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Sediment toxicity

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Reference
Endpoint:
sediment toxicity: long-term
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October 10 to November 13, 2014.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed in accordance with OECD & ASTM test guidelines in compliance with GLP.
Qualifier:
according to
Guideline:
OECD Guideline 218 (Sediment-Water Chironomid Toxicity Test Using Spiked Sediment)
Deviations:
no
Qualifier:
according to
Guideline:
other: ASTM Standard E 1706-05: Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates.
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
Not applicable.
Analytical monitoring:
yes
Details on sampling:
Samples of stock solutions were collected at preparation to confirm the concentrations used to dose the sediments and were held under refrigeration for 3 days prior to analysis. One of the additional replicate test chambers prepared for each treatment and control group was collected for analysis of overlying water, pore water and sediment on each of Days 0 and 7 and at test termination on Day 28, and were processed immediately for analysis.
Vehicle:
yes
Details on sediment and application:
Formulated sediment, as described in Kemble et al. was used as the test sediment. This formulated sediment is similar to that described in OECD Guideline 218, but uses alpha-cellulose as its source of organic matter instead of peat moss. Alpha-cellulose was selected by Kemble et al. as a more standardized source of organic matter than peat moss. The sediment was composed of 0.01% humic acid, 0.99% dolomite, 5% alpha-cellulose, 14% silt and clay (kaolin clay) and 80% industrial quartz sand. The dry constituents of the sediment were mixed in a PK Twinshell® mixer for 20 minutes, and the batch was stored under ambient conditions until used. The pH of the dry sediment was determined prior to use in the test and was within the desired range of 7.0 (± 0.5). The final pH of a 1:1 ratio of the sediment and water was 6.8. A sample of the formulated sediment used in the test was sent to Agvise Laboratories, Northwood, North Dakota, for characterization and analysis of total organic carbon (TOC). The percent organic carbon of the sediment was determined to be 2.9%.
Test organisms (species):
Chironomus riparius
Details on test organisms:
The midge, Chironomus riparius, was selected as the test species for this study. This species is representative of an important group of aquatic invertebrates and was selected for use in the study based upon past history of use in the laboratory.
Midges used in the test were obtained as egg masses from cultures maintained by Wildlife International, Easton, Maryland. The identity of the species was verified by the supplier of the original culture Environmental Consulting and Testing, Superior, Wisconsin.
Larvae used to start the test were collected from 18 separate egg masses and were hatched in water from the same source and at approximately the same temperature as was used during the test.
At the time of test initiation, the larvae were 1 to 4 days old. During the holding period, water temperatures ranged from 19.2 to 20.0°C, the pH of the water ranged from 8.3 to 8.7, and the dissolved oxygen concentrations were ≥89 mg/L (≥8.0% of saturation).
The larvae showed no signs of disease or stress prior to test initiation.
Feeding: During the holding period, the midge larvae were fed a suspension of TetraMin® flake food daily. During the test, the midges were fed ground TetraMin® flake food approximately three times per week beginning on Day 0, excluding the additional abiotic test chambers prepared for analytical sampling. Individual aliquots of approximately 20 to 30 mg of the ground food were supplied to each test chamber at each feeding.
Study type:
laboratory study
Test type:
static
Water media type:
freshwater
Type of sediment:
artificial sediment
Limit test:
no
Duration:
28 d
Exposure phase:
total exposure duration
Post exposure observation period:
No post exposure observation period specified in the study report.
Hardness:
144 – 220 mg/L as CaCO3
Test temperature:
Water temperatures were within the 20 ± 2°C range established for the test.
Temperature Range: 20.0 – 21.1°C
pH:
Measurements of pH ranged from 7.8 to 9.0 during the test.
Dissolved oxygen:
Dissolved oxygen concentrations remained ≥7.0 mg/L (≥78% of saturation) throughout the test.
Salinity:
Not applicable - freshwater study.
Ammonia:
<0.17 – 1.70 mg/L as NH3
Conductivity:
Not specified
Nominal and measured concentrations:
Nominal concentrations selected for use in this study were 63, 125, 250, 500 and 1000 mg/kg dry weight of sediment.
Measured concentrations ranged from approximately 90.4 to 98.0% of nominal.
Details on test conditions:
EXPERIMENTAL DESIGN
Midges were exposed to a geometric series of five test concentrations, a negative control (untreated formulated sediment), and a solvent control (acetone) for 28 days under static test conditions. Four replicate test chambers were maintained in each treatment and control group, with 20 midges in each test chamber, for a total of 80 midges per test concentration. Each test chamber contained sediment and overlying water. An additional four replicates were prepared in each treatment and control group for use in analytical sampling. No midges were placed in the additional replicates sampled on Day 0, but those sampled on Days 7 and 28 had midges added at the same time as the “biological” replicates on Day 0. These additional replicates were not used to evaluate the biological response of the test organisms.
Test concentrations in the sediment were prepared on a mg/kg dry weight basis. Nominal test concentrations were selected in consultation with the Sponsor based on exploratory range-finding toxicity data, and were 63, 125, 250, 500 and 1000 mg Reofos 35/kg of sediment. Test concentrations were measured in samples of overlying water, pore water, and sediment collected at the beginning of the test, on Day 7, and at the end of the test. The results of the study are based on mean measured test concentrations in the sediment.
The water/sediment systems in the test compartments were allowed to equilibrate for approximately 48 hours. First-instar, 1 to 4-day old midge larvae were impartially assigned to exposure chambers at test initiation. Observations of abnormal behavior were made daily throughout the test. During the period of emergence, the sex and number of fully emerged midges was recorded each day, and the total number of adults emerged by the end of the test period (Day 28) was determined. Percent emergence/mortality observed in the treatment groups was used to determine the 28-day EC50 value. Observations of the effects of the test substance on emergence and development data were used to determine the no-observed-effect concentration (NOEC) and the lowest-observed-effect concentration (LOEC).

