Anomalous relationship between free drug fraction and its total concentration in drug–protein systems: I. Investigation of propofol binding in model HSA solution

Abstract

According to contemporary knowledge, only the free form of a drug exhibits pharmacological activity due to its ability to cross the membranes and to interact with receptors. For the majority of drugs their free fraction in human plasma is usually constant, hence the therapeutic effect of a drug is most often correlated with its total concentration. However, in the case of certain drugs of narrow therapeutic window, for which a slight elevation of a free drug fraction (e.g., occurring in some pathological states) may exert a toxic effect, the therapeutic effect should be correlated with free concentration. As we reported earlier, at total propofol concentrations below 1 μg ml −1 the free fraction of the drug increases with the decrease of its total concentration. The presented paper discusses the results of several experiments performed in order to elucidate this anomalous phenomenon. The study is mainly focused on the investigation of the relationship between free drug percentage and total drug concentration in human serum albumin (HSA) solution, performed at different temperatures and in the presence of the salting-out agent. It was observed that a temperature increase leads to an increased propofol binding, which confirms the importance of hydrophobic interactions for the binding process. The addition of the salting-out agent significantly changing water structure of the drug–protein system is followed by disappearance of the anomalous relationship and results in constant value of the free drug fraction in the whole range of the studied total drug concentration. The results of the performed experiments indicate that the well-ordered water around a protein molecule (i.e., the hydration layer) constitutes the main factor responsible for the discussed anomalous free drug fraction changes.