title: Simultaneous determination of tryptophan, kynurenine, kynurenic and xanthurenic acids in honey by liquid chromatography with diode array, fluorescence and tandem mass spectrometry detection.
abstract: A liquid chromatography method using diode array-fluorescence detection and atmospheric pressure chemical ionization mass spectrometry (LC-DAD-FLD and LC-APCI-MS/MS) was developed to quantify the levels of tryptophan (TRP), kynurenine (KYN), kynurenic (KYNA) and xanthurenic (XA) acids in honey. This procedure involved isolating the compounds of interest via solid-phase extraction (SPE) with mixed-mode polymeric cartridges. Chromatographic separation of the analytes was performed in isocratic mode on a Synergi 4μ Hydro-RP 80 Å (150×4.60 mm i.d.) analytical column at 30 °C. The mobile phase of 20mM ammonium formate (pH 4) and methanol was passed at a flow rate of 0.5 mL/min. In replicate sets of spiked honey samples, the average analyte recoveries ranged from 60 to 98% for TRP, 55 to 120% for KYN, 65 to 106.5 for KYNA and 56 to 114% for XA. Detection limits ranged from 4 to 36 μg/kg for LC-DAD-FLD to 0.2 and 1.0 μg/kg for LC-APCI-MS/MS. A strong matrix effect was found when MS/MS was employed, necessitating calibration using the standard addition method on matrix-matched standards for each honey type. The method was used to quantify each of the compounds of interest in 17 honey samples of distinct botanical origins.
authors.affiliation: I.U.CINQUIMA, Analytical Chemistry Group, Faculty of Sciences, University of Valladolid, Dr. Mergelina s/n, 47011 Valladolid, Spain.
authors.forename: María E
authors.initials: ME
authors.lastname: Soto
authors.forename: Ana M
authors.initials: AM
authors.lastname: Ares
authors.forename: José
authors.initials: J
authors.lastname: Bernal
authors.forename: María J
authors.initials: MJ
authors.lastname: Nozal
authors.forename: José L
authors.initials: JL
authors.lastname: Bernal
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods; D005561:Formates; D006722:Honey / Q000032:analysis*; D007736:Kynurenic Acid / Q000032:analysis*; D007737:Kynurenine / Q000032:analysis*; D000432:Methanol; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013030:Spain; D053719:Tandem Mass Spectrometry / Q000379:methods; D014364:Tryptophan / Q000032:analysis*; D014977:Xanthurenates / Q000032:analysis*
chemical_list: D005561:Formates; D014977:Xanthurenates; C030544:formic acid; D007737:Kynurenine; C028330:xanthurenic acid; D014364:Tryptophan; D007736:Kynurenic Acid; D000432:Methanol