Experimental Exposure Assessment of Ionizable Organic Chemicals in In Vitro Cell-Based Bioassays

Chem Res Toxicol. 2020 Jul 20;33(7):1845-1854. doi: 10.1021/acs.chemrestox.0c00067. Epub 2020 May 19.

Abstract

Exposure assessment in in vitro cell-based bioassays is challenging for ionizable organic chemicals (IOCs), because they are present as more than one chemical species in the bioassay medium. Furthermore, compared to neutral organic chemicals, their binding to medium proteins and lipids is driven by more complex molecular interactions. Total medium concentrations (Ctotal,medium) and/or freely dissolved medium concentrations (Cfree,medium) were determined for one neutral chemical and 14 IOCs (acids, bases, multifunctional) at concentrations relevant for determination of cytotoxicity and effect. Cfree,medium was measured in two in vitro bioassays at the time of dosing and after 24 h of incubation using solid-phase microextraction. Cfree,medium was maximally 1.7 times lower than the nominal concentrations (Cnom) for the hydrophilic chemicals (caffeine and lamotrigine). For the organic acids (naproxen, ibuprofen, warfarin, and diclofenac), Cfree,medium was by a factor of 4 lower than Cnom at high concentrations, but the ratio was much higher at low concentrations, indicating a nonlinear binding behavior. The experimental Cfree,medium was also compared with Cfree,medium predicted with a mass balance model accounting for binding to medium proteins and lipids. The mass balance model performed well for five of the test chemicals (within a factor of 10), but it underestimated Cfree,medium by up to a factor of 1200 for chemicals that showed nonlinear binding to medium components. These findings emphasize that experimental exposure assessment is required for improved understanding of in vitro toxicity data.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biological Assay / methods*
  • Humans
  • Liposomes / chemistry
  • MCF-7 Cells
  • Organic Chemicals / toxicity*
  • Oxidative Stress / drug effects
  • PPAR gamma / metabolism
  • Serum Albumin, Bovine / chemistry
  • Solid Phase Microextraction
  • Water / chemistry

Substances

  • Liposomes
  • Organic Chemicals
  • PPAR gamma
  • Water
  • Serum Albumin, Bovine