. The "no-observed effect loading rate" (NOELR) was 2.2 mg/L.

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
21 October 1997 - 23 October 1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted in accordance with EC, OECD, and US EPA test guidelines, and in compliance with GLP.
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OTS 797.1300 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: Nominal loadings - 0 (control), 4.6, 10, 22, 46, and 100 mg/L.
- Sampling method: Duplicate pooled medium samples were taken at 0 and 48 hours; sampling methodology not specified.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The method of preparation was selected following advice given in ECETOC 1996 Monograph No 26 as the test material has poor water solubility.
Appropriate weights of the test substance 9.2, 20, 92 and 200 mg were dispersed into 2 litres of diluent water contained in a glass reagent bottle.
Each preparation was then stirred for at least 18 hours to give water accommodated fractions with initial loading rates of 4.6, 10, 22, 46 and 100 mg/L. After stirring, the solutions were allowed to settle for 1 hour before removing approximately 1 litre of the water accommodated fraction, from a mid water position, from each vessel to give the exposure solution.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: water flea
- Strain: Straus
- Source: Cultured in-house from a strain originating from the Institute National de Recherche Chimique Applique (IRChA), France.
- Age at study initiation (mean and range, SD): less than 24 hours old
- Method of breeding: The Daphnia used were bred under conditions which ensures that cultures reproduce by parthenogenesis

ACCLIMATION
- Acclimation period: Not applicable - the Daphnia used were all less than 24 hours old. Stock culture conditions are provided below.
- Type and amount of food: A suspension of the unicellular green alga, Chlorella vulgaris
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20 ± 1°C
pH:
7.6
Dissolved oxygen:
7.4 - 8.3 mgO2/L
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal loadings: 0 (control), 4.6, 10, 22, 46, and 100 mg/L.
Measured concentrations for triethylene glycol dibenzoate (expressed as % loading rate)
4.6 mg/L: 55 (Fresh prepared), 0 (expired / 48 hours old)
10 mg/L: 26, 0
22 mg/L: 37, 7
46 mg/L: 45, 29
100 mg/L: 28, 20
Measured concentrations for triethylene glycol monobenzoate (expressed as % loading rate in terms of triethylene glycol dibenzoate)
4.6 mg/L: 15 (Fresh prepared), 9 (expired / 48 hours old)
10 mg/L: 6, 0
22 mg/L: 7, 23
46 mg/L: 7, 22
100 mg/L: 4, 11
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type : closed, (loosely covered covered to minimise evaporative loss)
- Material, size, headspace, fill volume: 250 mL glass jars, filled with 200 mL test medium
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 40 mL test solution/ organism

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4 medium, prepared in house using analytical reagents and deionised water

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light, 8 hours dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : immobilisation

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
- Range finding study
- Test concentrations: A preliminary range finding study was conducted using initial loading rates of 1.0, 10 and 100 mg/L under similar exposure conditions to those subsequently used in the definitive test. The results of this preliminary range finding test indicated immobilisation occurred at loading rates above 10 mg/L.
- Results used to determine the conditions for the definitive study: Yes
Reference substance (positive control):
no
Duration:
24 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
26 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% confidence limits 20 - 35 mg/L
Dose descriptor:
NOELR
Effect conc.:
4.6 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: (10% immobilisation was observed in the 4.6 mg/L test group. This level of immobilisation is not considered significant
Details on results:
- Mortality of control: Two Daphnia in the control level were immobilised after 48 hours. As this corresponded to the no-effect level of 10%, the study results were still considered valid.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No precipitation or cloudiness was noted. The analytical method used to confirm the concentration of test material in solution determined concentrations of triethylene glycol monobenzoate TEGMB) as well as triethylene glycol dibenzoate (TEGDB); the concentration of TEGDB was found to be lower after 48 hours than at the start of the test in all cases, whereas the concentration of TEGMB had generally increased. This observation suggests that the monobenzoate was a degradation product of the dibenzoate.
Validity criteria fulfilled:
yes
Conclusions:
The 48 h EL50 (immobilisation value for TEGDB with Daphnia Magna) was determined to be 26 mg/L. The "No observed effect loading rate" (NOELR) was 4.6 mg/L.
Executive summary:

An acute Daphnia immobilisation study was performed to determine the acute toxicity to Daphnia Magna of the test material triethylene glycol dibenzoate (TEGDB). The study was conducted according to EC, OECD and US EPA test guidelines, and in compliance with GLP.

The 48 hour EL50 immobilisation value for TEGDB with Daphnia magna was determined to be 26 mg/L. The "No-observed effect loading rate" (NOELR) was 4.6 mg/L.

Description of key information

DEGDB:

Short-term toxicity to aquatic invertebrates:

- Key study, reliability 1, OECD 202, Daphnia magna - EL50 = 6.7 mg/L, NOELR = 1.0 mg/L

DPGDB:

Short-term toxicity to aquatic invertebrates:

- Key study, reliability 1, OECD 202, Daphnia magna - EL50 = 19.3 mg/L, NOELR = 2.2 mg/L

TEGDB

Short-term toxicity to aquatic invertebrates:

- Key study, reliability 1, OECD 202, Daphnia magna - EL50 (48h) = 26 mg/L, NOELR = 4.6 mg/L

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
6.7 mg/L

Additional information

This substance is a reaction mass of dipropylene glycol dibenzoate (DPGDB), diethylene glycol dibenzoate (DEGDB) and triethylene glycol dibenzoate (TEGDB). No testing has been performed on the reaction mass itself but data are available for DPGDB, DEGDB and TEGDB individually. All the studies were performed according to international test guidelines and in compliance with GLP. They are presented below, by component. 

The acute toxicity studies were carried out on Daphnia magna.  

These data demonstrate that DPDGB and DEGDB have similar environmental profiles with all EC/EL/LC/LL50 values in the range of >1.0 mg/L and <20 mg/L. TEGDB is less toxic with EC/EL/LL50 values in excess of 16 mg/L.

The most sensitive endpoint result obtained for these components has been used as the key value to evaluate hazard and is highlighted below. Where no data exist an overall statement that applies to the reaction mass has been made. No further testing is proposed.

The key value is:

Aquatic toxicity to algae and cyanobacteria EC50 = 6.7 mg/L

These values are from DEGDB, acute toxicity to daphnia magna performed on any of the components.

 

DEGDB

Short-term toxicity to aquatic invertebrates

The 48 hour EL50, immobilisation value withDaphnia Magnawas determined to be 6.7 mg/L. The NOELR was 1.0 mg/L (HLS 2001, VCL264/974222).

DPGDB

Short-term toxicity to aquatic invertebrates

The 48 hour EL50, immobilisation value withDaphnia Magnawas determined to be 19.3 mg/L. The "no-observed effect loading rate" (NOELR) was 2.2 mg/L (HLS 2001, VCL252/974221).

TEGDB

Short-term toxicity to aquatic invertebrates

The 48 hour EL50, immobilisation value withDaphnia Magnawas determined to be 26 mg/L. The NOELR was 4.6 mg/L (HLS 2001, VCL281/974224).