Residue Analysis of Glyphosate and Aminomethylphosphonic Acid (AMPA) in Soybean Using Liquid Chromatography Coupled with Tandem Mass Spectrometry

Abstract

496by mass or tandem mass spectrometry, especially by using electrospray ionization, because they are very polar and easily ionized on this technique. However, just a few methods using mass or tandem mass spectrometry coupled with liquid chromatography have been found in the literature for glyphosate and AMPA analysis. In most of published papers, pre and postcolumn derivatization procedures were employed to analyze the compound derivative by fluorescence detection (Sancho et. al. 1996; Hogendoon et. al. 1999). Vreeken and co-workers developed an analytical method to analyze glyphosate, AMPA and glufosinate in water samples using reversed phase liquid chromatography separation after pre-column derivatization with 9-fluorenylmethyl chloroformate (FMOC-Cl) and detection by tandem mass spectrometry (Vreeken et. al. 1998). Bauer and co-workers detected glyphosate and AMPA in water samples using ion chromatography followed by electrospray with single quadrupole mass spectrometry detection. (Bauer et. al. 1999). The ion chromatography separation without derivatization was also used by Granby and co-workers after clean-up on reversed phase column to analyze glyphosate and AMPA through electrospray ionization/tandem mass spectrometric analysis (Granby et. al. 2003). Goodwin and coworkers studied the electrospray negative ion fragmentation pathways of glyphosate and AMPA using an ion-trap mass spectrometer . Some other methods found in the literature to analyze glyphosate and AMPA include: capillary electrophoresis (Cikalo et. al. 1996), ion chromatography with conductivity detection (Zhu, et. al. 1999), ion chromatography with fluorescence detection (Patsias et. al. 2001), gas chromatography (Hudzin et. al. 2002), immunoassays (González-Martinez et. al. 2005), nuclear magnetic resonance (Deen et. al. 2002) and integrated pulse amperometry (Ji-Ye et. al. 2001). In this work, we investigated the potential of reversed phase liquid chromatography coupled with electrospray tandem mass spectrometry (LC-ESI/MS/MS) for the quantification of glyphosate and AMPA in soybean spiked samples. In this approach, the compounds were analyzed without derivatization procedures using calibration curves prepared in the matrix, after a simple sample extraction and liquid-liquid partition followed by protein precipitation step with organic solvent to minimize the complexity of the sample. The mobile phase composition and the matrix effects were also investigated to validate the method using a high flow gradient program in a total run time of four minutes for each analyte. Experimental Chemicals and standardsGlyphosate and AMPA standards were obtained from Sigma-Aldrich (Steinheim, Germany). Methanol and dichloromethane HPLC-grade solvents were purchased from J. T. Baker (Deventer, The Netherlands) and ammonium carbonate P.A. was obtained from Merck (Darmstadt, Germany). Purified water was obtained on EASYpure RF System from Barnstead (Dubuque, IA, USA). The stock solutions of glyphosate and AMPA at 500 mg L -1 were prepared by dissolution of t...