4,4,7,7-tetraethoxy-3,8-dioxa-4,7-disiladecane

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016-04-08 to 2016-08-05

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Test conducted using concentrations above the limit of solubility of the parent substance (predicted solubility is 20 mg/l – test concentrations range from 41-100 mg/l)

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Determination of the TOC:All concentration levels and the control were verified via analysis of the total organic carbon (TOC, according to DIN EN 1484), at the start of the exposure intervals (0 and 24 hours) and at the end of the exposure (24 and 48 hours).  
Sampling for the analytical monitoring: At the start of the exposure intervals (0 and 24 hours), sampling was carried out after preparation of the concentration levels. At the end of the exposure intervals (24 and 48 hours), samples were taken from additional replicates with test media, but without daphnids, which were incubated under test conditions until sampling.
Specific analysis was not possible.

Vehicle:

no

Remarks:

Solvent only used within the first non-GLP preliminary test

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A glass flask was filled with 1000 mL of ISO test water and an appropriate amount of the test item (density of the test item of 0.96 g/cm³ was taken into account) was slowly and steadily placed by pipette under the water surface to avoid concentration peaks leading to polymerization or other adverse effects.
A slow stirring procedure was applied. Gentle stirring was carried out for 2 hours with a magnetic stirrer at room temperature. Thereafter, the respective solution were allowed to stand for 1 hour for separation of the phases. Then, the respective test item concentration was removed from the approximate centre of the aqueous phase.
This procedure was carried out for each concentration level prior to the start of the test and prior to the water renewal after 24 hours.
The glass walls of the test vessels were saturated with the test solutions prior to initiating the exposure, to prevent possible adsorption of the test item to the glass surface during the exposure phase. For this purpose, the glass beakers were filled with the test solutions and allowed to stand for 15 minutes for saturation of the glass walls. Thereafter, the glass beakers were slightly swirled, emptied and refilled.

- Controls: Dilution water without test item incubated under the same conditions as the test groups

- Evidence of undissolved material (e.g. precipitate, surface film, etc.): All concentration levels were visually clear throughout the exposure intervals.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia Magna
- Strain/clone: STRAUS (clone 5)
- Source: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), 14195 Berlin, Germany
- Age of parental stock (mean and range, SD): 2 to 24 hours old daphnids
- Feeding during test : No

ACCLIMATION
- Acclimation period: At least 2 hours in dilution water
- Acclimation conditions (same as test or not): No differences stated
- Health during acclimation (any mortality observed): Not reported

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

Dilution water at start of the exposure intervals (0 and 24 hours): 269-260 mg CaCO3/L

Test temperature:

Dilution water at start of the exposure intervals (0 and 24 hours): 20.8 and 20.7°C
The incubator temperature was recorded throughout the period of the test. During the test period, the temperature in the incubator was 19 – 20 °C

pH:

Dilution water at start of the exposure intervals (0 and 24 hours): 7.86 (adjusted to 7.01 by addition of 1 M HCL) and 7.94 (adjusted to 7.07 by addition of 1 M HCL)
Test media at start of exposure intervals (fresh media) (0 and 24 hours): 7.14 - 7.20 and 7.12 - 7.20
Test media at end of exposure intervals (old media) (24 and 48 hours): 7.33 - 7.40 and 7.53 - 7.72

Dissolved oxygen:

Dilution water at start of the exposure intervals (0 and 24 hours): 9.28 and 9.35 mg/L
Test media at start of exposure intervals (fresh media) (0 and 24 hours): 8.15 - 8.37 mg/L and 7.92 - 8.38 mg/L
Test media at end of exposure intervals (old media) (24 and 48 hours): 7.24 - 8.21 mg/L and 7.44 - 8.46 mg/L

Conductivity:

Dilution water at start of the exposure intervals (0 and 24 hours): 645 µS/cm and 645 µS/cm

Nominal and measured concentrations:

Nominal concentration (mg/L): 41.0, 51.2, 64.0, 80.0, 100
Expected TOC based on nominal (mg C/L): 19.5, 24.3, 30.4, 38.0, 47.5
Measured TOC at start of exposure (mg C/L): 16.1, 22.3, 26.1, 31.3, 35.2
At the end of the exposure intervals (after 24 and 48 hours), the measured TOC concentrations were in the range of 96 - 106% of the initially measured values. Generally, the measured values were constant during the exposure period.

