Linker-Assisted Immunoassay and Liquid Chromatography/Mass Spectrometry for the Analysis of Glyphosate

Abstract

A novel, sensitive, linker-assisted enzyme-linked immunosorbent assay (L'ELISA) was compared to on-line solid-phase extraction (SPE) with high-performance liquid chromatography/mass spectrometry (HPLC/MS) for the analysis of glyphosate in surface water and groundwater samples. The L'ELISA used succinic anhydride to derivatize glyphosate, which mimics the epitotic attachment of glyphosate to horseradish peroxidase hapten. Thus, L'ELISA recognized the derivatized glyphosate more effectively (detection limit of 0.1 microg/L) and with increased sensitivity (10-100 times) over conventional ELISA and showed the potential for other applications. The precision and accuracy of L'ELISA then was compared with on-line SPE/HPLC/MS, which detected glyphosate and its degradate derivatized with 9-fluorenylmethyl chloroformate using negative-ion electrospray (detection limit 0.1 microg/ L, relative standard deviation +/- 15%). Derivatization efficiency and matrix effects were minimized by adding an isotope-labeled glyphosate (2-13C15N). The accuracy of L'EUSA gave a false positive rate of 18% between 0.1 and 1.0 microg/L and a false positive rate of only 1% above 1.0 microg/L The relative standard deviation was +/- 20%. The correlation of L'ELISA and HPLC/MS for 66 surface water and groundwater samples was 0.97 with a slope of 1.28, with many detections of glyphosate and its degradate in surface water but not in groundwater.

Environmental immunoassays

“…Yet, pesticides are probably still the most important analytes with novel applications like the stereoselective determination of enantiomers (of S-imidazolinone herbicides [1]) and with antibodies for the very small and charged herbicides glufosinate and glyphosate, the latter being detected after derivatization of the parent molecule ("linker-assisted enzyme-linked immunosorbent assay -L'ELISA" [2]). Immunoanalysis also conquers the realm of less favorable analytes that often have low solubility in water, e.g.…”

Liquid chromatographic determination of glyphosate and aminomethylphosphonic acid residues in rapeseed with MS/MS detection or derivatization/fluorescence detection

“…High-performance liquid chromatography with fluorescence detection (HPLC-FLD) or ultraviolet detection (HPLC-UV) [23,26,27] have an excellent track record in its determination. Other techniques include capillary electrophoresis [28,29], ion chromatography with conductivity [30] or fluorescence [31], gas chromatography [32,33], immunoassays [34,35], nuclear magnetic resonance [36], and integrated pulse amperometry [37].…”

“…LC-MS/MS has been used to detect glyphosate and AMPA [44,45] and glyphosate residues have been determined in a variety of environmental and food matrices after derivatization with FMOC [22,35,[44][45][46][47][48]. Over the last few years new methods have been reported which avoid derivatization [20,21,48].…”

Preconcentration of the herbicide glyphosate and its metabolite AMPA by Immobilised Metal Ion Affinity Chromatography (IMAC)

“…A great variety of methods for the determination of glyphosate and AMPA in environmental matrices have been applied [6]: gas chromatographic procedures using flame photometric [7], thermoionic [8], and electron capture [9,10] detectors; more sensitive and selective GC methods with MS and MS/MS detection [10 -13]; high performance liquid chromatographic (HPLC) methods with fluorescence detection involving either post-column or pre-column derivatisation with o-phthalaldehyde (OPA) [14 -17] and 9-fluorenylmethylchloroformate (FMOC) [18 -21], respectively; HPLC with mass detection [22]; and also ion chromatographic [23,24], capillary electrophoretic [25,26], and ELISA methods [27,28] have been reported to date.…”