Controlling the exposure of chemicals in in vitro mammalian cell assays is an important prerequisite for the application of in vitro methods in risk and hazard assessment of chemicals. Existing models require numerous physicochemical and system parameters to quantify the effective concentration in the assay. Synthesizing these studies, this article briefly communicates how the protein-rich supplement in the medium can be utilized to adjust constant and quantifiable exposure concentrations without the need for measurements and complex modeling. We present a simplified mass balance equation based on chemical properties and system parameters from openly accessible databases, which can be used to adjust the dose of chemicals in the exposure medium, leading to defined and stable freely dissolved concentrations (Cfree). The proposed framework prevents experimental artifacts associated with the use of cosolvents and medium oversaturation and enables the conversion of in vitro effect data to freely dissolved effect concentrations (ECfree), which can directly be applied in quantitative in vitro to in vivo extrapolation models and compared to other exposure scenarios.