Registration Dossier

Ecotoxicological information

Long-term toxicity to aquatic invertebrates

Currently viewing:

Administrative data

Link to relevant study record(s)

Referenceopen allclose all

Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Experimental starting: 2017-06-14; Experimental completion : 2017-08-03
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
The test was intended to focus on exposure to parent substance, however, analytical results showing loss of parent test material indicate that hydrolysis may have occurred and therefore exposure of the test organisms may have been to a mixture of hydrolysis products and parent substance. Measurement of the parent substance and subsequent geometric mean concentrations therefore may not give a true indication of which substance(s) the organisms were exposed to, or of the true toxicity of the parent substance.
Qualifier:
according to guideline
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
At the start of an exposure renewal interval (0 hours), samples of the fresh media were taken and analysed after preparation of the limit concentration and the solvent control.
At the end of an exposure renewal interval (24 hours), samples of the old media were taken for analysis directly from individual test vessels of the limit concentration, the solvent control and the control.
Sample storage: All samples were stored at 6 ± 2 °C until the start of the analysis, if necessary. Prepared samples were stored on an autosampler at room temperature until analysis.
Vehicle:
yes
Remarks:
acetone
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method:
Stock solution: A fresh stock solution of 1 g/L of the test item was daily prepared in acetone (solvent) before each renewal of the test solutions. The stock solution was prepared in a measuring flask (10 mL capacity) by directly weighing of 10 mg test item into the measuring glass flask, followed by filling up with acetone to 10 mL. The stock solution was used for the preparation of the limit concentration.

Preparation of the limit concentration: A limit concentration of 0.05 mg/L of the test item in Elendt M4 medium was freshly prepared before each renewal of the test solutions by adding 0.05 mL/L of the stock solution to the dilution water. The test item solution was mixed thoroughly by manual agitation.
The limit concentration was selected upon consultation with the sponsor based on the results of adequate non-GLP preliminary range finding tests.

- Controls: Control: Dilution water (Elendt M4) without addition of test item and solvent.
Solvent control: Additionally a solvent control, containing dilution water with 0.05 mL/L of acetone, was prepared and tested under the same conditions as the limit concentration and the control.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): 0.05 mL/L of acetone
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia
- Strain/clone: Clone 5
- Justification for species other than prescribed by test guideline: n/a
- Source: Origin: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), 14195 Berlin, Germany.
Breeder: Noack Laboratorien GmbH, Käthe-Paulus-Str. 1, 31157 Sarstedt, Germany

- Age of parental stock (mean and range, SD): Less than 24 hours old daphnids from a healthy stock were used for the study. Juvenile daphnids were removed from the culture vessels at the latest 24 hours before the start of the exposure and discarded. The juveniles born within this period of max. 24 hours preceding the exposure were used for the test. No first brood progeny was used for the test.

- Feeding during test
Daily; Pseudokirchneriella subcapitata (0.640 - 1.31 mL) and Desmodesmus subspicatus (0.539 - 0.921 mL) suspension was provided as food corresponding to 0.2 mg C per Daphnia and day.
There was variation according to the density of the algae suspension, but it was the same for all test groups on each feeding day.


ACCLIMATION
- Acclimation period: Acclimatisation was not necessary, because the composition of the dilution water is equivalent to the culture medium.
Culture conditions: Culture In glass vessels (2 - 3 L capacity) with approximately 1.8 L culture medium, at 20  2 °C, in an incubator, 16 hours illumination; light intensity of max. 1500 lx.
Culture medium: Elendt M4, according to OECD 211 modified to a total hardness of 160 to 180 mg CaCO3/L, is used.
Feeding of the culture stocks: The daphnids are fed at least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and Desmodesmus subspicatus, with an algae cell density of > 106 cells/mL. The algae are cultured at the test facility.
- Health during acclimation (any mortality observed): daphnids from a healthy stock were used for the study.


METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES:
10 daphnids in 10 replicates were used for the limit concentration, the solvent control and the control with one daphnid per replicate, which was held individually.
50 g test solution per replicate were weighed into each test vessel. This corresponds to 50 mL. The daphnids were transferred in a small amount of dilution water or test solution by pipette.
Renewal of the test solutions: The test solutions were renewed daily. For this purpose, a second set of test vessels were filled with the freshly prepared test solutions and the daphnids were transferred by pipette.
First appearance of juveniles was checked daily. The number of neonates (alive and dead progeny) was counted and dates of release were recorded once a day from the first day of appearance of neonates until the end of the test. The neonates (alive and dead) were removed after counting and before addition of algae to prevent them from consuming food intended for the adults. The number of aborted eggs or dead offspring and dates of observation were recorded.

Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
21 d
Hardness:
163 - 183 mg/L as CaCO3
Test temperature:
20.1 - 20.9 °C
pH:
In the limit concentration:
Day 0 fresh medium: 7.75; Day 1 Old medium: 7.96
Day 7 fresh medium: 8.58; Day 8 Old medium: 8.59
Day 14 fresh medium: 8.96; Day 15 Old medium: 8.53
Dissolved oxygen:
7.49 - 10.5 mg/l
Nominal and measured concentrations:
Nominal: 0.05 mg/l
Measured geo. mean: 0.00378 mg/l
The measured concentrations of the test item in fresh media at the start of the respective exposure intervals (0 hours) were in the range of 0.000656 and 0.0389 mg test item /L. At the end of the respective exposure intervals (24 hours), the measured test item concentrations in the old media were in the range of 0.000408 and 0.0107 mg test item /L. The geometric mean measured concentration is 0.00378 mg test item/L.
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass beakers (5 (ID) x 8 (H) cm), 100 mL capacity
- Type (delete if not applicable): open / closed; loosely covered with watch glasses
- Material, size, headspace, fill volume: Glass, 100 ml capacity, filled with 50 ml test solution.
- Aeration: No
- Type of flow-through (e.g. peristaltic or proportional diluter): n/a
- Renewal rate of test solution (frequency/flow rate): Daily renewal
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10
- No. of vessels per vehicle control (replicates): 10
- Biomass loading rate: Not reported

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4
- Culture medium different from test medium: No
- Intervals of water quality measurement: The water quality parameters (i.e. pH-values, dissolved oxygen concentration, total hardness and water temperature) were measured once within every 7 days of exposure, in fresh media at the start of an exposure interval (0 hours) and in old media at the end of an exposure interval (24 hours), in one replicate of the control, the solvent control and the limit concentration. The water quality parameters in fresh media at the start of an exposure interval were measured in an additional replicate without daphnids of the control, the solvent control and the limit concentration. At the end of an exposure interval, the water quality parameters of the old media were measured in a test vessel of the control, the solvent control and the limit concentration, which contained daphnids and food algae.
The temperature in the incubator was recorded throughout the test period with a thermo-hygrograph.


OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16/8 hours light/dark cycle
- Light intensity: Max. 1500 lx

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
Mortality of parental daphnids: The number of parental daphnids, which were immobilized or died during the test and the dates of death were observed and recorded once a day at least at the same time as the offspring was counted. Dead specimens were removed as soon as they were detected.

Neonates: First appearance of juveniles was checked daily. The number of neonates (alive and dead progeny) was counted and dates of release were recorded once a day from the first day of appearance of neonates until the end of the test. The neonates (alive and dead) were removed after counting and before addition of algae to prevent them from consuming food intended for the adults. The number of aborted eggs or dead offspring and dates of observation were recorded.

Additional observations / Condition of parental daphnida: Abnormalities (e.g. swimming behavior, number of males and winter eggs) were observed and recorded on each day of observation.

Total body length / Mean dry weight of parental daphnids: At the end of the test, the total length excluding the anal spine of each survived parental daphnid and the mean dry weight of the survived parental daphnids of the control, the solvent control and the limit concentration were determined. As stated in the OECD protocol, this may be useful additional information but is not used for determination of a NOEC.

