BACKGROUND AND OBJECTIVE In many clinical situations, measurement of the total drug concentration does not provide the needed information concerning the fraction of unbound drug in plasma, which is available for pharmacodynamic action. To address this, a 'normalized concentration' can be calculated on the basis of the observed total drug concentration and the serum protein level. Up to now, this method has only been applied to phenytoin. Several equations for calculating normalized concentrations of phenytoin have been published, many leading to different results. Regrettably, all of the equations in the current literature are based on an outdated model of drug binding to human serum albumin and are based on the fraction of unbound drug, which is known to depend on both protein and drug concentrations. In response to the relatively new scientific evidence about drug binding to human plasma proteins, the objective of the present study is to develop a general method for calculating normalized drug concentrations in the presence of altered plasma protein binding. METHODS When several drug molecules can be bound by a protein molecule, multiple equilibria are established; these equilibria may be formulated in terms of a stoichiometric analysis or a site-oriented analysis. Both models are currently encountered in the scientific literature, sometimes without clear identification of which model is used. The present study presents the basic equations for both models and shows how the normalized concentration can be calculated on the basis of the measured drug concentration, the protein level and the binding constants. RESULTS The normalized concentration can be calculated for any drug, using the same simple equation regardless of the binding model and the number of binding proteins. Explicit solutions are presented for particular cases of clinical importance. The new model is validated by comparison with the Winter-Tozer equation for calculating the normalized phenytoin concentration and is found to be equivalent for concentrations close to therapeutic concentrations. In the case of phenytoin, the main advantage of the new equation is that it also works outside the linear binding range. CONCLUSIONS A new comprehensive method for calculating normalized drug concentrations is developed, allowing drug concentrations to be interpreted correctly in cases of altered drug-protein binding. The calculations are based on binding constants and are applicable to any protein level and drug concentration, without being limited to linear binding of drugs to proteins. The new model is expected to become important in pharmacokinetic-pharmacodynamic modelling, allometric scaling and population pharmacokinetics because it provides the ability to accurately take into account physiological and pathological changes in protein binding. As a direct clinical application, the equations can be used to calculate normalized drug concentrations in patients with abnormal protein levels, such as the elderly, trauma patients and paediatric patients.