Simultaneous determination of organic acids and nitrate in xylem saps of the hyperaccumulator Alyssum murale by RP-HPLC after solid-phase extraction with nanosized hydroxyapatite

An analytical approach based on solid-phase extraction (SPE) with nanosized hydroxyapatite and reversed-phase high performance liquid chromatography (RP-HPLC) for the simultaneous determination of organic acids (oxalic, malic, malonic, citric, and fumaric) and nitrate in xylem saps of the hyperaccumulator Alyssum murale is presented in this study. The optimum experimental conditions for the separation and determination of organic acids and nitrate are studied. The interference from nitrate on oxalic acid in RP-HPLC is eliminated by SPE with nanosized hydroxyapatite, and the simultaneous determination of organic acids and nitrate is achieved by RP-HPLC at the optimum chromatographic conditions. The accuracy of the method is confirmed with an average recovery ranging between 95.2% and 99.8%, the relative standard deviations (RSD) are less than 2.0%. This method is successfully applied to determine the organic acids and nitrate in xylem saps of the hyperaccumulator Alyssum murale.