Gaining confidence in in vitro toxicity data with measured exposure concentrations

Area of Science:

• Toxicology
• Biochemistry
• Environmental Science

Background:

• Cell-based in vitro bioassays are vital New Approach Methodologies (NAMs) for chemical screening.
• Undefined chemical exposure is a major limitation hindering the acceptance of in vitro bioassays.
• Accurate exposure assessment is needed to complement cytotoxicity and specific endpoint testing.

Purpose of the Study:

• To measure freely dissolved concentrations (Cfree) of 24 test chemicals in cell-based bioassays.
• To assess the impact of chemical hydrophobicity on exposure concentrations over time.
• To provide measured exposure data for improved in vitro-to-in vivo extrapolation (IVIVE).

Main Methods:

• Quantified Cfree of 24 chemicals in four different cell-based bioassays using 96-well plates.
• Measured Cfree at the time of dosing and after 24 hours of exposure.
• Determined freely dissolved fractions (ffree) based on chemical hydrophobicity.

Main Results:

• Freely dissolved fractions (ffree) decreased with increasing chemical hydrophobicity.
• For 12 chemicals, Cfree decreased by over 20% during the bioassays.
• Loss of Cfree was attributed to sorption, metabolism, or degradation.

Conclusions:

• Simultaneous measurement of toxicity and exposure enhances the reliability of in vitro bioassays.
• Identified chemicals with unstable exposure concentrations and/or low bioavailability.
• Generated quantitative bioactivity data applicable for IVIVE.