Plant-based meat substitute analysis using microextraction with deep eutectic solvent followed by LC-MS/MS to determine acrylamide, 5-hydroxymethylfurfural and furaneol.

For the analysis of plant-based meat substitutes and the determination of Maillard reaction products such as acrylamide, 5-hydroxymethylfurfural and furaneol, a novel and effective procedure based on hydrophobic natural deep eutectic solvent and liquid chromatography coupled with tandem mass spectrometry was developed for the first time. The 49 compositions of the deep eutectic solvents were designed and screened to select the most suitable option. The terpenoids eugenol and thymol in a molar ratio of 2:1 were selected as precursors for solvent formation, allowing effective extraction of the target analytes. The developed procedure comprised two main steps: extraction - in which the analytes are isolated from the solid sample due to the salting-out effect and pre-concentrated in the deep eutectic solvent, and back-extraction - in which the analytes are re-extracted into the formic acid solution for subsequent mass spectrometric detection. As the density of the aqueous phases changed during the extraction and back-extraction steps, the phenomenon of inversion of the coalesced organic phase was observed, which simplified the withdrawing of the phases. The linear range was 1-50 ng/mL for acrylamide, 10-1000 ng/mL for 5-hydroxymethylfurfural and 200-1000 ng/mL for furaneol with coefficients of determination above 0.9952. The developed method was fully validated and found recoveries were in the range 83-120%, with CVs not exceeding 4.9%. The method was applied to real sample analysis of pea-based meat substitutes.