1,1,5,5,5-hexamethyl-3,3-bis[(trimethylsilyl)oxy]trisiloxane

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1st October to 30th November 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 204 (Fish, Prolonged Toxicity Test: 14-day Study)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Samples were collected from one test chamber of each treatment and control group two days prior to the start of the test after conditioning the diluter for approximately two days. Water samples also were collected from alternating replicate test chambers in each treatment and control group at the beginning of the test and on days 7 and 14 to measure concentrations of the test substance. The samples were collected from mid-depth into a 100-ml glass volumetric flask, and processed immediately for analysis.

Vehicle:

yes

Remarks:

DMF

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Individual stock solutions were prepared for each of the five concentrations tested. A primary stock solution was prepared by mixing a calculated amount of test substance into HPLC-grade dimethylformamide (DMF) at a nominal concentration of 10 µg/ml. An aliquot of the primary stock solution was diluted with DMF to create a secondary stock solution at a nominal concentration of 1510 ng/ml. Four additional stock solutions were prepared in DMF at nominal concentrations of 94.4, 189, 378 and 755 ng/ml by proportional dilution of the secondary stock. The stock solutions were mixed by inversion, and all appeared clear and colourless. Stock solutions were stored refrigerated in glass amber bottles or a 1000 mL graduated cylinder, and aliquots of each stock were placed in the syringe every two days during the study.
The five test substance stock solutions were injected into the diluter mixing chambers at a rate of 20 µl/minute where they were mixed with dilution water delivered at a rate of 200 ml/minute to achieve the desired test concentrations.
- Controls: The negative control received dilution water only.
The solvent control was prepared by delivering HPLC-grade DMF to the mixing chamber for the solvent control.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): DMF
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): The concentration of DMF in the solvent control and all M4Q treatment groups was 0.1 ml/l.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: Rainbow trout
- Strain: Not reported
- Source: The rainbow trout were received as juveniles from Thomas Fish Company of Anderson, California.
- Age at study initiation (mean and range, SD): Juveniles from the same source and year class
- Length at study initiation (length definition, mean, range and SD): Average of 10 fish: 4.9 cm, range of 4.5 - 5.3 cm.
- Weight at study initiation (mean and range, SD): Average wet weight (blotted dry) of 10 fish: 0.98 g, range of 0.70 - 1.3 g
- Method of breeding: not reported
- Maintenance of the brood fish: not reported

ACCLIMATION
- Acclimation period: 14 days
- Acclimation conditions (same as test or not): The fish were held in test water from the same source and at the same temperature as used during the test.
- Type and amount of food during acclimation: Commercially prepared diet supplied by Zeigler Brothers, Inc.
- Feeding frequency during acclimation: Daily
- Health during acclimation (any mortality observed): No mortality, signs of disease or stress.

FEEDING DURING TEST
- Food type: Commercially prepared diet supplied by Zeigler Brothers, Inc.
- Amount: A rate of 2% of body weight based on the initial weight for 10 negative control fish.
- Frequency: Daily

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

14 d

Hardness:

142 mg/l as CaCO3

Test temperature:

11.2 - 12.3 °C

pH:

8.0 - 8.3

Dissolved oxygen:

7.7 - 9.4 mg/l

Conductivity:

381 µS/cm

Nominal and measured concentrations:

Nominal: 9.44, 18.9, 37.8, 75.5 and 151 ng/l
Mean measured: 10.7, 21.7, 41.2, 82.3, 159 ng/l

Details on test conditions:

TEST SYSTEM
- Test vessel: The toxicity test was conducted using an exposure system consisting of a continuous-flow diluter used to deliver each concentration of the test substance, a solvent and a negative control (dilution water) to test chambers. Syringe pumps (Harvard Apparatus, were used to deliver test substance stock solutions or solvent to impartially assigned mixing chambers where the stocks or solvent were mixed with dilution water prior to delivery to the test chambers. The flow of dilution water into each mixing chamber was controlled using rotameters and was adjusted to provide approximately 10 volume additions of test water in each test chamber per day. After mixing, the flow from each mixing chamber was split to deliver test water to two replicate test chambers.
The syringe pumps used to deliver stock solutions or solvent to the mixing chambers, and the rotameters used to control the flow of dilution water to the mixing chambers were calibrated prior to the test. The proportion of the test water that was split into each replicate test chamber was checked prior to the test to ensure that flow rates varied by no more than 10% of the mean flow rate for the two replicates. Delivery of test solutions to the test chambers was initiated four days prior to test initiation in order to achieve equilibrium of the test substance. The general operation of the exposure system was checked visually at least once on the first and last days of the test and at least two times per day during the test.
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 25 litre stainless steel aquaria, filled with 15 litres test water. The depth of the test water was 18 cm.
- Aeration: No
- Type of flow-through (e.g. peristaltic or proportional diluter): Continuous flow diluter
- Renewal rate of test solution (frequency/flow rate): 10 volume additions of test water in each test chamber per day.
- No. of organisms per vessel: 10: Two replicate test chambers were maintained in each treatment and control group. Each replicate contained ten fish, resulting in a total of 20 fish per concentration.
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): 2
- Biomass loading rate: 0.072 g fish/l/day at test initiation; 0.087 g fish/l/day at test termination

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
The water used for culturing and testing was freshwater obtained from a well approximately 40 meters deep located on the Wildlife International Ltd. site. The well water is characterized as moderately-hard water. 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, the water was filtered to 0.45 µm to remove fine particles.
- Total organic carbon: Not reported
- Alkalinity: 174 mg/l as CaCO3
- Conductivity: 381 µS/cm
- Culture medium different from test medium: No
- Intervals of water quality measurement: Temperature, dissolved oxygen and pH were measured at the beginning of the test, at least twice per week and at the end of the test.
Hardness, alkalinity and specific conductance were measured at the beginning of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light, 8 hours dark, 30-minute transition period.
- Light intensity: Fluorescent light bulbs that emit wavelengths similar to sunlight.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): Observations of mortality and signs of toxicity or abnormal behaviour were made daily during the test. A representative sample of the test population was measured for total length and wet weight (blotted dry) prior to test initiation. At test termination, total length and wet weight (blotted dry) was measured for all surviving fish in each treatment and control group.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: Approximately 2
- Justification for using less concentrations than requested by guideline: n/a
- Range finding study
- Test concentrations: Not reported
- Results used to determine the conditions for the definitive study: Concentrations were based on the results of exploratory range finding toxicity data and the reported water solubility of the test substance (150.7 ng/l).

Reference substance (positive control):

not specified

Duration:

14 d

Dose descriptor:

EC50

Effect conc.:

> 159 ng/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks:

and sublethal effects

Remarks on result:

other: highest concentration tested

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

159 ng/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks:

and sublethal effects

Remarks on result:

other: highest concentration tested

Details on results:

- Behavioural abnormalities: All fish in the negative control, solvent control and treatment groups appeared normal throughout the test.
- Observations on body length and weight: There were no statistically significant differences in any of the growth parameters between the negative and solvent control groups (p > 0.05). Therefore, the control data were pooled for comparisons with the treatment groups. Dunnett's test indicated there were no statistically significant reductions in mean total length or mean wet weight among fish in any of the treatment groups in comparison to the pooled controls (p >0.05).
The threshold level for lethal and sublethal effects was >159 ng/L, the highest concentration tested. The NOEC was 159 ng/L.
- Other biological observations: No
- Mortality of control: None
- Other adverse effects control: No
- Abnormal responses: No
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No

Sublethal observations / clinical signs:

Table: Mean lengths and wet weights of rainbow trout exposed to M4Q for 14 days

Mean measured concentration (ng/l)	Mean total length (± SD) (mm)	Mean wet weight (± SD) (g)
Negative control	50.9 ± 0.21	1.23 ± 0.007
Solvent control	49.6 ± 0.99	1.13 ± 0.071
Pooled control	50.2 ± 0.93	1.18 ± 0.068
10.7	50.8 ± 0.21	1.23 ± 0.007
21.7	50.1 ± 0.07	1.22 ± 0.000
41.2	49.7 ± 1.20	1.18 ± 0.078
82.3	50.8 ± 0.07	1.24 ± 0.014
159	49.1 ± 0.28	1.17 ± 0.028

Validity criteria fulfilled:

yes

Conclusions:

A 14-day EC50 value of >159 ng/l and a NOEC value of 159 ng/l (highest concentrations tested), based on mean measured concentrations, have been determined for the effects of the test substance on sublethal effects and mortality of Oncorhynchus mykiss, in accordance with OECD Guideline 204.