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers, loosely covered with watch glasses
- Material, size, headspace, fill volume: 4 (ID) x 7 (H) cm), 50 mL capacity. Test volume 20 ml. 20 g test solution per replicate were weighed into the test vessel. This corresponds to 20 mL per test vessel. The daphnids were inserted with a small amount of dilution water (start of the exposure) or test solution (water renewal) by a pipette.
- Aeration: No
- Renewal rate of test solution (frequency/flow rate): The study was performed with a semi-static test design (with a renewal of the test solutions after 24 hours) as a worst case exposure, because the test item undergoes rapid hydrolysis and the unspecific analytical method which was used for the analytical monitoring cannot confirm the chemical stability of the test item. The test solutions were renewed after 24 hours. For this purpose, a second set of test vessels was filled with the freshly prepared test solutions and the daphnids were transferred by pipette
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO test water, according to OECD 202, Annex 3. The dilution water was adjusted to pH 7 ± 0.2 by addition of 1 M HCl
- Total organic carbon: Not reported
- Particulate matter: Not reported
- Metals: Not reported
- Pesticides: Not reported
- Chlorine: Not reported
- Alkalinity: Not reported
- Ca/mg ratio: Not reported
- Culture medium different from test medium: No differences stated
- Intervals of water quality measurement: Dilution water: Prior to the start of the exposure (0 hours) and the water renewal (24 hours) (pH-value, dissolved oxygen concentration, temperature and conductivity). Test media: At the start of the exposure intervals (0 and 24 hours) (fresh media) (pH-value, dissolved oxygen concentration) (via one additional replicate per concentration level and control) and at the end of the exposure intervals (24 and 48 hours) (old media) (pH-value, dissolved oxygen concentration) (via all replicates per concentration level and control).

OTHER TEST CONDITIONS
- Adjustment of pH: The pH was measured directly after addition of the test item. Since the measured pH-values were within pH 7 ± 0.2, no re-adjustment of the pH was carried out.
- Photoperiod: 16/8 hours light/dark cycle
- Light intensity: Diffuse light, light intensity of max. 20 µEm-2s-1 (max. 1340 lx)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobilisation was determined in all groups after 24 and 48 hours. An animal was considered immobile, if it was not able to swim in the water phase within 15 seconds after gentle agitation of the test vessel. Other observations were not made.

VEHICLE CONTROL PERFORMED: N/A

RANGE-FINDING STUDY
- Test concentrations: First non-GLP range finder (static, with 100 µL/L of the Solvent THF, test solutions stirred 24hrs prior to test): 1, 10 and 50 mg/L (nominal); Second non-GLP range finder (semi-static - reneval at 24 hours, without solvent, test solutions stirred 24 hours with 2 hours to settle prior to test): 100 mg/l (nominal)
- Results used to determine the conditions for the definitive study: First range finder (1, 10, 50 mg/l): 0% immobilisation at 24 and 48 hours. Second range-finder (100 mg/l): 5% immobilisation at 24 hours; 48% immobilisation at 48 hours.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate p.a. (SIGMA)

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

72.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: (95% CL 65.9 - 81.3 mg/L)

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

43.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: (95% CL < 41.0 - 53.6 mg/l)

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Since no specific analysis was carried out, the effect values were based on the nominal concentration levels of the test item.

Results with reference substance (positive control):

The percentage immobility for the reference item was determined after 24 hours. The EC50-value with 95% confidence limits (CI) was calculated by sigmoidal dose-response regression. The EC50-value for the most recent of the monthly performed reference tests was:
EC50: 2.08 mg/L (Cl: 1.00 - 4.00 mg/L)
The EC50-value of the reference item potassium dichromate after 24 hours is within the required concentration range of 0.6 - 2.4 mg/L of quality criteria according to AQS P 9/2 (02/2000) for daphnids clone 5 cultured in Elendt M4 medium. The EC50-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD-Guideline 202.

In the control group, no daphnids were immobilised or showed any signs of disease or stress, e.g. discoloration or unusual behaviour such as trapping on the surface of the water, during the -hour test period (required: not more than 10% of the daphnids in the control).

 

The dissolved O2 concentration at the end of the exposure intervals was ≥ 7.24 mg/L (required: ≥ 3 mg/L at the end of the exposure intervals) in the test vessels of all tested concentration levels and the control.