Test Water
The water used for holding and testing was freshwater obtained from a well approximately 40 meters deep located on the Wildlife International site. The well water was passed through a sand filter to remove particles greater than approximately 25 μm, and pumped into a 37,800-L storage tank where the water was aerated with spray nozzles. Prior to use in the test system, the water was filtered to 0.45 μm to remove fine particles and was passed through an ultraviolet (UV) sterilizer.

Preparation of Test Concentrations
The test substance was administered to the test organism in sediment. This route of administration was selected because it represents the most likely route of exposure to sediment dwelling organisms.
Individual stock solutions were prepared for use in spiking the sediment to prepare each of the five concentrations tested. Test concentrations were not adjusted for the active ingredient of the test substance during preparation, and are based on the test substance as received. A 50-mL primary stock solution was prepared by mixing a calculated amount of test substance into HPLC-grade acetone at a nominal concentration of 100 mg/mL. The primary stock solution was mixed by inversion, and appeared clear and colorless with no visible precipitates. Four secondary stock solutions (50 mL each) were prepared in acetone at nominal concentrations of 6.30, 12.5, 25.0 and 50.0 mg/mL by serial dilution of the primary or previous stock. The secondary stock solutions were mixed by inversion, and were all clear and colorless, with no evidence of precipitates.
To prepare a batch of sediment for each treatment level, a 14-mL volume of the appropriate stock solution was added to 70 grams of sand in a labeled glass beaker and was stirred with a glass stir rod until homogenous. This dosed “sand premix” was placed under a fume hood and the acetone was allowed to evaporate for approximately 1.5 hours. The 70-gram sand premix was added to 630 grams of untreated formulated sediment in a 2000-mL plastic Nalgene® bottle and mixed on a rotary mixer for approximately 1.5 hours. Additional formulated sediment (700 g) was added to the premix to achieve a final weight of 1400 grams. This 1400-g batch sediment was mixed on a rotary mixer for approximately 66 hours prior to transfer of the dry sediment to the test compartments. Since a solvent (acetone) was used in the preparation of the test sediments, a solvent control was included in the test design. The solvent control sediment was prepared using 14 mL of acetone, with the same mixing procedures as the treated sediments, but with no test substance added. The negative control sediment was prepared without the addition of test substance or solvent.

