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Ecotoxicological information

Sediment toxicity

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Reference
Endpoint:
sediment toxicity: long-term
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 218 (Sediment-Water Chironomid Toxicity Test Using Spiked Sediment)
Version / remarks:
adopted April 2004
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
The concentrations of the test item were determined in the overlying water, the pore water and the sediment of all test concentration of nominal: 62.5, 125, 250, 500 and 1000 mg test item/kg dry sediment and of the controls (solvent control and control) from test start and test end.
Vehicle:
yes
Remarks:
acetone
Details on sediment and application:
Formulated Sediment:
The dry sediment was prepared according to OECD test guideline 218:
- 4.5 % sphagnum moss peat, finely ground (particle size <= 1 mm) and air dried
- 74.75 % quartz sand (grain size: > 50 % of the particles in the range of 50 - 200 µm)
- 20 % kaolinite clay (kaolinite content  30 %)
- 0.75 % CaCO3, of chemically pure quality was added to adjust the pH of the final mixture of the sediment to 6.7.
- The organic carbon content (without the food source) of the final mixture was 1.8 %.
- 1 % Urtica powder was added as food source in addition to the dry sediment components.
- The dry constituents (peat, sand and kaolin clay) were mixed together and stored dry until use.
- Four days before the application deionised water was added to obtain homogeneous sediment with a moisture content of 44 % of dry weight of the sediment.
- Small samples of the sediment were taken to determine the moisture content of the final mixture (after addition of the quartz sand). The mean moisture content was 38.7%.


