Registration Dossier

Ecotoxicological information

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:
23 March - 10 April 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Reliability 1 is assigned because the study was conducted according to OECD TG 202 in compliance with GLP, without deviations that influence the quality of the results.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
(2004)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
(2008)
Deviations:
no
Qualifier:
according to
Guideline:
other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23, December 14, 2000
Deviations:
not applicable
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: Liquid
Details on test material:
- Name of test material in report: IFF 09-0035
- Physical state: Clear liquid
- Storage condition of test material: At room temperature protected from light

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Samples for possible analysis were taken from all test concentrations and the control according to the schedule below.

Frequency: at t=0 h and t=48 h
Volume: 3.0 mL from the approximate centre of the test vessels
Storage: Samples were stored in a freezer until analysis

At the end of the exposure period, the replicates were pooled at each concentration before sampling.

Additionally, reserve samples of 3.0 mL were taken for possible analysis (samples were taken from freshly prepared solutions at the start of the test and from one replicate of each concentration at the end of the test). If not used, these samples were stored in a freezer for a maximum of three months after delivery of the draft report, pending on the decision of the sponsor for additional analysis.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

The batch of IFF 09-0035 tested was a clear liquid and the substance was not completely soluble in test medium at the concentrations tested. No correction was made for the purity/composition of the test substance.

Preparation of test solutions started with a loading rate of 100 mg/L applying 19.5 hours of magnetic stirring to ensure reaching maximum dissolution in test medium. The resulting emulsion was subsequently left to settle for 3 hours resulting in a clear and colourless solution with a floating layer of undissolved test item. The Water Accomodated Fraction (WAF) was then collected by means of siphoning. The lower test concentrations were prepared by subsequent dilutions of the WAF in test medium. The final test solutions were all clear and colourless.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Dapnia magna
- Source: In-house laboratory culture with a known history, at least third generation, obtained by acyclical parthenogenesis under specified breeding conditions.
- Age at study initiation: young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old.
- Method of breeding: start of each batch with newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium M7 in an all-glass culture vessel. Renewal of the cultures: after 7 days of cultivation half of the medium twice a week.
- Feeding during test: no

ACCLIMATION
- Acclimation period: no

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
none

Test conditions

Hardness:
180 mg CaCO3/L
Test temperature:
19.5 -19.9°C
pH:
7.9 - 8.3
Dissolved oxygen:
9.2 - 9.9 mg O2/L
Nominal and measured concentrations:
Based on the results of the range-finding test the following test concentrations were assigned to the definitive test:
Nominal test concentrations: 4.6, 10, 22, 46 and 100% of a WSF prepared at a loading rate of 100 mg/L
Mean measured test concentrations: 1.6, 3.3, 8.8, 19 and 33 mg/L.
Details on test conditions:
TEST SYSTEM
- Test vessel: 100 mL, all-glass, closed airtight
- Aeration: None
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 5 daphnids per vessel containing 100 mL test solution

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Adjusted ISO medium
- Culture medium different from test medium: Yes, culture medium had extra trace elements, macro nutrients and vitamins added as specified in Medium M7.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16h

EFFECT PARAMETERS MEASURED: mobility at 24 and 48 hours ; pH and dissolved oxygen at the beginning and at the end of the test. Temperature continuously.
Reference substance (positive control):
yes
Remarks:
potassium dichromate

Results and discussion

Effect concentrations
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
5.4 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% CI: 3.3 - 8.8 mg/L
Details on results:
No effects were observed in the control and the two lowest concentrations, whereas at the three highest concentrations all daphnids were affected at the end of the exposure period (see Table 1 below).

Analytical results:
Measured concentrations are presented in Table 2 below.

Individual pH, temperature and dissolved oxygen values remained within acceptable limits throughout the duration of the study.
Results with reference substance (positive control):
48h-EC50: 0.32 mg/L (95% confidence interval between 0.29 and 0.37 mg/L)

Any other information on results incl. tables

Table 1: Number of introduced daphnids and incidence of immobility in the final test

Time (h)

Replicate

Concentration IFF 09-0035 (mg/L)

Control

1.6

3.3

8.8

19

33

0

A

5

5

5

5

5

5

B

5

5

5

5

5

5

C

5

5

5

5

5

5

D

5

5

5

5

5

5

Total introduced

20

20

20

20

20

20

24

A

0

0

0

1

4

5

B

0

0

0

0

4

5

C

0

0

0

1

5

5

D

0

0

0

0

5

5

Total immobilised

0

0

0

2

18

20

Effect %

0

0

0

10

90

100

 

 

 

 

 

 

48

A

0

0

0

5

5

5

B

0

0

0

5

5

5

C

0

0

0

5

5

5

D

0

0

0

5

5

5

Total immobilised

0

0

0

20

20

20

Effect %

0

0

0

100

100

100

Table 2            Average exposure concentration versus nominal concentration

Test group

 IFF 09-0035 (%WSF)

Measured concentrations (mg/l)

Average exposure (mg/l)

t=0h

t=48h

4.6

1.74

1.46

1.6

10

3.35

3.26

3.3

22

9.49

8.10

8.8

46

20.0

17.8

19

100

31.8

33.4

33

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
See Overall remarks
Conclusions:
The 48h-EC50 value to Daphnia magna is 5.4 mg/L based on mean measured concentrations.
Executive summary:

A study was performed to assess the acute toxicity of the substance to Daphnia magna. The study was conducted in accordance with OECD Guideline for Testing of Chemicals No. 202.

Based on the results of a preliminary range-finding test, Water Soluble Fractions (WSFs) were prepared with individual loading rates of 1.6, 3.3, 8.8, 19 and 33 mg/L.

20 daphnids less than 24 hours old) were exposed for 48 hours in the dark to a control group and each prepared concentration. In view of the volatile properties of the test substance, the test vessels were closed air-tight. After 24 and 48 hours, any incidence of immobility was recorded in each vessel.

Samples taken from all concentrations were analysed with a validated GC-FID method.

Analysis of the samples taken at the start of the final test showed measured concentrations of 1.74, 3.35, 9.49, 20.0 and 31.8 mg/L in the test groups representing 4.6, 10, 22, 46 and 100% WAF, respectively. Measured concentrations after 48 hours were at 84 to 105% relative to the start concentrations.Based on these results, the average exposure concentrations were 1.6, 3.3, 8.8, 19 and 33 mg/L.

The 48h-EC50 value to Daphnia magna was 5.4 mg/L based on mean measured concentrations.