Preparation of Test Chambers
Eight replicate test chambers were prepared for each treatment and control group. Four replicates per group were used for biological observations. An additional four replicates per group were prepared for use, as needed, in analytical confirmation of concentrations on Days 0, 7 and 28. After mixing the batch sediments, approximately 2 cm of the appropriate dosed sediment was placed in the bottom of each test chamber (glass jar) on a top-loading balance, and the weight of the sediment was recorded. Approximately 600 mL of overlying water was slowly added to each test chamber, while avoiding disturbance of the sediment, and each test chamber was loosely covered. After preparation, the test chambers were transferred to a temperature-controlled environmental chamber, gentle aeration was applied to each test chamber, and the sediment/water mixtures were allowed to acclimate under static conditions for approximately 48 hours.

Test Apparatus
The test chambers were glass jars containing approximately 2 cm of sediment and approximately 600 mL of overlying water. The depth of the sediment measured in a representative chamber was 1.8 cm, and the depth of the overlying water in a representative compartment was 8.4 cm. The approximate 1:4 ratio of sediment depth to overlying water depth was maintained throughout the test by periodically replacing water lost due to evaporation with reverse-osmosis water. Loose plastic covers were placed over each test chamber during the test, and aeration was applied to each test chamber through a glass pipette that extended to a depth not closer than 2 cm to the surface of the sediment. Air was bubbled into the test chambers at a rate that did not disturb the sediment. The test chambers were randomly arranged in a temperature-controlled environmental chamber during the test, and were labeled with the project number, test concentration and replicate designation. The general operation of the test apparatus was checked visually at least once each day during the test.

Environmental Conditions
The test systems were illuminated using fluorescent tubes that emit wavelengths similar to natural sunlight. The lights were controlled by an automatic timer to provide a photoperiod of 16 hours of light and 8 hours of darkness. A 30-minute transition period of low light intensity was provided when lights went on and off to avoid sudden changes in light intensity. Light intensity was measured at the water surface of one representative test chamber at the beginning of the test using a SPER Scientific Model 840006 light meter.
The test was conducted at a target water temperature of 20 ± 2°C. Temperature was measured in the overlying water of one replicate test chamber of each treatment and control group daily during the test using a liquid-in-glass thermometer. Measurements typically rotated among replicate test chambers in each group at each measurement interval. Water temperature also was monitored continuously in a container of water placed adjacent to the test chambers using a validated environmental monitoring system (AmegaView Central Monitoring System). The system measurements were verified prior to exposure initiation with a digital thermometer.
Dissolved oxygen concentrations were measured in samples of overlying water from one replicate test chamber of each treatment and control group daily during the test using a Thermo Orion Model 850Aplus dissolved oxygen meter. Measurements of pH were made in samples of overlying water from one replicate test chamber of each treatment and control group at test initiation, once each week during the test, and at test termination using a Thermo Orion Model 525Aplus pH meter. Measurements of dissolved oxygen and pH typically rotated among replicate test chambers in each group at each measurement interval.
Hardness, alkalinity, specific conductance and ammonia were measured in composite samples of overlying water from the negative and solvent control group replicates and the highest concentration treatment group replicates at the beginning and end of the test. Hardness and alkalinity measurements were made by titration based on methods in Standard Methods for the Examination of Water and Wastewater. Specific conductance was measured using an Acorn Series Model CON6 conductivity-temperature meter. Ammonia was measured using a Thermo Scientific Orion Dual Star pH/ISE meter.

