High-performance liquid chromatography-tandem mass spectrometry method for simultaneous detection of ochratoxin A and relative metabolites in Aspergillus species and dried vine fruits

A simple, sensitive and reliable quantification and identification method was developed for simultaneous analysis of ochratoxin A (OTA) and its related metabolites ochratoxin alpha (OTα), ochratoxin B (OTB) and mellein. The method was assessed by spiking analytes into blank culture media and dried vine fruits. Performance was tested in terms of accuracy, selectivity and repeatability. The method involves an ultrasonic extraction step for culture samples using methanol aqueous solution (7:3, v/v); the mycotoxin is quantified by high-performance liquid chromatography coupled with electrospray ionisation and triple quadrupole mass spectrometry (LC-ESI-MS/MS). The recoveries were 74.5–108.8%, with relative standard deviations (RSDs) of 0.4–8.4% for fungal culture. The limits of detection (LODs) were in the range of 0.03–0.87 μg l^–1, and the limits of quantification (LOQs) ranged from 0.07 to 2.90 μg l^–1. In addition, the extraction solutions and clean-up columns were optimised specifically for dried vine fruit samples to improve the performance of the method. Methanol–1% sodium bicarbonate extraction solution (6:4, v/v) was determined to be the most efficient. Solid-phase extraction (SPE) was performed as a clean-up step prior to HPLC-MS/MS analysis to reduce matrix effects. Recoveries ranged from 80.1% to 110.8%. RSDs ranged from 0.1% to 3.6%. LODs and LOQs ranged from 0.06 to 0.40 μg kg^–1 and from 0.19 to 1.20 μg kg^–1, respectively. The analytical method was established and used to identify and quantify OTA and related compounds from Aspergillus carbonarius and Aspergillus ochraceus in cultures and dried vine fruits. The results showed that A. carbonarius produced OTα, OTB and OTA, whereas A. ochraceus produced OTB, OTA and mellein after 7 days of cultivation. Of 30 commercial samples analysed, 10 were contaminated with ochratoxins; OTB, OTα and mellein were also detected in different samples.