Incipient cytotoxicity: A time-independent measure of cytotoxic potency in vitro

Influence of in Vitro Assay Setup on the Apparent Cytotoxic Potency of Benzalkonium Chlorides

“…Membrane affinity is higher for BACs with longer alkyl chain lengths. , Therefore, the hydrophobicity is likely higher and diffusion through aqueous solutions and cellular uptake may be slower, compared to surfactants with shorter alkyl chain lengths, as is observed for other chemicals . Considering this, the equilibrium time of the chemical distribution in the wells and particularly into the cells, will likely be slower for surfactants with long alkyl chain lengths . In this case, 48 h might not be enough for BAC18 to become equilibrated with all assay components and medium constituents.…”

“…Understanding the effect of exposure duration is important to achieve accurate and reproducible cytotoxic potency determinations in vitro. Gülden et al 54 noted that using the incipient EC 50 or at least 72 h EC 50 , instead of variable or arbitrary exposure times, will make comparisons to other in vitro assays, as well as extrapolations to in vivo experiments more meaningful. However, depending on the cell type and assay, it may not always be possible to expose the cells for 72 h or longer, which is also the case for the RTgill-W1 studied.…”

“…EC 50 values after 48 h of exposure are lower than after 24 h exposure. These findings agree with those of Gülden et al , where EC 50 values of various chemicals decreased with increasing exposure time in cultures of C6 glioma or Balb/c 3T3 cells until an “incipient” EC 50 is reached. The incipient EC 50 remains constant with longer exposure times.…”

Biokinetics and xenobiotic bioavailability

“…The field of genetic analysis is progressing rapidly and interested parties should maintain knowledge of current best scientific practice in this area as next generation sequencing begins to become a routine tool. Doing so problems can be avoided early in the process and not jeopardise the cell lines used for regulatory purposes as has happened for the Bhas 42 cell line in a cell transformation in vitro method (OECD, 2016 [42] ) where issues related to misidentification arose at a late stage.…”

“…Thus, in an in vitro system, the freely dissolved concentration of the test item in the medium or in the cell (being as close to the target as possible) is the central parameter. Some processes (Groothuis et al, 2015 [42] ; Heringa et al, 2006 [43] ); depicted schematically in , result in a freely dissolved concentration that is not the same as the nominal concentration (i.e., the added concentration).…”