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Long-term toxicity to fish

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Reference
Endpoint:
fish early-life stage toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 210 (Fish, Early-Life Stage Toxicity Test)
Deviations:
yes
Remarks:
one day oxygen contents were below 60% in several test replicates
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Remarks:
HPLC-MS/MS
Details on sampling:
- freshly prepared stock solution on day: 0, 7, 16, 23, 33
- control: at end of test (day 35)
- test solution on day: 0, 2, 9, 19, 26, 35
Vehicle:
no
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
- Species: Danio rerio Hamilton-Buchanan 1822 (Zebrafish)
- Origin: Fertilised eggs were collected from adult zebrafish from in-house cultures, which have been maintained and bred at ECT since December 2015
- Number of animals: 6 replicates of 20 fertilized eggs per concentration level
- Holding conditions and fertilisation of test organisms: In the culture tanks, the fish were kept in reconstituted water (OECD guideline 203, 1992) diluted with deionised water (1:1; v/v) and supplemented with 1% of artificial seawater (Tropic Marin, Dr. Biener GmbH Aquarientechnik, Wartenberg, Germany; salinity 28 ± 2‰).- Temperature: 26 ± 2 °C
- Photoperiod: 12/12 hours light/dark cycle
- Feeding: a combined diet of newly hatched nauplii of Artemia sp. (Sanders Brine Shrimp Co., Morgan, UT 84050, U.S.A.) and TetraMin (Tetra Werke, Melle, Germany)
Test type:
flow-through
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
35 d
Hardness:
9.1 - 10.0 °dH
Test temperature:
25.2 - 26.1 °C
pH:
7.5 - 7.6
Dissolved oxygen:
4.1 - 8.1 mg/L
Conductivity:
767 - 810 µS/cm
Nominal and measured concentrations:
nominal: 10 mg/L
measured: < LOQ - 1.6 µg/L (LOQ: Limit of quantification: 0.1 µg/L)
Details on test conditions:
Test medium: Reconstituted water (OECD guideline No. 203) mixed with deionised water (1:1; v/v), supplemented with 1% artificial seawater
Endpoints: NOEC, LOEC
Biological parameters: Hatching success; Number of larvae hatching each day; Time to start of hatching and end of hatching; Cumulative mortality (Post-Hatch Success (Survival)); Number of deformed or abnormally appearing larvae; Number of organisms exhibiting abnormal behaviour; Length of the surviving fish; Wet and dry weight of the surviving fish.
Test duration: 35 days
Temperature (target): 26 ± 1.5 °C
Photoperiod: 12/12 hours light/dark cycle
Light intensity (target): 500 - 1000 lx
Test units: Glass vessels, 0.8 L volume
Test concentration (limit concentration): 100% of a filtrate at a loading rate of 10 mg test item/L plus a control
No. of replicates per control: 6
No. of replicates per treatment: 6
Renewal of test solution during exposure: Permanent (flow-through)
Chemical analysis of test concentrations: Control and limit test solutions
Data evaluation: Shapiro-Wilk’s test on normal distribution, Levene’s test on variance homogeneity, Welch-t test, Student-t test
Reference substance (positive control):
no
Duration:
35 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
number hatched
Duration:
35 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
35 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
behaviour
Duration:
35 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
length
Duration:
35 d
Dose descriptor:
NOEC
Effect conc.:
>= 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
weight
Duration:
35 d
Dose descriptor:
LOEC
Effect conc.:
> 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
number hatched
Duration:
35 d
Dose descriptor:
LOEC
Effect conc.:
> 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
35 d
Dose descriptor:
LOEC
Effect conc.:
> 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
behaviour
Duration:
35 d
Dose descriptor:
LOEC
Effect conc.:
> 0.43 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
weight
Validity criteria fulfilled:
yes
Remarks:
dissolved oxygen content were measured below 60% in several replicates of the control, however, this minor deviation had no influence on the reliability of the test data and the integrity of the study
Conclusions:
No chronic toxic effect of MBTS to Danio renio at a limit concentration of 10 mg/L (nominal) could be observed.
Executive summary:

A study was conducted according to OECD Guideline for Testing of Chemicals, No. 210 "Fish, Early-life Stage Toxicity Test" to determine lethal and sub-lethal effects of the test item MBTS on the early-life stages of Danio rerio (zebrafish). To achieve this, a limit concentration of the test item in aqueous solution was prepared using a filtrate at a loading rate of 10 mg test item/l (mean measured >0.43 µg/L). The test organisms were exposed to this limit concentration as well as to control without the test item. During the test period the test solutions were be renewed in a flow-through system.

 

Newly fertilised eggs of the test species were exposed to this test concentration until 30 days post-hatch. Lethal and sub-lethal effects of the test item on eggs, larvae and juvenile fish were assessed in comparison with control values to determine the lowest observed effect concentration (LOEC) and the no observed effect concentration (NOEC). The following biological parameters were investigated: hatching success; number of larvae hatching each day; time to start of hatching and end of hatching; cumulative mortality, post-hatch success (survival); number of deformed or abnormally appearing larvae; number of organisms exhibiting abnormal behaviour; length of the surviving fish; wet and dry weight of the surviving fish. 6 replicates of 20 fertilized eggs per concentration and control were exposed.

As no chronic toxic effects were observed, a NOEC of >= 10 mg/L (nominal) equaling >= 0.43 µg/L (mean measured) and a LOEC of >10 mg/L (nominal) equaling >0.43 µg/L (mean measured) were determined for MBTS for hatching success, post-hatch success (survival), number of healthy fish, length of surviving fish, wet and dry weight of the surviving fish, respectively.

Description of key information

No chronic toxic effect of MBTS to Danio renio in a limit test at a test item concentration of 10 mg/L (nominal) were observed during the experimental period of 35 days (ECT 2017).

Key value for chemical safety assessment

Additional information

The derived 35d-NOEC is found to be >= 10 mg/L (nominal) and the 35d-LOEC >10 mg/L (nominal).