Simultaneous determination of procaine and para-aminobenzoic acid by LC–MS/MS method

A sensitive high performance liquid chromatography tandem mass spectrometry (LC–MS/MS) method has been developed for simultaneous determination of procaine and its metabolite p-aminobenzoic acid (PABA). N-Acetylprocainamide (NAPA) was used as an internal standard for procaine and PABA analysis. This assay method has also been validated in terms of linearity, lower limit of detection, lower limit of quantitation, accuracy and precision as per ICH guidelines. Chromatography was carried out on an XTerra™ MS C18 column and mass spectrometric analysis was performed using a Quattro Micro™ mass spectrometer working with electro-spray ionization (ESI) source in the positive ion mode. Enhanced selectivity was achieved using multiple reaction monitoring (MRM) functions, m/z 237 → 100, m/z 138 → 120, and m/z 278 → 205 for procaine, PABA and NAPA, respectively. Retention times for PABA, procaine and NAPA were 4.0, 4.7 and 5.8 min, respectively. Linearity for each calibration curve was observed across a range from 100 nM to 5000 nM for PABA, and from 10 nM to 5000 nM for procaine. The intra- and inter-day relative standard deviations (RSD) were <5%.