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Ecotoxicological information

Long-term toxicity to fish

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Administrative data

Endpoint:
fish early-life stage toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Mar. 24, 2011 to May. 18, 2011
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: OECD guideline test with GLP compliance, but without GLP certificate. Furthermore prior to initiating the definitive test, a preliminary test was performed without GLP compliance.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 210 (Fish, Early-Life Stage Toxicity Test)
Deviations:
yes
Remarks:
As the stock solution in the pre-test was enough for the definitive test and the concentration was verified by HPLC. Therefore, the stock solution for the definitive test adopted stock solution prepared in pre-test.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Type:
Constituent
Test material form:
other: liquid
Details on test material:
Internal No. 0002T0001
Batch No. 20091104

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
During the test, the concentrations of the test substance were determined once every 6 days both in fresh and old solution. Approximate 200 mL samples of freshly prepared solutions were taken from each test concentration and solvent control. After 48 h renewal, approximate 200 mL (3 x approximate 70 mL per concentration and solvent control) of old solutions were taken from the center of test vessels. When samples were not analyzed immediately, they were frozen (≤ -20 ℃). During the test, dissolved oxygen, pH, total hardness and temperature were measured in all test vessels.

Test solutions

Vehicle:
yes
Details on test solutions:
The preliminary test was conducted at the concentration of 0.4 mg/L. Based on the results of the pre-test (Non-GLP state), test concentrations of
0.4 mg/L were selected.
As the stock solution in the pre-test was enough for the definitive test and the concentration was verified by HPLC (Nominal concentration was 10.0g/L; measured concentration was 8.76 g/L).Therefore, the stock solution prepared in the preliminary test was used in the definitive test. Detailed preparation was as followed: 250.5 mg test substance was fully dissolved and made up to 25 mL with dimethylformamide (DMF) to obtain the stock solution of 10 g/L. 120 µL stock solution was taken and made up to a total volume of 3000 mL with dilution water to obtain test solution. The solution was equally transferred into three test vessels. If the concentration analysis was needed, 128 µL stock solution was taken and made up to a total volume of 3200 mL with dilution water.

Blank control: Dilution water only
Solvent control: Dilution water with 0.04 mL /L DMF

Test organisms

Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
Zebra-fish (Brachydanio rerio) aged about 6 months purchased from a commercial supplier was used as the brood fish. At the start of the test, fertilized eggs produced by the brood fish were exposed to the test solutions. The Water used for acclimation was the same as that used in the test. For spawning, the male and female brood fish at the ratio of 1: 2 were transferred to fish tank with scoop net, taking care to minimize possible stress due to handling.

Study design

Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
30 d
Post exposure observation period:
no data

Test conditions

Hardness:
114-155 mg/L (CaCO3)
Test temperature:
23.2-24.1°C
pH:
7.54-8.17
Dissolved oxygen:
82.9-99.2 %
Salinity:
no data
Nominal and measured concentrations:
nominal concentration: 0.4 mg/l
measured concentration: 0.37 mg/l and standard deviation was 0.03
Details on test conditions:
Sixty fertilized eggs were selected and randomly distributed into three the test vessels each treatment, 20 eggs for each test vessel. The test begins when all the eggs were placed in the test vessels, which means three replicates each test concentration and controls. Twenty eggs per test vessels were exposed. Dechlorinated tap water filtered through the quartz sand, activated carbon and precision filter was used as test dilution water in the test. The test duration was 30 days post-hatch. The test solutions were renewed every two days. Two renewal procedures were used during the exposure. Before post-hatching: the embryos were transferred into additional test vessels containing new test solutions by glass tubes (equipped with a rubber bulb) or the embryos were retained in the test vessels, the old test solution (at least two thirds) was removed by siphoning and then new solution was supplemented. After post-hatching: the larvae and juvenile fish were transferred into additional test vessels containing new test solutions by siphoning or the test organisms were retained in the test vessels while two thirds of the test water was renewed by siphoning. First feeding was on the 3th day post-hatch, food was supplied once a day. Yolk was used as food for larvae, 40- 60 µL yolk solution was added into each test vessel. BSN48 was for juveniles, the BSN48 was filtered to concentrate and then diluted with dilution water. About 60 µL was added into each test vessel.
Reference substance (positive control):
not required

Results and discussion

Effect concentrationsopen allclose all
Duration:
30 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
30 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
morphology
Remarks:
Abnormality rate
Duration:
30 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
length
Duration:
30 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
weight
Details on results:
No significant difference was found for the number of larvae hatching, cumulative mortality and numbers of deformed larvae between the treatment group (0.4 mg/L) and controls. There was an increasing trend in the lengths, weight of test fish at the end of the test among blank control, solvent control, and treatment group (0.4 mg/L). There was significant difference (P < 0.01) for the length and weight of fish between the treatment group and solventNo significant difference was found for the number of larvae hatching, cumulative mortality and numbers of deformed larvae between the treatment group (0.4 mg/L) and controls. There was an increasing trend in the lengths, weight of test fish at the end of the test among blank control, solvent control, and treatment group (0.4 mg/L). There was significant difference (P < 0.01) for the length and weight of fish between the treatment group and solvent
Results with reference substance (positive control):
not determined
Reported statistics and error estimates:
Results of the toxicity test were interpreted using the function of data analysis in EXCEL Version 2003. Analysis of variance was used to analyse variations within each set of replicates. The NOEC was estimate by One Way Analysis of Variance (One Way ANOVA).

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
fulfilled validity criteria
Conclusions:
In the fish early life stage toxicity test, based on the nominal concentration, the NOEC of BDP to Zebra-fish (Brachydanio rerio) was greater than or equal to 0.4 mg/L, and which was above the water solubility of BDP.
Executive summary:

Early-life Stage Toxicity Test (Semi-Static System) of BDP to Zebra-fish was conducted in accordance with OECD test guideline 210 with GLP compliance. The study was conducted under semi-static conditions with the nominal concentration of 0.4 mg/L and controls (including blank control and solvent control) for a period of 30 days after post-hatch. 60 eggs were used per concentration and controls. The eggs were divided equally into three test chambers per treatment. The test solution was renewed every two days. The mortality/survival at embryo, larval and juvenile stages were observed and recorded once a day during exposure. At the end of the exposure, the total dry weight of all surviving fish was determined and the length of surviving fish was measured. Compared with control group, the NOEC was calculated to be greater than 0.4 mg/l.