Additional endpoints: The time to first brood, the intrinsic rate of population increase and the number and size of first brood per animal were reported, but not used for endpoint calculations.


TEST CONCENTRATIONS
-Spacing factor for test concentrations: n/a
-Justification for using less concentrations than requested by guideline: The limit concentration was selected upon consultation with the sponsor based on the results of adequate non-GLP preliminary range finding tests. Test item concentration and application for the definitive study was based on the observation that the maximum water solubility seems to between 0.01 – 0.05 mg/L, based on the measured water solubility of the parent substance and results from the 1st preliminary range finding test (no mortality up to 1.0 mg/l prepared without solvent). Recoveries obtained from the 2nd preliminary range finding test were considered as a possible overestimation of solubility due to use of the solvent acetone. Based on this, the application system with solvent acetone with a test item concentration of 0.05 mg/L was chosen to combine the findings of the two preliminary range finding tests and obtain results inside the range of the true water solubility of the test item under test conditions. (See further details on the non-GLP preliminary studies in the Section “Any other information on materials and methods” below).
-Range finding study
-Test concentrations: 1st preliminary range finding test: 0.01, 0.1 and 1 mg/l slow stirring.2nd preliminary range finding test: 0.01, 0.1 and 1 mg/l with acetone - Results used to determine the conditions for the definitive study:
1st preliminary range finding test: No mortalities
2nd preliminary range finding test: Mortalities in the 1 mg/l concentration (above the LoS)
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.05 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Remarks:
and mortality
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.004 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Remarks:
but exposure is also to hydrolysis products
Basis for effect:
reproduction
Remarks:
and mortality
Details on results:
- Behavioural abnormalities: None
- Observations on body length and weight:
- Other biological observations:
- Mortality of control: Not significant
- Other adverse effects control:
- 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
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- Relevant effect levels: EC50 1.31 mg/L

Table: Effects on Reproduction for all Introduced and for the Surviving Parents (based on the nominal / geometric mean measured concentration of the test item)

Nominal test item

concentration [mg/L]

Geometric mean measured test item

concentration [mg/L]

Mean number of offspring per

introduced parental daphnid

   

Mean number of offspring

per survived parental daphnid

   
     Mean  SD  CV  Mean  SD  CV
 0.05  0.00378  96.8  26.9  27.8  103.7  16.9  16.3
 Solvent control  -  90.5  27.4  30.2  102.4  8.7  8.5
 Control  -  80.1  30.0 37.4  91.5  12.8  14.0

Table: Mortality [%] of the Adult Daphnids after 7, 14 and 21 Days of Exposure (n = 10)

Nominal test item

concentration [mg/L]

Geometric mean measured test item

concentration [mg/L]

Adult mortality (%)

   
     7 days  14 days  21 days
 0.05  0.00378  0  0  10
 Solvent control  -  0  10  20
 Control  -  0  10 20

Table: First Appearance of Living Juveniles and Mean Number of Broods in the Individual Groups

Nominal test item

concentration [mg/L]

 

Geometric mean measured test item

concentration [mg/L]

 

 Day of first appearance of living juveniles

at the parental daphnid in replicate no.

                   First appearance
     1  2  3  4  5  6  7  8  9  10  Mean day
 0.05  0.00378  9  10  10  9  8  8  8  8  8  8  8.6
 Solvent control  -  9  8  9  8  8  8  10  9  9  10  8.8
 Control  -  9  9  8  9  8  10  8  9  9  9  8.8

Table: Stillborn Juveniles and Aborted Eggs produced by the Parental Daphnids after 21 Days

Nominal test item

concentration [mg/L]

Geometric mean measured test item

concentration [mg/L]

Number of stillborn juveniles

 Number of aborted eggs  Total number

Number of adults [N]

