Determination of nitric oxide in hydrophytes using poly(methacrylic acid-ethylene glycol dimethacrylate) monolith microextraction coupled to high-performance liquid chromatography with fluorescence detection

Nitric oxide (NO) is a bioactive molecule that has recently emerged as a cellular messenger in numerous physiological processes in plants. A novel high-performance liquid chromatography (HPLC) method combined with poly(methacrylic acid-ethylene glycol dimethacrylate) (MAA-EGDMA) monolith microextraction (PMME) is developed for sensitive determination of NO in hydrophytes. NO is derivatized using a fluorescent probe, 1,3,5,7-tetramethyl-8-(3′,4′-diaminophenyl)-difluoroboradiaza-s-indacene (DAMBO), and then the derivatives are extracted with PMME and analyzed by high-performance liquid chromatography (HPLC) with fluorescence detection. The conditions for the derivatization and the subsequent extraction of NO derivatives are optimized in detail. The detection limit (S/N = 3) of NO is determined to be 2 × 10−12 mol L−1. Close correlation coefficient and excellent method reproducibility are obtained for the analyte over a linear range of 9 × 10−11–4.5 × 10−8 mol L−1. The inter- and intraday relative standard deviations (R.S.D.s) are less than 5%. The proposed method is successfully applied to the determination of NO levels in hydrophytes samples.