Application of the Test Item and the Controls
Pre-Experiments:
- Pre-experiments were performed to determine a suitable concentration range and to establish suitable methods to prepare the test solutions.
- The pre-experiments were not performed in compliance with the GLP-Regulations and will be excluded from the Statement of Compliance in the final report, but the raw data of these tests will be archived under the study number of the present study.
Test Concentrations:
- 1000, 500, 250, 125 and 62.5 mg test item/kg dry sediment (spacing factor 2), solvent a control and a control. The test concentrations are equivalent to arithmetic mean measured concentrations of 43.8, 83.1, 162, 374 and 761 mg test item/kg dry sediment.
Dosage of Test Item:
- Before test start, a concentrated stock solution was prepared by dissolving 1.41 mL (equivalent to 1395.9 mg) in 50 mL acetone. Then appropriate volumes of this stock solution were serially diluted with acetone. A volume of 20 mL of the stock solution or the corresponding dilution was applied to a fraction of 70 g quartz sand for each treatment. Then, the solvent was allowed to evaporate for 130 minutes. Afterwards, the spiked quartz sand was added to the pre-moistened sediment equivalent to 490 g dry weight to result in a total sediment amount of 560 g dry weight for each test item group to achieve the desired test concentrations and mixed as homogeneously as possible. Portions of these sediment fractions were used to set up the test units.
- After an acclimatisation period of 2 days after set up of the test units the larvae were introduced. The day of introduction of the larvae was designated as Day 0 of the exposure (= start of the test).
Control:
- In the control, a sediment-water system was used without addition of the solvent or the test item.
Solvent Control:
- In the solvent control, a sediment-water system was used without addition of the test item but with the same amount of quartz sand and solvent per g dry sediment as used for the test concentrations.
Test organisms (species):
Chironomus riparius
Details on test organisms:
- Age at Test Start: Only fresh egg masses were used as source for the test animals, which were taken from the culture 4 and 5 days before adding the test organisms to the test vessels. The larvae began to hatch 2 days after the eggs were laid and were 2 to 3 days old (first instar) at test start (Day 0).
- Sex: Male and female
- Origin: Chironomus riparius in-house laboratory culture
- Holding Conditions: In accordance with the test guidelines, Chironomus riparius is bred in the laboratories of ibacon under standardised conditions. The larvae are held at 20 ± 2 °C in the same kind of water that is used for the test (see 6.5). They are fed ad libitum with commercial fish food (TETRA MIN Hauptfutter, TETRA-Werke, 49324 Melle, Germany).
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
Hardness:
258.1 to 267.0 mg/L as CaCO3 at test start, 258.1 to 382.7 mg/L as CaCO3 at test end.
Test temperature:
18.3 to 20.3 °C
pH:
7.4 to 8.8
Dissolved oxygen:
17 to 99 %
The O2 content at test start was measured to be very low at two replicates at the concentration of 43.8 test item/kg d.s. and one replicate at the concentration of 162 test item/kg d.s.. It cannot be excluded that the test animals were affected by this lack of oxygen. Therefore, these replicates were excluded from the evaluation.
Ammonia:
0.1 mg NH4+ /L at test start, 0.0 to 0.2 mg NH4+ /L at test end
Nominal and measured concentrations:
nominal: 62.5, 125, 250, 500 and 1000 mg test item/kg dry sediment
Arithmetic mean measured concentrations: 43.8, 83.1, 162, 374 and 761 mg test item/kg dry sediment.
Details on test conditions:
- Test Units: Glass beakers of 600 mL volume. The glass beakers have a diameter of 7.5 cm and a height of 17.5 cm. Each glass beaker was covered with a glass plate to prevent the escape of emerged test animals and evaporation of the test water.
- Medium: Reconstituted Water (Elendt "M4"), formulated sediment
- Introduction of Sediment: 100 +- 1 g moist artificial sediment was added to each test vessel to a depth of 1.5 cm.
- Introduction of Test Water: 280 mL reconstituted water was added to each test vessel to a depth of approximately 6 cm. Hence, the ratio of the depth of the sediment layer to the depth of the overlying water was approximately 1:4.
The level of test water was marked on the outside of the test vessel. The water levels were maintained with deionised water during the study to the original starting volume to prevent the concentration of the test item changing. Water levels did not change by more than 10 % during the whole test.
- Sediment-Water Systems: The test vessels were prepared 2 days before the start of the test and were allowed to acclimate under test conditions. During this period, the sediment-water system was left under gentle aeration. Aeration of the water was stopped for approximately 24 hours after addition of the larvae to the test vessel. The test vessels were covered with glass plates.
- Sampling of Egg Masses: Five and four days before adding the test organisms to the test vessels, egg masses were taken from the cultures and placed in small vessels filled with culture medium.
- Introduction of Larvae: Twenty first instar larvae (2 to 3 days post hatching) were allocated randomly to each test vessel with a blunt pipette and a watch glass. The larvae were added 2 days after the application of the test item. The day of insertion of the larvae was designated as day 0 of exposure (= start of the test).
- Replicates: 4 replicates per treatment, 4 replicates for the control and 6 replicates for the solvent control. For analytical sampling on test start (Day 0), 2 additional vessels were prepared for each test concentration, for the control and for the solvent control.
- Test Procedure: A static exposure system was chosen. Two days after setup of the sediment-water system with spiked sediment, the larvae were added. Subsequently, aeration of the sediment-water system was stopped for approximately 24 hours. During the exposure period, the water levels were topped up to the original volume in order to compensate water evaporation.
- Feeding: The sediment contained a food source i.e. Urtica powder. Therefore, it was not necessary to feed the larvae during the test.
- Light Regime: 16 h light : 8 h dark
- Light Intensity: 541 to 785 lux. The light intensity was measured once during the test.
- Exposure Duration: 28 days
- Endpoints: Total number of adults emerged (emergence rate) and the time to emergence (development rate).
Reference substance (positive control):
yes
Remarks:
For the evaluation of the quality of the chironomids and the consistency of the experimental conditions, the reference item potassium dichromate is tested periodically to assure that the test protocol and test conditions are reliable.
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
83.1 mg/kg sediment dw
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
emergence rate
Duration:
28 d
Dose descriptor:
EC50
Effect conc.:
183 mg/kg sediment dw
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
emergence rate
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
83.1 mg/kg sediment dw
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
development rate
Duration:
28 d
Dose descriptor:
EC50
Effect conc.:
860 mg/kg sediment dw
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
development rate
Details on results:
The NOEC for emergence rate was calculated using the Step-down Rao-Scott-Cochran-Armitage Test Procedure. For the evaluation of the NOEC for development rate the Williams t-test was used.
The EC values for emergence rate were determined by Weibull analysis.
The EC values for development rate were calculated by the probit analysis using linear maximum likelihood regression.
The software used to perform the statistical analysis was ToxRat Professional, Version 3.3.0, ToxRat Solutions GmbH.
Validity criteria fulfilled:
yes
Remarks:
emergence in control >= 70 %, main emergence to adults in control between 12 and 23 days after insertion, oxygen concentration >= 60 %, pH of overlying water 6 - 9, water temperature should not differ by more than +- 1.0 °C between the test vessels
Conclusions:
The influence of Tris(2-ethylhexyl) phosphate on the survival and development of Chironomus riparius was determined during an exposure period of 28 days. The 28 day NOEC for the emergence rate was determined to be 83.1 mg test item/kg dry sediment. The NOEC for the development rate was determined to be 83.1 mg test item/kg dry sediment. The EC50 for the emergence rate was calculated to be 183 mg test item/kg dry sediment and 860 mg test item/kg dry sediment for development rate.
All reported results refer to arithmetic mean values.
Executive summary:

