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Endpoint:
sediment toxicity: short-term
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions
Remarks:
acceptable for assessment
Qualifier:
equivalent or similar to guideline
Guideline:
other: ASTM 1992 E 1383-92: ASTM 1995 E 1706-00; SETAC 1993
GLP compliance:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): Phenanthrene
- Source of test material: no data
- Analytical purity: > 99 %
Analytical monitoring:
yes
Details on sampling:
SEDIMENT
- Concentrations: all nominal concentration levels
- Sampling interval: at start and end of exposure
- Method: samples from 5 replicates were pooled

OVERLYING WATER
- Concentrations: all nominal concentration levels
- Sampling interval: at start and end of exposure
- Method: samples from 5 replicates were pooled
Vehicle:
yes
Remarks:
acetone
Details on sediment and application:
PREPARATION OF SPIKED SEDIMENT
- Details of spiking: spiking method according to Ditsworth et al. (1990)
- Method of mixing: stock solutions of test substance in acetone were evaporated in 2-L glass jars. Sediment (800 g ww, 23 % water) was added and rolled (12 h, 10 rpm at room temp. in the dark).
- Equilibration time: 24 h
- Equilibration conditions: 20 °C in the dark
- Controls: vehicle controls (acetone without test substance); control samples were prepared in the same manner as described above, but without test item
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Evaporation of vehicle before use: yes
Test organisms (species):
Chironomus riparius
Details on test organisms:
TEST ORGANISM
- Common name: harlequin fly
- Source: TEPE laboratory cultures, Savoy University, France
- Keeping of stock: in aquaria (5 L, 20 ± 2 °C)
- Food type: TetraMin
- Age of animals at beginning of exposure: 48 h old larvae
- Feeding during test: No extra food supplied (TetraMin and cellulose present in formulated sediment was the only food source during exposure period)
In a part of the experiment, addtional TetraMin was daily applied during the exposure period in order to check if addition of uncontaminated food influences toxicity results.
Study type:
laboratory study
Test type:
static
Water media type:
freshwater
Type of sediment:
artificial sediment
Limit test:
no
Duration:
10 d
Exposure phase:
total exposure duration
Post exposure observation period:
no
Hardness:
no data
Test temperature:
20 ± 2 °C
pH:
no data
Dissolved oxygen:
close to saturation level
Salinity:
no data
Ammonia:
no data
Nominal and measured concentrations:
Nominal: 0, 10, 30, 100, 300 mg/kg (dw)
For all exposure levels, the recovery was higher than 77 %; no change of test substance concentration was observed between initiation and termination of the experimen.
Details on test conditions:
TEST SYSTEM
- Test container (material, size): 250 mL glass beaker
- Weight of wet sediment: 100 g wet sediment
- Overlying water volume: 200 mL synthetic freshwater (modified OECD medium - OECD, 1993 - with moderate nutrient contents)
- Aeration: yes
- Replacement of evaporated test water: water loss (ca. 5 mL/wk) was compensated by adding distilled water twice per week.
- Diet: TetraMin and cellulose present in artificial sediment (see below) were the only food sources, no additional food applied.

EXPOSURE REGIME
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 5
- No. of replicates per control / vehicle control: 5
- Type and preparation of food: the only food source during the exposure period was TetraMin and cellulose present in the formulated sediment.

CHARACTERIZATION OF ARTIFICIAL SEDIMENT (all percentages relative to dry weight)
- Composition: 65 % sand (washed and sieved to 500 µm - 2 mm)
30 % kaolin clay
4.85 % alpha-cellulose
0.15 % TetraMin
0.1 % calcium carbonate
- Method of preparation (if artificial substrate): moistened and vigorously mixed with artificial freshwater (see above)
- Total organic carbon (TOC): 2 %

OTHER TEST CONDITIONS
- Light quality: White light (10 % UV)
- Photoperiod: 16 h light, 8 h dark
- Light intensity: 2500 lux

EFFECT PARAMETERS MEASURED: mortality, growth

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3 to 3.33
- Range finding study: no data
Reference substance (positive control):
not specified
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
68 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value (13.6 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
13.6 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: 95%-C.I.: 12.1 - 15.37
Key result
Duration:
10 d
Dose descriptor:
NOEC
Effect conc.:
50 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value (10 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Key result
Duration:
10 d
Dose descriptor:
NOEC
Effect conc.:
10 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Key result
Duration:
10 d
Dose descriptor:
LOEC
Effect conc.:
30 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
No effect on growth was observed for all conditions tested.
Three experiments were conducted: one with and two without additional food. No significant differences could be observed. The lowest observed LC50 (basis mortality) is listed above.
Reported statistics and error estimates:
Calculation of EC50, LC50, 95% confidence limits: Spearman-Karber method
Data analysed with Statview F-4.5, significant differences (p < 0.05) between means were determined using t-test

As a consequence of the high spiking efficiency and stability of exposure, toxicity data were calculated with nominal concentrations (see above: nominal and measured concentrations).

NOEC recalculated to a sediment with 10 % organic carbon: 50 mg/kg dw

Endpoint:
sediment toxicity: short-term
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material is phenanthrene a polycyclic aromatic hydrocarbon. It consists of a three-ring system and is beside others a major constituent of the target substance. The target substance anthracene oil (benzo[a]pyrene < 50 ppm, AOL) is composed of a broad range of PAH predominantly consisting of two up to four aromatic rings.
Toxic effects of anthracene oil on sediment organisms will be characterised by the range of PAH that constitute its composition. Therefore, results obtained from sediment toxicity tests with the AOL constituent phenanthrene can be used in order to characterise the toxic potential of anthracene oil on sediment dwelling organisms.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source test material is the substance phenanthrene. It was purchased from a commercial source and the analytical purity of the test substance is reported to be > 99 %. Hence, the study results are related to effects caused by phenanthrene.
The target material anthracene oil is a UVCB substance produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. PAH contained in AOL range from naphthalene up to pyrene and benzofluorenes. Two- and three-ring aromatics amount to about 50 % (typical concentration). Main constituent is phenanthrene present in a typical concentration of ca. 25 to 31 % (composite sample 7). PAH with four rings accumulate to about 10 %.