 0.05  0.00378  0  0  0  9
 Solvent control  -  0  0  0  8
 Control  -  0  0 0  8

N = number of the parental daphnids

Table: Measured Concentrations of 2,4,6,8-tetramethyl-2,4,6,8-tetravinylcyclotetrasiloxane during the Definitive Test (Days 0, 1, 7, 8, 14 and 15)

 Sampling Date

 Day 0 fresh

media

 

 Day 1 old

media

 

 Day 7 fresh

media

 

 Day 8 old

media

 

 Day 14

fresh media

 

 Day 15 old media

 

 Geometric mean

 Nominal test item concentration

[mg/l]

 Meas. conc.

[mg/l]

 %

Meas. conc.

[mg/l] 

 %

 Meas. conc.

[mg/l]

 %

 Meas. conc.

[mg/l]

 %

 Meas. conc. [mg/l]

 %

 Meas. conc [mg/l]

 Meas. conc. [mg/l]

0.0500 

 0.0389

 78

 0.0107

 21

 0.01211)

 24

 0.00215

 4

 0.000656

 1

 0.0004082)

 1

 0.00378

 Solvent control

 <LOQ

 -

 <LOQ

 -

 <LOQ

-

 <LOQ

 -

 <LOQ

 -

 <LOQ

 -

 <LOQ

 Control

 <LOQ

 <LOQ

 <LOQ

 -

 <LOQ

 -

 <LOQ

 -

 <LOQ

 -

 <LOQ

Meas. conc. = measured concentration of the test item, enrichment and dilution factors taken into account

% = percent of the nominal concentration of the test item

LOQ = limit of quantification of the analytical method (0.5 µg/L of the test item)

1) = value does not include two separate replicates which were measured on 2017-07-20, since they showed a significant decrease in the concentration (0.00211 mg/L), no relative standard deviation was calculated

2) = measured value > 70% LOQ, therefore taken into account

Decrease of the test item recoveries over the course of the study may be explained by the increase of pH in the test vessels throughout exposure. While the pH was within the valid range throughout exposure, elevated pH levels may have affected the stability of the test item.

The reason for this elevation of pH during the exposure period is unclear, but reliable und uniform pH adjustment after application of the test solutions in the test vessels was not technically feasible.

As reported in IUCLID Section 5.1.2, hydrolysis of the test substance may be acid or base catalysed; the rate of reaction is expected to be slowest at around pH 7 and increase as the pH is raised or lowered. The predicted hydrolysis half-life of the substance at pH 7 and 20-25°C is approximately 63 hours; at pH 9 and 20-25°C the hydrolysis half-life is 0.8 hours. Consequently, the increasing pH of the culture media/dilution water over the course of the study may have increased the degree of hydrolysis of the test item, causing the decrease of the test item recoveries.

The time interval between sampling and extraction of the sample with cyclohexane is not reported. Further hydrolysis of the test item may have occurred in the time between sampling and extraction, and it is possible that this may also have impacted the test item recoveries reported although it is likely to have less impact overall than possible pH-mediated effects.

 

Table: Water quality parameters: pH-Values

Nominal

test item

concentration

[mg/L]

Geometric

mean measured

test item

concentration

[mg/L]

Day 0

Day 1

Day 7

Day 8

Day 14

Day 15

Fresh medium

Old medium

Fresh medium

Old medium

Fresh medium

Old medium

0.05

0.00378

7.75

7.96

8.58

8.59

8.96

8.53

Solvent control

7.66

8.04

8.58

8.43

8.96

7.57

Control

7.92

7.90

8.51

8.45

8.92

7.71

 

 

