Registration Dossier

Administrative data

Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
26 March - 26 October 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study conducted under GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
Guideline:
other: European Commission Regulation (EC), EC No. 440/2008, Part C, PubNo. L142, C.1.
Deviations:
no
Guideline:
other: ISO International Standard 7346-2.
Deviations:
no
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:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): MTDID 7831
- Physical state: Waxy solid
- Analytical purity: 93.96 w%
- Storage condition of test material: At room temperature in the dark

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: 2 mL aliquots
- Sampling method: During the limit test single 2 mL samples were taken from the center of the test vessels from the test solution and control vessels. Samples were taken at the start of the test and after 72 hours from the freshly prepared solutions. Samples were also taken at the first renewal (24 hours) and at the end of the test from the 24 hour solutions.
- Sample storage conditions before analysis: Samples were stored in a freezer (≤ 15 °C) until analysis

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Fresh solutions were prepared daily. Loading rates of 6 mg/l were prepared for the first two days (inadvertently) and at 5 mg/L for the last two days. The pH of the test medium was not adjusted. Maximum solubility was achieved by a 30-minute treatment period with ultrasonic waves followed by 1 day of magnetic stirring. The resulting dispersions were left to settle for a maximum of 30 minutes after which the clear Water Accommodated Fraction (WAF) was collected by siphoning. The final test solutions were all clear and colourless.
- Controls: Test medium without test substance or other additives.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no visible undissolved test substance was observed at the test initiation.

Test organisms

Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
TEST ORGANISM
- Common name: Zebra-fish
- Strain: Zodiac
- Source: Wageningen University and Research Centre, The Netherlands.
- Length at study initiation: rangefinder, 3.1 ± 0.3 cm; limit test, 3.7 ± 0.2 cm
- Weight at study initiation: rangefinder, 0.44 ± 0.13 g; limit test, 0.92 ± 0.25 g
- Feeding during test: No

ACCLIMATION
- Acclimation period: At least 12 days after receipt by laboratory delivery.
- Acclimation conditions (same as test or not): unknown
- Type and amount of food: Pelleted fish food (Cyprico Crumble Excellent (300-500 um), Coppens International bv, Helmond, The Netherlands)
- Feeding frequency: Daily until 24 hours prior to the start of the test.
- Health during acclimation (any mortality observed): less than 5% mortality in the seven days prior to the test

Study design

Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Remarks on exposure duration:
daily renewal of test medium

Test conditions

Hardness:
180 mg/L as CaCO3
Test temperature:
21.1 - 21.7 °C
pH:
7.4 - 8.0
Dissolved oxygen:
7.3 - 9.3 mg/L
Nominal and measured concentrations:
Nominal: Water Accommodated Fraction at a loading rate of 5 and 6 mg/L
Mean measured: 0.32 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 40 L, all-glass container. Fill volume was 19 L.
- Aeration: None
- Renewal rate of test solution (frequency/flow rate): Daily
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.34 g fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO medium prepared using reverse-osmosis purified water (GEON Waterbehandeling, Berkel-Enschot, The Netherlands)
- Culture medium different from test medium: No
- Intervals of water quality measurement: Daily in all vessels with surviving fish, beginning at the start of the test (day 0)

OTHER TEST CONDITIONS
- Adjustment of pH: None
- Photoperiod: 16 hours light; 8 hours dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortality and/or severely distressed fish at 2.5, 24, 48, 72, and 96 hours.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: One concentration tested
- Justification for using less concentrations than requested by guideline: Limit test
- Range finding study: Yes
- Test concentrations: WAFs based on 1.0 mg/L, 10 mg/L, and 100 mg/L loading rates and ten fold dilution of 1.0 mg/L WAF to prepare a 0.1 mg/L group.
- Results used to determine the conditions for the definitive study: Yes. No mortality or clinical effects were observed at any of the test concentrations. The 100 mg/L loading rate had a measured concentration had an initial concentration (0h) of 2.48 mg/L and an end concentration (96h) of 0.38 mg/L. In the final test, a loading rate of 5 or 6 mg/L was used and the test solution was renewed daily.
Reference substance (positive control):
no

Results and discussion

Effect concentrations
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
> 0.32 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: No mortality or clinical effects were observed during the test period. WAF loading rate: 5 or 6 mg/L
Details on results:
Measured concentrations are shown in Table 1. Mortality results are shown in Table 2.
- Behavioural abnormalities: None noted
- Observations on body length and weight: None noted
- Other biological observations: No mortality or clinical effects were observed
- Mortality of control: 0
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: C4 Acrylate has a limited water solubility. No undissolved test substance was visible in final test solutions.
- Effect concentrations exceeding solubility of substance in test medium: Yes. WAFs were also prepared at a loading rate of 5 mg/L in the study with Daphnia magna. The resulting mean measured concentration was considerably higher (1.4 mg/L) in the Daphnia WAFs despite the fact that the same medium was used as the fish test. This difference was likely caused by the difference in the volume of the medium used for the preparation of test solutions (1 vs 25 liters) and the lower stirring force used to prepare the fish test solutions. It is believed by the testing laboratory that the maximum solubility in this medium was reached in the zebra-fish study and that likely the higher concentrations in the Daphnia study were above the medium solubility although no undissolved test substance was visible.

Any other information on results incl. tables

Table 1. Concentrations of test substance in the test medium

TIme of sampling

[hours]

                              Concentration

Nominal

C4 Acrylate

(mg/I)

Measured

C4 Acrylate

(mg/l)

Percent of initial concentration

0

0

n.d.

[fresh]

6.0

0.395

24

0

n.d.

[old]

6.0

0.224

57%

72

0

n.d.

[fresh]

5.0

0.418

96

0

n.d.

[old]

5.0

0.269

64%

n.d. Not detected

Table 2. Acute mortality of zebra-fish in the final test

 

Measured Concentration

C4 Acrylate

(mg/L)

Initial No. of fish

Cumulative mortality

Total mortality (%)

2.5 hr

24 hr

48 hr

72 hr

96 hr

Control

7

0

0

0

0

0

0

0.32

7

0

0

0

0

0

0

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
No control mortality, DOC > 60% of saturation
Conclusions:
C4 acrylate is not acutely toxic to zebra-fish at the limit of water solubility.
Executive summary:

The toxicity of C4 acrylate to the zebra-fish, Brachydanio rerio, was assessed in a 96-hour toxicity test conducted according to the OECD 203 (1992) method.

The 96-hour LC50 of C4 acrylate to Brachydanio rerio was > 0.32 mg/L. C4 acrylate induced no visible or lethal effects in zebra-fish at a WAF prepared at a loading rate of 5 or 6 mg/L, which corresponded to an average measured exposure concentration of 0.32 mg/L.

The study was performed in accordance with internationally-accepted test guidelines and Good Laboratory Practice (GLP) standards. Therefore, this study is reliable without restrictions and the results are suitable for purposes of Risk Assessment, Classification & Labeling, and PBT Analysis.

Result Synopsis

Test type: Semi Static (based on data obtained using OECD 203 methodology)

96-hr LC50 > 0.32 mg/L