3. ANALOGUE APPROACH JUSTIFICATION
Properties of the target substance anthracene oil, relevant for environmental distribution and/or ecotoxicological effects, will be determined by the properties of the PAH that are constituents of anthracene oil. Under environmental conditions or during processing of the substance, environmentally available (volatile or water soluble) components of anthracene oil can be released. These will be PAH (mainly compounds consisting of two to four fused rings). In combination, they will specify the environmental distribution properties and ecotoxicological effects of anthracene oil as a whole. In addition, phenanthrene is considered to adequately represent the PAH-related environmental toxicity of AOL. It is one of the most abundant components in AOL with a high toxicity on sediment dwelling organisms. Therefore, phenanthrene is selected as marker substance to represent the environmental toxicity of AOL (see below Chapter 7.1.4. - Discussion). For these reasons, it is justified to use data determined for phenanthrene as a constituent of anthracene oil to characterise ecotoxicological effects (toxicity to soil dwelling organisms) of anthracene oil itself.
Reason / purpose for cross-reference:
read-across source
Principles of method if other than guideline:
Read-across to preceding entry:
Source test material: phenanthrene (generic, commercial product);
Reference: Verrhiest et al. 2001
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
68 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value (13.6 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Key result
Duration:
10 d
Dose descriptor:
NOEC
Effect conc.:
50 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value (10 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Details on results:
No effect on growth was observed for all conditions tested.
Three experiments were conducted: one with and two without additional food. No significant differences could be observed. The lowest observed LC50 (basis mortality) is listed above.
Endpoint:
sediment toxicity: long-term
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions
Remarks:
acceptable for assessment
Qualifier:
equivalent or similar to guideline
Guideline:
other: ASTM 1992 E 1383-92: ASTM 1995 E 1706-00; SETAC 1993
GLP compliance:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): Phenanthrene
- Source of test material: no data
- Analytical purity: > 99 %
Analytical monitoring:
yes
Details on sampling:
SEDIMENT
- Concentrations: all nominal concentration levels
- Sampling interval: at start and end of exposure
- Method: samples from 5 replicates were pooled

OVERLYING WATER
- Concentrations: all nominal concentration levels
- Sampling interval: at start and end of exposure
- Method: samples from 5 replicates were pooled
Vehicle:
yes
Remarks:
acetone
Details on sediment and application:
PREPARATION OF SPIKED SEDIMENT
- Details of spiking: spiking method according to Ditsworth et al. (1990)
- Method of mixing: stock solutions of test substance in acetone were evaporated in 2-L glass jars. Sediment (800 g ww, 23 % water) was added and rolled (12 h, 10 rpm at room temp. in the dark).
- Equilibration time: 24 h
- Equilibration conditions: 20 °C in the dark
- Controls: vehicle controls (acetone without test substance); control samples were prepared in the same manner as described above, but without test item
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Evaporation of vehicle before use: yes
Test organisms (species):
Hyalella azteca
Details on test organisms:
TEST ORGANISM
- Source: TEPE laboratory cultures, Savoy University, France
- Keeping of stock: in aquaria (5 L, 20 ± 2 °C)
- Food type: TetraMin
- Age of animals at beginning of exposure: 2 - 3 weeks
- Size at beginning of exposure: 0.5 - 1 mm
- Feeding during test: No extra food supplied (TetraMin and cellulose present in formulated sediment was the only food source during exposure period)




Study type:
laboratory study
Test type:
static
Water media type:
freshwater
Type of sediment:
artificial sediment
Limit test:
no
Duration:
14 d
Exposure phase:
total exposure duration
Post exposure observation period:
no
Hardness:
no data
Test temperature:
20 ± 2 °C
pH:
no data
Dissolved oxygen:
close to saturation level
Salinity:
no data
Ammonia:
no data
Nominal and measured concentrations:
Nominal: 0, 10, 30, 100, 300 mg/kg (dw)
For all exposure levels, the recovery was higher than 77 %, no change of test substance concentration was observed between initiation and termination of the experiment.
Details on test conditions:
TEST SYSTEM
- Test container (material, size): 250 ml glass beaker
- Weight of wet sediment: 100 g wet sediment
- Overlying water volume: 200 ml synthetic freshwater (modified OECD medium - OECD, 1993 - with moderate nutrient contents)
- Aeration: yes
- Replacement of evaporated test water: Water loss (ca. 5 mL/wk) was compensated by adding distilled water twice per week.
- Diet: TetraMin and cellulose present in artificial sediment (see below) were the only food sources, no additional food applied.

EXPOSURE REGIME
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 5
- No. of replicates per control / vehicle control: 5
- Type and preparation of food: the only food source during the exposure period was TetraMin and cellulose present in the formulated sediment

CHARACTERIZATION OF ARTIFICIAL SEDIMENT (all percentages relative to dry weight)
- Composition: 65 % sand (washed and sieved to 500 µm - 2 mm)
30 % kaolin clay
4.85 % alpha-cellulose
0.15 % TetraMin
0.1 % calcium carbonate
- Method of preparation (if artificial substrate): moistened and vigorously mixed with artificial freshwater (see above)
- Total organic carbon (TOC): 2 %

OTHER TEST CONDITIONS
- Light quality: White light (10 % UV)
- Photoperiod: 16 h light, 8 h dark
- Light intensity: 2500 lux

EFFECT PARAMETERS MEASURED: mortality, growth

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3 to 3.33
- Range finding study: no data
Reference substance (positive control):
not specified
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
50 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value recalculated to a standard sediment with 10 % organic carbon
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
10 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value
Key result
Duration:
14 d
Dose descriptor:
LC50
Effect conc.:
20.5 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value; 95% C.I.: 17.82 - 23.68 mg/kg dw
Key result
Duration:
14 d
Dose descriptor:
LOEC
Effect conc.:
30 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value
Remarks on result:
other: measured concentrations were ≥ 77 % of nominal concentrations; therefore nominal values are reported
Details on results:
No effect on growth was observed for all conditions tested.
Reported statistics and error estimates:
Calculation of EC50, LC50, 95% confidence limits: Spearman-Karber method
Data analysed with Statview F-4.5, significant differences (p < 0.05) between means were determined using t-test.

As a consequence of the high spiking efficiency and stability of exposure, toxicity data were calculated with nominal concentrations (see above: nominal and measured concentrations).

NOEC recalculated to a sediment with 10 % organic carbon: 50 mg/kg (dw).

Endpoint:
sediment toxicity: long-term
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material is phenanthrene a polycyclic aromatic hydrocarbon. It consists of a three-ring system and is beside others a major constituent of the target substance. The target substance anthracene oil (benzo[a]pyrene < 50 ppm, AOL) is composed of a broad range of PAH predominantly consisting of two up to four aromatic rings.
Toxic effects of anthracene oil on sediment organisms will be characterised by the range of PAH that constitute its composition. Therefore, results obtained from sediment toxicity tests with the AOL constituent phenanthrene can be used in order to characterise the toxic potential of anthracene oil on sediment dwelling organisms.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source test material is the substance phenanthrene. It was purchased from a commercial source and the analytical purity of the test substance is reported to be > 99 %. Hence, the study results are related to effects caused by phenanthrene.
The target material anthracene oil is a UVCB substance produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. PAH contained in AOL range from naphthalene up to pyrene and benzofluorenes. Two- and three-ring aromatics amount to about 50 % (typical concentration). Main constituent is phenanthrene present in a typical concentration of ca. 25 to 31 % (composite sample 7). PAH with four rings accumulate to about 10 %.

