1-Ethoxy-2,3-difluor-4-(trans-4-propylcyclohexyl)-benzene

Long-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Jul 24, 2013-Mar 04, 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was performed according to GLP and the methods applied are fully compliant with OECD TG 211.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

1-methoxy-4-(trans-4-propylcyclohexyl)-benzene

EC Number:

617-272-7

Cas Number:

81936-32-5

Molecular formula:

C16H24O

IUPAC Name:

1-methoxy-4-(trans-4-propylcyclohexyl)-benzene

Test material form:

solid: crystalline

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

The saturated solution of the test item, all further dilution levels and the control were analysed via GC-FID at the start of the exposure intervals (0 h) on days 0, 9, 14, 19 and at the end of the exposure intervals (48 or 72 h) on days 2, 12, 16, 21.For the analyses at the end of the exposure intervals, separate replicates without daphnids and algal food were prepared and stored under test conditions until sampling.

Test solutions

Vehicle:

no

Details on test solutions:

The test item is a white, crystalline solid and is insoluble in water. The preliminary testing has shown a high variation of analytical and biological results. Producing of constant and equal concentrations of the saturated solution of the insoluble test item was difficult (e.g. complete removal of undissolved test item). This results in varying effects concerning the observed adult mortality. Therefore, it was considered, that a full dose-response test should be carried out with a saturated solution of the test item and four further dilution levels of 1:2- 1:4 - 1:8 - 1:16. The saturated solution was prepared according to the OECD guidance document no. 23 (2000).

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

Test species Daphnia magna STRAUS (Clone 5)Reason for the selection of the test speciesDaphnia magna is the preferred species in accordance with the test guideline and is bred at the test facility. Origin lnstitut fur Wasser-, Boden- und Lufthygiene (WaBolu)Breeder DR.U. NOACK-LABORATORIEN,Kathe-Paulus-Str. 1, 31157 Sarstedt, Germany

Test conditions

Hardness:

> 140 mg/L

Test temperature:

20 ± 2°C

pH:

7-8

Dissolved oxygen:

4,2 - 8,6 mg/L

Nominal and measured concentrations:

13.1 µg/L (nominal)

Details on test conditions:

Culture In glass vessels (2- 3 L capacity) with approximately 1.8 L culture mediumTemperature 20 ± 2°C Environmentin an incubatorPhotoperiod16 h illumination Light Intensity max. 20 µE m-2 s-1Culture medium Elendt M4, according to ELENDT ( 1990), modified to a total hardness of 160 to 180 mg CaC03/LCulture feeding 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> 10^6 cells/ml

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

> 13.1 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

reproduction

Remarks on result:

other: NOEC = solubility limit

Effect concentrations 2

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

> 13.1 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality

Remarks on result:

other: NOEC = solubility limit

Details on results:

The coefficient of variation of the number of living offspring produced per parental daphnid was 11 % in the control. At the geometric mean measured concentrations of 0.763 to 13.1 µg/L, the coefficient of variation was in the range of 10 to 15 %. Generally, in a well-run test, the coefficient of variation around the mean number of living offspring per parent animal should be larger or equal to 25 %.The intrinsic rates of natural increase of the surviving parental daphnids accounting for generation time and number of offspring were used for calculation of population growth and maintenance. No biologically significant adult mortality (20 %) was observed during the test period of 21 days. In the geometric mean measured concentrations of 0.763 to 5.28 µg/L and in the control, all parental daphnids survived until the end of the test after 21 days. At the geometric mean measured concentration of 13.1 µg/L (saturated solution), one of ten daphnids did not survive until the end of the test after 21 days. The first brood was released between days 8 and 11 by all parental daphnids of the control and the geometric mean measured concentrations of 0.763 to 13.1 µg/L.Four broods were produced by all parental surviving daphnids of the geometric mean measured concentrations of 0.763 to 13.1 µg/L and the control.No stillborn juveniles or aborted eggs were observed in the control during the exposure period of 21 days. Related to the total number of produced juveniles (dead + alive) the percentage of dead juveniles was smaller or equal than 2 % in the geometric mean measured concentrations of 0. 763 to 13.1 µg/L. The mean values of the body length of the surviving parental daphnids in the geometric mean measured concentrations of 0.763 to 13.1 µg/L were determined to be in the range of 5.08 to 5.20 mm per daphnid and 5.13 mm per daphnid in the control group.The mean dry body weight of the surviving parental daphnids was determined to be in the range of 0.79 to 0.91 mg per daphnid in the geometric mean measured concentrations of 0.763 to 13.1 µg/L and 0.63 mg per daphnid in the control.No males or ephippia (winter eggs) were observed in the control or in the geometric mean measured concentrations of 0.763 to 13.1 µg/L.Water quality parameters (i.e. pH-value, dissolved oxygen concentration, total water hardness and temperature) were within the acceptable limits.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

