Characterization of drug-protein interactions by capillary electrophoresis hyphenated to mass spectrometry

The demand for analytical techniques to evaluate and measure drug–plasma protein interactions continues to increase. The binding of drugs to plasma proteins is an important parameter to determine during the drug development process because it impacts both pharmacokinetics and pharmacodynamics. Among the numerous methods that have been proposed to perform such studies, CE in frontal analysis mode (CE/FA) is attractive because it consumes a relatively low amount of samples, is fast, and enables analyses under near-physiological conditions. Most CE/FA applications have been performed with UV detection and often lack sensitivity. In this study, CE was hyphenated to MS to enhance the sensitivity of the method and to evaluate strong drug–plasma protein interactions. To adapt the previously developed CE/FA-UV method to CE/FA-MS, different parameters were considered, such as the buffer composition, the rinsing step, and the ESI and MS parameters. The most critical aspect involved obtaining stable MS signals. Good results were achieved due to careful optimization of the ESI and MS parameters, among which the sheath liquid composition appeared to be the most significant. Interactions between six drugs and 1-acid glycoprotein and three drugs and BSA, including basic, neutral, and acidic drugs, were measured with the optimized CE/FA-MS method. The obtained affinity constants ranged from 1·10−4 M−1 to 2·10−5 M−1 and were in good agreement with the results that were obtained by CE/FA-UV and equilibrium dialysis.