3. ANALOGUE APPROACH JUSTIFICATION
Properties of the target substance anthracene oil, relevant for environmental distribution and/or ecotoxicological effects, will be determined by the properties of the PAH that are constituents of anthracene oil. Under environmental conditions or during processing of the substance, environmentally available (volatile or water soluble) components of anthracene oil can be released. These will be PAH (mainly compounds consisting of two to four fused rings). In combination, they will specify the environmental distribution properties and ecotoxicological effects of anthracene oil as a whole. In addition, phenanthrene is considered to adequately represent the PAH-related environmental toxicity of AOL. It is one of the most abundant components in AOL with a high toxicity on sediment dwelling organisms. Therefore, phenanthrene is selected as marker substance to represent the environmental toxicity of AOL (see below Chapter 7.1.4. - Discussion). For these reasons, it is justified to use data determined for phenanthrene as a constituent of anthracene oil to characterise ecotoxicological effects (toxicity to soil dwelling organisms) of anthracene oil itself.
Reason / purpose for cross-reference:
read-across source
Principles of method if other than guideline:
Read-across to preceding entry:
Source test material: phenanthrene (generic, commercial product);
Reference: Verrhiest et al. 2001
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
50 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value (10 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Key result
Duration:
14 d
Dose descriptor:
LC50
Effect conc.:
ca. 100 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: original value (20.5 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Details on results:
No effect on growth was observed for all conditions tested.
Effect concentrations are reported as nominal values because recovery of the test substance (measured values) was higher than 77 % at all exposure levels and the test substance concentrations were stable during exposure
Endpoint:
sediment toxicity: long-term
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
acceptable for assessment
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 218 (Sediment-Water Chironomid Toxicity Test Using Spiked Sediment)
Version / remarks:
Draft test guideline, 2001
GLP compliance:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): phenanthrene
- Source of test material: Sigma-Aldrich
- Analytical purity: 99.5 %
Analytical monitoring:
yes
Details on sampling:
SEDIMENT
- Concentrations: all concentrations
- Sampling interval: at day 0, 14, and 28 of the experiment (day 0 = addition of larvae)

PORE WATER
- Concentrations: all concentrations
- Sampling interval: at day 0, 14, and 28 of the experiment (day 0 = addition of larvae)
- Sample volume: 2.5 mL (after centrifugation of the wet sediment)

OVERLYING WATER
- Concentrations: all concentrations
- Sampling interval: at day 0, 14, and 28 of the experiment (day 0 = addition of larvae)
- Sample volume: 2.5 mL directly from the surface water
Vehicle:
yes
Remarks:
acetone
Details on sediment and application:
PREPARATION OF SPIKED SEDIMENT
- Materials mixed: natural sediment (source: lake Drontermeer (reference location, The Netherlands)), test material dissolved in acetone, Dutch standard water (DSW; pH = 8.4, alkalinity 1.2 meq/L, hardness 200 mg/L CaCO3), midge food (Trouvit (Trouw, France) and Tetraphyll (Tetrawerke, Melle, Germany)
- Method of mixing: Borosilicate serum bottles (3 L) were filled with 630 g wet weight sediment and 2.1 L Dutch standard water and spiked using the appropriate stock solutions of acetone. A surplus of midge food, 0.15 g of a Trouvit and Tetraphyll mixture (20/1 w/w), was added to each serum bottle. For a thorough mechanical mixing of the spiked sediment suspension, the bottles were incubated on a rolling device in water at 20 ºC with a rolling speed of 25 rpm for one day.
- Details of spiking: the test material was dissolved in acetone as carrier solvent; for each test material concentration, a separate stock solution was made, resulting in equivalent solvent concentrations in all treatments (0.3 % v/v).
- Test setup: the definitive test was performed in 500 mL glass beakers. These were filled with 350 ml of the spiked sediment suspension as prepared above.
- Equilibration time in test vessels: 4 days
- Equilibration conditions: test conditions
- Controls: vehicle control (0.3 % acetone without test substance)
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium (stock solution and final test solution): 0.3 % (v/v; final test solution)
- Evaporation of vehicle before use: not specified
Test organisms (species):
Chironomus riparius
Details on test organisms:
TEST ORGANISM
- Source: no data
- Age of animals at beginning of exposure: first instar larvae (less than 24 h old)
- Feeding during test: no additional food applied (food is already incorporated in the spiked sediment, see above)
Study type:
laboratory study
Test type:
static
Water media type:
freshwater
Type of sediment:
natural sediment
Limit test:
no
Duration:
28 d
Exposure phase:
total exposure duration
Post exposure observation period:
no
Hardness:
no data
Test temperature:
20 °C
pH:
no data
Dissolved oxygen:
no data
Salinity:
no data
Ammonia:
no data
Nominal and measured concentrations:
Nominal concentrations: not reported
Measured concentrations (initial concentrations):
Sediment: 96.73, 237.7, 424.8, 503.3 µmol/kg dw
17.24, 42.37, 76.43, 89.7 mg/kg dw
At start, after 14 d and at the end (28 d) of the experiment, samples of sediment, overlying water, and pore water were collected to measure the actual test substance concentration. Detailed data are not reported.
Details on test conditions:
TEST SYSTEM
- Test container: Glass beaker (500 mL) covered with transparent foil
- Test suspension volume: 350 mL (sediment and overlying water); equilibration and settling of sediment: 4 d (start of exposure)
- Weight of wet sediment with and without pore water: no data
- Overlying water volume: no data
- Depth of sediment and overlying water: no data
- Aeration: gently aerated using a glass Pasteur pipette
- Replacement of evaporated test water, if any: yes

EXPOSURE REGIME
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 5 (2 additional replicates (without test organisms) were used for test substance analysis at day 0 and day 14)
- No. of replicates per control / vehicle control: 5
- Feeding regime: sediment was already spiked with food, no additional food applied

RENEWAL OF OVERLYING WATER: no

OVERLYING WATER CHARACTERISTCS
- Type of water: Dutch standard water (DSW) (reconstituted water: deionised water with 200 mg/L CaCl2 * 2 H2O, 180 mg/L MgSO4 * H2O, 100 mg/L NaHCO3, and 20 mg/L KHCO3; pH ca. 8.2)
- Properties of DSW: alkalinity 1.2 meq/L, hardness 200 mg/L CaCO3)

SOURCE OF NATURAL SEDIMENT
- Location and description of sampling site: Lake Drontermeer (reference location in The Netherlands)
- Contamination history of site: as reference location, no significant contamination to be expected (individual PAH concentrations below 0.01 mg/kg dw)

HANDLING OF NATURAL SEDIMENT: the sediment was provided by AquaSense/Grontmij (Amsterdam, The Netherlands). After sampling, it was homogenised and frozen to eliminate indigenous fauna, and kept at -20 °C until use. Three days before the spiking procedure was started, the sediment was thawed at 4 °C.

CHARACTERIZATION OF SEDIMENT: Dutch reference sediment (12 - 14 % organic matter determined by loss on ignition, 15.5 % clay)

OTHER TEST CONDITIONS
- Test beakers were placed in an incubator equipped with a mercury light source that contains a UV spectrum simulating that of natural light (50 μmol·m2·s-1 of visible light; HPI-T 400 W, Philips, Amsterdam, The Netherlands).
- Photoperiod: light/dark 16:8 h

EFFECT PARAMETERS MEASURED: mortality (28 day survival), delay of development from first instar to imagine stage; from week 2 onwards, the beakers were checked daily to remove, count and sex the emerging imagines.

