Development of a liquid chromatography–tandem mass spectrometry method for the determination of puberulic acid in red yeast rice supplements

The recent outbreak resulting in over 80 deaths and 500 hospitalizations has raised significant concerns regarding puberulic acid, a potentially nephrotoxic contaminant found in red yeast rice (RYR) supplements. This incident highlights the urgent need for a reliable analytical method to assess contamination levels and guide regulatory actions. Our study presents the first method for the direct and accurate quantification of puberulic acid in RYR using liquid chromatography–tandem mass spectrometry (LC–MS/MS). Mass spectrometric parameters were optimized using negative electrospray ionization and selected reaction monitoring (SRM). Chromatographic separation was achieved with a hydrophilic interaction liquid chromatography (HILIC) column and a basic mobile phase. A three-cycle extraction process with 90 % methanol yielded optimal recovery rates. The calibration curve demonstrated excellent linearity ( r ² = 0.999), with detection and quantification limits of 1.95 ng/mL and 6.55 ng/mL, respectively. Method validation confirmed its accuracy and precision, establishing this approach as a crucial tool for detecting puberulic acid in RYR supplements. • A direct and reliable analytical method for nephrotoxic contaminant puberulic acid determination. • Addresses recent outbreak-associated morbidity and mortality. • Highly accurate and precise method for regulatory use.