Procedures and Biological Observations and Measurements
The test was initiated following the equilibration period for the water/sediment systems. Aeration to the test chambers was stopped prior to adding the organisms and was restarted approximately 24 hours later. To initiate the test, one to two first-instar larvae (1-4 days old) were added to a test chamber until it contained 20 individuals; this was repeated until all chambers contained 20 larvae. Test chambers were initiated one at a time and in sequential order starting with the controls then proceeding from the lowest treatment group to the highest treatment group in order to prevent contamination of test material into lower treatment groups or the controls. Bias was eliminated by making sure there was a large pool of healthy, appropriately aged organisms to choose from. The test chambers prepared for analytical sampling on Day 0 did not contain midges. All transfers were made below the water surface using wide-bore pipettes.
The test chambers were observed daily during the test to make visual assessments of any abnormal behavior (e.g., leaving the sediment, unusual swimming). During the period of expected emergence, the sex and number of fully emerged midges were recorded on a daily basis. After identification, the midges were removed from the test chambers. When the total number of adults emerged in each replicate at the end of the test (Day 28) was less than the number initially placed in each replicate, then those individuals not accounted for were considered dead.
Reference substance (positive control):
no
Duration:
28 d
Dose descriptor:
EC50
Effect conc.:
87 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
emergence rate
Remarks on result:
other: 95% confidence interval of 73.6 to 102 mg/kg.
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
< 37 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
development rate
Duration:
28 d
Dose descriptor:
LOEC
Effect conc.:
37 mg/kg sediment dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
development rate
Details on results:
Measurement of Test Concentrations
Nominal concentrations selected for use in this study were 63, 125, 250, 500 and 1000 mg/kg dry weight of sediment. During the course of the test, the appearance of the overlying water was observed in the test chambers. At test initiation, the overlying water appeared clear and very light tan. At test termination, the overlying water appeared clear and light yellow.
Measured concentrations ranged from approximately 90.4 to 98.0% of nominal.
All reported sediment concentrations are expressed on a dry weight basis with an LOQ of 12.5 mg/kg sediment. The LOQ for overlying and pore water analyses was 0.100 mg/L. Measured concentrations of the test substance in negative and solvent control sediment and water samples on Days 0, 7 and 28 were below the LOQ. Measured concentrations in the sediment samples collected from the treatment groups ranged from approximately 49.7 to 92.6% of nominal. Measured concentrations in the overlying water and pore water samples ranged from approximately 0.471 to 2.28 mg/L and 2.70 to 7.42 mg/L, respectively. When measured concentrations of the sediment samples collected during the test were averaged, the mean measured test concentrations for this study were 37, 79, 187, 366 and 688 mg/kg, representing 59, 63, 75, 73 and 69% of nominal concentrations, respectively. The results of the study were based on the mean measured concentrations in sediment.
Mass balance of the test substance indicates the concentration in the total test system remained fairly consistent. Reofos 35 remained at fairly consistent concentrations in the sediment during the study and had a slight increase in concentration in the overlying water and pore water over the course of the study. At study start approximately 72% of the total amount of Reofos 35 was in the sediment, and by the end of the study the mean amount in the sediment decreased to 63%. Approximately 2.2% of the total amount was found in the overlying water at study start, increasing to approximately 3.2% at the end of the study. Only a small percentage of material was measured in the pore water during the study.

