Challenges regarding analysis of unbound fraction of highly bound protein antiretroviral drugs in several biological matrices: lack of harmonisation and guidelines

Drug Discov Today. 2015 Apr;20(4):466-74. doi: 10.1016/j.drudis.2014.11.010. Epub 2014 Nov 20.


The unbound drug concentration in plasma is usually considered the only active fraction; thus the binding of a drug to a protein limits its pharmacological actions. This is of special importance for those highly bound drugs. Therefore, binding studies can be of great utility for those drugs where relationship between free and total drug concentration is variable among patients, or it can be altered by some condition or disease, or even by interactions with other drugs. However, there is a lack of validation guidelines for the determination of unbound concentrations. Antiretroviral drugs (ARVs), protease inhibitors (PIs), efavirenz and nevirapine are highly bound to proteins. Here, we present a review on the overall methods for the study of unbound fractions of highly bound plasma protein ARVs. We also provide a critical evaluation of the methods applied, their differences and the main points to be controlled and validated.

Publication types

  • Review

MeSH terms

  • Animals
  • Anti-Retroviral Agents / blood*
  • Anti-Retroviral Agents / cerebrospinal fluid
  • Anti-Retroviral Agents / pharmacokinetics
  • Benzoxazines / blood*
  • Benzoxazines / cerebrospinal fluid
  • Benzoxazines / pharmacokinetics
  • Drug Monitoring / methods
  • Drug Monitoring / standards*
  • Humans
  • Nevirapine / blood*
  • Nevirapine / cerebrospinal fluid
  • Nevirapine / pharmacokinetics
  • Practice Guidelines as Topic / standards*
  • Predictive Value of Tests
  • Protein Binding
  • Reproducibility of Results


  • Anti-Retroviral Agents
  • Benzoxazines
  • Nevirapine
  • efavirenz