Aim: Development of a high-sensitivity chiral LC–MS/MS method was required to evaluate a combination of pramipexole (S-PPX) and its enantiomer dexpramipexole (R-PPX) in a proposed clinical trial. The previously available methods suffered from low sensitivity for the (S)-enantiomer in the presence of the more abundant (R)-enantiomer. Based on the projected dosing regimen in the clinical trial, a 5000-fold improvement in sensitivity was required for the (S)-enantiomer. Methodology: Spiked human plasma samples were extracted by liquid–liquid extraction using ethyl acetate and injected onto a CHIRALPAK ID column under pH gradient conditions. Conclusion: An improved analytical method was developed and validated with a final LLQ for (S)-PPX of 0.1 ng/ml in the presence of 2000 ng/ml of (R)-PPX.
Keywords:
- bioanalysis
- chiral chromatography
- dexpramipexole
- high sensitivity
- human plasma
- LC–MS
- liquid–liquid extraction
- method development
- pramipexole
- validation