Development and validation of a rapid method for the determination of natamycin in wine by high-performance liquid chromatography coupled to high resolution mass spectrometry

Abstract

A selective, sensitive and rapid procedure based on liquid chromatography coupled to electrospray ionisation in positive mode high resolution mass spectrometry (LC-ESI-HR-MS) has been developed and single laboratory validated for the determination of natamycin, a naturally occurring fungicide, in white and red wine. This technique allowed accurate masses to be measured and interferences excluded from the extracted ion chromatograms. Natamycin measured at a high mass accuracy of m/z 666.31069 and a confirmatory ion at m/z 503.22672 facilitated the use of minimal sample preparation procedures. This allowed for the direct injection (20 µL) of wine, thereby minimising the photosensitive analyte degradation. Chromatographic separation was achieved on a C₁₈ column using a mobile phase gradient of 10% to 90% methanol : acetic acid (97 : 3, v/v) over 25 minutes with a total run time of 30 minutes. The limits of detection and quantification were 3 and 5 µg L⁻¹ respectively. The method was successfully validated to show the standard range linearity, sensitivity, accuracy and precision in the matrices tested. The method was applied to 190 commercial wines, with 50 (26%) positive results (>5 µg L⁻¹) over a period of three months between December 2009 and February 2010, with concentrations ranging from 7 to 2134 µg L⁻¹. Natamycin is not permitted for use in wine within the European Union and this method may be used to aid enforcement of regulations.