VEHICLE CONTROL PERFORMED: yes
Reference substance (positive control):
not specified
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
ca. 100 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
development time
Remarks:
average day of emergence
Remarks on result:
other: original data recalculated to a standard sediment with 10 % organic carbon
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
76 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
development time
Remarks:
average day of emergence
Remarks on result:
other: original value (sediment with 12 to 14 % organic matter)
Key result
Duration:
28 d
Dose descriptor:
LOEC
Effect conc.:
ca. 90 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
development time
Remarks:
average day of emergence
Remarks on result:
other: highest dose tested; delay in emergence is > 10 % compared to the control
Key result
Duration:
28 d
Dose descriptor:
LC50
Effect conc.:
107 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks:
emergence ratio
Remarks on result:
other: 95% C.I.: 88 - 125 mg/kg dw; original values are given in µmol/kg dw

Table 3.1. LC50 values for the effect of phenanthrene on Chironomus riparius survival (emergence ratio) after 28 days exposure in sediment, expressed as sediment concentrations (μmol/kg dw), pore water concentrations (μM) and water concentrations (μM).

LC50 (95% c.i.)

sediment (μmol/kg dw)

pore water (μM)

water (μM)

Phenanthrene

600 (496 – 704)

0.44 (0.37 – 0.52)

0.31 (0.24 – 0.37)

Table 3.2. Lowest observed effect concentrations (LOEC) for the effect of phenanthrene on the average day of emergence of Chironomus riparius, expressed as sediment concentrations (μmol/kg dw), pore water concentrations (μM) and water concentrations (μM); m: males, f: females.

LOEC

Sediment (μmol/kg dw)

pore water (μM)

water (μM)

m

f

m

f

m

f

Phenanthrene

503.3

503.3

0.34

0.34

0.24

0.24

At lower concentrations, no statistically significant effects were observed.

Validity criteria fulfilled:
not specified
Conclusions:
The long-term (28 d) LC50 of phenanthrene to Chironomus riparius was determined to be 107 mg/kg dw (ca. 140 mg/kg dw when normalised to 10 % organic carbon) in a Sediment-Water Chironomid Toxicity Test using spiked sediment similar to OECD TG 218 (draft version). In the same test, the NOEC for the effect of phenanthrene on the average day of adult midge emergence was found to be 76 mg/kg dw (normalised value ca. 100 mg/kg dw).
Endpoint:
sediment toxicity: long-term
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material is phenanthrene a polycyclic aromatic hydrocarbon. It consists of a three-ring system and is beside others a major constituent of the target substance. The target substance anthracene oil (benzo[a]pyrene < 50 ppm, AOL) is composed of a broad range of PAH predominantly consisting of two up to four aromatic rings.
Toxic effects of anthracene oil on sediment organisms will be characterised by the range of PAH that constitute its composition. Therefore, results obtained from sediment toxicity tests with the AOL constituent phenanthrene can be used in order to characterise the toxic potential of anthracene oil on sediment dwelling organisms.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source test material is the substance phenanthrene. It was purchased from a commercial source (Sigma-Aldrich) and the analytical purity of the test substance is reported to be 99.5 %. Hence, the study results are related to effects caused by phenanthrene.
The target material anthracene oil is a UVCB substance produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. PAH contained in AOL range from naphthalene up to pyrene and benzofluorenes. Two- and three-ring aromatics amount to about 50 % (typical concentration). Main constituent is phenanthrene present in a typical concentration of ca. 25 to 31 % (composite sample 7). PAH with four rings accumulate to about 10 %.

3. ANALOGUE APPROACH JUSTIFICATION
Properties of the target substance anthracene oil, relevant for environmental distribution and/or ecotoxicological effects, will be determined by the properties of the PAH that are constituents of anthracene oil. Under environmental conditions or during processing of the substance, environmentally available (volatile or water soluble) components of anthracene oil can be released. These will be PAH (mainly compounds consisting of two to four fused rings). In combination, they will specify the environmental distribution properties and ecotoxicological effects of anthracene oil as a whole. In addition, phenanthrene is considered to adequately represent the PAH-related environmental toxicity of AOL. It is one of the most abundant components in AOL with a high toxicity on sediment dwelling organisms. Therefore, phenanthrene is selected as marker substance to represent the environmental toxicity of AOL (see below Chapter 7.1.4. - Discussion). For these reasons, it is justified to use data determined for phenanthrene as a constituent of anthracene oil to characterise ecotoxicological effects (toxicity to soil dwelling organisms) of anthracene oil itself.
Reason / purpose for cross-reference:
read-across source
Principles of method if other than guideline:
Read-across to preceding entry:
Source test material: phenanthrene (generic, commercial product);
Reference: Bleeker et al. 2003
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
ca. 100 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
development time
Remarks:
average day of emergence
Remarks on result:
other: original value (76 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Key result
Duration:
28 d
Dose descriptor:
LC50
Effect conc.:
ca. 140 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks:
emergence ratio
Remarks on result:
other: original value (107 mg/kg dw) recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Conclusions:
The long-term (28 d) LC50 of phenanthrene to Chironomus riparius was determined to be ca. 140 mg/kg dw (normalised to 10 % organic carbon) in a Sediment-Water Chironomid Toxicity Test using spiked sediment similar to OECD TG 218 (draft version). In the same test, the NOEC for the effect of phenanthrene on the average day of adult midge emergence was found to be ca 100 mg/kg dw (normalised to 10 % organic carbon).
The test results of the source substance phenanthrene (marker substance) are adopted as representative for the target substance anthracenen oil.
Endpoint:
sediment toxicity: short-term
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions
Qualifier:
equivalent or similar to guideline
Guideline:
other: US EPA/600/R-94/025
Version / remarks:
June 1994
Qualifier:
equivalent or similar to guideline
Guideline:
ASTM E1367 (10-day Static Sediment Toxicity Test (Media: Sediment-estuarine))
Version / remarks:
1992
Principles of method if other than guideline:
Besides phenanthrene, three other PAH were tested in this study. Substances were applied in individual tests and combined as approx. equitoxic mixture in order to evaluate, whether effects observed were additive or deviating from additivity. In addition, the effect of UV radiation on the toxicity of the individual compounds was investigated.
GLP compliance:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): Phenanthrene
- no further information reported
Analytical monitoring:
yes
Details on sampling:
SEDIMENT
- Concentrations: all concentrations (taken from chemistry replicates)
- Sampling interval: at initiation of the test
- Sample storage before analysis: yes, after pretreatment
Vehicle:
yes
Remarks:
acetone
Details on sediment and application:
PREPARATION OF SPIKED SEDIMENT
- Pooling or mixing of different substrates: no
- Details of spiking: a stock solution was prepared by adding test material to 200 mL of acetone resulting in a concentration of 5.79 phenanthrene/mL. A volume of the stock solution, necessary to produce the desired sediment concentration was added to a 1-galon (3.785 L) glass rolling jar. The acetone was evaporated by directing a stream of clean, dry air into the mouth of the jar while it was slowly rolled on its side leaving a uniform layer of test material deposited on the side of the jar. A total of 1080 mL of sediment was added to the jar. The jar was sealed with a Teflon-lined lid, rolled for 2 h at 15 °C stored overnight at 4 °C, rolled for an additional 2 h at 15 °C and stored for 27 d at 4 °C bevor the initiation of the toxicity tests.
- Equilibration time: 27 d
- Equilibration conditions: closed vessels stored at 4 °C (see above)
- Controls: solvent controls, chemical controls
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium (stock solution and final test solution): not applicable
- Evaporation of vehicle before use: yes, see above
Test organisms (species):
Rhepoxynius abronius
Details on test organisms:
TEST ORGANISM
- Common name: freeburrowing, marine infaunal amphipod
- Source: lower Yaquina Bay, Oregon, USA
- Details on collection: 4 to 6 days before the experimet with a small biological dredge
- Age of parental stock (mean and range, SD): not applicable
- Breeding conditions: not applicable
- Age of animals at beginning of exposure: adult animals
- Size of animals at beginning of exposure: 3 - 4 mm in length
- Feeding during test: no

