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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:
25 Jul. - 19 Aug. 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Principles of method if other than guideline:
open test system, testing of water-accommodated fractions (WAF)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
- Name of test material (as cited in study report): Novares L 800 (former name Hydrocarbons, C9- unsaturated, polymerized, CAS-no. 71302-83-5)
- Composition of test material, percentage of components: see Section 1.2 Composition
- Stability under test conditions: no measured data; based on chemical structure assumed to be stable
- Storage condition of test material: room temperature, exclusion of light
Analytical monitoring:
yes
Remarks:
determination of Total Organic Carbon (TOC)
Details on sampling:
Samples for TOC-analysis were taken at 0 h and 48 h
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water Accommodated Fractions (WAFs) were prepared by stirring various amounts of the test item in dilution water overnight at room temperature with magnetic stirrers. Prior, the TS was dissolved in dichloromethane, and aliquots of the stock solution were added to the flasks. After evaporation of the solvent, dilution water was added.
After equilibration overnight under stirring under closed conditions, the suspensions were filtered through glass fibre filters. The filtrates were used for the test without further dilution.

- Pre-conditioning: The test vessels of the Daphnia-Test were rinsed with control media and WAFs for ca. 1 h before the definite start of the tests to facilitate adsorption of the test item to the glass walls of the beakers. Control media and WAFs were then discarded, and the test vessels were refilled again with control media and WAFs, respectively.
- Differential loading: control, 5, 10 and 100 mg/L (range finding)
control, 10, 20, 40, 80, and 100 mg/L (main test)
- Controls: without test material (blank; Daphnia medium)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): none
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia
- Strain: STRAUS (clone 5)
- Source: derived from Dr. M. Bergtold, BASF, Limburgerhof
- Age at study initiation (mean and range, SD): ≤ 24 h
- Method of breeding: The animals are cultivated by transferring them at intervals of 1 to 3 d into new test medium followed by feeding.
- Feeding during test
- Food type: algae (Desmodesmus subspicatus) and/or Chlorella cells

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
250 mg/L (as CaCO3)
Test temperature:
20.7 – 21.8 °C (20 ± 1 °C) at 0 and 48 h
pH:
7.9 - 8.2 at 0 and 48 h
Dissolved oxygen:
7.2 - 8.3 mg/L at 0 and 48 h
Nominal and measured concentrations:
Main test series:
Time [h] Control 10 mg/L 20 mg/L 40 mg/L 80 mg/L 100 mg/L (loading)
===============================================================================
0 0.16 0.0 0.01 0.05 19.78 0.40 mg C/L (TOC of WAF), mean of 4 measurements
48 0.0 0.0 0.0 0.23 15.17 0.30 mg C/L (TOC of WAF), mean of 4 measurements
==============================================================================
The TS-related TOC was indistinguishable from the background, i.e. the water solubility is not measureable (<0.5 mg/L).
The high concentrations at 80 mg/L are considered to be an artefact due to contamination.
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): closed with glass stopper
- Material, size, headspace, fill volume: glass beaker, 100 mL, test volume: 100 mL
- Aeration: none
- Renewal rate of test solution (frequency/flow rate): none
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): not applicable

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted, according to DIN 38412, part 11
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light, 8 h dark
- Light intensity: no data, Gro-Lux fluorescent tubes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study
- Test concentrations: 5, 10, and 100 mg/L (loading)
- Results used to determine the conditions for the definitive study: immobilisation 100%
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
EL50
Remarks:
effective loading
Effect conc.:
54 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% C.I. = 45.0 - 64.7 mg/L
Duration:
48 h
Dose descriptor:
EL10
Remarks:
effective loading
Effect conc.:
27.5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% C.I. = 20.2 - 37.6 mg/L
Duration:
48 h
Dose descriptor:
NOELR
Remarks:
No-observable effect loading rate
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks:
0/20 immobile
Details on results:
- Other biological observations: none
- Mortality/immobilisation in control: 0/20
- Other adverse effects control: none
- Abnormal responses: none
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none
- Effect concentrations exceeding solubility of substance in test medium: none - WAF
Reported statistics and error estimates:
Determination of the EL (EL= Effective Loading) and NOELR / LOELR using Probit-analysis (Finney-method, lognormal distribution; confidence limit 95 %, variance analysis