No significant effects on the reproductive output or adult mortality were observed in this study. The No Observed Effect Concentration (NOEC) after 21 days was assessed at 13.1 µg/L, which is the maximum solubility level of the test item in the dilution water.

Executive summary:

Study Design

This study was performed according to GLP and the methods applied are fully compliant with OECD TG 211. In the assay semi static conditions have been applied. The concentrations of the test item were analytically verified via GC-FID of samples taken at the start of the exposure intervals on days 0, 9, 14, 19 (0 h) and at the end of the exposure intervals on days 2, 12, 16 and 21 (48 or 72 h) of the saturated solution, all tested dilution levels and the control.

Results

The measured concentrations of the test item at the start of the exposure intervals were in the range of < LOQ to 39.2 µg/L. At the end of the exposure intervals (48 or 72 h), the measured concentrations of the test item were in the range of < LOQ to 14.7 µg/L.
The measured values below 70 % of the LOQ (1.00 µg/L) were taken into account with half of the LOQ (= 0.500 µg/L). The geometric mean measured concentration  of the test item were calculated to be: 0. 763 - 1.23 - 2.40 - 5.28 - 13.1 µg/L and used for the assessment of the effect levels.

The average number of living juveniles per surviving parental daphnid at the end of the test after 21 days was 89 in the control group. The reproductive output was comparable to the control at the geometric mean measured test item concentrations of 0.763 to 13.1  µg/L (One Analysis of Variance, DUNNETT's method, p = 0.05). An EC10- or EC50 could not be calculated, because no effects were observed.

The coefficient of variation of the number of living offspring produced per parental daphnid was 11 % in the control. At the geometric mean measured concentrations of 0.763 to 13.1 µg/L, the coefficient of variation was in the range of 10 to 15 %. Generally, in a well-run test, the coefficient of variation around the mean number of living offspring per parent animal should be larger or equal to 25 %.
The intrinsic rates of natural increase of the surviving parental daphnids accounting for generation time and number of offspring were used for calculation of population growth and maintenance.

No biologically significant adult mortality (20 %) was observed during the test period of 21 days. In the geometric mean measured concentrations of 0.763 to 5.28 µg/L and in the control, all parental daphnids survived until the end of the test after 21 days. At the geometric mean measured concentration of 13.1 µg/L (saturated solution), one of ten daphnids did not survive until the end of the test after 21 days.
The first brood was released between days 8 and 11 by all parental daphnids of the control and the geometric mean measured concentrations of 0.763 to 13.1 µg/L.

Four broods were produced by all parental surviving daphnids of the geometric mean measured concentrations of 0.763 to 13.1 µg/L and the control.
No stillborn juveniles or aborted eggs were observed in the control during the exposure period of 21 days. Related to the total number of produced juveniles (dead + alive) the percentage of dead juveniles was smaller or equal than 2 % in the geometric mean measured concentrations of 0. 763 to 13.1 µg/L.
The mean values of the body length of the surviving parental daphnids in the geometric mean measured concentrations of 0.763 to 13.1 µg/L were determined to be in the range of 5.08 to 5.20 mm per daphnid and 5.13 mm per daphnid in the control group.

The mean dry body weight of the surviving parental daphnids was determined to be in the range of 0.79 to 0.91 mg per daphnid in the geometric mean measured concentrations of 0.763 to 13.1 µg/L and 0.63 mg per daphnid in the control.

No males or ephippia (winter eggs) were observed in the control or in the geometric mean measured concentrations of 0.763 to 13.1 µg/L.
Water quality parameters (i.e. pH-value, dissolved oxygen concentration, total water hardness and temperature) were within the acceptable limits.

Conclusion

No significant effects on the reproductive output or adult mortality were observed in this study. The No Observed Effect Concentration (NOEC) after 21 days was assessed at 13.1 µg/L, which is the maximum solubility level of the test item in the dilution water.