title: Development and validation of a liquid chromatography-tandem mass spectrometry method to determine intact glucosinolates in bee pollen.
abstract: A new method was developed to determine twelve intact-glucosinolates (GLSs) (glucoiberin, GIB; glucoraphanin, GRA; glucoerucin GER; gluconapin, GNA; glucotropaeolin, GTL; glucobrassicin, GBC; gluconasturtiin, NAS; glucoalyssin, ALY; 4-hydroxyglucobrassicin, 4OH; 4-methoxyglucobrassicin, 4ME; neoglucobrassicin, NEO; sinigrin, SIN) in bee pollen, by means of liquid chromatography tandem mass spectrometry (LC-MS/MS) with electrospray ionization (ESI). An efficient extraction procedure was proposed (average analyte recoveries were between 85% and 96%); this involved a solid-liquid extraction (SLE) with heated water, followed by a solid phase extraction (SPE) with a weak anion exchange (NH2) sorbent. Chromatography was performed on a Gemini(®) C18 analytical column with a mobile phase of formic acid in water (0.5%,v/v) and formic acid in acetonitrile (0.5%,v/v), in gradient elution mode at 1mL/min, resulted in baseline-separated peaks and a run time of 30min. The method was fully validated in terms of selectivity, limits of detection (LOD) and quantification (LOQ), linearity, carry-over effect, reinjection reproducibility, precision and accuracy. A good selectivity, low LODs and LOQs, ranging from 1 to 16μg/kg, wide linear ranges from LOQ to 1000μg/kg, and satisfactory reinjection reproducibility, precision and accuracy with relative standard deviation and relative error values lower than or equal to 9%; meanwhile, results indicates a negligible carry-over effect. The proposed method was applied to analyze intact-GLSs in bee pollen. Nine of the GLSs studied were identified in certain samples analyzed over a wide concentration range (LOQ-2226μg/kg), and significant differences in GLS content were observed among the samples.
authors.affiliation: IU CINQUIMA, Analytical Chemistry Group, University of Valladolid, 47011 Valladolid, Spain.
authors.forename: Ana M
authors.initials: AM
authors.lastname: Ares
authors.affiliation: IU CINQUIMA, Analytical Chemistry Group, University of Valladolid, 47011 Valladolid, Spain.
authors.forename: María J
authors.initials: MJ
authors.lastname: Nozal
authors.affiliation: IU CINQUIMA, Analytical Chemistry Group, University of Valladolid, 47011 Valladolid, Spain. Electronic address: jose.bernal@qa.uva.es.
authors.forename: José
authors.initials: J
authors.lastname: Bernal
mesh_terms: D000818:Animals; D001516:Bees; D002853:Chromatography, Liquid / Q000379:methods*; D005961:Glucosinolates / Q000032:analysis* / Q000737:chemistry; D057230:Limit of Detection; D016014:Linear Models; D011058:Pollen / Q000737:chemistry*; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D005961:Glucosinolates
keywords: Glucosinolates; Keywords: Bee pollen; LC–MS/MS; Solid phase extraction; Solid–liquid extraction