Validity criteria fulfilled:
yes
Conclusions:
A 21-day No-Observed-Effect Concentration (NOEC) of ≥0.00378 mg/L based on geometric mean measured concentrations has been determined for the effects of the test substance on survival and reproduction of Daphnia magna, in accordance with OECD guideline 211. No effects were observed at the maximum achievable concentration in test medium.
Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
2003-04-09 to 2003-04-30
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
Triplicate 100-µL aliquots of the stock solution used to prepare the test media were assayed.
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: Stock solution preparation (vehicle, solvent, concentrations) and stability:  The primary stock solution for [14C] D5 was prepared by quantitatively transferring the entire amount of test substance received to a 100-mL volumetric flask with acetone and bringing to final volume with acetone.  Triplicate 100-µL aliquots of the stock solution were then assayed by liquid scintillation counting (LSC).  Based on this analysis and the Study Sponsor supplied specific activity of 5.758 µCi/mg  (12783 dpm/µg), the stock solution was determined to have a concentration of 256.9 µg/mL.  Secondary dosing stock solutions (11, 21, 43, 85 and 170 µg/mL) were prepared weekly by adding the appropriate amount of the 256.9 µg/mL primary stock solution to a 5-mL volumetric flask and bringing it to volume with acetone as follows:

Stock Volume of 256.9 ug/mL Volume diluted to
Conc. Primary Stock Used
(µg/mL) (mL) (mL)
11 0.215 5.0
21 0.410 5.0
43 0.840 5.0
85 1.660 5.0
170 3.310 5.0

The individual exposure solutions were prepared at test initiation and at each renewal interval (daily) using the following preparation scheme:

Nominal Test Stock Sol. Volume of Stock Volume of
Conc. Used Solution Dilution
(ug/L) (ug/mL) Added(uL) Water (L)
Solvent control NA (acetone) 6.5 0.065
1.1 11 6.5 0.065
2.1 21 6.5 0.065
4.3 43 6.5 0.065
8.5 85 6.5 0.065
17 170 6.5 0.065
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM

- Source: Laboratory cultures maintained at Springborn Smithers. Test organisms were < 24 hours old at the initiation of the test.  

- Age of parental stock (mean and range, SD): Test organisms were not from the first brood of culture organisms (first brood occurred on March 25,2003) and were derived from healthy culture stock.

- Type and amount of food: During culture, parental daphnids were fed 2.0 mL of a unicellular green algae (Ankistrodesmus falcatus) once daily.  The food solution contained approximately 4 x 10E7 cells/mL of algae.  In addition, daphnids were fed 0.5 mL of YCT suspension (yeast, cereal leaves and digested flaked fish food) once daily

- Culture medium: The culture water was prepared by fortifying well water based on the formula for hard water (U.S. EPA, 1975) and filtering it through Amberlite XAD-7 resin column to remove any potential organic contaminants.  This water had total hardness and total alkalinity ranges as calcium carbonate (CaCO3) of 160 to 170 mg/L and 110 to 120 mg/L, respectively, a pH range of 7.4 to 8.2, a temperature of 19 to 21 ºC, a dissolved oxygen concentration of 6.4 to 9.4 mg/L, and a specific conductivity of 500 micromhos per centimeter. 

ACCLIMATION

- Acclimation conditions: The daphnid culture area received a regulated photoperiod of 16 hours of light and 8 hours of darkness.  Light intensity of 70 to 90 foot candles at the surface of the culture solution was provided by fluorescent bulbs.  Culture vessels were maintained in a temperature-controlled water bath designed to maintain the culture solution temperature at 20 ± 2 ºC. 
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d
Hardness:
Total hardness range as calcium carbonate (CaCO3): 160 to 170 mg/L
Test temperature:
20 to 23ºC for test solutions (water bath temperature ranged from 20 to 21 ºC)
pH:
6.6-8.5
Dissolved oxygen:
Dissolved oxygen concentration range: 3.9-9.4 mg/L
Salinity:
not applicable
Nominal and measured concentrations:
Nominal test concentrations were 1.1, 2.1, 4.3, 8.5 and 17 µg/L.  The nominal concentrations were selected to equal, but not to exceed, the reported water solubility of D5 (17 µg/L, Chandra, 1997).