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

11th May to 21st May 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1075 (Freshwater and Saltwater Fish Acute Toxicity Test)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Samples were collected from one test chamber of each treatment and control group four days prior to the start of the test after conditioning the diluter for approximately 18 hours. Water samples also were collected from alternating replicate test chambers in each treatment and control group at the beginning of the test and at 48 and 96 hours (+- 1 hour) to measure concentrations of the test substance. The samples were collected from mid-depth into a 100-ml glass volumetric flask, and processed immediately for analysis.

Vehicle:

yes

Remarks:

DMF

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Individual stock solutions were prepared for each of the five concentrations tested. A primary stock solution was prepared by mixing a calculated amount of test substance into HPLC-grade dimethylformamide (DMF) at a nominal concentration of 10 μg/ml. An aliquot of the primary stock solution was diluted with DMF to create a secondary stock solution at a nominal concentration of 1510 ng/ml. Four additional stock solutions were prepared in DMF at nominal concentrations of 94.4, 189, 378 and 755 ng/ml by proportional dilution of the secondary stock. The stock solutions were mixed by inversion, and all appeared clear and colourless. Stock solutions were stored refrigerated in glass amber bottles or a 1000 mL graduated cylinder, and aliquots of each stock were placed in the syringe every two days during the study.
The five test substance stock solutions were injected into the diluter mixing chambers at a rate of 20 μl/minute where they were mixed with dilution water delivered at a rate of 200 ml/minute to achieve the desired test concentrations.
- Controls: The negative control received dilution water only.
The solvent control was prepared by delivering HPLC-grade DMF to the mixing chamber for the solvent control.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): DMF
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): The concentration of DMF in the solvent control and all M4Q treatment groups was 0.1 ml/l.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

- Common name: Rainbow trout
- Strain: Not reported
- Source: The rainbow trout were received as juveniles from Thomas Fish Company of Anderson, California.
- Age at study initiation (mean and range, SD): Juveniles from the same source and year class.
- Length at study initiation (length definition, mean, range and SD): not reported
- Weight at study initiation (mean and range, SD): not reported
- Method of breeding: Not reported
- Maintenance of the brood fish: Not reported
ACCLIMATION
- Acclimation period: 14 days
- Acclimation conditions (same as test or not): The fish were held in test water from the same source and at the same temperature as used during the test.
- Type and amount of food during acclimation: Commercially prepared diet supplied by Zeigler Brothers, Inc.
- Feeding frequency during acclimation: Daily
- Health during acclimation (any mortality observed): No mortality, signs of disease or stress.
FEEDING DURING TEST - None

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

144 mg/l as CaCO3

Test temperature:

11.7 - 12.4 °C

pH:

8.1 - 8.3

Dissolved oxygen:

7.4 - 8.7 mg/l

Conductivity:

369 μS/cm

Nominal and measured concentrations:

Nominal: 9.44, 18.9, 37.8, 75.5 and 151 ng/l
Mean measured: 8.68, 24.5, 47.4, 101 and 182 ng/l

Details on test conditions:

TEST SYSTEM
- Test vessel: The toxicity test was conducted using an exposure system consisting of a continuousflow diluter used to deliver each concentration of the test substance, a solvent and a negative control (dilution water) to test chambers. Syringe pumps (Harvard Apparatus, Massachusetts) were used to deliver test substance stock solutions or solvent to impartially assigned mixing chambers where the stocks or solvent were mixed with dilution water prior to delivery to the test chambers. The flow of dilution water into each mixing chamber was controlled using rotameters and was adjusted to provide approximately 10 volume additions of test water in each test chamber per day. After mixing, the flow from each mixing chamber was split to deliver test water to two replicate test chambers.
The syringe pumps used to deliver stock solutions or solvent to the mixing chambers, and the rotameters used to control the flow of dilution water to the mixing chambers were calibrated prior to the test. The proportion of the test water that was split into each replicate test chamber was checked prior to the test to ensure that flow rates varied by no more than 10% of the mean flow rate for the two replicates. Delivery of test solutions to the test chambers was initiated five days prior to test initiation in order to achieve equilibrium of the test substance. The general operation of the exposure system was checked visually at least once on the first and last days of the test and at least two times per day during the test.
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 25 litre stainless steel aquaria, filled with 15 litres test water.
The depth of the test water was 18 cm.
- Aeration: No
- Type of flow-through (e.g. peristaltic or proportional diluter): Continuous flow diluter
- Renewal rate of test solution (frequency/flow rate): 10 volume additions of test water in each test chamber per day.
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): 2
- Biomass loading rate: 0.044 g fish/l/day

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The water used for culturing and testing was freshwater obtained from a well approximately 40 meters deep located on the Wildlife International Ltd. site. The well water is characterized as moderately-hard water. 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, the water was filtered to 0.45 μm to remove fine particles and passed through an ultraviolet (UV) sterilizer to remove microorganisms.
- Total organic carbon: Not reported
- Alkalinity: 184 as CaCO3
- Ca/mg ratio: not reported
- Culture medium different from test medium: No
- Intervals of water quality measurement: Temperature was measured at the beginning of the test and at the end of the test.
Dissolved oxygen and pH were measured at the beginning of the test, at approximately 24-hour intervals, and at the end of the test.
Hardness, alkalinity and specific conductance were measured at the beginning of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light, 8 hours dark, 30-minute transition period.
- Light intensity: Fluorescent light bulbs that emit wavelengths similar to sunlight.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): Observations of mortality and signs of toxicity or abnormal behaviour were made approximately 7, 24, 48, 72 and 96 hours after test initiation.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: Approximately 2
- Justification for using less concentrations than requested by guideline: n/a
- Range finding study
- Test concentrations: Not reported
- Results used to determine the conditions for the definitive study: Results used to determine the conditions for the definitive study: Concentrations were based on the results of exploratory range finding toxicity data and the reported water solubility of the test substance (150.7 ng/l).

Reference substance (positive control):

not specified

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 182 ng/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: highest concentration tested

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

182 ng/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: highest concentration tested

Details on results:

- Behavioural abnormalities: None
- Observations on body length and weight: Not reported
- Other biological observations: None
- Mortality of control: None
- Other adverse effects control: No
- Abnormal responses: None
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No

Validity criteria fulfilled:

yes

Conclusions:

A 96-hour LC50 value of >182 ng/l and a NOEC value of 182 ng/l (highest concentrations tested), based on mean measured concentrations, have been determined for the effects of the test substance on
mortality of Oncorhynchus mykiss, in accordance with OECD Guideline 203. All fish in the control and treatment groups appeared normal throughout the test and showed no overt signs of toxicity.

Description of key information

14-d EC₅₀: >159 ng/l; 14-d NOEC 159 ng/l (highest concentrations tested) (mean measured concentrations): effects of the test substance on sublethal effects and mortality of Oncorhynchus mykiss.

96-h LC₅₀: >182 ng/l; 96-h NOEC 182 ng/l (highest concentrations tested) (mean measured concentrations): effects of the test substance on mortality of Oncorhynchus mykiss.

Additional information

Two toxicity studies are available with fish:

A 14-day EC₅₀ value of >159 ng/l and a NOEC value of 159 ng/l (highest concentrations tested), based on mean measured concentrations, have been determined for the effects of the test substance on sublethal effects and mortality of Oncorhynchus mykiss, in accordance with OECD Guideline 204.

The test solution preparation method and test system design indicate that the test organisms were predominantly exposed to the parent substance.

A 96-hour LC₅₀ value of >182 ng/l and a NOEC value of 182 ng/l (highest concentrations tested), based on mean measured concentrations, have been determined for the effects of the test substance on mortality of Oncorhynchus mykiss, in accordance with OECD Guideline 203. All fish in the control and treatment groups appeared normal throughout the test and showed no overt signs of toxicity.

The test solution preparation method and test system design indicate that the test organisms were predominantly exposed to the parent substance.