Table 1: Immobilisation rate in dependence of time and loading rate

[number per total animals, %] (Report 10.1.4)

Control

10 mg/L

20 mg/L

40 mg/L

80 mg/L

100 mg/L

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Immobilisation 24 h

0/20

0

0/20

0

0/20

0

0/20

0

3/20

15

14/20

70

Immobilisation 48 h

0/20

0

0/20

0

1/20

5

5/20

25

14/20

70

19/20

95

Validity criteria fulfilled:
yes
Remarks:
- Immobilisation in the control: 0 % (nominal value ≤10 %) - Dissolved oxygen in all vessels: >= 7.2 mg/L (nominal value >= 5 mg/L)
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 Jun. - 14 Jul. 2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Principles of method if other than guideline:
open test system, testing of water-accommodated fractions (WAF)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
- Name of test material (as cited in study report): Novares TL 10; CAS-no. 71302-83-5 (Hydrocarbons, C9- unsaturated, polymerized)
- Composition of test material, percentage of components: see Section 1.2 Composition
- Lot/batch No.: 26697
- Stability under test conditions: no measured data; based on chemical structure assumed to be stable
- Storage condition of test material: room temperature, exclusion of light
Analytical monitoring:
yes
Remarks:
determination of Total Organic Carbon (TOC)
Details on sampling:
Samples for TOC-analysis were taken at 0 h and 48 h
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water Accommodated Fractions (WAFs) prepared by stirring various amounts of the test item in Daphnia-medium for 24 h with magnetic stirrers (open conditions). The WAFs were prepared in 2-L glass beakers. The beakers were equipped with a glass tube that allowed the separation of the water phase by siphoning.
The test loading rates of the TS were weighed in weighing scoops that afterwards were placed into the glass beakers. The beakers were then filled with Daphnia-medium. Mixing was carried out at a speed that was slow enough not to cause dispersion or emulsification of the undissolved fraction of the test item. To ensure this, the vortex developed at the surface by stirring was set at ~ 10 % of the water depth.
After stirring for 24 h the WAFs were allowed to stand for 1 h before use to facilitate phase separation. The extracts gained with this method were clear.
- Pre-conditioning: The test vessels of the Definite Daphnia-Test were rinsed with control media and WAFs for ca. 0.5 h before the definite start of the tests to facilitate adsorption of the test item to the glass walls of the beakers. Control media and WAFs were then discarded, and the test vessels were refilled again with control media and WAFs, respectively.
- Differential loading: control, 1, 10 and 100 mg/L (range finding)
control, 100 mg/L
- Controls: without test material (blank; Daphnia medium)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): none
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia
- Strain: --
- Source: origin German Environmental Ministry
- Age at study initiation (mean and range, SD): ≤ 24 h
- Method of breeding: The animals are cultivated in a climate-controlled room (light/dark = 14 h/10 h; illumination: Gro-Lux fluorescent tubes).
- Feeding during test
- Food type: algae (Desmodesmus subspicatus)

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
see data under "salinity"
Test temperature:
21.5 – 21.8 °C (20 °C ± 2 °C)
pH:
7.9 - 8.2 at 0, 24 and 48 h
Dissolved oxygen:
7.5 - 7.9 mg/L at 0, 24 and 48 h
Salinity:
The total of Ca- and Mg-ions was 2.5 mmol/L, the ratio of Ca/Mg was 4:1, the ratio of Na/K was 10:1.
The total alkalinity of the solution was 0.8 mmol/L, the pH-value was 7.8 ± 0.2.
Nominal and measured concentrations:
Main test series:
Time [h] Control 100 mg/L (loading)
================================
0 0.3 1.0 mg C/L (TOC of WAF)
48 0.3 0.6 mg C/L (TOC of WAF)
=====================================

Range finding:
Time [h] Control 1 mg/L 10 mg/L 100 mg/L (Loading)
=======================================================
0 0.3 0.5 0.2 0.2 mg C/L (TOC of WAF)
48 0.5 1.0 0.6 0.7 mg C/L (TOC of WAF)
=========================================================
The TS-related TOC was indistinguishable from the background, i.e. the water solubility is not measureable.
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: glass beaker, 50 mL, test volume: 20 mL
- Aeration: none
- Renewal rate of test solution (frequency/flow rate): none
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): not applicable