Time-weighted mean measured concentrations: 1.0, 1.7, 3.5, 7.2 and 15 µg/L

The results are interpreted with reference to measured concentrations
Details on test conditions:
TEST SYSTEM

- Test vessel: The test vessels used to expose the test organisms were 65-mL screw top GC vials.  Test vessels were filled completely (no headspace) and sealed with the screw top cover to minimize the loss of D5 from the test solution.  The screw top covers contained a Teflon-lined septum through which each stock solution was added.  

- No. of vessels per concentration (replicates): Eleven replicate test vessels were established for each nominal concentration and the controls (i.e., replicates A through J for the biological measurements and replicate K for confirmation of test substance concentration in newly prepared test solutions).  

- Test design (number of replicates, individuals per replicate, concentrations):  The test was initiated when daphnids, < 24 hours old, were impartially distributed to each of the eleven replicates for each nominal concentration and the controls and vessels were dosed.  Daphnids were added impartially to an intermediate beaker by adding no more than one daphnid to each vessel until all vessels contained one daphnid.  This procedure was repeated until each beaker contained eleven daphnids.  The test was initiated when each intermediate beaker of eleven daphnids was distributed (one daphnid per replicate vessel) to an impartially selected treatment level or control.  Ten replicate vessels (A through J) were used to monitor the biological response of the organisms.  The eleventh replicate vessel was established for each treatment level (replicate K) and contained one daphnid as well as food to mimic exposure conditions.  Two additional replicates containing no food or daphnids (replicates L and M) were prepared for the 17 µg/L treatment level for analytical purposes.  In addition, two more replicate vessels (replicates N and O) were prepared for the control and 17 µg/L treatment level for determination of total hardness, total alkalinity and specific conductance on test day 0.  These replicates were prepared only once at the beginning of the study.

TEST MEDIUM / WATER PARAMETERS

- Source/preparation of dilution water: Fortified well water. Dilution water chemistry (hardness, alkalinity, pH, specific conductivity):  The water prepared during the definitive exposure was characterized as having a total hardness range as calcium carbonate (CaCO3) of 160 to 170 mg/L, a total alkalinity (as CaCO3) of 110 mg/L, a pH of 7.9, a dissolved oxygen concentration range of 9.4 to 9.8 mg/L, and a specific conductivity of 500 µmhos/cm.  
FEEDING DURING TEST

- Feeding during test: During the definitive exposure, the test organisms were fed a diet consisting of a suspension of green algae (Ankistrodesmus falcatus; 4 x 10E7 cells/mL), introduced at a rate of 0.200 mL of green algae suspension and 0.050 mL of YCT suspension per test vessel, once daily.
OTHER TEST CONDITIONS

LIGHTING

- Lighting (quality, intensity, and periodicity):  The test area was illuminated with fluorescent bulbs at an intensity of 8 to 12 µEom-2os-1 at the solutions' surface.  Light intensity was measured using a LI-COR photometer (Model LI-189).  The photoperiod during the test was the same as in the culture area.  Sudden transitions from light to dark and vice versa were avoided.  


EFFECT PARAMETERS MEASURED:  21-day survival, reproduction and growth (dry weight and total body length)
Reference substance (positive control):
no
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 15 µg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
act. ingr.
Basis for effect:
other: 21-day survival, reproduction and growth (dry weight and total body length)
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
> 15 µg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
act. ingr.
Basis for effect:
other: mortality, reproduction and growth
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 15 µg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
act. ingr.
Basis for effect:
other: mortality, reproduction and growth
Remarks on result:
other: not calculable
Details on results:
After 21 days of exposure, survival among the dilution water control and solvent control daphnids averaged 90% and 100%, respectively. The mean number of offspring released by each female daphnid of the dilution water control and the solvent control during the 21-day test was 148 and 145 offspring per female. Statistical analysis (t-Test) determined that there was no significant difference between the dilution water control and the solvent control survival, reproduction, length and dry weight; therefore the dilution water control and solvent control data were pooled for each endpoint. The coefficient of variation around the number of offspring per female in the pooled control of this study was 7.5%. As demonstrated by the performance of the dilution water control, the exposure system provided conditions that were appropriate for promoting acceptable survival and reproduction of the test species and met the minimum standard criteria established by OECD guidelines (i.e., > or = 80% survival and > or = 60 offspring per female). The dilution water control daphnids released their first brood of offspring on test day 8. Time of first brood release was within normal performance expectation for this species.

