| In this study, an ultrasonic and heat-assisted liquid–liquid extraction method combined with deep eutectic solvent-based
dispersive liquid–liquid microextraction procedure was developed for the extraction of four phytosterols from cow milk
butters. The analytes were determined by gas chromatography-fame ionization detector without derivatization. Initially, the
sample was transferred into a falcon tube and placed into a sonication bath adjusted at 50 °C. Afterward, ethanol was mixed
with the butter and the mixture was sonicated. After centrifugation, the ethanolic phase collected on top of the sample was
withdrawn. After addition of a few microliters, ethyl (methyl) ammonium chloride/pivalic acid deep eutectic solvent, the
mixture was sucked into a glass syringe and injected hastily into deionized water. The turbid solution was centrifuged and
the sedimented phase was removed. Then, it was contacted with sodium hydroxide solution to remove the extracted fatty
acids. The proposed method presented high enrichment factors (340–425) and extraction recoveries (68–85%), low limits
of detection (0.51–1.3 ng g−1) and quantifcation (1.7–4.3 ng g−1) and acceptable relative standard deviations (≤8.4%).
The analytes contents of several cow milk butter samples were extracted and quantifed with the developed method and the
results were compared with ISO 23349. |