Observations and Measurements
All water quality measurements were within the desired ranges. Water temperatures were within the 20 ± 2°C range established for the test. Measurements of pH ranged from 7.8 to 9.0 during the test. Dissolved oxygen concentrations remained ≥7.0 mg/L (≥78% of saturation) throughout the test.
Measurements of specific conductance, hardness and alkalinity were comparable between the control and treatment groups. The Day 0 measurements were consistent with typical water quality of Wildlife International well water, while the Day 28 measurements were higher than typical water quality of Wildlife International well water. The higher measurements at the end of the test may be a result of the static nature of the test. The departure from typical measurements did not have a negative impact on the results of the test as is demonstrated by the control groups meeting validity criteria established for the test by the OECD 218 guideline. Measurements of ammonia in the overlying water at the beginning and end of the test were below the limit of quantitation (LOQ of <0.17 mg/L, the lowest calibration standard) except in the 688 mg/kg treatment group on Day 28 which was 1.7 mg/L. Light intensity at test initiation was 533 lux at the surface of the water of one representative test chamber.
No unusual observations of organisms avoiding the sediment occurred during the test. There were a few observations of larvae on the surface of the sediment or swimming in the water column during the study, but these did not appear to be dose-responsive, were comparable between the control and treatment groups, and were not considered to be treatment-related. There also were a few observations of dead pupae or larvae and adults that emerged and died. These numbers were concentration-responsive, and were considered to be treatment-related.
There was an apparent treatment-related effect on emergence in the 79, 187, 366 and 688 mg/kg treatment groups. Emergence was first noted on Day 12 of the test and continued through Day 27. The mean emergence ratios in the negative control, solvent control and the 37, 79, 187, 366 and 688 mg/kg treatment groups were 0.99, 0.96, 0.94, 0.89, 0.26, 0.08 and 0.03, respectively. The 28-day EC50 for emergence was determined to be 87 mg/kg, with a 95% confidence interval of 73.6 to 102 mg/kg. Dunnett’s test indicated that there was a statistically significant difference in emergence in the 79, 187, 366 and 688 mg/kg treatment groups in comparison to the pooled control (p ≤ 0.05). Therefore, the LOEC for emergence ratio was determined to be 79 mg/kg and the NOEC was 37 mg/kg.
There were apparent treatment-related effects on development times and development rates (defined as the portion of larval development which takes place per day) in the 187, 366 and 688 mg/kg treatment groups. The mean development times in the negative control, solvent control and the 37, 79, 187, 366 and 688 mg/kg treatment groups were 14.1, 14.1, 14.8, 15.3, 17.3, 23.2 and 22.0, respectively. The mean male development rates were 0.0771, 0.0771, 0.0721, 0.0710, 0.0661, 0.0377 and 0.0465, respectively. The mean female development rates were 0.0713, 0.0717, 0.0689, 0.0646, 0.0584, 0.0519 and 0.0465, respectively. Dunnett’s test indicated that there were statistically significant differences in development times in the 187, 366 and 688 mg/kg treatment groups. Therefore the NOEC for development time is 79 mg/kg and the LOEC is 187 mg/kg. Dunnett’s test also indicated that there were statistically significant differences in development rates in the 79, 187, 366 and 688 mg/kg treatment groups in comparison to the pooled control (p ≤ 0.05) for the females and in all treatment groups for the males. Therefore, the LOEC for development rates was determined to be 37 mg/kg and the NOEC was <37 mg/kg.
Results with reference substance (positive control):
Postive control not used in this study.
Reported statistics and error estimates:
Dunnett’s test indicated that there was a statistically significant difference in emergence in the 79, 187, 366 and 688 mg/kg treatment groups in comparison to the pooled control (p ≤ 0.05).
Dunnett’s test indicated that there were statistically significant differences in development times in the 187, 366 and 688 mg/kg treatment groups.
Dunnett’s test also indicated that there were statistically significant differences in development rates in the 79, 187, 366 and 688 mg/kg treatment groups in comparison to the pooled control (p ≤ 0.05) for the females and in all treatment groups for the males.

Measured Concentrations of Reofos 35 in Sediment Samples

Nominal Test Concentration (mg/kg)

Rep.

Sample Number1(616A-119-)

Sampling Time (Days)

Measured Concentration (mg/kg)2,3

Percent of Nominal4

Mean Measured Concentration (mg/kg)

