A simple and rugged method of analysis for grayanotoxins I and III in honey using liquid chromatography triple quadrupole mass spectrometry with electrospray ionization was developed. This paper describes the first LC-MS/MS method for the quantitation and confirmation of the grayanotoxins in honey using "dilute-and-shoot" sample preparation approach. Honey sample was diluted 10-fold in methanol-water (1:4 v/v) prior to analysis. Chromatographic separation was achieved on a reversed phase HPLC column using a water-methanol gradient with 0.1% acetic acid. The method was fully validated for quantitative purposes. Overall recoveries, selectivity, overall intraday and interday repeatability, decision limit, and detection capability of the analytes was determined. The matrix effects, ruggedness, and analyte stability in standards and samples were studied. Ten real honey samples were successfully analyzed using the developed method. All the samples were found to contain residues of GTXs ranging from 0.1 to 39 mg/kg.
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