ACCLIMATION: no data
Study type:
laboratory study
Test type:
static
Water media type:
saltwater
Type of sediment:
natural sediment
Limit test:
no
Duration:
10 d
Exposure phase:
total exposure duration
Post exposure observation period:
no
Hardness:
no data
Test temperature:
15 °C
pH:
no data
Dissolved oxygen:
no data
Salinity:
28 ppt
Ammonia:
no data
Nominal and measured concentrations:
Nominal: 1.1, 1.53, 2.18, 3.15, 4.45, 6.36, 9.09 mg/g OC (organic carbon)
Measured: 0 (vehicle control) 0.8, 1.26, 1.69, 2.08, 3.65, 4.17, 6.94 mg/g OC (organic carbon)
Details on test conditions:
TEST SYSTEM
- Test container: Glass beakers (1 L) incubated in water bath (15 °C)
- Test sediment depth: 2 cm (volume ca. 175 ml)
- Weight of wet sediment with and without pore water: no data
- Overlying water: sediment was covered with 775 mL seawater
- Aeration: through 1-mL pipettes

EXPOSURE REGIME
- No. of organisms per container: 20
- No. of replicates per treatment group: 1
- No. of replicates per control / vehicle control: 5 (vehicle control), 1 chemistry replicate (for sediment analysis)
- Feeding regime: no feeding

OVERLYING WATER CHARACTERISTCS
- Type of water: seawater (interstitial salinity adjusted to 28 ppt)

SOURCE OF NATURAL SEDIMENT
- Location and description of sampling site: uncontaminated sediment from McKinney Slough, Oregon, USA

HANDLING OF NATURAL SEDIMENT
- Time of collection: no data
- Core depth: no data
- Water depth: no data
- Treatment: in order to remove macrobiota and debris, sieved through 0.5 mm mesh size into seawater with a salinity of 28 ppt; alowed to settle for 24 h before spiking
- Storage conditions: no data
- Storage duration (prior to test): no data

CHARACTERIZATION OF SEDIMENT
- Particle size distribution: poorly sorted medium silt
- Solid content: 40.6 %
- Presence of macrophytes/animals: removed by sieving (see above)
- Sediment sieved: yes (see above)
- Total organic carbon: 2.9 % (measured in test samples)

OTHER TEST CONDITIONS
- Photoperiod: no data, according to US EPA guideline 24 h darkness

EFFECT PARAMETERS MEASURED: Mortality

VEHICLE CONTROL PERFORMED: yes

PHOTOTOXICITY TEST (from main experiment)
Surviving individuals from each replicate (main experiment) were placed into watch glasses (diameter: 110 mm, 30 ml clean 28 ppt seawater):
- Exposition to UV radiation: 1 h, growth chamber, 15 °C
- UV radiation and visible light: simulated UV radiation in full sun light (combination of UV-A 340 nm and UV-B 313 nm fluorescent lamps).
- Vessels: watch glasses covered with nylon window screening and an additional cellulose acetate layer to avoid UV-C radiation.
- Light quality: UV-B (280-320 nm): 58 ± 2 µW/cm²; UV-A (321-400 nm): 167 ± 11 µW/cm²; visible light (401-800 nm): 1517 ± 77 µW/cm²; no UV-C (below 280 nm).
- Effect parameter: mortality and ability to rebury

TEST CONCENTRATIONS
- Spacing factor for test concentrations: ca. 1.42
- Range finding study: yes
- Test concentrations: 1.14, 2.00, 4.85, 9.32 mg/g OC
- Results used to determine the conditions for the definitive study: concentrations in the main were based on the LC50 value obtained in the preliminary study.
Reference substance (positive control):
yes
Remarks:
Cadmium (applicated as CdCl2 in control experiment)
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
ca. 220 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: recalculated to a standard sediment with 10 % organic carbon
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
64.4 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: recalculated from the original value to a sediment with 2.9 % organic carbon
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
2.22 other: mg/g OC (organic carbon content)
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: 95% C.I.: 3.00 - 3.17; original value reported in the publication
Key result
Duration:
10 d
Dose descriptor:
LC10
Effect conc.:
ca. 204 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: determined from data within the publication using a log-logistic dose-response relationship; original value normalised to a standard sediment with 10 % organic carbon
Details on results:
- Mortality of test animals at end of exposure period: 100 % in the three highest test concentrations, 20 and 0 in the two lowest test concentration
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Test result of reference substance: 0.8 mg Cd/L (main test), 1.35 mg Cd/L (preliminary test)
- Acceptable effect levels: range from 0.19 to 2.00 Cd mg/L
Reported statistics and error estimates:
Probit analysis or the trimmed Spearman–Karber method were used to calculate the 10-d LC50 value based on measured concentrations of each of the four compounds in treatments spiked with individual chemicals

Photoinduced toxicity: 6.5 % dead or unable to rebury, i.e. reburial was not strongly influenced by irradiation with UV.

