Development of\na Specific Fluorescence Post-column\nDerivatization Method Coupled with Ion-Pair Chromatography for Phytate\nAnalysis in Food

Phytate in plants (inositol phosphates, InsPs) affects\nmineral\nbioavailability. However, methods for their quantification may lead\nto variable results, and some are nonspecific (spectrophotometric\ntechniques). In this study, ion-pair high-performance liquid chromatography\n(HPLC) was coupled with post-column derivatization to allow fluorescence\ndetection (FLD, λ<sub>excitation</sub>324/λ<sub>emission</sub>364 nm) of InsPs. The fluorescence derivatization reaction, the main\ninput of this study, was based on a ternary complex among phytate,\niron(III), and 1,10-phenanthroline. Phytic acid (InsP<sub>6</sub>)\nand three other InsPs (InsP<sub>3–5</sub>) were analyzed in\npeanuts and soybean products after extraction in HCl 0.66 M, followed\nby purification on strong anion exchange\ncartridges. The novel method, named IP-HPLC–FLD, selectively\nseparated InsP<sub>3–6</sub> with linear ranges between 600\nand 2000 mg 100 g<sup>–1</sup> (<i>R</i><sup>2</sup> > 0.99). The limits of detection and quantification were between\n120–180 and 340–540 mg 100 g<sup>–1</sup>, respectively.\nAs the relative standard deviations were under 10%, the IP-HPLC–FLD\nmethod is suitable for phytate analysis.