Mean Measured Percent of Nominal

Negative Control

E

S-1A

0

<LOQ

--

--

--

F

S-8

7

<LOQ

--

G

S-15

28

<LOQ

--

Solvent Control

E

S-2A

0

<LOQ

--

--

--

F

S-9

7

<LOQ

--

G

S-16

28

<LOQ

--

63

E

S-3A

0

41.4

65.7

37 ± 5.23

CV = 14.1%

59

F

S-10

7

38.7

61.5

G

S-17

28

31.3

49.7

125

E

S-4A

0

97.5

78.0

79 ± 17.2

CV = 21.9%

63

F

S-11

7

74.5

59.6

G

S-18

28

63.8

51.1

250

E

S-5A

0

163

65.2

187 ± 21.0

 CV = 11.2%

75

F

S-12

7

203

81.3

G

S-19

28

194

77.5

500

E

S-6A

0

463

92.6

366 ± 88.5

CV = 24.2%

73

F

S-13

7

290

58.1

G

S-20

28

344

68.8

1000

E

S-7Re5

0

597

59.7

688 ± 95.3

 CV = 13.9%

69

F

S-14

7

787

78.7

G

S-21

28

679

67.9

1Due to unacceptable R2values, day 0 sediment samples were reanalysed.

2The limit of quantification (LOQ) was 12.5 mg/kg, calculated as the product of the concentration of the lowest calibration standard (0.100 mg/L) and the dilution factor of the matrix blank samples (125).

3Analytical results were generated using wet weights. The tabulated values are reported on a dry weight basis.

4Results were generated using Excel 201 in the full precision mode. Manual calculations may differ slightly.

5Sample 616A-119-S-7A was reanalysed.

 

Measured Concentrations of Reofos 35 in Overlying Water Samples

Nominal Test Concentration (mg/kg)

Rep.

Sample Number (616A-119-)

Sampling Time (Days)

Measured Concentration (mg/L)1,2

Negative Control

E

OW-1

0

<LOQ

F

OW-8

7

<LOQ

G

OW-15

28

<LOQ

Solvent Control

E

OW-2

0

<LOQ

F

OW-9

7

<LOQ

G

OW-16

28

<LOQ

63

E

OW-3

0

0.471

F

OW-10

7

0.734

G

OW-17

28

0.759

125

E

OW-4

0

0.848

F

OW-11*

7

0.979

G

OW-18

28

1.06

250

E

OW-5*

0

1.16

F

OW-12*

7

1.44

G

OW-19

28

1.83

500

E

OW-6*

0

1.64

F

OW-13*

7

1.71

G

OW-20

28

1.71

1000

E

OW-7*

0

1.68

F

OW-14*

7

2.17

G

OW-21

28

2.28

1The limit of quantification (LOQ) was 0.100 mg/L, calculated as the product of the concentration of the lowest calibration standard (0.100 mg/L) and the dilution factor of the matrix blank samples (1.00).

2Results were generated using Excel 2010 in the full precision mode.

*Samples reanalysed.

 

Measured Concentrations of Reofos 35 in Pore Water Samples

Nominal Test Concentration (mg/kg)

Rep.

Sample Number (616A-119-)

Sampling Time (Days)

Measured Concentration (mg/L)1,2

Negative Control

E

PW-1

0

<LOQ

F

PW-8

7

<LOQ

G

PW-15

28

<LOQ

Solvent Control

E

PW-2

0

<LOQ

F

PW-9

7

<LOQ

G

PW-16

28

<LOQ

63

E

PW-3*

0

3.10

F

PW-10*

7

2.70

G

PW-17

28

3.06

125

E

PW-4*

0

3.76

F

PW-11*

7

3.46

G

PW-18

28

4.13

250

E

PW-5*

0

4.17

F

PW-12*

7

5.14

G

PW-19

28

6.45

500

E

PW-6*

0

3.97

F

PW-13*

7

6.57

G

PW-20

28

7.07

1000

E

PW-7*

0

4.77

F

PW-14*

7

7.42

G

PW-21

28

6.50

1The limit of quantification (LOQ) was 0.100 mg/L, calculated as the product of the concentration of the lowest calibration standard (0.100 mg/L) and the dilution factor of the matrix blank samples (1.00).

2Results were generated using Excel 2010 in the full precision mode.

*Samples reanalysed.

