Simultaneous determination of caffeic acid and its major pharmacologically active metabolites in rat plasma by LC-MS/MS and its application in pharmacokinetic study.

A simple, sensitive and selective high-performance liquid chromatography electrospray ionization tandem mass spectrometry (LC-MS/MS) method was developed for simultaneous determination and pharmacokinetic study of caffeic acid (CA) and its active metabolites. The separation with isocratic elution used a mobile phase composed of methanol and water (containing 0.1% formic acid) at a flow rate of 0.2mL/min. The detection of target compounds was done in selected reaction monitoring (SRM) mode. The SRM detection was operated in the negative electrospray ionization mode using the transitions m/z 179 ([M-H](-))135 for CA, m/z 193 ([M-H](-))134.8 for ferulic acid and isoferulic acid and m/z 153 ([M-H](-))108 for protocatechuic acid. The method was linear for all analytes over the investigated range with all correlation coefficients 0.9931. The lower limits of quantification were 5.0ng/mL for analytes. The intra- and inter-day precisions (relative standard deviation) were <5.86 and <6.52%, and accuracy (relative error) was between -5.95 and 0.35% (n=6). The developed method was applied to study the pharmacokinetics of CA and its major active metabolites in rat plasma after oral and intravenous administration of CA. Copyright (c) 2014 John Wiley & Sons, Ltd.