Abstract
A new method was developed and in-house validated to determined ethoxyquin (EQ), butylated hydroxyanisole (BHA) and propyl gallate (PG) in fish silage and fish oil used for production of animal feed. Sample pre-treatment comprises use of QuEChERS (quick, easy, cheap, effective, rugged and safe) extraction and gives extracts for liquid chromatography-triple quadrupole mass spectrometry analysis without matrix interferences. The results allow quantification of the antioxidants in all matrices by using deuterated internal standards and calibration curves made in solvent, as well as establishing low detection limits (0.012–0.015 mg/kg) and quantification limits (0.040–0.050 mg/kg) needed regarding the recent suspension of EQ as feed additive within the European Union. The proposed method was validated in terms of linearity, accuracy (relative error, ±1.4%), precision (RSD, ≤2.7%) and recovery (matrix effect, 97–101%) where all the parameters show acceptable results according to recognized guidelines (including EuraChem) within method validation. Combined expanded measurement uncertainty was estimated with a coverage factor of 2 by including all contributors to analytical variation, showing results of ±15% or better. The combined QuEChERS and LC-MS/MS method was successfully applied to salmon silage samples and their corresponding aqueous and oil fractions.
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