Abstract
Unbound volume of distribution is often treated as an intrinsic, species-invariant property. However, mechanistic analysis demonstrates that it is influenced by plasma protein binding, particularly when plasma and tissue binding are coupled through shared binding components and their relative compartmental distribution. Evidence across rat strains suggests that plasma protein binding significantly modulates half-life and hepatic extraction, thereby impacting in vivo efficacy. These insights argue for revisiting common cross-species scaling practices and for considering plasma protein binding as an explicit design parameter.