Confocal Raman microscopy for simultaneous monitoring of partitioning and disordering of tricyclic antidepressants in phospholipid vesicle membranes

Characterization of drug-membrane interactions is important in order to understand the mechanisms of action of drugs and to design more effective drugs and delivery vehicles. Raman spectra provide compositional and conformational information of drugs and lipid membranes, respectively, allowing membrane disordering effects and drug partitioning to be assessed. Traditional Raman spectroscopy and other widely used bioanalytical techniques such as differential scanning calorimetry (DSC) and nuclear magnetic resonance (NMR) typically require high sample concentrations. Here, we describe how temperature-controlled, optical-trapping confocal Raman microscopy facilitates the analysis of drug-membrane interactions using micromolar concentrations of drug, while avoiding drug depletion from solution by working at even lower lipid concentrations. The potential for confocal Raman microscopy as an effective bioanalytical tool is illustrated using tricyclic antidepressants (TCAs), which are cationic amphiphilic molecules that bind to phospholipid membranes and influence lipid phase transitions. The interaction of these drugs with vesicle membranes of differing head-group charge is investigated while varying the ring and side-chain structure of the drug. Changes in membrane structure are observed in Raman bands that report intra- and intermolecular order versus temperature. The partitioning of drugs into the membrane can also be determined from the Raman scattering intensities. These results demonstrate the usefulness of confocal Raman microscopy for the analysis of drug-membrane systems at biologically relevant drug concentrations. Effective tools for monitoring drug-membrane interactions are crucial for rational design of new drugs. Copyright (C) 2009 John Wiley & Sons, Ltd.