A high throughput passive dosing format for the Fish Embryo Acute Toxicity test.

Area of Science:

• Environmental Toxicology
• Aquatic Toxicology
• Chemical Safety Assessment

Background:

• High-throughput testing using the Fish Embryo Acute Toxicity (FET) test (OECD TG 236) faces challenges with hydrophobic substances due to sorptive and evaporative losses.
• These losses lead to declining and poorly controlled exposure concentrations, compromising test reliability.

Purpose of the Study:

• To enhance exposure conditions in FET tests for hydrophobic chemicals.
• To evaluate a passive dosing system utilizing silicone O-rings in standard 24-well polystyrene plates.

Main Methods:

• Zebrafish embryos were exposed to phenanthrene concentrations using silicone O-rings in 24-well plates until 120 hours post-fertilization (hpf).
• Exposure concentrations, lipid-normalized body residues, and chemical activity were measured.
• Mortality (LC50) and sublethal morphological effects (e.g., swim bladder inflation, swimming activity) were assessed.

Main Results:

• A linear dilution series of phenanthrene was achieved, with LC50 at 120 hpf determined as 310 μg/L.
• Critical body residue (CBR50) and lethal chemical activity (La50) were 2.72 mmol/kg and 0.047, respectively, consistent with baseline toxicity.
• Impaired swim bladder inflation and reduced swimming activity were observed, with reduced activity attributed to baseline toxicity.

Conclusions:

• Silicone O-rings facilitate a linear dilution series and stable aqueous concentrations for phenanthrene in FET tests.
• This passive dosing method allows for reliable effect concentration determination and mechanistic toxicity insights.
• The silicone O-rings themselves showed no toxicity, indicating their suitability for testing hydrophobic chemicals in FET assays.