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water
- Total organic carbon: no data
- Alkalinity: 0.8 mmol/L
- Ca/Mg ratio: 4:1
- Conductivity: no data
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light, 8 h dark
- Light intensity: no data, Gro-Lux fluorescent tubes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: not applicable (limit test)
- Range finding study
- Test concentrations: 1, 10, 100 mg/L (loading)
- Results used to determine the conditions for the definitive study:
immobilisation 0 %
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
EL50
Remarks:
effective loading
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
NOELR
Remarks:
No-observable effect loading rate
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks:
0/20 immobile
Details on results:
- Other biological observations: none
- Mortality/immobilisation in control: 0/20
- Other adverse effects control: none
- Abnormal responses: none
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none
- Effect concentrations exceeding solubility of substance in test medium: none
Reported statistics and error estimates:
Determination of the EL50 (EL= Effective Loading) and NOELR was accomplished using Probit-analysis (Finney-method, lognormal distribution; confidence limit 95 %, significance level: p <0.05) (not relevant in this case).

Table 1: Immobilisation rate in dependence of time and loading rate, number of carapace moults, and

number of living animals trapped at the surface [number per total animals, %] (Report Tab. 8)

Control

100

mg/L

number

[%]

number

[%]

Immobilisation 24 h

0/20

0

0/20

0

Immobilisation 48 h

0/20

0

0/20

0

Moults 48 h

17/20

85

18/20

90

Living animals at the surface 48 h

0/20

0

0/20

0

Validity criteria fulfilled:
yes
Remarks:
- Immobilisation in the control: 0 % (nominal value ≤10 %) - Dissolved oxygen in all vessels: >= 7.5 mg/L (nominal value >= 3 mg/L)
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
04 Jun. - 25 Jul. 2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Principles of method if other than guideline:
open test system, testing of a water-accommodated fractions (WAF)
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
- Name of test material (as cited in study report): Novares L 100; CAS-no. 71302-83-5 (Hydrocarbons, C9- unsaturated, polymerized)
- Composition of test material, percentage of components: see Section 1.2 Composition
- Lot/batch No.: 24106
- Stability under test conditions: no measured data; based on chemical structure assumed to be stable
- Storage condition of test material: room temperature, exclusion of light
Analytical monitoring:
yes
Remarks:
determination of Total Organic Carbon (TOC)
Details on sampling:
Samples for TOC-analysis were taken at 0 h and 48 h
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Water Accommodated Fractions (WAFs) prepared by stirring various amounts of the test item in Daphnia-medium for 24 h with magnetic stirrers (open conditions). The WAFs were prepared in 2-L glass beakers. The beakers were equipped with a glass tube that allowed the separation of the water phase by siphoning.
The test loading rates of the TS were weighed in weighing scoops that afterwards were placed into the glass beakers. The beakers were then filled with Daphnia-medium. Mixing was carried out at a speed that was slow enough not to cause dispersion or emulsification of the undissolved fraction of the test item. To ensure this, the vortex developed at the surface by stirring was set at ~ 10 % of the water depth.
After stirring for 24 h the WAFs were allowed to stand for 1 h before use to facilitate phase separation. The extracts gained with this method were clear.
- Pre-conditioning: The test vessels of the Definite Daphnia-Test were rinsed with control media and WAFs for ca. 0.5 h before the definite start of the tests to facilitate adsorption of the test item to the glass walls of the beakers. Control media and WAFs were then discarded, and the test vessels were refilled again with control media and WAFs, respectively.
- Differential loading: control, 1, 10 and 100 mg/L (range finding)
control, 25 mg/L, 50 mg/L, 100 mg/L, 250 mg/L, 500 mg/L, and 1000 mg/L
- Controls: without test material (blank; Daphnia medium)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): none
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia
- Strain: --
- Source: origin German Environmental Ministry
- Age at study initiation (mean and range, SD): ≤ 24 h
- Method of breeding: The animals are cultivated in a climate-controlled room (light/dark = 14 h/10 h; illumination: Gro-Lux fluorescent tubes).
- Feeding during test
- Food type: algae (Desmodesmus subspicatus)

