Reaction mass of bis(2-ethylhexyl) adipate and cyclohexyl 2-ethylhexyl adipate and dicyclohexyl adipate

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

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 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: #30869
- Expiration date of the lot/batch: 26 Nov 2019
- Purity: 99.8 area-% (GC)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test substance is a poorly water-soluble mixture; therefore, a water accommodated fraction (WAF) was prepared using a liquid-liquid saturator technique following general guidance provided in OECD 23. A glass aspirator bottle (2-L) with a bottom side-outlet attached to a stopcock is filled with 2-L test media. The test substance was homogenized by shaking the container before use. The test solution was prepared separately (differential loading) by directly adding test substance to test medium. 20.4 μL test substance (considering density = 0.9802 g/cm3) were pipetted carefully with a positive displacement pipette (Gilson Microman M10) on the water surface. The bottle was closed, and the solution stirred slowly at approx. 100 rpm on a magnetic stir plate for approx. 2 days. The undissolved test substance remained on the surface and the required volume of the saturated solution was removed from the bottom of the bottle. The aqueous fraction was inspected visually for the presence of any undissolved test substance by observing the scattering of a laser light through the test solution (Tyndall effect) and turned out negative. The aqueous fraction of the test solution was considered the water accommodated fraction (WAF) in test media.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): The test solutions were visibly colorless and clear after preparation.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Age at study initiation (mean and range, SD): < 24h
- Method of breeding: Daphnia brood stock are kept in mass cultures consisting of approx. 20–30 individuals for a maximum of 4 weeks. All individuals in the mass culture originate from a single female. The Daphnia are cultured under the identical conditions as the test including test media (Elendt M4), water quality, temperature (20 ±1°C), and diet.
- Source: inhouse breeding; clone supplied by the Institut National de Recherche Chimique Appliquée, France, in 1978.
- Age of parental stock (mean and range, SD):
- Feeding during test
- Food type: live green algae Desmodesmus subspicatus
- Amount: 0.1 mg TOC/parent animal and day: days 0-3; 0.12 mg TOC/parent animal and day: days 4-5; 0.14 mg TOC/parent animal and day: days 6-7; 0.2 mg TIOC/animal and day: days 8 -21
- Frequency: daily

ACCLIMATION
- Acclimation period: None, since the Daphnia are cultured under the identical conditions as the test including test media (Elendt M4), water quality, temperature (20 ±1°C), and diet.
- Acclimation conditions (same as test or not): same
- Health during acclimation (any mortality observed): Only young Daphnia from healthy cultures without signs of stress are used for testing

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

21 d

Hardness:

2.43 - 2.46 mmol/L

Test temperature:

19 - 20 °C

pH:

7.8 - 8.0

Dissolved oxygen:

8.5 - 9.0 mg/L

Salinity:

-

Conductivity:

-

Nominal and measured concentrations:

Nominal: 10 mg/L as loading rate WSF

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL glass beakers covered with glass Petri plates and 50 mL test solution
- Aeration: none (medium aerated for approx. 24h prior to use)
- Renewal rate of test solution (frequency/flow rate): daily
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10
- Biomass loading rate: 0.02 animals/mL

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4
- Total organic carbon: < 2 mg/L
- Ca/mg ratio: 4:1
- Conductivity: 550–650 μS/cm
- Culture medium different from test medium: no
- Intervals of water quality measurement: Dissolved oxygen, pH and temperature were measured in the old and in the freshly prepared test solution in replicate 1 of each test group for one interval per week. In addition, temperature was measured continuously during the whole exposure period in a separate vessel filled with water proximal to the test vessels. Hardness was measured in the freshly prepared test solution in an additional replicate of each concentration for one interval per week and in the old test solution from the combined replicates of each test group for one interval per week.

OTHER TEST CONDITIONS
- Photoperiod: 16h light: 8h dark
- Light intensity: 676–711 lux at a wave length of 400–750 nm

EFFECT PARAMETERS MEASURED: Mortality, reproduction (as number of living young), % aborted eggs, time to first brood and growth as length in mm were recorded throughout the test period.

Reference substance (positive control):

yes

Remarks:

sodium chloride

Key result

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

>= 10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

reproduction

Remarks on result:

other:

Remarks:

mean living young per surviving adult

Details on results:

- Behavioural abnormalities: no
- Observations on body length and weight: no differences between treatment and control
- Mortality of control: One mortality was observed in the control and 10 mg/L test groups (10% mortality). This minor level of effect is within the accepted tolerance for control performance (OECD 211) and since no more mortality was observed the following days the effect was thus not toxicologically relevant. These accidental mortalities were excluded from the calculation of % Mortality.

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: EC50(48h) = 4.9 mg/L
- Other: This result is within the range of 3.88–7.22 g/L, which represents ±2 standard deviations from the published EC50(48 h) of 5.55 g/L and indicates that the culture of Daphnia magna used in this study is responding normally to toxic stress.

Validity criteria fulfilled:

yes

Conclusions:

There was no significant effect on Daphnia reproduction in the treatment group with loading rate 10 mg/L.

Description of key information

NOELr(21d) >= 10 mg/L (loading rate WSF, nominal) for reproductive output of Daphnia magna (OECD 211)

Additional information

One study investigating the long-term toxicity of the test substance to aquatic invertebrates is available. The key study was performed according to GLP and OECD guideline 211 using Daphnia magna as test organism (BASF 2019). The test substance is a poorly water-soluble mixture; therefore, a water soluble fraction (WSF) of a 10 mg/L loading rate was prepared using a liquid-liquid saturator technique following general guidance provided in OECD 23. Concentration control analyses were not performed, because a specific method for analyses in the required concentration range is not available. No inhibition of reproductive output was observed in the treatment in comparison to the control. Thus, the 72h-NOELr is determined to be >= 10 mg/L as loading rate WSF based on nominal concentrations. Based on this result, the test substance is considered to be not harmful to aquatic invertebrates at long-term exposure up to the saturation limit in water.