Validity criteria fulfilled:
yes
Remarks:
mortality in control sediments ≤ 1.7 %
Conclusions:
In a 10 day toxicity test similar/according to US EPA guideline EPA/600/R-94/025 with the freeburrowing marine infaunal amphipod Rhepoxynius abronius, a LC50 of 220 mg/kg sediment dw was determined (value normalised to a sediment withn 10 % organic carbon). Applying a log-logistic dose-response relationship on the available data, a LC10 value of 204 mg/kg sediment dw was estimated (value normalised to 10 % organic carbon).
Endpoint:
sediment toxicity: short-term
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material is phenanthrene a polycyclic aromatic hydrocarbon. It consists of a three-ring system and is beside others a major constituent of the target substance. The target substance anthracene oil (benzo[a]pyrene < 50 ppm, AOL) is composed of a broad range of PAH predominantly consisting of two up to four aromatic rings.
Toxic effects of anthracene oil on sediment organisms will be characterised by the range of PAH that constitute its composition. Therefore, results obtained from sediment toxicity tests with the AOL constituent phenanthrene can be used in order to characterise the toxic potential of anthracene oil on sediment dwelling organisms.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source test material is the substance phenanthrene. Analytical purity of the material is not reported but considered to be high, as a commercial product is used in the study. Hence, the study results are related to effects caused by phenanthrene.
The target material anthracene oil is a UVCB substance produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. PAH contained in AOL range from naphthalene up to pyrene and benzofluorenes. Two- and three-ring aromatics amount to about 50 % (typical concentration). Main constituent is phenanthrene present in a typical concentration of ca. 25 to 31 % (composite sample 7). PAH with four rings accumulate to about 10 %.

3. ANALOGUE APPROACH JUSTIFICATION
Properties of the target substance anthracene oil, relevant for environmental distribution and/or ecotoxicological effects, will be determined by the properties of the PAH that are constituents of anthracene oil. Under environmental conditions or during processing of the substance, environmentally available (volatile or water soluble) components of anthracene oil can be released. These will be PAH (mainly compounds consisting of two to four fused rings). In combination, they will specify the environmental distribution properties and ecotoxicological effects of anthracene oil as a whole. In addition, phenanthrene is considered to adequately represent the PAH-related environmental toxicity of AOL. It is one of the most abundant components in AOL with a high toxicity on sediment dwelling organisms. Therefore, phenanthrene is selected as marker substance to represent the environmental toxicity of AOL (see below Chapter 7.1.4. - Discussion). For these reasons, it is justified to use data determined for phenanthrene as a constituent of anthracene oil to characterise ecotoxicological effects (toxicity to soil dwelling organisms) of anthracene oil itself.
Reason / purpose for cross-reference:
read-across source
Principles of method if other than guideline:
Read-across to preceding entry:
Source test material: phenanthrene (generic, commercial product);
Reference: Swartz et al. 1997
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
ca. 220 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: recalculated to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Key result
Duration:
10 d
Dose descriptor:
LC50
Effect conc.:
2.22 other: mg/g OC (organic carbon content)
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: 95% C.I.: 3.00 - 3.17; original value reported for the source substance
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Key result
Duration:
10 d
Dose descriptor:
LC10
Effect conc.:
ca. 204 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: determined from data within the publication using a log-logistic dose-response relationship; original value normalised to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Details on results:
Photoinduced toxicity: 6.5 % dead or unable to rebury, i.e. reburial was not strongly influenced by irradiation with UV.
Conclusions:
In a 10 day toxicity test similar/according to US EPA guideline EPA/600/R-94/025 with the freeburrowing marine infaunal amphipod Rhepoxynius abronius, a LC50 of 220 mg/kg sediment dw was determined (value normalised to a sediment with 10 % organic carbon). Applying a log-logistic dose-response relationship on the available data, a LC10 value of 204 mg/kg sediment dw was estimated (value normalised to 10 % organic carbon).
The test results of the source substance phenanthrene (marker substance) are adopted as representative for the target substance anthracenen oil.
Endpoint:
sediment toxicity, other
Remarks:
short-term and long-term
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
no guideline available
Principles of method if other than guideline:
Sediment toxicity tests with an estuarine copepod using two different PAH (phenanthrene and fluoranthene) and diesel fuel. PAH effects on reproduction were determined besides 96h LC50 values and effects on grazing rates.
GLP compliance:
not specified
Specific details on test material used for the study:
- Name of test material (as cited in study report): Phenanthrene, PHN
- Source of test material: Aldrich Chemical Co., Milwaukee, WI, USA)
- Analytical purity:98%
Analytical monitoring:
yes
Details on sampling:
SEDIMENT
- Concentrations: all concentrations
- Sampling interval: at the start of the experiments
- Sample storage before analysis: no data
Vehicle:
yes
Remarks:
acetone
Details on sediment and application:
PREPARATION OF SPIKED SEDIMENT
- Pooling or mixing of different substrates: no
- Details of spiking: stored sediment was fully homogenised with the overlaying water and 133 g of wet sediment (20 g dry wt) was transferred to 250 mL beakers and vigorously stirred under vortex. The appropriate amount of phenanthrene carried in 0.2 mL of acetone was spiked on the slurry and stirred for 4 h.
- Equilibration time: 3 to 5 weeks
- Equilibration conditions: in the dark at 4 °C
- Controls: negative control and solvent control
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium: 0.2 mL
- Evaporation of vehicle before use: no
Test organisms (species):
other: Schizopera knabeni Lang
Details on test organisms:
TEST ORGANISM
- Common name: estuarine copepod
- Strain/clone:
- Justification for species other than prescribed by test guideline:
- Source: mono-specific laboratory culture started with a stock collected in October 1993 form the surface sediment from an intertidal mud flat of a Spartina alterniflora salt-marsh at Port Fourchon, Louisiana.
- Details on collection: copepods were harvested by sieving the culture medium through a 125 µm mesh. Retained copepods were sorted under a stereo microscope.
- Age of parental stock (mean and range, SD):
- Breeding conditions: animals were cultured sediment-free in 500 mL Erlenmeyer flasks at room temperature with 25‰ artificial seawater (ASW) fully renewed every 14 days. Copepods were fed twice a week with a mixture of Chaetocerous muelleri (a planktonic diatom) and Microfeast Plus Larval Diet.
- Age of animals at beginning of exposure: adult
- Feeding during test: see below (Details on test conditions)

ACCLIMATION
- no data
Study type:
laboratory study
Test type:
static
Water media type:
saltwater
Remarks:
artificial saltwater (ASW)
Type of sediment:
natural sediment
Remarks:
mudflat marsh sediment
Limit test:
no
Duration:
4 d
Exposure phase:
total exposure duration
Remarks:
4 d toxicity test (short-term exposure)
Duration:
14 d
Exposure phase:
reproduction phase
Remarks:
reproductive output bioassay (long-term exposure)
Post exposure observation period:
no
Test temperature:
25 °C
Dissolved oxygen:
4 d toxicity test: 4.8 - 4.9 mg/L (end and start of the experiment)
Reproduction output bioassay: 4.36-4.18 mg/L (end and start of the experiment)
Salinity:
25 ‰
Nominal and measured concentrations:
Only measured concentrations reported:
Total number of sediment concentrations: 56 ± 4, 126 ± 7, 261 ± 15, 514 ± 21, 1030 ± 52 mg/kg sediment dw
4 day toxicity test: 126, 261, 514, 1030 mg/kg sediment dw
reproductive output bioassay: 56, 126, 261 mg/kg sedimentn dw
Details on test conditions:
4-DAY SHORT-TERM TEST
TEST SYSTEM
- Test container (material, size): carrier glass vials (28x45 mm)
- Sediment volume: 1 mL
- Weight of wet sediment with and without pore water: sediment dw: ca. 160 mg
- Overlying water volume: 10 mL
- Depth of sediment and overlying water: 2-3 mm
- Aeration: no

