3,9-dimethyl-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane 3,9-dioxide

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2016-08-30 to 2016-09-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

3,9-dimethyl-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane 3,9-dioxide

EC Number:

221-088-9

EC Name:

3,9-dimethyl-2,4,8,10-tetraoxa-3,9-diphosphaspiro[5.5]undecane 3,9-dioxide

Cas Number:

3001-98-7

Molecular formula:

C7H14O6P2

IUPAC Name:

3,9-dimethyl-2,4,8,10-tetraoxa-3λ⁵,9λ⁵-diphosphaspiro[5.5]undecane-3,9-dione

Test material form:

solid: particulate/powder

Details on test material:

AFLAMMIT® PCO 962

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: At the start of the exposure intervals (0 and 24 h), sampling was carried out after preparation of the limit concentration. At the end of the exposure intervals (24 and 48 h), samples were taken directly from the test vessels.
- Sample storage conditions before analysis: All original samples were stored at room temperature until preparation. Prepared samples were stored in an autosampler at room temperature until analysis.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The limit concentration (100 mg/L of the test item were weighed out) was freshly prepared with dilution water before the start of the exposure (0 hours) and the water renewal (24 hours). The limit concentration was mixed thoroughly by manual agitation until the test item was completely dissolved.
- Controls: Dilution water without test item incubated under the same conditions as the limit test item concentration
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): no

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Strain/clone: Clone 5
- Source: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), D-14195 Berlin, Germany
- Age of parental stock (mean and range, SD): not stated
- Age at study initiation (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 the following 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: The daphnids were not fed during the study.
- Food type: The culture daphnids were fed with a mix of unicellular green algae with an algae cell density of > 10^6 cells/ml.
- Amount: ad libitum
- Frequency: at least 5 times per week
- Method of breeding: In glass vessels (2 - 3 L capacity) with approximately 1.8 L culture medium, at 20 ± 2°C, in an incubator, 16 h illumination, light intensity of max. 1500 lx

ACCLIMATION
Acclimatization was not necessary, because the dilution water was equivalent to the culture medium.

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

not applicable

Test conditions

Hardness:

176-178 mg CaCO3/L

Test temperature:

20.5-20.7°C

pH:

7.50-7.56

Dissolved oxygen:

8.02-8.28 mg/L

Salinity:

not applicable

Conductivity:

476-491 mg/L

Nominal and measured concentrations:

100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers (4 cm ID x 7 cm H), 50 mL capacity, loosely covered
- Material, size, headspace, fill volume: 20 mL
- Aeration: no
- No. of organisms per vessel: 5 daphnids
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4, according to OECD 202, Annex 3 (2004), modified to a total hardness of 160 to 180 mg CaCO3/L, is used.
- Culture medium different from test medium: no
- Intervals of water quality measurement: Prior to the start of the exposure (0 h) and the water renewal (24 h), the water quality parameters (i.e. pH value, dissolved oxygen concentration, temperature, conductivity, alkalinity and total hardness) of the dilution water were measured.

OTHER TEST CONDITIONS
- Photoperiod: 16/8 h light/dark cycle
- Light intensity: Diffuse light, light intensity of max. 1500 lx

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Immobilization was determined in all replicates after 24 and 48 h.

VEHICLE CONTROL PERFORMED: no

RANGE-FINDING STUDY
- Spacing factor for test concentrations: limit test
- Test concentrations: 0.1, 1.0, 10.0, 100 mg/L
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Key result

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: Immobilization was determined in all groups after 24 and 48 h. One animal was found to be immobilized in the limit test concentration..
- Observations on body length and weight: not determined
- Other biological observations:no
- Mortality of control: 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 48 h test period
- Other adverse effects control: no
- Abnormal responses: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: The EC50-value of the reference item is also within the recommended range of 0.6 - 2.4 mg/L 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.
- ECx: EC50: 1.77 mg/L (Cl: 1.00 - 4.00 mg/L)

Reported statistics and error estimates:

An EC50-value was calculated for the reference item by sigmoidal dose-response regression with the software GraphPad Prism5. The respective 95% confidence limits were estimated directly from the observation data, since only one partial effect was observed. The highest reference item concentration resulting in 0% immobilization and the lowest reference item concentration resulting in 100% immobilization were used as 95% confidence limits.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

In the limit concentration 100 mg/L of the test item AFLAMMIT® PCO 962, no effects on Daphnia magna were observed.