Table 1. Immobilisation Rates after 24 and 48h of Exposure during definitive test

Nominal test concentration (mg/L)	Percentage immobility after 24 hours (Mean)	Percentage immobility after 48 hours (Mean)
0 (Control)	0	0
41.0	0	5
51.2	0	25
64.0	0	30
80.0	0	65
100	0	90

 

Table 2. Measured TOC concentrations during definitive test

Nominal test concentration (mg/L)	Expected TOC value^2) (mg C/L)	Measured concentration^1)at start of the exposure interval 0 hour (mg/L)	Measured concentration^1)at the end of the exposure interval 24 hours (mg/L)	Measured concentration^1)at the start of the exposure interval 24 hours (mg/L)	Measured concentration^1)at the end of the exposure interval 48 hours (mg/L)
0 (Control)	< 2.00	< 2.00*	2.39*	< 2.00*	2.85
41.0	19.5	16.1	16.2*	18.9*	19.9
51.2	24.3	22.3	22.5*	24.7*	24.5
64.0	30.4	26.1	25.8*	28.4*	28.8
80.0	38.0	31.3	30.1*	25.0*	25.0
100	47.5	35.2	34.3*	39.6*	40.1

 

1)    = measured TOC concentration, mean value of 2 injections, dilution factor taken into account

2)    = calculatedwith factor 0.475 carbon content

* = re-analysed

The limit of quantification of the analytical method is set to 2.00 mg/L.

Validity criteria fulfilled:

yes

Conclusions:

A 48 hour EC50 value for Daphnia magna of 72.6 mg/L (nominal) (95% confidence limits: 65.9 - 81.3 mg/L) was determined in a reliable study conducted according to an appropriate OECD test guideline, with acceptable restrictions. The restrictions were that the test was conducted using concentrations above the limit of solubility of the parent substance (predicted solubility is 20 mg/l – test concentrations range from 41-100 mg/l). Test conditions and preparation of test solutions was chosen to keep the parent substance as stable as possible, but hydrolysis cannot be foreclosed. Therefore, all effect values are based on the complete mixture of parent and hydrolysis products, expressed as nominal values of the test item.
The study was performed with a semi-static test design (with a renewal of the test solutions after 24 hours) as a worst case exposure, because the test item undergoes rapid hydrolysis and the unspecific analytical method which was used for the analytical monitoring cannot confirm the chemical stability of the test item. The test solutions were renewed after 24 hours. No trapping on the surface was recorded for any replicate during the exposure phase.

Description of key information

Short-term toxicity to invertebrates: 48 hr EC50 72.6 mg/L (nominal) (OECD 202), in terms of the substance as tested. The observations in this study are attributed to the exposure of test organisms to the parent substance in the test system. The parent substance hydrolyses at a moderate rate in water but only non-specific analytical evidence is available. In the absence of substance-specific analytical evidence, the EC50 in terms of the parent substance can be estimated as approximately 57 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

72.6 mg/L

Additional information

A 48-h EC50 value of 72.6 mg/L and EC10 of 43.6 mg/L (nominal) have been determined for the effects of the registered substance on mobility of Daphnia magna. In view of the test media preparation method (shortened stirring time) and semi-static exposure regime (24-hourly media renewal) it is likely that the test organisms were exposed predominantly to the parent substance. The nominal test concentrations are supported by DOC analysis which showed that the total dissolved organic compounds in the test medium (parent substance plus all hydrolysis products) were well maintained throughout the exposure period. However, it is known that the parent substance undergoes hydrolysis in water at a moderate rate (conditions in the test were pH 7.0 - 7.7 in test media and temperature was 20-21ºC; half-life under these conditions is not known but a half-life of 36 h at pH 7 and 25ºC is available). The geometric mean concentration over time, for the two test concentrations between which the EC50 lies, can be estimated as 51 and 63 mg/L. The EC50 in terms of the parent substance can be estimated to lie within this range and an approximate value is likely to be around 57 mg/L. It is not appropriate to use this value directly in the chemical safety assessment, but it is useful for comparison of sensitivity with the available short-term aquatic results.

Above a concentration of approximately 1000 mg/L, oligomeric, cyclic or cross-linked condensation products could potentially form over time in the aqueous test media. No evidence of undissolved material is indicated in the study report for this test and in view that the maximum concentration was 100 mg/L of test substance (equivalent to 53 mg/L as silanol hydrolysis product) condensation phenomena are not expected to be a major issue. The effects observed in the test are therefore attributed to parent substance in the test system. Although the predicted limit of solubility has apparently been exceeded, undissolved material was not observed and significant physical fouling is not anticipated in this concentration range.