At test termination (day 21) survival of 100% was observed among daphnids exposed to all treatment levels tested (1.0 to 15 µg/L). Statistical analysis determined that no significant difference in survival was observed in any of the treatment levels tested when compared to the pooled control data (95%). The 21-day EC50 for survival was empirically estimated to be > 15 µg/L.

Following 21 days of exposure, daphnids exposed to the 1.0, 1.7, 3.5, 7.2 and 15 µg/L treatment levels had released a mean cumulative number of offspring per female of 150, 148, 145, 138 and 139, respectively, and were not significantly different from the mean cumulative number of offspring
released among the pooled control organisms (146 offspring per female). First brood release in all treatment levels and controls tested occurred on test day 8. The 21-day EC50 for reproduction was empirically estimated to be > 15 µg/L.

At test termination (day 21), the mean total body length of daphnids exposed to the control and solvent control averaged 5.01 and 4.97 mm, respectively. Statistical analysis (t-Test) determined that there was no significant difference between control and solvent control; therefore control and solvent control data were pooled (pooled control = 4.99 mm). The mean total body length of daphnids exposed to the 1.0, 1.7, 3.5, 7.2 and 15 µg/L treatment levels averaged 5.00, 4.99, 4.99, 5.03 and 5.05 mm, respectively, and were not significantly different from the mean total body length of the pooled control daphnids (4.99 mm).

At test termination (day 21), the mean dry weights of daphnids exposed to the control and solvent control averaged 1.16 and 1.08 mg, respectively. Statistical analysis (t-Test) determined that there was no significant difference between control and solvent control; therefore control and solvent control data were pooled (pooled control = 1.12 mg).

The mean dry weights of daphnids exposed to the 1.0, 1.7, 3.5, 7.2 and 15 µg/L treatment levels averaged 1.15, 1.19, 1.17, 1.19 and 1.20 mg, respectively, and were not significantly different from the mean dry weight of the pooled control daphnids (1.12 mg).

The data generated during the 21-day static-renewal exposure demonstrated that concentrations up to 15 µg/L of D5 had no adverse effects on survival, reproduction or growth of Daphnia magna. The 21-day Lowest-Observed-Effect Concentration (LOEC) and No-Observed-Effect Concentration (NOEC) for D5 and D. magna was > 15 µg/L and ≥15µg/L, respectively.
Reported statistics and error estimates:
 The 21-day EC50 values for survival, reproduction and growth were empirically estimated.

Biological observations:

Survival, reproduction, total body lengths and dry weights of parental daphnids at 

termination of the full life-cycle exposure with D5 and daphnids (Daphnia magna):
                                
                          Test Day 21
Time-Weighted   Mean    Mean Cumulative  Mean Total  Mean Dry
Mean            %       # of Offspring   Body        Weight
Conc.          Survival    Released      Length      in mg
(ug/L)                  /Female(SD)      in mm(SD)    (SD)  
  
Control         90         148(13)      5.01(0.09)   1.16(0.05)
Solvent Control 100        145(10)      4.97(0.09)   1.08(0.16)
Pooled Control   95        146(11)      4.99(0.09)   1.12(0.13)
1.0             100        150(11)      5.00(0.06)   1.15(0.04)
1.7             100        148(13)      4.99(0.07)   1.19(0.04)
3.5             100        145(12)      4.99(0.07)   1.17(0.04)
7.2             100        138(8)       5.03(0.08)   1.19(0.06)
15              100        139(18)      5.05(0.08)   1.20(0.05)
                                