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
see data under "salinity"
Test temperature:
20.0 – 21.0 °C (20 °C ± 2 °C)
pH:
7.8 - 8.1 at 0, 24 and 48 h
Dissolved oxygen:
8.0 - 8.1 mg/L at 0, 24 and 48 h
Salinity:
The total of Ca- and Mg-ions was 2.5 mmol/L, the ratio of Ca/Mg was 4:1, the ratio of Na/K was 10:1.
The total alkalinity of the solution was 0.8 mmol/L, the pH-value was 7.8 ± 0.2.
Nominal and measured concentrations:
Main test series:
Time [h] Control 25 mg/L 50 mg/L 100 mg/L 250 mg/L 500 mg/L 1000 mg/L (loading)
==================================================================
0 0.4 0.9 n.d. n.d. 0.7 n.d 0.8 mg C/L (TOC of WAF)
48 0.1 0.3 n.d n.d. 0.3 n.d 0.5 mg C/L (TOC of WAF)
=======================================================================

Range finding:
Time [h] Control 1 mg/L 10 mg/L 100 mg/L (Loading)
=======================================================
0 0.3 0.5 0.4 0.3 mg C/L (TOC of WAF)
48 0.5 0.6 0.5 0.6 mg C/L (TOC of WAF)
=========================================================
The TS-related TOC was indistinguishable from the background, i.e. the water solubility is not measureable.
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: glass beaker, 50 mL, test volume: 20 mL
- Aeration: none
- Renewal rate of test solution (frequency/flow rate): none
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): not applicable

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water
- Total organic carbon: no data
- Alkalinity: 0.8 mmol/L
- Ca/Mg ratio: 4:1
- Conductivity: no data
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light, 8 h dark
- Light intensity: no data, Gro-Lux fluorescent tubes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2 - 2.5
- Range finding study
- Test concentrations: 100 and 1000 mg/L (loading)
- Results used to determine the conditions for the definitive study:
immobilisation 100 %
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
EL50
Remarks:
effective loading
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
NOELR
Remarks:
No-observable effect loading rate
Effect conc.:
1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks:
0/20 immobile
Details on results:
- Other biological observations: none
- Mortality/immobilisation in control: 0/20
- Other adverse effects control: none
- Abnormal responses: none
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none
- Effect concentrations exceeding solubility of substance in test medium: none
Reported statistics and error estimates:
Determination of the EL50 (EL= Effective Loading) and NOELR was accomplished using Probit-analysis (Finney-method, lognormal distribution; confidence limit 95 %, significance level: p <0.05) (not relevant in this case).

Table 1: Immobilisation rate in dependence of time and loading rate, number of carapace moults, and

number of living animals trapped at the surface [number per total animals, %] (Report Tab. 12)

Control

25 mg/L

50 mg/L

100 mg/L

250 mg/L

500 mg/L

1000 mg/L

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Number

[%]

Immobilisation 24 h

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

Immobilisation 48 h

0/20

0

0/20

0

0/20

0

1/20

5

0/20

0

0/20

0

0/20

0

Moults 48 h

18/20

90

16/20

80

18/20

90

8/20

40

8/20

40

16/20

80

14/20

70

Living animals at
the surface 48 h

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

0/20

0

Validity criteria fulfilled:
yes
Remarks:
- Immobilisation in the control: 0 % (nominal value ≤10 %) - Dissolved oxygen in all vessels: >= 7.5 mg/L (nominal value >= 3 mg/L)

Description of key information

Short-term toxicity to daphnia was investigated in three studies using three different types/technical products of NAF-AO. One of this tests was a limit test. EL 50 ranged from 54 mg/L to > 1000 mg/L. No data is available for marine water invertebrates.

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
54 mg/L

Additional information

Based on differences in the technical production process, different types of NAF-AO can be obtained. Three of this technical products (Novares L 100, L 800, and TL 10) were tested in daphnia short-term toxicity studies according to OECD TG 202 performed under GLP conditions. Concentrations reported for the aqueous test media relate to loadings.

EL50 values obtained in the three studies range from 54 mg/L to > 1000 mg/L. A limit test with Novares TL 10 provided an EL50 value of > 100 mg/L (NOELR 100 mg/L). For Novares L 100, an EL 50 of > 1000 mg/L was determined (NOELR 1000 mg/L, highest concentration tested). In the test with Novares L 800, the EL50 was shown to be 54 mg/L. There is no obvious explanation for the variety of the different products.