Simultaneous determination of gallic acid and gentisic acid in organic anion transporter expressing cells by liquid chromatography–tandem mass spectrometry
2013
Wang, Li | Halquist, Matthew S. | Sweet, Douglas H.
In order to elucidate the role of organic anion transporters (OATs) in the renal elimination of gallic acid and gentisic acid, a new, rapid, and sensitive liquid chromatography-tandem mass spectrometry (LC–MS/MS) method has been developed and validated for the simultaneous determination of gallic acid and gentisic acid in cell lysate, using Danshensu as the internal standard (IS). After a simple liquid–liquid extraction, the analytes were detected in negative ESI mode using selected reaction monitoring. The precursor-to-product ion transitions (m/z) were 169.0→125.0, 153.1→108.0, and 196.8→135.2 for gallic acid, gentisic acid, and the IS, respectively. Chromatographic separation was achieved on a C18 column using mobile phases consisting of water with 0.1% acetic acid (A) and acetonitrile with 0.05% formic acid. (B) The total run time was 3min and calibration curves were linear over the concentrations of 0.33–2400ng/mL for both compounds (r2>0.995). Good precision (between 3.11% and 14.1% RSD) and accuracy (between −12.7% and 11% bias) was observed for quality controls at concentrations of 0.33 (lower limit of quantification), 1, 50, and 2000ng/mL. The mean extraction recovery of gallic acid and gentisic acid was 80.7% and 83.5%, respectively. Results from post-column infusion and post-extraction methods indicated that the analytical method exhibited negligible matrix effects. Finally, this validated assay was successfully applied in a cellular uptake study to determine the intracellular concentrations of gallic acid and gentisic acid in OAT expressing cells.
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