2-[10-(acetyloxy)-9a,11a-dimethyl-7-oxo-3H,3aH,3bH,4H,5H,7H,9aH,9bH,10H,11H,11aH-cyclopenta[a]phenanthren-1-yl]-2-oxoethyl acetate

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:

26 January 2018 till 07 Feb 2018

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)

Version / remarks:

2004

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult items and mixtures, OECD series on testing and assessment number 23

Version / remarks:

December 14, 2000

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2-[10-(acetyloxy)-9a,11a-dimethyl-7-oxo-3H,3aH,3bH,4H,5H,7H,9aH,9bH,10H,11H,11aH-cyclopenta[a]phenanthren-1-yl]-2-oxoethyl acetate

EC Number:

607-277-2

Cas Number:

23825-05-0

Molecular formula:

C25H30O6

IUPAC Name:

2-[10-(acetyloxy)-9a,11a-dimethyl-7-oxo-3H,3aH,3bH,4H,5H,7H,9aH,9bH,10H,11H,11aH-cyclopenta[a]phenanthren-1-yl]-2-oxoethyl acetate

Test material form:

solid: particulate/powder

Details on test material:

- Appearance: Off-white powder
- Storage condition of test material: At room temperature

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Samples for possible analysis were taken from all test concentrations and the control at t=0 and t=48 hours.

Test solutions

Vehicle:

no

Details on test solutions:

The batch of PREDIAC-Z tested was not completely soluble in test medium at the loading rate initially prepared. Preparation of test solutions started with a loading rate of 100 mg/L applying a three-day period of magnetic stirring to ensure maximum dissolution of the test item in medium. Thereafter, the aqueous Saturated Solution (SS) was collected by filtration through a 0.45 µm membrane filter and used as the highest test concentration. Lower test concentrations (1% and 10% of the SS) were prepared by subsequent dilutions of the SS in test medium. All test solutions were clear and colorless at the end of the preparation procedure. Hence tested concentrations were PREDIAC-Z Solutions containing 1.0, 10 and 100% of a Saturated Solution prepared at a loading rate of 100 mg/L. As controls test medium without test item or other additives was used.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

Species: Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by a cyclical parthenogenesis under specified breeding conditions.
Source: In-house laboratory culture with a known history.
Validity of batch: Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20%, presence of males, ephippia or discoloured animals and there was no delay in the production of the first brood.
Characteristics: For the test selection of young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old.
Feeding during test: none.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

180 mg/L expressed as CaCO3

Test temperature:

20-21°C

pH:

Range at start and end in control and the test concentration of 1.7 mg/L: 7.9-8.1

Dissolved oxygen:

Range at start and end in control and the test concentration of 1.7 mg/L: 8.6-9.2 mg/L

Nominal and measured concentrations:

Nominal concentrations were PREDIAC-Z Solutions containing 1.0, 10 and 100% of a Saturated Solution (SS) prepared at a loading rate of 100 mg/L. Samples taken from the control and the highest test concentration were analysed. At the start of the test, a concentration of 1.67 mg/L (rounded to 2 significant figures: 1.7 mg/L) was measured in the undiluted SS. During the exposure period, this concentration remained stable, i.e. was 90% of the initially measured concentration at the end of the test (measured concentration 1.51 mg/L). Based on these results, the effect parameters were based on initially measured concentrations (i.e. 1.7 mg/L).

Details on test conditions:

Test vessels: 60 mL, all-glass;
Number of daphnids: 20 each in the control and the undiluted SS, 10 per intermediate test concentration;
Loading: 5 per vessel containing 50 mL of test solution;
Light: 16 hours photoperiod daily;
Feeding: No feeding;
Aeration: No aeration of the test solutions;
Introduction of daphnids: Within 15 minutes after preparation of the test solutions.
Immobility (including mortality): Recorded at 24 hours and at 48 hours.
Dissolved oxygen & pH: Measured at the beginning of the test, for the highest concentration and the control.
Temperature of medium: Measured continuously in a temperature control vessel, beginning at the start of the test.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7)

Results and discussion

Effect concentrations

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 1.7 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Remarks on result:

other: See remark

Remarks:

The measured concentration of 1.7 mg/L represents the maximum soluble concentration of the test item in test medium at a loading rate of 100 mg/L.

Details on results:

In the control and in the PREDIAC-Z Solutions containing 1.0, 10 and 100% of a Saturated Solution prepared at a loading rate of 100 mg/L, 0% immobility was recorded after 24 and 48 hours. This etablishes the 48-h EC50 as >1.7 mg/L based on measured concentrations.

Results with reference substance (positive control):

Potassium dichromate (K2Cr2O7) was tested in a separate test on 02-04 January 2018. The 24h-EC50 was 0.87 mg/L with a 95% confidence interval between 0.80 and 0.95 mg/L. This is within the expected range according to OECD 202 (2012): 06-2.1 mg/L, confirming the sensitivity of the test organisms.

Reported statistics and error estimates:

Not applicable.

Any other information on results incl. tables

Small responses at the retention time of the test item were detected in the chromatograms of the blank QC samples. It was considered to derive from carry-over in the analytical system since similar responses were found in the analytical blanks. The maximum contribution to the QC samples at the lowest concentration level (0.001 mg/L) was 15%. It was considered that it has no significant impact on the results of the study.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

1. In the control: 0% immobility, no signs of disease or stress. 2. The oxygen concentration at the end of the test was ≥3 mg/L in control and test vessels.

Conclusions:

The 48h-EC50 for Daphnia magna exposed to PREDIAC-Z was beyond the tested range, i.e. exceeded a measured concentration of 1.7 mg/L being considered to be the maximum soluble concentration of the test item in test medium at a loading rate of 100 mg/L.

Executive summary:

The objective of the study was to evaluate PREDIAC-Z for its ability to generate acute toxic effects on the mobility of Daphnia magna during an exposure period of 48 hours and, if possible, to determine the EC₅₀ at 24 and 48 hours of exposure.

The study procedures described in this report were based on the OECD guideline No. 202, 2004. In addition, procedures were based on the test methods described in the OECD series on testing and assessment number 23, 2000.

A Saturated Solution (SS) was prepared at a loading rate of 100 mg/L and used as the highest concentration. Lower concentrations (1.0 and 10% of the SS) were prepared by diluting the highest concentration in test medium.

A combined limit/range-finding test was performed. Twenty daphnids per group (5 per replicate, quadruplicate) were exposed to an untreated control and to an undiluted SS prepared at a loading rate of 100 mg/L, in a limit test. In addition ten daphnids per group (5 per replicate, duplicate) were exposed to solutions containing 1.0 and 10% of the SS prepared at a loading rate of 100 mg/L in the combined range-finding test. The total exposure period was 48 hours and samples for analytical confirmation of exposure concentrations were taken at the start and at the end of the test.

No immobility was observed in the control or at any of the test concentrations throughout the exposure period.

Samples taken from the control and the highest test concentration were analysed. At the start of the test, a concentration of 1.7 mg/L was measured in the undiluted SS. During the exposure period, this concentration remained stable, i.e. was 90% of the initially measured concentration at the end of the test.

The study met the acceptability criteria prescribed by the study plan and was considered valid.

In conclusion, the 48h-EC₅₀ for Daphnia magna exposed to PREDIAC-Z was beyond the tested range, i.e. exceeded a measured concentration of 1.7 mg/L being considered to be the maximum soluble concentration of the test item in test medium at a loading rate of 100 mg/L.