A study was conducted according to OECD 218 in a static sediment-water system to assess the toxicity of Tris(2 -ethylhexyl) phosphate towards Chironomus riparius.

This study encompassed 5 test concentrations, a solvent control and a control, with 4 replicates for each test concentration and the control and with 6 replicates for the solvent control, each containing 20 larvae. The 5 test concentrations were: 1000, 500, 250, 125 and 62.5 mg test item/kg dry sediment. The test concentrations are equivalent to arithmetic mean measured concentrations of 43.8, 83.1, 162, 374 and 761 mg test item/kg dry sediment. Two days after setup of the sediment-water system with spiked sediment, the larvae were added. The day of insertion of the larvae was designated as day 0 of exposure. The following test conditions were measured: Water temperature: 18.3 to 20.3 °C; pH value:7.4 to 8.8; dissolved oxygen concentration:17 to 99 %; photoperiod: 16 h light - 8 h dark; light intensity: 541 to 785 lux.

The larvae were exposed for 28 days.

The influence of Tris(2-ethylhexyl) phosphate on the survival and development of Chironomus riparius was determined during an exposure period of 28 days. The 28 day NOEC for the emergence rate was determined to be 83.1 mg test item/kg dry sediment. The NOEC for the development rate was determined to be 83.1 mg test item/kg dry sediment. The EC50 for the emergence rate was calculated to be 183 mg test item/kg dry sediment and 860 mg test item/kg dry sediment for development rate.

All reported results refer to arithmetic mean values.

Description of key information

A study was conducted according to OECD 218 in a static sediment-water system to assess the toxicity of Tris(2 -2thylhexal) phosphate towards Chironomus riparius.

This study encompassed 5 test concentrations, a solvent control and a control, with 4 replicates for each test concentration and the control and with 6 replicates for the solvent control, each containing 20 larvae. The 5 test concentrations were:1000, 500, 250, 125 and 62.5 mg test item/kg dry sediment. The test concentrations are equivalent to arithmetic mean measured concentrations of 43.8, 83.1, 162, 374 and 761 mg test item/kg dry sediment. Two days after setup of the sediment-water system with spiked sediment, the larvae were added.The day of insertion of the larvae was designated as day 0 of exposure.The following test conditions were measured:Water temperature:18.3 to 20.3 °C; pH value:7.4 to 8.8; dissolved oxygen concentration:17 to 99 %; photoperiod: 16 h light - 8 h dark; light intensity: 541 to 785 lux.

The larvae were exposed for 28 days.

The influence of Tris(2-ethylhexyl) phosphate on the survival and development of Chironomus riparius was determined during an exposure period of 28 days. The 28 day NOEC for the emergence rate was determined to be 83.1 mg test item/kg dry sediment. The NOEC for the development rate was determined to be 83.1 mg test item/kg dry sediment. The EC50 for the emergence rate was calculated to be 183 mg test item/kg dry sediment and 860 mg test item/kg dry sediment for development rate.

All reported results refer to arithmetic mean values.

Key value for chemical safety assessment

EC50 or LC50 for freshwater sediment:
183 mg/kg sediment dw
EC10, LC10 or NOEC for freshwater sediment:
83.1 mg/kg sediment dw

Additional information