 

Percentage of Reofos 35 in Sediment, Overlying Water and Pore Water Based on Measured Concentration Mass Balance Calculations

Nominal Test Concentration (mg/kg)

Study Day

% in Sediment

% in Overlying Water

% in Pore Water

Total %

63

0

65.7

3.6

0.5

69.8

7

61.4

5.5

0.5

67.5

28

49.7

5.7

0.5

55.9

125

0

78.0

2.6

0.3

80.9

7

59.6

3.8

0.4

63.9

28

51.0

4.0

0.4

55.5

250

0

65.2

2.2

0.2

67.6

7

81.2

2.8

0.3

84.3

28

77.6

3.5

0.4

81.5

500

0

92.6

1.6

0.1

94.3

7

58.0

1.6

0.1

59.7

28

68.8

1.6

0.1

70.6

1000

0

59.7

0.8

0.1

60.5

7

78.7

1.0

0.1

79.8

28

67.9

1.1

0.1

69.1

Calculated using Excel 2010 in full precision mode. Manual calculations may differ slightly.

 

Means and Ranges of Water Quality Measurements Taken in Overlying Water During the 28-Day Exposure to Sediment-Incorporated Reofos 35

Mean Measured Sediment Concentration (mg/kg)

Mean ± SD and Range of Measured Parameters

Temperature1

DO2(mg/L)

pH

Hardness (mg/L as CaCO3)3

Alkalinity (mg/L as CaCO3)3

Conductivity3(μS/cm)

Ammonia (mg/L as NH3)4

Negative Control

20.7 ± 0.18 (20.4 – 21.1)

8.6 ± 0.23 (8.2 – 9.1)

8.3 ± 0.17 (8.2 – 8.6)

1845

(148 – 220)

1855

(176 – 194)

3775

(326 – 428)

<LOQ5

(<0.17 - <0.17)

Solvent Control

20.5 ± 0.20 (20.0 – 20.9)

8.5 ± 0.26 (8.1 – 8.9)

8.5 ± 0.25 (8.3 – 8.9)

1725

(144 – 200)

1755

(174 – 176)

3825

(333 – 430)

<LOQ

(<0.17 - <0.17)

37

20.6 ± 0.13 (20.4 – 20.9)

8.4 ± 0.27 (7.8 – 8.8)

8.5 ± 0.19 (8.2 – 8.7)

--

--

--

--

--

--

--

--

79

20.6 ± 0.15 (20.3 – 20.9)

8.4 ± 0.23 (8.0 – 8.9)

8.5 ± 0.19 (8.3 – 8.8)

--

--

--

--

--

--

--

--

187

20.6 ± 0.16 (20.1 – 20.9)

8.4 ± 0.25 (8.0 – 8.9)

8.4 ± 0.17 (8.3 – 8.7)

--

--

--

--

--

--

--

--

366

20.5 ± 0.21 (20.1 – 20.9)

8.5 ± 0.24 (7.7 – 8.8)

8.5 ± 0.27 (8.3 – 9.0)

--

--

--

--

--

--

--

--

688

20.6 ± 0.19 (20.1 – 20.9)

8.4 ± 0.36 (7.0 – 8.8)

8.4 ± 0.35 (7.8 – 8.7)

1785

(148 – 208)

1805

(176 – 184)

3985

(341 – 455)

0.9

(<0.17 – 1.7)

1Manual temperature measurements. Temperature monitored continuously during the test ranged from approximately 19.57 to 20.32°C, measured to the nearest 0.01°C.

2A dissolved oxygen concentration of 5.4 mg/L represents 60% saturation at 20°C in freshwater.

3-- = no measured scheduled.

4The LOQ for ammonia analyses was set at 0.17 mg/L, the concentration of the lowest calibration standard.

5No standard deviation is reported when only two measurements are recorded Day 0 and Day 28).

