Glyphosate – Determination of glyphosate and AMPA in urine by LC-MS/MS

Biomonitoring Method – Translation of the German version from 2025

Laura Kenny¹ (Method development)
Craig Sams¹ (Method development)
Kate Jones¹ (Method development)
Elisa Polledri² (External verification)
Rosa Mercadante² (External verification)
Silvia Fustinoni² (External verification)
  Thomas Göen³ (Head of the working group “Analyses in Biological Materials” of the Permanent Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area, Deutsche Forschungsgemeinschaft)
  Andrea Hartwig⁴ (Chair of the Permanent Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area, Deutsche Forschungsgemeinschaft)
  MAK Commission<sup>5

1</sup> Health and Safety Executive (HSE) Science and Research Centre, Harpur Hill, SK17 9JN Buxton (Derbyshire)
² Laboratory of Environmental and Occupational Toxicology, Department of Clinical Sciences and Community Health, University of Milano and Fondazione IRCCS Ca Granda Ospedale Maggiore Policlinico, Via Francesco Sforza 35, 20122 Milano
³ Friedrich-Alexander-Universität Erlangen-Nürnberg, Institute and Outpatient Clinic of Occupational, Social, and Environmental Medicine, Henkestraße 9–11, 91054 Erlangen, Germany
⁴ Institute of Applied Biosciences, Department of Food Chemistry and Toxicology, Karlsruhe Institute of Technology (KIT), Adenauerring 20a, Building 50.41, 76131 Karlsruhe, Germany
⁵ Permanent Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area, Deutsche Forschungsgemeinschaft, Kennedyallee 40, 53175 Bonn, Germany

Abstract

The working group “Analyses in Biological Materials” of the German Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area developed and verified the presented biomonitoring method. The aim of this method is the selective and sensitive quantitation of glyphosate (N-phosphonomethylglycine) and its only metabolite, aminomethylphosphonic acid (AMPA), in urine. Samples undergo solid-phase extraction prior to liquid chromatography-tandem mass spectrometry using glyphosate-2-¹³C,¹⁵N and AMPA-¹³C,¹⁵N,D₂ as internal standards. Calibration is carried out with urine from persons with no known exposure to glyphosate and AMPA. The procedure has been comprehensively validated and the reliability data have been confirmed by replication and verification of the procedure in a second, independent laboratory. Good precision data with standard deviations of 1.3–9.8% for glyphosate and 1.9–5.4% for AMPA, as well as good accuracy data with mean relative recoveries in the range of 91–102% for glyphosate and 100–106% for AMPA, show that the method provides reliable and accurate analytical results. The method is both selective and sensitive, and the limits of quantitation of 0.1 μg/l for glyphosate and 0.5 μg/l for AMPA are sufficient to determine occupational exposure as well as some of the background exposure in the general population.

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