EXPOSURE REGIME
- No. of organisms per container (treatment): 15
- No. of replicates per treatment group: 4
- No. of replicates per control / vehicle control: 4 / 4
- Feeding regime: no feeding

REPRODUCTIVE OUTPUT BIOASSAY
TEST SYSTEM
- Test container (material, size): carrier glass vials (28x45 mm)
- Sediment volume: 1 mL
- Weight of wet sediment with and without pore water: sediment dw: ca. 160 mg
- Overlying water volume: 10 mL
- Depth of sediment and overlying water: 2-3 mm
- Aeration: no

EXPOSURE REGIME
- No. of organisms per container (treatment): one copulating pair - an adult male clasping a pre-adult female (copepodeite V) or. less freqently , an adult female
- No. of replicates per treatment group: 10
- No. of replicates per control / vehicle control: 10 / 10
- Feeding regime: once at the start of the test
- Type and preparation of food: commercial feed, Microfeast Plus Larval Diet
- Amount of food: 0.1 mg, single dose

RENEWAL OF OVERLYING WATER: no

OVERLYING WATER CHARACTERISTCS
- Type of water (e.g. deionized, ground water, sea water, Elendt medium acc. to OECD 219): artificial salt water

SOURCE OF NATURAL SEDIMENT
- Location and description of sampling site: mudflat in a Spartina alterniflora salt marsh near Cocoldrie, Louisiana
- Contamination history of site: typical total PAH concentration is 0.26 mg/kg sediment dw

HANDLING OF NATURAL SEDIMENT
- Time of collection: no data
- Core depth: no data
- Water depth: no data
- Storage conditions: stock test-sediment was prepared by sieving the mud through a 45-µm mesh. The sediment that went through the sieve was allowed to settle overnight at 4 °C. The supernatant was removed by aspiration, and the sediment was autoclaved. In order to mimic the mudflat superficial layer, a liquid slurry (dry-t-wet ration = 0.15 was created by homogenising the autoclaved sediment with the appropriate volume of 25 ‰ ASW. The stock sediment was stored at 4 °C.
- Storage duration (prior to test): no data

CHARACTERIZATION OF SEDIMENT
- Total organic carbon (%): 1.5

OTHER TEST CONDITIONS
- Photoperiod: no lighting, test vessels were kept in the dark.
- Storage conditions of test vials: vials were placed in moisture chambers (loosely covered plastic containers underlined with soaked paper towels, which created a humid environment to retard evaporation form experimental dishes). Chambers were kept in an incubator with no illumination.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): 4 d toxicity test: mortality, at the end of experiment; reproduction output bioassay: mortality, realised offspring (nauplii and copepodites), total offspring (eggs still present in females and realise offspring), each at the end of the experiment

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: ca. 2 to 2.2
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
3 493 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks:
4 d toxicity test
Remarks on result:
other: original value normalised to a standard sediment with 10 % organic carbon
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
524 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks:
4 d toxicity test
Remarks on result:
other: original value
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
< 373 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
14 d reproduction test
Remarks on result:
other: original value normalised to a standard sediment with 10 % organic carbon
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
< 56 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
14 d reproduction test
Remarks on result:
other: original value, lowest concentration tested; the data reported do not allow a valid extrapolation to effect levels at lower concentrations (e.g. EC10)
Key result
Duration:
14 d
Dose descriptor:
EC50
Effect conc.:
347 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
14 d reproduction test
Remarks on result:
other: original value normalised to a standard sediment with 10 % organic carbon
Key result
Duration:
14 d
Dose descriptor:
EC50
Effect conc.:
52 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
14 d reproduction test
Remarks on result:
other: original value
Details on results:
10 d toxicity test
- Mortality of test animals at end of exposure period: in control replicates for all experiments, mortality ranged from 0 to 13 %. At the lowest concentration, mortality just above background was observed. At the next higher concentration, about 10 % of the test animals had died. At the two highest test concentrations, mortality was ca. 60 and 80 %.

Reproduction output assay
- Mortality of test animals at end of exposure period: no adult mortality was observed in the controls. Adult survival was 70 % in all phenanthrene treatments (three different concentrations).
- No. of offspring produced: in the three test concentrations, the mean realised offspring production was significantly reduced to 58, 35, and 33 % compared to uncontaminated control sediments
- Egg production: The mean number of eggs carried by surviving females at test termination was not significantly different among treatments. Mean clutch size determined for surviving ovigerous females was 17.5, 19.1, and 19.3 for control, solvent control, and PAH exposed females, respectively. Clutch size was not significantly different among all treatments.
Reported statistics and error estimates:
Mortality, grazing rate, number of offspring, and clutch size were analysed using on-way ANOVA. Mortality data were arcsine-square-root transformed prior to analysis so that the data conformed to the assumption of homogeneity of variances.
LC50 values were computed using probit analysis. Point estimates for sublethal effects (grazing rate and reproductive output) were calculated using linear interpolation combined with bootstrapping, or ICp method (US EPA, 1994a), IC50 estimates the contaminant concentrations causing a 50% reduction in the measured endpoint in relation to the control.
All the analyses were performed using TOXCALC version 5.0 (Tidepool Software; McKinleyvill, CA, USA).
Validity criteria fulfilled:
yes
Remarks:
mortality in controls ≤ 13 %, dissolved oxygen above 4.2 % (ca. 69 % of water saturation)
Conclusions:
Phenanthrene showed a low acute toxicity to the marine infaunal amphipod Schizopera knabeni (LC50 = 3493 mg/kg sediment dw, normalised to an organic carbon content of 10 %). In a 14 day reproduction output experiment based on the number of live offspring, already the lowest test concentration (373 mg/kg sediment dw, normalised value) resulted in a significant impairment in the number of realised offspring (number of nauplii and copepodites) (reduction to 58 %). EC50 was determined to be 208 mg/kg sediment dw (value normalised to 10 % organic carbon). As the lowest tested concentration is even higher than the EC50 value, an extrapolation to effect levels at lower concentrations (e.g. EC10) will provide not reliable values due to a high level of uncertainty.
Endpoint:
sediment toxicity, other
Remarks:
short-term and long-term
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material is phenanthrene a polycyclic aromatic hydrocarbon. It consists of a three-ring system and is beside others a major constituent of the target substance. The target substance anthracene oil (benzo[a]pyrene < 50 ppm, AOL) is composed of a broad range of PAH predominantly consisting of two up to four aromatic rings.
Toxic effects of anthracene oil on sediment organisms will be characterised by the range of PAH that constitute its composition. Therefore, results obtained from sediment toxicity tests with the AOL constituent phenanthrene can be used in order to characterise the toxic potential of anthracene oil on sediment dwelling organisms.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source test material is the substance phenanthrene. It was purchased from a commercial source (Aldrich Chemical Co.) and the analytical purity of the test substance is reported to be 98 %. Hence, the study results are related to effects caused by phenanthrene.
The target material anthracene oil is a UVCB substance produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. PAH contained in AOL range from naphthalene up to pyrene and benzofluorenes. Two- and three-ring aromatics amount to about 50 % (typical concentration). Main constituent is phenanthrene present in a typical concentration of ca. 25 to 31 % (composite sample 7). PAH with four rings accumulate to about 10 %.

