Ethoxyquin – Bestimmung von 2,2,4-Trimethyl-6(2H)-chinolinon in Urin mittels UPLC-MS/MS

Biomonitoring-Methode

Gerhard Scherer¹ (Methodenentwicklung)
Nikola Pluym¹ (Methodenentwicklung)
Max Scherer¹ (Methodenentwicklung)
Nadine Rögner¹ (Methodenentwicklung)
Markus Stoeckelhuber¹ (Methodenentwicklung)
Thomas Jäger² (Methodenprüfung)
  Thomas Göen³ (Leitung der Arbeitsgruppe „Analysen in biologischem Material“ der Ständigen Senatskommission zur Prüfung gesundheitsschädlicher Arbeitsstoffe, Deutsche Forschungsgemeinschaft)
  Andrea Hartwig⁴ (Vorsitz der Ständigen Senatskommission zur Prüfung gesundheitsschädlicher Arbeitsstoffe, Deutsche Forschungsgemeinschaft)
  MAK Commission<sup>5

1</sup> ABF Analytisch-biologisches Forschungslabor GmbH, Semmelweisstraße 5, 82152 Planegg, Deutschland
² BASF SE, Corporate Health Management, Carl-Bosch-Straße 38, 67056 Ludwigshafen, Deutschland
³ Friedrich-Alexander-Universität Erlangen-Nürnberg, Institut und Poliklinik für Arbeits-, Sozial- und Umweltmedizin, Henkestraße 9–11, 91054 Erlangen, Deutschland
⁴ Institut für Angewandte Biowissenschaften, Abteilung Lebensmittelchemie und Toxikologie, Karlsruher Institut für Technologie (KIT), Adenauerring 20a, Geb. 50.41, 76131 Karlsruhe, Deutschland
⁵ Ständige Senatskommission zur Prüfung gesundheitsschädlicher Arbeitsstoffe, Deutsche Forschungsgemeinschaft, Kennedyallee 40, 53175 Bonn, Deutschland

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 (MAK Commission) developed and verified this biomonitoring method for the determination of the most important urinary metabolite of ethoxyquin – 2,2,4‑trimethyl‑6(2H)-quinolinone (EQI). Ethoxyquin is a quinoline-based synthetic antioxidant. Its use as active substance in pesticides was prohibited in 2011. Nevertheless, it continues to be used as a feed additive and is mainly introduced into the environment by feeding treated fishmeal to fish in aquaculture. A human metabolism study showed that ethoxyquin is first metabolised to 1,2‑dihydro-2,2,4‑trimethyl-6‑quinolinol (hydroxyquin) and is then further oxidised to the more stable EQI. In the biomonitoring method presented here, the glucuronide of hydroxyquin in the urine sample is enzymatically hydrolysed using β‑glucuronidase from E. coli. Thereafter, hydroxyquin oxidises spontaneously to EQI. The hydrolysate is subject to salt-assisted liquid-liquid extraction (SALLE) with ethyl acetate. Analysis is performed by UPLC‑MS/MS after positive electrospray ionisation (ESI+). Calibration is performed using standards prepared in pooled urine and processed analogously to the samples. EQI‑D₁₀ is applied as an internal standard. The method provides reliable and accurate analytical results, as shown by the good precision data with standard deviations no greater than 6%. Good accuracy data were obtained with mean relative recovery values in the range of 103–110%. The method is both selective and sensitive, whereby a quantitation limit of 0.03 μg EQI/l was achieved.

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