SD = standard deviation

Established endpoints for the full life-cycle exposure with D5 and daphnids (Daphnia magna):
Endpoint        Concentration(µg/L)        95%C.I.(µg/L)
EC50 (survival)      >15                    NA
EC50 (reproduction)  >15                    NA
EC50 (growth)        >15                    NA
21-day NOECa         
15                    NA
21-day LOEC          >15                    NA

NA = not applicable

EC50, NOEC and LOEC at 21 days: The data generated during the 21-day static-renewal exposure demonstrated that concentrations up to 15 µg/L of D5 had no adverse effects on survival, reproduction or growth of Daphnia magna.

The 21-day Lowest-Observed-Effect Concentration (LOEC) and No-Observed-Effect-Concentration (NOEC) for D5 and Daphnia magna was >15 µg/L and  15 µg/L, respectively. 

The 21-day EC50 value for survival, reproduction and growth was empirically estimated to be greater than the highest concentration tested (i.e., 15 µg/L).

Validity criteria fulfilled:
yes
Conclusions:
A 21-day No-Observed-Effect Concentration (NOEC) of ≥15µg/L has been determined for the effects of the test substance on survival, reproduction and growth of Daphnia magna.

Description of key information

Long-term toxicity to invertebrates: 21 day NOEC ≥0.00378 mg/l (guideline OECD 211 (Daphnia magna Reproduction Test)).    

Key value for chemical safety assessment

Additional information

A reliable long-term invertebrate toxicity study is available for the registration substance, 2,4,6,8-tetramethyl-2,4,6,8-tetravinyltetrasiloxane (Vi4-D4, CAS 2554-06-5).

A 21 day No-Observed-Effect Concentration (NOEC) of ≥0.00378 mg/l based on geometric mean measured concentrations, highest concentration tested, has been determined for the effects of Vi4-D4 on survival and reproduction of Daphnia magna, conducted in accordance with OECD TG 211 and in compliance with GLP (Noack, 2018). No effects were observed at the maximum achievable concentration of the test substance in test media.

The hydrolysis half-life of the substance and test media preparation methods indicate that the test organisms will have been exposed to the parent substance.

The data above are supported by read-across evidence relevant to the impurities. The registration substance has an average purity of >70% Vi4-D4, with <20% 2,4,6,8,10-pentamethyl-2,4,6,8,10-pentavinylcyclopentasiloxane Vi5-D5 (CAS 17704-22-2; Impurity 1) and <10% 2,4,6-trimethyl-2,4,6-trivinylcyclotrisiloxane Vi3-D3 (CAS 3901-77-7; Impurity 2) present as impurities. After due consideration of the properties, the presence of these impurities is not expected to affect the overall hazard profile of the substance.

Read-across studies are in place as supporting studies, to consider the properties of the impurities. Data for Vi5-D5 are read-across from decamethylcyclopentasiloxane D5 (CAS 541-02-6). These siloxanes have similar properties with regard to aquatic ecotoxicity.  Further information is given in a supporting report (PFA, 2017) attached in Section 13 of the IUCLID dossier.

A reliable long-term invertebrate study is available for the siloxane D5 (CAS 541-02-6) and is read-across to Impurity 1 (Vi5-D5) (Springborn Smithers, 2003). This study (21 day reproduction test with Daphnia magna) gave a result 21 day NOEC value of ≥15 µg/l. Effects were not seen within the limit of solubility in test media.

No reliable long-term invertebrate study is available to read-across to Impurity 2. Effects are not anticipated at the limit of solubility and the available data for the main data set are conservative in respect of Impurity 2, which has a much shorter hydrolytic half-life and lower log Kow. The chemical safety assessment is therefore not sensitive to this data gap.

The available long-term aquatic toxicity data indicate that there are no effects on aquatic organisms at the maximum achievable concentration of the test substance in test media.