Determination of Bongkrekic Acid and Iso-Bongkrekic Acid in Red Yeast Rice and Its Related Products by Liquid Chromatography-Tandem Mass Spectrometry

Bongkrekic acid (BA) is a metabolite that is lethal to humans and is produced by Burkholderia gladioli pathovar cocovenenans (BGC) containing the bon gene cluster. An analytical method for BA and its stereoisomer, iso-bongkrekic acid (IBA), was established for deeply fermented rice products, such as red yeast rice and its related products, by liquid chromatography–tandem mass spectrometry (LC-MS/MS). BA and IBA in samples were extracted with 80% methanol/water containing 1% ammonia by ultrasonic assistance, cleaned by solid-phase extraction (SPE) with an anion-exchange cartridge, separated with an XBridgeTM BEH C18 chromatographic column (150 mm × 3.0 mm, 2.5 μm), measured by MS/MS, and quantified using an isotope-labeled internal standard. The influencing factors such as extraction solvents, ultrasonic extraction time and loading volume during SPE clean-up were optimized to improve the recoveries of BA and IBA. Also, the extraction efficiency between the simulated quality control and directly spiked samples, the matrix effects with the use of different chromatographic columns, the impact of co-eluting interferences under different mobile phase compositions and the conversion of BA to IBA are discussed. The linear range of BA and IBA was 0.1–50 ng/mL (equivalent to 1–500 μg/kg in the sample), with the correlation coefficient exceeding 0.999. The limits of detection and quantification were 0.3–0.5 μg/kg and 1.0–1.5 μg/kg, respectively. The average recoveries of the analytes were 88.4–107% with relative standard deviations (RSDs, n = 6) between 2.9% and 11.8% in red yeast rice and its related products under three spiking levels. This optimized method was applied to study the contamination of BA and IBA in commercially available samples. This study can provide technical support for the supervision and risk assessment of BA and IBA in red yeast rice and its related products.