5 Exposure assessment in bioassays

The CITEPro module “Exposure assessment in bioassays” aims at the direct combination of the bioassay testing in module 1 with instrumental analysis. The effects measured in in vitro bioassays are typically reported as nominal effect concentrations. However, various processes, e.g. sorption to plastic well plates, binding to medium components, metabolism, and volatilization can influence the bioavailability of the test chemicals in in vitro bioassays. The free concentration (Cfree) in the assay medium is regarded a better dose metric than the nominal concentration. In serum-free media Cfree can be determined directly, while a solid phase microextraction (SPME) method is used for the cell-based bioassays of module 1 to remove the sample matrix. All samples are analyzed using an LC-MS/MS system from Agilent. The experimental exposure assessment is accompanied by physicochemical characterization of the test chemicals. A Sirius T3 from Pion Inc. will be used for the determina-tion of acidity constants, pH dependent solubility and octanol-water distribution ratios of ionizable test chemicals.

Application: Free concentration measurement in cell-based bioassays for quantitative in vitro to in vivo extrapolation and determination of partition constants

Selected literature references
 

Henneberger, L., J. Huchthausen, N. Wojtysiak and B. I. Escher (2021). "Quantitative In Vitro-to-In Vivo Extrapolation: Nominal versus Freely Dissolved Concentration." Chemical Research in Toxicology 34(4): 1175-1182.

Henneberger, L., M. Mühlenbrink, M. König, R. Schlichting, F. C. Fischer and B. I. Escher (2019). "Quantification of freely dissolved effect concentrations in in vitro cell-based bioassays." Arch Toxicol 93(8): 2295-2305.