3. ANALOGUE APPROACH JUSTIFICATION
Properties of the target substance anthracene oil, relevant for environmental distribution and/or ecotoxicological effects, will be determined by the properties of the PAH that are constituents of anthracene oil. Under environmental conditions or during processing of the substance, environmentally available (volatile or water soluble) components of anthracene oil can be released. These will be PAH (mainly compounds consisting of two to four fused rings). In combination, they will specify the environmental distribution properties and ecotoxicological effects of anthracene oil as a whole. In addition, phenanthrene is considered to adequately represent the PAH-related environmental toxicity of AOL. It is one of the most abundant components in AOL with a high toxicity on sediment dwelling organisms. Therefore, phenanthrene is selected as marker substance to represent the environmental toxicity of AOL (see below Chapter 7.1.4. - Discussion). For these reasons, it is justified to use data determined for phenanthrene as a constituent of anthracene oil to characterise ecotoxicological effects (toxicity to soil dwelling organisms) of anthracene oil itself.
Reason / purpose for cross-reference:
read-across source
Principles of method if other than guideline:
Read-across to preceding entry:
Source test material: phenanthrene (generic, commercial product);
Reference: Lotufo 1997
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
3 493 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks:
4 d toxicity test
Remarks on result:
other: original value (524 mg/kg dw) normalised to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted as representative for the target substance anthracene oil
Key result
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
< 373 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
14 d reproduction test
Remarks on result:
other: original value (< 56 mg/kg dw) normalised to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Key result
Duration:
14 d
Dose descriptor:
EC50
Effect conc.:
347 mg/kg sediment dw
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
14 d reproduction test
Remarks on result:
other: original value (52 mg/kg bw) normalised to a standard sediment with 10 % organic carbon
Remarks:
the test result of the source substance is adopted for the target substance anthracene oil
Conclusions:
Phenanthrene showed a low acute toxicity to the marine infaunal amphipod Schizopera knabeni (LC50 = 3493 mg/kg sediment dw, normalised to an organic carbon content of 10 %). In a 14 day reproduction output experiment based on the number of live offspring, already the lowest test concentration (373 mg/kg sediment dw, normalised value) resulted in a significant impairment in the number of realised offspring (number of nauplii and copepodites) (reduction to 58 %). EC50 was determined to be 347 mg/kg sediment dw (value normalised to 10 % organic carbon). As the lowest tested concentration is even higher than the EC50 value, an extrapolation to effect levels at lower concentrations (e.g. EC10) will provide not reliable values due to a high level of uncertainty.
The test results of the source substance phenanthrene (marker substance) are adopted as representative for the target substance anthracene oil.

Description of key information

No data on anthracene oil (benzo[a]pyrene < 50 ppm, AOL) as such is available. Phenanthrene, a key constituent of anthracene oil, has been adopted as marker substance. Toxicity values (short- and long-term) have been located for freshwater and estuarine/marine sediment organisms. The values displayed below (all normalised to 10 % organic carbon) are the lowest for the respective endpoint.

Key value for chemical safety assessment

EC50 or LC50 for freshwater sediment:
68 mg/kg sediment dw
EC50 or LC50 for marine water sediment:
220 mg/kg sediment dw
EC10, LC10 or NOEC for freshwater sediment:
50 mg/kg sediment dw
EC10, LC10 or NOEC for marine water sediment:
373 mg/kg sediment dw

Additional information

Studies on sediment toxicity with anthracene oil itself could not be identified. But data obtained with phenanthrene as test substance are available. These will be used to characterise the toxicity of AOL to sediment dwelling organisms.

AOL contains mainly three-ring aromatic compounds and to a lesser extent PAHs with four rings (see Chapter 1.). Two-ring aromatics are minor. In combination, these substances will constitute the sediment toxicity of AOL.

Main constituent of AOL is phenanthrene. It is present in AOL in concentrations up to 31 % (average 28 %). Toxicity to sediment dwelling organisms of phenanthrene is comparable to the sediment toxicity of other constituents of AOL. Even if some other AOL constituent can have a somewhat higher toxicity to sediment organisms, this is counterbalanced by the much higher concentration of phenanthrene. Therefore, the sediment toxicity of phenanthrene is considered to be representative for total AOL integrating the toxic effects of all its other PAH constituents. Thus, phenanthrene can be used as substitute and marker substance to characterise the sediment toxicity of total AOL.

Sediment toxicity of phenanthrene has extensively been evaluated in the EU Risk Assessment Report of pitch, coal tar, high-temp. (EU 2008). The sediment toxicity of phenanthrene reported here is based on this evaluation.

There are freshwater test results available for three different taxonomic groups (annelids, crustaceans, and insect larvae) representing different habitats and living conditions. Short-term result were obtained in two tests with annelids and insect larvae, while long-term tests have been conducted with annelids, crustaceans, and insect larvae. Studies with annelids are not included in IUCLID, because test results are higher compared to the other test species (annelids seem to be less susceptible to the sediment toxicity of phenanthrene). Sediment tests in saltwater (two short-term and one long-term test) were performed with crustaceans as test species.

The NOEC/LC10 values (normalised to standard condition of 10 % organic carbon) for freshwater long-term tests ranged from 50 (crustacean) to above 1000 (annelid) mg/kg sediment dw. Taking all tests into account, crustaceans and insect larvae appear to be the most sensitive species. The lowest long-term NOEC (50 mg/kg sediment dw) resulting from a test with Hyalella azteca (Verrhiest et al. 2001) is used for the derivation of the PNEC(freshwater sediment).

The results of the three toxicity tests with sediment organisms in saltwater gave evidence, that the sediment toxicity of phenanthrene to saltwater organisms is not higher than to freshwater organisms. Effect concentrations determined with the marine infaunal amphipod Rhepoxynius apronius (short-term tests) were all above 200 mg/kg sediment dw (values normalised to standard condition of 10 % organic carbon). The long-term NOEC (reproduction) determined with the sediment dwelling copepod Schizopera knabeni (Lotufo 1997) was shown to be lower than 370 mg/kg sediment dw (value normalised to 10 % organic carbon, lowest concentration tested). As the marine sediment toxicity of phenanthrene appears to be not higher than freshwater sediment toxicity, the NOEC of 50 mg/kg dw for freshwater sediment is used as starting point for the derivation of the PNEC(saltwater sediment).

Reference:

EU (2008): European Union Risk Assessment Report - Coal-Tar Pitch, high temperature. The Netherlands (rapporteur) (URL:https://echa.europa.eu/documents/10162/433ccfe1-f9a5-4420-9dae-bb316f898fe1)