 

Summary of Emergence and Development of Midges (Chironomus riparius) During the 28-Day Exposure to Sediment-Incorporated Reofos 35

Mean Measured Sediment Concentration (mg/kg)

Number Emerged1

Mean Emergence Ration2,4

Mean Development Time (Days)4

Mean Development Rate3,4

Males

Females

Total

(Males)

(Females)

Negative Control

34

45

79

0.99

14.1

0.0771

0.0713

Solvent Control

35

42

77

0.96

14.1

0.0771

0.0717

37

38

37

75

0.94

14.8

0.0721*

0.0689

79

39

32

71

0.89*

15.3

0.0710*

0.0646*

187

7

14

21

0.26*

17.3*

0.0661*

0.0584*

366

2

4

6

0.08*

23.2*

0.0377*

0.0519*

688

1

1

2

0.03*

22.0*

0.0465*

0.0465*

1Each replicate contained 20 midge larvae at test initiation, for a total of 80 larvae per control and treatment group.

2Emergence ratio is calculated as the number of emerged midges divided by the initial number exposed, and corresponds to percent emergence.

3The development rate represents that portion of larval development which takes place per day.

4Calculated using SAS. Manual calculations may differ slightly.

*Indicates a statistically significant difference in comparison to the pooled control (p≤0.05) using Dunnett’s test.

28-Day EC50 for emergence: 87 mg/kg, with a 95% confidence interval of 73.6 to 102 mg/kg.

Validity criteria fulfilled:
yes
Conclusions:
Midges (Chironomus riparius) were exposed for 28 days to five mean measured concentrations of sediment-incorporated Reofos 35 ranging from 37 to 688 mg/kg. There were treatment-related effects observed on emergence in the 79, 187, 366 and 688 mg/kg treatment groups and development at all tested concentrations. Based on the mean measured concentrations in sediment, the 28-day EC50 value for emergence was 87 mg/kg, with a 95% confidence interval of 73.6 to 102 mg/kg. The slope of the concentration-response curve was 2.336. Based on the effects observed on development rate, the LOEC for the study was 37 mg/kg and the NOEC was <37 mg/kg.
Executive summary:

The objective of this study was to determine the effects of sediment-incorporated Reofos 35 on the midge, Chironomus riparius, during a 28-day exposure period under static test conditions. The endpoints evaluated for the test were total number of adults emerged and the development time. Emergence ratios and development rates also were determined.

The study was conducted according to the procedures outlined in the protocol, “Reofos 35: A Prolonged Sediment Toxicity Test with the Midge (Chironomus riparius) Using Spiked Sediment”. The protocol was based on procedures in the OECD Guidelines for the Testing of Chemicals, Guideline 218: Sediment-Water Chironomid Toxicity Test Using Spiked Sediment and ASTM Standard E 1706-05: Standard Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater Invertebrates.

 

Midges (Chironomus riparius) were exposed for 28 days to five mean measured concentrations of sediment-incorporated Reofos 35 ranging from 37 to 688 mg/kg. There were treatment-related effects observed on emergence in the 79, 187, 366 and 688 mg/kg treatment groups and development at all tested concentrations. Based on the mean measured concentrations in sediment, the 28-day EC50 value for emergence was 87 mg/kg, with a 95% confidence interval of 73.6 to 102 mg/kg. The slope of the concentration-response curve was 2.336. Based on the effects observed on development rate, the LOEC for the study was 37 mg/kg and the NOEC was <37 mg/kg.

Description of key information

Sediment Toxicity

Key value for chemical safety assessment

EC50 or LC50 for freshwater sediment:
87 mg/kg sediment dw
EC10, LC10 or NOEC for freshwater sediment:
37 mg/kg sediment dw

Additional information

Midges (Chironomus riparius) were exposed for 28 days to five mean measured concentrations of sediment-incorporated Reofos 35 ranging from 37 to 688 mg/kg. There were treatment-related effects observed on emergence in the 79, 187, 366 and 688 mg/kg treatment groups and development at all tested concentrations. Based on the mean measured concentrations in sediment, the 28-day EC50 value for emergence was 87 mg/kg, with a 95% confidence interval of 73.6 to 102 mg/kg. The slope of the concentration-response curve was 2.336. Based on the effects observed on development rate, the LOEC for the study was 37 mg/kg and the NOEC was <37 mg/kg.