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

Biomonitoring Method – Translation of the German version from 2023

Hans-Wolfgang Hoppe¹ (Method development)
Ute Fest¹ (Method development)
Elisabeth Eckert² (External verification)
Johannes Müller² (External verification)
  Thomas Göen² (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 ForschungsgemeinschaftExternal verification; )
  Andrea Hartwig³ (Chair of the Permanent Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area, Deutsche Forschungsgemeinschaft)
  MAK Commission<sup>4

1</sup> MVZ Medical Laboratory Bremen GmbH, Haferwende 12, 28357 Bremen, Germany
² Friedrich-Alexander-Universität Erlangen-Nürnberg, Institute and Outpatient Clinic of Occupational, Social, and Environmental Medicine, Henkestraße 9–11, 91054 Erlangen, Germany
³ Institute of Applied Biosciences, Department of Food Chemistry and Toxicology, Karlsruhe Institute of Technology (KIT), Adenauerring 20a, Building 50.41, 76131 Karlsruhe, Germany
⁴ 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 Permanent Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area developed and verified the presented biomonitoring method. Glyphosate (N‑phosphonomethylglycine) is a synthetic phosphonic acid derivative which has been used as a broad-spectrum herbicide since 1974. Its only known metabolite is aminomethylphosphonic acid (AMPA). Exposure in occupational settings is predominantly due to inhalation and dermal contact with glyphosate. The general population is exposed to glyphosate and AMPA via both dietary (plant and animal products) and environmental (soils, surface water, and groundwater) exposure.

The aim of this work was to develop a selective method for the determination of glyphosate and AMPA in urine. The method has been comprehensively validated, and the reliability data have been confirmed by replication and verification of the procedure in a second, independent laboratory. The analytes are directly derivatised in the dried urine sample with trifluoroacetic anhydride and trifluoroethanol without an initial extraction step. Calibration is performed using aqueous calibration standards processed analogously to the samples. As internal standards, glyphosate‑d₂ and ¹³C,¹⁵N‑AMPA are added to the urine samples and calibration standards. The derivatives are measured after capillary gas-chromatographic separation with tandem mass-spectrometric detection (GC‑MS/MS) using negative chemical ionisation (NCI).

The good precision and accuracy data show that the method provides reliable and accurate analytical results. The method is both selective and sensitive, and the quantitation limit of 0.1 μg/l urine for glyphosate and AMPA is sufficient to determine occupational exposure as well as higher background levels in the general population.

---

---